Structure and Function of the Skin Flashcards
_________ are a gene family critical for normal embryologic development; they likely play a very important role in development of skin adenexa, pigment system, and stratified epithelium during embryogenesis.
Homeobox (hox) genes
What are the three layers of the ectodermal covering?
- The basal layer (stratum germinativum)
- The outer layer (periderm)
- Then 3 layers forms in between the two layers (stratum intermedium)
Dermal development is characterized by an _____ in thickness and number of fibers, a decrease in ground substance, and the transition of ____ cells to _______.
increase
mesenchymal cells to fibroblasts.
Fetal skin contains a large percentage of type____ collagen compared with the skin of an adult, which contains a large proportion of type ____collagen.
III
I
Embryonal stratum germinativum differentiates into hair germs (primary epithelial germs) that give rise to what three structures?
hair follicles, sebaceous glands, and epitrichial (apocrine) sweat glands.
The epidermal placode develops into what three epithelial cylinders?
Innermost = hair shaft
Outermost cylinder = outer root sheath (ORS)
Middle = inner root sheath
As the hair peg lengthens, and develops into a hair follicle and hair, list the three bulges that appear?
Lowest (deepest) = develops into the attachment for the arrector pili muscle
Middle = develops into the sebaceous gland
Uppermost = develops into the epitrichial sweat gland
Secondary hairs develop on the caudal or cranial side of the primary hair follicle.
caudal
What are the six major morphogenic family systems have been recognized to be important
Genes of the Hox cluster
Fibroblast growth factor (FGF)
Transforming growth factor (TGF-B)
Sonic hedgehog (shh)
Wingless or wnt pathway
Neurotrophins
What is a marker of early anagen hair follicles and it is important in the protein cross-linking that contributes to the shape and strength of hair?
Transglutaminase
The diameter of the hair shaft is determined by the volume of ______, and the length of the hair shaft is determined by both the rate of the hair growth and duration of the anagen.
the hair matrix epithelium
The hairs of the cat have been divided into three types based on the gross appearance:
Guard hairs (thickest, straight, evenly tapered to a fine tip). Awn hairs (thinner, possessing subapical swelling below the hair tip). Down hairs (thinnest, evenly crimped or undulating.)
***The secondary hairs of the cat are far more numerous than primary***
All hair follicles grow in what direction in relation to the epidermis?
obliquely (30-60 degrees)
During what phase of the hair cycle do large numbers of keratinocytes in the outer root sheath undergo apoptosis and the epithelial strand regress?
catagen
_____ are thought to play a fundamental role in induction of epithelial differentiation. They are the primary target cells that respond to hormones and mediate growth-stimulating signals to the follicular epithelial cells.
Dermal papilla cells (mesenchymal component of hair bulb)
Alterations in ____ that control hair follicle structure result in follicular dysplasia.
morphogens
What is the normal coat comprised of?
The normal coat is see in the GSD, Welsh Corgi and wild dogs. It is composed of primary hairs (course guard hairs or bristles) and secondary hairs (fine hairs or undercoat).
What are the reported ultrastructural changes reported in non-lesional AD skin?
Widening of the intercellular spaces, retention of lamellar bodies in the corneocytes and irregularities and fragmentation of lipid lamellae.
Similar to humans with AD
Colonization of staphylococcus sp can result in an increase in ____ and ____ from cutaneous T-cells. This increase in ___ can induce the production of fibronectin, which increases the adherence of staph to keratinocytes.
IL-4/IL-13
IL-4
***Dupulimab binds to the alpha subunit which is shared by IL-13***
Colonization by Staph spp. further damages atopic skin as bacteria produce ______ and _____, which additionally decrease ceramides in the SC.
ceramidases and proteases
What gene family encodes information critical for normal embryologic development of skin adenexa, pigment system and stratified epithelium during embryogenesis?
Homeobox (Hox) genes
Encode a sequence of 60 amino acids that bind to DNA and regulate transcription (expression of genes into proteins)
Embryonic skin consists of a single layer of _____ cells and a dermis containing loosely arranged _____ cells embedded in an interstitial ground substance.
ectodermal
mesenchymal
Melanocytes (neutral crest origin) and Langerhans cells (bone marrow origin) become identifiable during what period of embryonic maturation?
ectodermal
Dermal development is characterized by an increase in thickness and number of fibers, a decrease in ____ and a transition of mesenchymal cells to _____.
ground substance
fibroblasts
Fetal skin contains a large percentage of type ___ collagen compared to the skin of adults which contains a large amount of type ___ collagen.
III
I
The embryonal stratum germinativum differentiates into hair germs (primary epithelial germs) that give rise to ____, _____ and _____.
hair follicles
sebaceous glands
apocrine glands
Hair germs initially consist of an area of crowding or deeply basophilic cells in the basal layer of the epidermis. The areas of crowding becomes buds known as______ that protrude into the dermis. Each bud lies a group of mesencymal cells, referred to as _____ from which the dermal papilla is later formed.
epidermal placodes
dermal condensate
What three epithelial cylinders develop from the dermal placode?
innermost = hair shaft middle = IRS outermost = ORS
As the hair peg lengthens and develops into a hair follicle and hair, three bulges appear? What are the three bulges and what do they develop into?
The lowest (deepest) of the bulges develops into the attachment for the arrector pili muscle; the middle bulge differentiates into the sebaceous glands and the uppermost bulge evolves into the epitrichial sweat gland. These appendages develop on the cranial side of primary hair follicles; secondary hair follicles develop on the caudal side.
Where do first hairs appear on the fetus?
vibrissae and tactile or sinus hairs that develop on chin, eyebrows and upper lip as slightly raised dots.
At birth, the majority of hair follicles or dogs are ____ ones; ______ develop caudal to the primary hairs during the first 12- 28 weeks of life.
primary
secondary follicles
_____ are substances that control hair follicle development.
morphogens
The skin, hair and subcutis of a newborn puppy represent XX% of its body weight, these structures represent approximately XX% of body weight
24
12
The pH of normal canine skin is ranges from __ to __.
4.84 - 9.95
What enzyme is a marker of early anagen hair follicles, and it is important in the protein cross-linking that contributes to the shape and remarkable physical strength of hairs?
Transglutaminase
What are the known growth factors involved in hair follicle induction?
EGF, TGFB1, TGFB2, neurotrophic 3. These growth factors control cellular proliferation and collagenase release from cultured hair follicles.
What cell types are involved in the regulation of hair growth, especially during catagen phase?
Interplay between MHC I, chondroitin proteoglycans and activated macrophages.
What are intrinsic factors involved in hair cycle?
growth factors, cytokines, dermal papilla, inflammatory cells int he immediate environment
The hair cycle and haricot are also affected by hormonal changes. In general, anagen is initiated and advanced by _____ and ____ hormone. Excessive amounts of ____ or ____ inhibit anagen and suppress hair growth rate.
Thyroid, Growth
GC, estrogens
____ cells, which are mesenchymal component of the hair bulb, are considered to play a fundamental role in the induction of epithelial differentiation.
Dermal papilla, These cells are morphologically and functionally differentiated from dermal fibroblasts and are thought to be the primary target cells that respond to hormones and mediate growth-stimulating signals to the follicular epithelial cells.
What is a ‘normal coat’?
Typified by that seen in GSD, Corgi and wild dogs.
It is composed of primary hairs (course guard hairs or bristles) and secondary hairs (fine hairs or undercoat).
What is a ‘short’ coat?
The coarse short coat is typified by the Rottweiler and many of the terriers. This type of coat has a strong forth of primary hairs and a much lesser growth of secondary hairs.
The fine short coat is exemplified by the boxer, dacshunds, and miniature pinchers. This type of coat has the largest number of hairs per uint area. the secondary hairs are numerous and well developed, and the primary hairs are reduced in size compare dot those of the normal coat.
What is the ‘long coat’?
The long coat can also be arranged into two subdivisions: the fine long coat and the woolly or course long coat. The fine long coat is found int he cocker spaniel, the Pomeranian and the chow chow.
The woolly or course long coat is found in the Poodle, Bedlington terrier and Kerry blue terrier.
Secondary hairs make up 70% of the total weight of these coats.
Describe the classic ‘agouti-type’ hair
Ex are the GSD and Norwegian Elkhound
The tip is white or light, the heavy body is pigmented brown or black, and the base is a light yellow or red-brown.
Pigment cells in the bulb or the hair deposit pigment in or between the cortical and medullary hair cells.
The black-brown pigment is _____; yellow-red pigment is called _____.
eumelanin
pheomelanin
Activation of melanocortin 1 receptors (MCR1 gene) on melanocytes leads to production of ______; inhibition or MCR1 results in the production of _____.
eumelanin
pheomelanin
What gene controls the shade of eumelanin?
Tyrosine related protein gene 1 (TYRP-1)
The generally recognized gene series involved in the coat color of dogs are called A, B, C, D, E, G, H, I, K, M, P, R, S and T.
- Agouti (ASIP)
- brown (TYRP1)
- C (colored/albino)
- D (blue dilution, melanophilin (MLPH)
- extension (MC1R)
- G (graying)
- H (Harlequin)
- I (phaeomelanin dilution)
- K (black, beta0defensin 103)
- M (merle, SILV)
- P (dilute)
- S (spotting, microphthalmia-associated transcription factor, MITF)
- T (ticking)
What does the ‘tabby’ coloration consist of?
The underlying pattern is agouti - characterized by hairs with a blueish base and a black tip separated by yellow banding.
The tabby genes determine whether the cat has narrow, vertical, gently curving stripes (mackerel), larger patches (blotch) or Abyssinian pattern.
How does temperature play a role in ‘tipped hair coats’?
Pointed hair coats are characterized by pale-colored hair not he body with darker hairs on the extremities or points. Points drive through a temperature-dependent enzyme that converts melanin precursors into melanin by the process of oxidation. Higher temperatures = lighter hair.
Inflammation and hyperemia result in more lightly colored new hair.
Expression of this gene is C, tyrosinase gene, TYR
What gene determines the multicolored coats in tortoiseshell and piebald spotting patterns?
White gloves gene (KIT)
What gene dilutes black to blue (gray), orange to cream, and seal-point ti blue-point (Siamese) AND is inherited as an autosomal recessive trait?
The dilution gene, melanophilin, MLPH
What is the difference between Cornish Rex and Devon Rex?
The Cornish Rex lacks primary hairs; The Devon Rex has primary hairs that resemble secondary hairs with absent or stubbled whiskers.
What are the boundaries of the areas of distribution of the main cutaneous nerve stems?
Voight lines
What lines reflect the course of blood vessels or lymphatics?
Langer lines
What lines form the pattern assumed by many different nevoid and acquired skin diseases?
Blashko Lines
These lines reflect a mosaic condition deriving either from a single mutated clone of cells originating from a post zygotic mutation or from an X-linked mutation made evident by lyonization.
What are the FOUR distinct cell types within the epidermis?
keratinocytes, melanocytes, Langerhan cells, and Merkel cells.
What layer of the epidermis consists of cuboidal to columnar cells resting on the basement membrane zone that separates the epidermis from the dermis?
Basal layer, most of these cells are keratinocytes that are constantly reproducing and pushing upward to replenish the epidermal cells above,
Cells of the stratum basale contain K5 and K14 keratin filaments.
What major cell adhesion molecule provides an anchor for keratin filaments connecting the nuclear envelop to the cell membrane and also anchors adjacent cells together to stabilize the structure of the epidermis?
Desmosomes
What are the three major gene families the molecular components of desmosomes belong to?
plakins (desmoplakin)
armadillo proteins (plakophilin, plakoglobin)
cadherins (desmogleins, desmocollins)
What structures are junctional complexes distributed along the inner aspect of basal keratinocytes; their major role is epidermal-dermal adhesion.
hemidesmosomes
What structures are a large family of cell surface adhesive receptors. These cell surface glycoproteins are important in cell-cell and cell-matrix interactions, and also act as signal transducers through which extracellular and intracellular compartments can influence and modify each other?
Integrins
Each intern consists of a heterodimer of an a and b subunit which are noncovalently associated. In the epidermis, integral expression is confined to the basal layer.
Which integrins are most abundant in the epidermis?
a2, a3,b1,a6 and b4
a5b1 function?
mediates keratinocyte adhesion to fibronectin
a2b1 function?
mediates keratinocyte adhesion to collagens type 1 and type iV and laminin
a3b1 function?
receptor for epiligrin and is involved in adhesion to laminin
a1b5 function?
mediates keratinocyte adhesion to vitronectin
a6b4 function?
mediates keratinocyte adhesion to laminin
What layer of the epidermis is composed of daughter cells from the stratum basale?
Stratum Spinosum
What are the four major types of adhesive and communicative structures the mediate keratinocyte adhesion?
desmosomes, hemidesmosomes, adherens junctions and focal adhesions,
What structures serve primarily as intercellular route of chemical communication?
gap junctions
What are the three types of cytoplasmic filaments that make up the keratinocyte cytoskeleton?
cytokeratin
actin
microtubules (tubulin)
_____ and _____ are crucial for desmosome and hemidesmosome formation.
calcium and calmodulin
What three keratinocyte derived calmodulin-binding proteins participate in a flip-flop regulation of calcium-calmodulin interactions?
caldesmon, desmocalmin and spectrin
All epithelia express a keratin pair; which oncists of what structures?
One keratin chain from the acidic subfamily (type 1 keratin, cytokeratins 9-20) and one chain from the neutral-basic subfamily (type II keratins, cytokeratins 1-8).
What keratins make up the spinous layer?
K1/K10 AND retain K5/14 synthesized in the basal layer
Keratinocytes of the stratum spinosum synthesize what products? These are secreted into the intercellular spaces of the stratum granulosu and are important in the barrier function of the epidermis.
synthesize lamellar granules (keratinosomes, membrane-coating granules, Odland bdies) which contain glycolipids, glycoproteins, phosholipids, free sterols, glucosylceramides and acid hydrolyses.
Keratinocytes in stratum spinosum synthesize lamellar granules (Odland bodies) which consist of what substances?
profillagrin, keratin filaments, loracrin
Where are keratohyalin granules synthesized?
Keratohyalin granules are synthesized in the stratum granulosum; however, they are not true granules because they lack a membrane and are more accurate described as insoluble aggregates.
What are profillagrin degradation productions?
urocanic acid, carboxylic acid, lactic acid; important for normal stratum corneum hydration and also help filter UV radiation.
What is a cysteine-rich protein synthesized in the stratum granulosum in association with keratinohyalin granules?
loracrin (synthesized in stratum granulosum) involved in binding keratin filaments together in the corneocyte and anchoring them to the cross-linked envelope.
What are the two morphologic forms of keratihyalin granules in the rodent?
The P-F granule - irregularly shaped and contains profillagrin; whereas, the L granule is smaller, rounded and contains loricrin.
What are the protein components of the CE?
lorcrin, involucrin, filaggrin, elfin, cystitis A, cornifelin, small proline-rich proteins and “late envelope” proteins
______ are a superfamily of enzymes that are important in apoptosis, keratinization and hair follicle formation.
Transglutaminases; keratinocytes and epidermal transglutaminase mediate the sequential cross-linking of the cornfield cell envelop precursor proteins.
Cheifly expressed in the stratum granulosum and upper stratum spinosum.
The most important product of the epidermis is ____; a highly stable disulfide condition-containing fibrous protein.
keratins
alpha - keratin = skin/hair
beta keratins - feathers/scale
What is the function of ornithine decarboxylase?
This enzyme is essential for biosynteisis of polyamides (putrescine, spermidine, and spermine); which encourage epidermal proliferation.
What are the four distinct cellular events in the process of cornification?
- keratinization - synthesis of the principle fibrous proteins of the keratinocytes
- keratohyaline synthesis - including the histidine-rich protein filaggrin
- formation of the highly cross-linked insoluble stratum corneum
- generation of neutral lipid-enriched intercellular domains resulting from the secretion of distinctive lamellar granules.
Where are lamellar bodies primarily synthesized?
keratinocytes in the stratum spinosum
What are amide-linked fatty acids containing a long-chain amino alcohol (spinchoid base).
ceramides; they are the most important lipid component for lamellar arrangement in the stratum corneum and barrier function.
Linoleic acid, an essential fatty acid for dogs and cats, found in which ceramides?
Ceramides 1, 4 and 9.
Which ceramides bind to involucrin and other proteins of the CE, forming a scaffold for the binding of tother lipids into intercellular lipid bilayer?
Cermides 1 and 2
______ play a crucial role plasticizing the horney layer to allow stretching and bending by fluidizing barrier lipids.
ceramides
What are transient amplifying cells?
Proliferating keratinocytes that have limited capacity for self renewal & will after a couple of divisions terminally differentiate
Both stem cells and amplifying cells lives in basal layer & lower spinous layer
Divide rapidly—have an avid uptake but short retention of thymidine
What are the three basal cell states?
- Holoclones: correspond to stem cells
Colonies of small, regular cells
Give rise to many other colonies
A single stem cell from holoclone can give rise to up to 4 million keratinocytes.
2.Meroclones: correspond to the transient amplifying cells
Produce colonies, but fewer in number than holoclones
Located in the basal and suprabasal layers
Provide the bulk of cell divisions needed for stable self-renewal
- Paraclones: correspond to differentiated, postmitotic cells; Undergo terminal differentiation
Yield only very small number of colonies
List characteristics of the basal cell layer?
Contains mitotically active, columnar shaped keratinocytes
Contain pigmented melanosomes transferred via phagocytosis from melanocytes
Attach via keratin filaments (K5 & K14) to basement membrane zone at hemidesmosomes
Also express K19 & 15
Attach to other surrounding cells through desmosomes
Major function: provided mechanical attachement to underlying basement membrane through hemidesmosomes & integrins
Desmosomes
Ca-dependent cell surface modifications that promote adhesion of epidermal cells and resistance to mechanical stress
Site where the metabolic barrier of the epidermis occurs
Most of the process of terminal differentiation is determined by ______ levels
Low calcium maintains keratinocytes in a high proliferation/low differentiation state
High calcium promotes terminal differentiation of keratinocytes
_____ promotes transition from spinous to granular layer cells
Protein kinase C
Recessive X-linked ichthyosis
mutation of steroid sulfatase
Results in retention hyperkeratosis
Profilaggrin
(F granules)
1st appearing in the granular layer & diffusely distributed w/n cytoplasm
Profliaggrin: large Ca binding phosphoprotiein of 10-12 repeating copies of filaggrin protein
Release from the keratohyalin granules results in a Ca2+ dependent cleavage (dephosphorylation & proteolysis) into filaggrin monomers
Occurs upon entering the transition zone between granular layer & stratum corneum
Filaggrin
aggregates with keratin to form macrofilaments
A cationic protein, rich in histidine residues
Function: an intermediate filament-associated protein, organizing & aggregating keratin into macrofibrils
Degraded into free amino acids & molecules including:
Urocanic acid & pyrrolidone carboxylic acid
Contribute to hydration of stratum corneum & help filter UV radiation
Mutation in Transglutaminase 1 gene
lamellar ichthyosis
Autosomal recessive condition characterized by large scales and disruption in the uppermost differentiating layers of the epidermis
Loricrin abnormalities lead to what two diseases in humans?
Vohwinkel syndrome with ichthyosis
progressive symmetric keratodermia
Clear Layer contains refractile droplets and a semifluid called what?
eleidin
Stratum lucidum, stratum conjunctum
Fully keratinized, compact, thin layer of dead cells
Anuclear, homogeneous, and hyaline-like
Differs histochemically from SC by being rich in protein-bound lipids
Best developed in food pads; less developed in nasal planum and absent in all other places
What layer of the epidermis is characterized by stacked layers of anucleate, flattened, eosinophilic cornified cells?
Stratum Corneum - Drastic remodeling occurs during the transition from granular cell layer to the stratum corneum
Formed by a two-compartment system of lipid-depleted, protein-enriched corneocytes surrounded by a continuous extracellular lipid matrix
Primary functions of the extracellular lipid matrix:
Regulation of permeability, desquamation, antimicrobial peptide activity, toxin exclusion, and selective chemical absorption
Primary functions of the corneocytes:
Mechanical reinforcement
hydration
cytokine-mediated initiation of inflammation
protection from UV damage
The cytoskeleton of all cells of mammals are composed of three filament system:
Microfilaments (6nm)
Intermediate filaments (8-10nm)
microtubules (25nm)
All intermediate filaments contain what 3 structures?
- Central α helical rod
- Conserved secondary structure
- Flanked by distinct amino head & carboxyl terminal end domains
Intermediate filaments are subdivided into what 6 groups based on tissue specificity and amino acid sequence?
Acidic type I——-epithelial keratin (9-23)
Basic type II—–epithelial keratin (1-8)
Type III: vimentin (mesenchymal cells), desmin (muscle), peripherin (peripheral nervous system), & glial fibrillary acidic protein (GFAP) (Glial cells and astrocytes)
Type IV: α-internexin (neurofilaments)
Type V: lamins (nucleus)
Type VI: nestin (stem cells of CNS and skeletal muscle)
What are the 3 types of cytoplasmic filament that keratinocyte cytoskeleton consists of?
Actin, microtubules and cytokeratin
Largest group of intermediate filaments
Keratins
Hallmark of all epithelial cells, including keratinocytes
Account for up to 85% of total protein of fully differentiated cells
Serve a predominately structural role in the cells
Describe Type 1 keratin filaments
Acidic
K9-K20
Describe Type 2 keratin filaments
Basic or neutral
K 1-8
Primary keratin structure
Two different chains assemble in pairs to create dimmers
Consist of an acidic & a basic keratin subunit
Composed of two α-helical chains oriented in a parallel & interwined in a coiled-coil rod formation
Describe formation of the cornified cell envelope
As intracellular Ca2+ levels rise in suprabasal cells, envoplakin, periplakin and involucrin are expressed (PIE) and fuse to the plasma membrane via the action of TG1 and 5 (calcium dependent)
Lamellar granules fuse to the plasma membrane then get lipid excretion into the intercellular space.
TG1 crosslinks ceramide to proteins
TG3 crosslinks Loricrin (most abundent) & SPR (small proline rich peptides) - Now PILES)
Where are intercellular lipids initially stored?
Intercellular skin lipids are initially stored in the lamellar granules which get extruded into the extracellular space in the upper granular layer during cornification
What is the precursor molecule to ceramides?
glucosylceramides (precursor to ceramides, dominant component of the stratum corneum lipids)
Where do lipids originate from?
Lipids originate in the Golgi apparatus & first seen in the upper spinous layer (abundant in the granular layer)
Lamellar granules fuse with the plasma membrane of keratinocytes & secrete their contents releasing stacks of membranous disks into the intercellular spaces
Role in synthesis & storage of cholesterol
Vital for normal adhesion & desquamation of cornified cells
Abnormal steroid and lipid metabolism
Recessive X-linked ichthyosis – mutation of steroid sulfatase
Results in retention hyperkeratosis
Phospholipids become _____ cholesterol becomes ____, & spingolipids become _____.
Phospholipids become free fatty acids, cholesterol becomes cholesterol esters, & spingolipids become ceramides.
**β glucocerebroside, acid sphingomyelinase & phospholipase A are involved in this modification***
List proteins in the cornfield envelope
- loricrin
- involucrin
- filaggrin
- elafin
- cystatin A
- cornifelin
- small proline rich proteins
- “late envelope proteins’
Trichohyalin
Significant component of the keratohyalin granules
Also part of the granular layer of the epidermis
Involucrin
Common component of the cornified envelope
Becomes cross-linked to other proteins and is thought to be part of the early component in the assembly of cornified envelope—scaffolding
Adjacent to the cell membrane & as the cell membrane becomes replaced, involucrin thought to be the preferred substrate to which external lipids (ceramides) are attached to the surface of the CE
Small Proline-rich proteins (SPRs)
Consist of a family of related small proline rich proteins
Serve as cross bridging proteins
The amount of SPR compared to Loricrin allows for the flexibility, rigidity or the toughness of a composite material (tissue)
Envoplakin
Expressed at desmosomal plaques
Homologous to desmoplakin
Link the CE to desmosomes and keratin filaments attachments for ceramides
Periplakin
Expressed along with envoplakin at desmosomal plaques
Serve as attachments for ceramides
Envoplakin & periplakin → heterotetramers.
Associate with the plasma membrane in a Ca2+ dependent manner
Profilaggrin
Formed initially in the granular layer & initially highly phosphorylated & accumulated in the keratohyaline granules
During cornification dephosphorylated & cleaved into filaggrin
Filaggrin
Becomes cross-linked to the epidermal CE & coordinates the structure of the cornified cells
Aggregates keratin intermediate filaments into tight bundles—this allows for the collapse of the cells
Filaggrin half life is 6 hours & degraded mostly into free amino acids
High concentration of hydrophilic a.a. is essential for the retention of water & contributes to the osmolarity &flexibility of the cornified layer
Three main functions of filaggrin?
- Alignment of the keratin intermediate filaments
- Controls cell shape (contracts)
- Maintenance of epidermal texture
Cystatin A
Keratolinin
Expressed in the spinous layer
Substrate for epidermal TGases & a minor component of the CE
Cysteine protease inhibitor - Might be relevant for the bacteriostatic properties of the skin
Elafin
SKALP; skin derived anti-leukopeptidase
A minor component of CE from normal adult epidermis
A potent inhibitor of elastase & proteinase 3
Highly expressed in wounded or psoriatic skin
Transglutaminases (TG)
Superfamily of Ca dependent enzymes important for apoptosis, keratinization, and hair follicle formation
Present in all stratified epithelia and hair follicles
Mainly expressed in the granular to upper spinous layers
Catalyze formation lysine bonds between proteins
Catalyze the covalent incorporation of biogenic polyamines into proteins
Transglutaminase I
Expressed primarily in keratinocytes
Membrane bound, processed form accounts for most of the activity in differentiating keratinocytes
Catalyzes cross-linking of involucrin at the plasma membrane & cornifin, elafin and the SPR proteins are added subsequently
Tranglutaminase 2
Vit A increases activity
Important role in would healing – involved in matrix repair & remodeling
Transglutaminase 3
cytoplasmic enzyme
Initially cross-links loricrin and SPRs together to form small interchain oligomers, which are then permanently affixed to the developing cornified cell envelope via further cross-linking by TGase1
Transglutaminase 5
Present in the upper epidermal layers (spinous & granular layers)
Induced in the early stages of keratinocyte differentiation
Can cross link Loricrin
Which enzymes are responsible for desquamation?
Acid phosphatase, Cathepsin-B, Carboxypeptidase, Sterol sulfatase
Corneocyte shedding/Desquamation
The final products of differentiation, the squames, finally reach the edge of the epidermis they are mummified remnants composed of cornified envelope surrounded by stacks of lipids packed with keratin macrofilament —Squames are then shed
The corneocytes are held together by ________
corneodesmosomes
Modified desmosomes structures
Located in the upper spinous and granular layers
Corneodesmosin
After transport and release from the lamellar bodies, corneodesmosin appears in the extracellular space & become associated with the desmosomal desmogleins & desmocollins
Corneodesmosin is cleaved by the stratum corneum ___ and ____ enzymes causing separation of adjacent corneocytes with cell shedding
tryptic and chymotryptic enzymes
Saturated FA
contain no carbon-carbon double bonds
Unsaturated FA
contain carbon-carbon double bonds
Polyunsaturated fatty acids (PUFAs)
Unsaturated fats with two or more double bonds
Cats are unique: lack _____, have limited ____ activity; unable to synthesize arachidonic acid – must obtain it from the diet
lack Δ-6-desaturase
Δ-5-desaturase
Mammals are unable to synthesize _____ and ______
linoleic acid or α-linolenic acid
Can make arachidonic acid from linoleic acid and eicosapentaenoic acid from α-linolenic acid
Essential fatty acid
a PUFA that cannot by synthesized by animals but is required for normal physiologic function
n-3 family
provided by the diet in green leafy veggies
Eicosapentaenoic acid can be obtained from marine fish oil
n-6 family
derived from veggies and present in seed oils (safflower & sunflower)
γ-Linolenic acid (product of linoleic acid) is found in seed oils of borage, evening primrose, & black currant
n-9 FAs
(oleic acid) – can ben synthesized de novo from acetyl coenzyme A by liver & tissue microsomes therefore is non-essential
EFA Deficiency
Causes renal & reproductive abnormalities, decreased growth rate, immunologic abnormalities, hypotrichosis, alopecia, scaling skin, weak cutaneous blood vessels and an increases tendency to bruise, decrease wound healing, sebaceous gland hypertrophy accompanied by increased sebum viscosity, and increased transepidermal water loss. Transdermal drug delivery systems use this intercellular lipid portion of the epidermis to allow penetration into the underlying dermis and vasculature
Arachidonic acid
most important of all of the EFAs; Fundamental component of cell membranes
Skin is unable to convert linoleic acid to AA; lacks Δ-6-DES & Δ-5-DES activity
Most AA is present in the phospholipids of the cell membrane
Major biologic function: parent to leukotrienes (LTs), prostaglandins (PGs), thromboxanes (TXs), & hydroxyl fatty acids (hydroxyeicosatetraenoic acids (HETEs).
What is the rate limiting step in eicosanoid synthesis?
Release of AA from membrane store; Requires liberation by a phospholipase
Activated by trauma, thermal injury, ischemia, endotoxin release, antigen-antibody interactions, increased intracellular calcium, cytokine production, or histamine release
PGD2
Major site: Mast cells, Langerhans Cells
Major biologic activity: neutrophil chemotaxis; induces vasodilation, relaxes smooth muscle, inhibits platelet aggregation, bronchoconstriction
PGE2
Major site: Keratinocytes, Fibroblasts & Macrophages
Major biologic activity: plasma exudation, stimulates cell proliferation, hyperalgesia, relaxes smooth muscle increases vasodilation
PGF2a
Major site: Keratinocytes
Major biologic activity: vasoconstriction, bronchoconstriction and smooth muscle contraction, stimulates cell proliferation
PGH2
Major biologic activity: precursor to thromboxanes A2 and B2, induction of platelet aggregation and vasoconstriction
PGI2
Major site: Vascular Endothelial Cells, Fibroblasts, Macrophages
Major biologic activity: inhibits platelet and leukocyte aggregation, induces vasodilation relaxes smooth muscle
TXA2
Major site: Platelets, Macrophages
Major biologic activity: induces platelet aggregation, vasoconstriction
TXB2
Major site: Platelets
Major biologic activity: induces vasoconstriction
LTB4
Major site: Monocytes, Basophils, Neutrophils, Eosinophils, Mast Cells, Keratinocytes
Major biologic activity: induces neutrophil and eos chemotaxis and aggregation/activations, vascular permeability, enhances C3B receptor expression, neutrophil degranulation
LTC4
Major site: Mast cells, Keratinocytes
Major biologic activity: Plasma exudation, mucus secretion, component of SRS-A, skin vasodilation, increases vascular permeability and bronchoconstriction
LTD4
Major site: Mast cells, Keratinocytes
Major biologic activity: component of SRS-A, skin vasodilation, mucus secretion increases vascular permeability and bronchoconstriction
LTE4
Major site: Mast Cells & Basophils
Major biologic activity:component of SRS-A, microvascular vasoconstrictor and bronchoconstriction
What are the changes in lipids from the granular to the cornified cell layers?
- Replacement of phospholipids by ceramides
- Increase in free sterols
- Large increase in free fatty acids at the expense of triglycerides and phospholipid
Transformation of these lipids provides the outer epidermal layers with a more stable, waxy, and impermeable barrier. Lipids also allow for water-holding in the stratum corneum, cohesion and desquamation of corneocytes, and control of epidermal proliferation and differentiation.
What are the two primary sources of lipids in the SC?
corneocyte envelope & intercellular lamellae
Lipids on the outer surface of the SC originate from:
Cat & Dog: predominately epidermal origin
Human: predominately sebaceous origin
What makes up the lipid portion of the CCE?
Majority are ceramides, cholesterol, fatty acids & cholesterol esters
Intercellular skin lipids are initially stored in the lamellar granules which get extruded into the extracellular space in the upper granular layer during cornification
Lamellar granules
Membrane-coating granules, Odland bodies, or keratinosomes
Secretory granules (type of lysosome) that deliver precursors of stratum corneum lipids into the intracellular space
Contain glycoproteins, glycolipids, phospholipids, free sterols, and a number of acid hydrolases, including lipases, proteases acid phosphatases, and glycosidases
Glucosylceramides are the precursors to what?
precursor to ceramides, dominant component of the stratum corneum lipids
Lipids originate in the XXX & first seen in the upper spinous layer (abundant in the granular layer)
Golgi apparatus
Complex changes in lipid composition occurs after the _______ of the granules by the enzymatic action of acid hydrolases
extrusion of the contents
Phospholipids become free fatty acids, cholesterol becomes cholesterol esters, & spingolipids become ceramides.
β glucocerebroside, acid sphingomyelinase & phospholipase A are involved in this modification
What is the intracellular lipid content?
15-25% free fatty acids
40-50% ceramides
20-25% cholesterol
5-10% cholesterol sulfate
The precursors to XXX are glucosylceramides which are located within the lamellar granules
ceramides
ceramides
the most important lipid component for lamellar arrangement in the stratum corneum & in barrier function.Plasticize the horny layer to allow stretching & bending by fluidizing barrier lipids (absence results in dissolution of water barrier)
Primary carriers of long change fatty acids and linoleic acid
Covalently linked to the corneocyte envelope
____ are incorporated into the ceramides
PUFA
Linoleic acid specifically
Free fatty acids cause an XX pH optimal for certain key enzymes
acidic
_____ and ____ are activated at an acidic pH & in turn generate ceramides
β-glucocerebrosidase and acidic sphingomyelinase (aSMase)
Acidification also regulates stratum corneum integrity/cohesion, thereby restricting desquamation
Corneodesmosomes are degraded by _____ & _____ enzymes, which exhibit optimal activity in neutral-basic pH
chymotryptic & tryptic enzymes
Lipid envelope
is located exterior to the protein envelope & is covalently attached via ester bonds to the protein envelope via TGase 1. Consists of a monomolecular layer of ω-hydroxy-ceramides which interdigitate with the intercellular lipid lamellae in a Velcro-like fashion.
Most cholesterol comes from _______ unchanged
Steroid sulfatase decreases the amount of cholesterol sulfate rather than adding cholesterol to the lipid barrier
Increases in cholesterol sulfate leads to an excess of scale (X-linked ichthyosis in humans).
Turnover rates for keratinocyte proliferation
Normal dog 22 days
Seborrheic Cocker 8 days
Horse 17 days
Goat 26 days
Pig 15 days
Human 26-42 days
16-30 from s. basale to s. spinosum
14 days in s. corneum
Desmosomes
Transmembrane Protein: Desmosomal cadherins Dsg I, II, III Dsc I, II, II
Plaque Protein: Plakoglobin, desmoplakin I, II, IV; desmocalmin; Plakophilin
Cytoskeleton Filaments: Cytokeratin
Function and Location: Cell-cell adhesion, all keratinocytes
Desmosomes
“Spines” of the spinous layer
Ca-dependent cell surface modifications that promote adhesion of epidermal cells and resistance to mechanical stress
Mechanically hold together the keratinocytes together
Physiologic communication between keratinocytes
Mediators of intercellular adhesion
Desmosomal plaque
Plakoglobin, desmoplakins I & II, keratocalmin, desmoyokin and plakophilin
Desmoplakin 1 binds directly to type ___ keratins (K1, 2, 5 & 6)
II
Keratocalmin/Desmocalmin
Calmodulin-binding protein
Promotes calcium dependent assembly and function of desmosomes
Desmogleins 1 and 3
part of the transmembrane portion of the desmosomal plaque
Calcium-dependent adhesion molecule (cadherins)
Desmocollins
part of the transmembrane portion of the desmosomal plaque
Calcium-dependent adhesion molecule (cadherins)
Cadherins
transmembrane glycoproteins
Mediate cell-cell adhesion in a Ca-dependent manner
Major adhesion molecules in desmosomes and adherens junctions
_________ is important as a mediator of adhesion
Ca
Exemplified by Darier and Hailey-Hailey disease which exhibit characteristic epidermal dyscohesion
Both are caused by mutations in genes that regulate Ca transport
Model of hemidesmosome
Keratin intermediate filaments insert into the plate portion (or inner plaque), which consists of bullous pemphigoid anitgen 230 (BP 1) and plectin.
The transmembrane component (or outer plaque) consists of bullous pemphigoid anitgen 180 (BP 2) and α6β4 integrin.
The extracellular domain of BP 180 has been localized to the extracellular space beneath the hemidesmosome (the lamina lucida) and has been identified as type XVII collagen. The intracellular domain of PB180 is localized in the hemidesmosomal plaque.
What are the extracellular components of hemidesmosomes?
Anchoring filaments originate at the hemidesmosomes and insert into the lamina densa. The major component of these filaments is laminin 5 which is localized mainly to the lamina densa and the lower lamina lucida and is associated with α6β4 in hemidesmosomes.
Plectin attaches intermediate filaments to
- HDs
- plasma membranes of basal keratinocytes
- cross-links other proteins associated with the cytplasmic plaques of HDs
Bullous pemphigoid antigen 1 (BP230, BPAG1)
First of two autoantigens in the hemidesmosones of basal keratinocytes detected using circulating autoanitbodies from patients with bullous pemphigoid (BP)
Plakin, 230 kDa; non-collagenous protein localized to the cytoplasmic plaque of HDs
Bullous pemphigoid antigen 180 (BP 2)
Type II transmembrane collagen (specifically, type XVII collagen)
Associated with HD-anchoring filament complexes in basal keratinocytes
NC16A domain of BP180
Large, non-collagenous region neat the transmembrane region
Binds extracellular N-terminus of α6-integrin
Contains the epitopes recognized by BP autoantibodies
Target of autoantibodies in a feline, porcine, and equine model of BP
α6β4 integrin
Hemidesmosome-associated integrin; receptor for Laminin 5
Large cytoplasmic tail of integrin subunit β4 contains sequences that are required for localization of in HDs
The membrane-proximal region of integrin subunit β4 directly associates with plectin, while its distal carboxyl-terminal region binds BPAG1
The proximal extracellular domain of integrin subunit α6 binds the NC16A region of BPAG2 (collagen XVII).
Laminins 1 and 5 serve as the preferred ligand for this integrin.
Mutation in either α6 or β4 integrin subunits develop Junction Epidermolysis Bullosa associated with pyloric atresia.
α5β1
mediates keratinocyte adhesion to fibronectin
α2β1
mediates keratinocyte adhesion to collagens type I and IV and laminin
α3β1
receptor for epiligrin; involved in adhesion to laminin
α1β5
mediates keratinocyte adhesion to vitronectin
α6β4
mediates keratinocytes adhesion to laminin
β2 integrins
Leukocyte functional antigen-1 (CD11a/CD18)
Adhesion molecule on T lymphocytes
Essential for most adhesion-dependent lymphocyte function
MAC-1, CR3, CD11b/CD18 and CD11c/CD18 (CR4)
Found on mononuclear phagocytes, neutrophils, NK cells
Adherens junctions
Small punctuate cell-cell attachment points
Connect actin filaments to adjacent cells
Major players: E and P cadherins
E-cadherins
Associated with actin filaments via interactions with catenins, and may regulate the orgainziaiton of adherens junctions and influence epidermal stratification
Expressed in keratinocytes of all epidermal layers
P-cadherins
expressed only in basal keratinocytes
Cytoplasmic domains of the cadherins connects with α- and β-catenin, and plakoglobin (γ-catenin).
Catenins connect cadherins to the cytoskeletal network of β-catenin & plakoglobin; α-catenin directly associates with acitn
Tight Junctions
An intercellular junction composed of a series of fusions of the junctional membrane, forming a continuous seal
Serves as a selective barrier to small molecules and as a total barrier to large molecules
Also known as impermeable junction; occluding junction; zonula occludens
Most important tight junction proteins in the human epidermis:
Occludin, claudins, and zonal occluding proteins
Occludin: restricted to the stratum granulosum
Zonal occluding protein-1 & claudin-4: found in the suprabasal layers
Claudins-1 & -7: found in all epidermal layers
Gap junctions
Sites of physiologic communication; while desmosomes/hemidesmosomes are sites of mechanical coupling
More abundant in more differentiated keratinocytes
Consist of 2 connexons from adjacent cell membranes
Connexins
Intracellular gatekeepers
Transmembrane proteins
Connexons on adjoining cells associate to form gap junctions
Different connexin based on layer of epidermis
Basal: Connexin 43
Spinous: Connexin 26
Cornified cells: Connexons 31 and 31.1
These changes relate to intracellular ionic signaling (specifically calcium)
Connexin-26 is one of the most highly upregulated genes in human psoriatic plaques
Missense mutations of connexin-26 result in 5 distinct ichthyosis-like skin disorders
Skin-associated lymphoid tissue
Langerhans’ cells, keratinocytes, epidermotrophic T lymphocytes, & draining peripheral lymph nodes
List 5 immunologic role of keratinocytes
Produce IL-1
Produce various cytokines (IL-3, protaglandins, leukotrienes, interferon)
Are phagocytic
Can express antigens associated with immune response gene in a variety of lymphocyte-mediated skin diseases (presumably as a result of interferon-γ secretion by activated lymphocytes)
Immunologic activity of unperturbed skin
produce only a few mediators
Cytokines: IL-1, IL-7 & transforming growth factor-β (TGF-β)
Contain large quantities of pre-formed & biologically active IL-1α and immature IL-1β
immediate initiator of inflammatory and repair processes after epidermal injury
IL-7
important lymphocyte growth factor
TGF-β
growth-regulating effects on keratinocytes
Modulates inflammatory/immune response
Important for Langerhans’ cell development
Antimicrobial peptides
an important evolutionary conserved innate host defense mechanism in many organisms
Keratinocytes produce cathelicidins (IL-37) and β-defensins
MOA may relate to membrane insertion and pore formation
β-defensins
Cysteine-rich cationic low-molecular-weight antimicrobial peptides
Essential part of cutaneous innate immunity
Attract dendritic cells and memory T cells via CC cehmokine receptor 6 (CCR6)
Cathelicidins:
cationic peptides with a structurally variable antimicrobial domain at the C-terminus
In mammals, pigs & cattle – a variety of cathelicidin genes exists
Pronounced antibacterial, antifungal, and antiviral activities
Attracts mast cells and neutrophils
Secreted into human sweat
Down regulated in atopic dermatitis
Psoriasin (S100A7)
secreted predominately by keratinocytes
plays a major role in killing E. coli
Keratinocytes synthesize and release complement components ___ and factor ___.
C3 and factor B
Eicosanoids
an ensemble of lipid mediators regulating inflammatory and immunologic reactions
Can produce and release the cylcooxygenase product prostaglandin E2, (both pro-inflammatory and immunosuppressive proteins)
When acting on dendritic cells, promotes the development of IL-4 dominated type 2 T cell responses
Things that stimulate keratinocyte growth
1) EGF
2) Transforming growth factor α
3) KGF
4) IL-1 α
5) IL-6 (increasing UV light)
6) β-FGF
Difference between neoplastic and regular keratinocytes
1) Neoplastic cell cycle shorter
2) Neoplastic cells have whorls
3) Anisocytosis
4) Lacks cells surface antigens
5) Growth faster without Cholera toxin and EGF
Integrins
Large family of cell surface adhesive receptors (surface glycoproteins)
Important in cell-cell and cell-matrix interactions
Facilitate association of matrix molecules and basal keratinocytes
Act as a link for collagen, laminin, fibronectin, thrombospondin, and vitronectin to the cytoskeleton of keratinocytes
Vital for communication
β1 integrins
found on the lateral surfaces of keratinocytes
Downregulated during differentiation, thought to be important for extracellular matrix attachment
β4 integrins
found associated with hemidesmosomes; mediates station adhesion
During wound healing, ____ integrins are redistributed to promote keratinocyte migration on matrix molecules like fibronectin.
B1 integrins
Integrins α5β1 and ανβ1 are also involved with wound healing
Difference between Epidermal growth factor (EGF) and TGF
EGF is not produced by keratinocytes; TGF is produced by keratinocytes
Both bind to the same tyrosine kinase receptor in the basal cell and suprabasal cell layers to stimulate proliferation
Over-expression leads to hyperplasia, hyperkeratosis, formation of spontaneous papillomas and hyper proliferative disorders
Keratinocyte-derived growth factor (KGF)
dermal transcripts with receptors on keratinocytes
member of the fibroblast growth factor family
Potent mitogen for keratinocytes
Ectopic expression will lead to epidermal thickening
Transforming growth factor β (TGF-β)
causes a decrease in DNA synthesis and mitosis of keratinocytes, promotes differentiation
TGF-β1 and TGF-β2 are expressed in the upper epidermal layers;
upregulated by TGF-β as well as other agents that promote differentiation (ie Ca)
Important role in homeostatic regulation of the epidermis
Other growth factors produced by keratinocytes include Basic fibroblast growth factor (influence on the proliferation of melanocytes) & platelet-derived growth factor (functions in the dermis)
Vitamin D3 effects on proliferation and differentiation?
Causes a decrease in keratinocyte proliferation
Causes an increase in keratinocyte differentiation
Terminal Epidermal Differentiation Processes that are Calcium-Dependent
Intercellular structure formation – Desmosome assembly & function, Gap junction assembly & function
Protein Expression – Involucrin, Keratins 1, 2e, 9, & 10, Loricrin, Profilaggrin, Small proline-rich peptides, Transglutaminase
Release of keratohyaline granule contents – Release of loricrin & profilaggrin
Enzyme function – Cleave of profilaggrin to filaggrin, Transglutaminase activity
Optimal water concentration of SC
20-35%
Must be at least 10% to look & feel normal
Atopy and filaggrin defect in humans
Atopic dermatitis (eczema) in humans affects ~20% population Two independent loss-of-function genetic variants in the gene encoding filaggrin (FLG) are very strong predisposing factors for atopic dermatitis
Origin of melanocytes?
Neural Crest
Location of melanocytes?
Basal layer of the epidermis
outer root sheath and hair matrix of hair follicles
ducts of sebaceous and sweat glands
perivascularly in the superficial dermis
Eye, ear, and CNS
There are no histochemical stains the cane performed on routinely process skin biopsy specimens that exclusively stain melanin. What stains rely oj the ability o melanin to reduce silver from a silver solution.
Do not stain readily with hematoxylin eosin and actually show up clear
Argentaffin stains rely on the ability of melanin to reduce silver from a silver solution. Examples of argentaffin stains include Fontana Masson and GMS. Argyrophil are similar stains but use an external suaver reducer to produce elemental silver (Grimelous stain).
These agents also stain neurosecretory granules and formalin pigment.
IHC of Melanocytes?
Positive for vimentin and S-100 protein
Melanocytes produce what cytokines in response to inflammation?
Secrete IL-8, IL-1α, TNFα, MCF in response to inflammation
Functions of melanocytes:
cosmetic entity: protective coloration (camouflage and mimicry), social communication (sexual attraction)
barrier against ionizing radiation: protection against UV light
scavenger of cytotoxic radicals and intermediates: esp. during rapid proliferation and differentiation of the hair follicle
additional roles include: restrain keratinocyte proliferation, calcium homeostasis
secrete IL-8, IL-1α, TNFα, MCF in response to inflammation
Pigmentation: Steps (4 steps)
The synthesis and distribution of melanin in the epidermis, involves several steps:
Transcription of proteins required for melanogenesis
Melanosome biogenesis
Transport of melanosomes to the tips of melanocyte dendrites
Transfer of melanosomes to keratinocytes
Constitutive pigment
the pigmentation that is genetically determined in the absence of stimulatory influences
Facultative pigment
the pigmentation which occurs with various stimuli (ex UV light, inflammation, hormones)
Normal human skin colors and pigments (4)
Red: Dermis by oxygenated hemoglobin
Yellow: Epidermis by exogenously produced carontenoids
Brown: Endogenously produced melanin
Blue: Dermis by unoxygenated hemoglobin in venules
What is the rate limiting step in melanogenesis?
Tyrosinase is the rate limiting enzyme in the melanin pathway
Melanogensis place exclusively within melanocytes and on the specialized organelle, the melanosome
Where are the majority of melanocytes present?
Largest number of melanocytes is present in the skin and hair
Present in the epidermal basal layer
Approx 1 melanocyte per 10-20 basal keratinocytes in dogs
Nevi and melanomas
mislocation of melanocytes in the dermis.
Constant need for synthesis and transfer of melanosomes from melanocytes to keratinocytes to maintain continuous pigmentation
Where is the only site of pigment production for the hair shaft? How are these different that epidermal melanocytes?
Hair bulb: only site of pigment production for the hair shaft
Located in the proximal hair bulb during anagen phase
Differ from epidermal melanocytes by being larger, more dendritic, with more extensive Golgi and RER, and by producing larger melanosomes.
Melanocytes proliferate, migrate, & undergo maturation during _____
early to mid-anlagen
Depends on c-Kit expression by melanocytes and SF synthesis by keratinocytes
Melanogenesis & melanin transfer to keratinocytes occur throughout anagen phase
Melanocytes apoptose during late catagen
Melanogenically active melanocytes are restricted to the upper hair matrix of the anagen hair follicle, below the pre-cortical keratinocyte population, correlating with the anagen transfer of melanin predominately to the hair shaft cortex, less to the medulla, & only rarely to the hair cuticle
Melanocytes determine hair color by the amount of ______ transferred as well as by the ____ of eumelanin (black-brown) to pheomelanin (red-yellow)
melanin
ratio
Define Graying
the gradual decline in the melanogenically active melanocytes in the hair bulb (determined by age and genetics); there is also a component of ineffective melanosome transfer
Where do ocular melanocytes reside? What is their main function?
Reside in the cochlea; important for hearing
Reside as intermediate cells of the stria vascularis
A loss of otic melanocytes leads to deafness.
Help in the maintenance of the endolymph through the regulation of potassium transport
Endolymphatic fluid of the cochlea maintain high concentrations of positively charged ______.
high concentrations of positively charged potassium
In the absence of cochlear melanocytes, the endolymphatic potential is low, leading to deafness.
Congenital Hereditary Sensorineural Deafness - what are the genes involved in cats and dogs?
Usually seen in dog and cat breeds with white pigmentation
Dogs carrying the piebald or merle genes & cats carrying the white gene, the hair cell loss is secondary to degeneration of the cochlear blood supply
studies demonstrate an absence of strial melanocytes
In the Dalmatian, postnatal auditory function development has been shown to proceed normally up until 3 wks at which point the strial degeneration produces rapid loss of hair cell function
In general, loss of melanocytes at any age — deafness
Waardenburg syndrome type 2
deaf due to lack of melanocytes within the inner ear (like the Dalmatian)
***However there is a preservation of hearing in albinos, which indicates that melanin production within otherwise viable melanocytes are not essential for hearing. BUT albinos are more susceptible than the normal population to hearing loss from noise and/or exposure to toxic agents, therefore melanin provides some protective effect.
Where are cephalic melanocytes located?
Dispersed throughout the meninges; particularly dense in the leptomeninges above the pons and medulla oblongata.
Function as scavengers for toxic cations and ROS
Melanocytogenesis
- Precursor cell (melanoblasts) development and migration from the neural crest to peripheral sites
- Differentiation of melanoblasts into melanocytes
- Survival and proliferation of melanocytes
- Synthesis of melanosomes and melanin
How do melanoblasts development and migration from the neural crest to peripheral sites
Derived from neuroectoderm; from the dorsal portions of the closing neural tube.
Migrate along a dorsolateral path between the mesodermal and ectodermal layer.
Final destinations include: hair follicles, skin, inner ear cochlea, choroids, ciliary body, and iris
Defects in melanocyte migration typically appear most prominently on the ventral surface as ‘white spotting’, as this is the greatest distance (watershed zone) form the dorsum
Factors that influence the development and migration of melanoblasts
Wnt
ET3 (endothelin-3)
BMPs (bone morphogentic proteins)
steel factor (SF; stem cell factor SCF, c-Kit ligand)
Hepatocyte growth factor (HGC)/scatter factor
Wnt glycoproteins play an important role in melanogenesis via what ways?
Expressed in the dorsal neural tube during neural crest cell migration
Directs the maturation of pluripotent neural crest cells into melanoblasts
Bind and activate Frizzled, a transmembrane heptahelical G protein-linked receptor for Wnt
Induce the accumulation of β-catenin
Regulation of microphthalmia-associated transcription factor (Mitf) transcription
Mift (microphthalmia transcription factor) is the primary regulator of melanogenesis. Mift induces the transcription of what 4 factors to allow melanogenesis to occur?
- tyrosinase
- tyrosinase-related protein-1 (TRP-1)
- 3,4 dihydrozyphenylalanin (DOPA)
- chrome tautomerase (TRP-2)
Bone Morphogenetic Proteins: (BMPs):
Belong to the transforming growth factor-β family
Suppresses neural crest cell differentiation into melanoblasts:
• Wnt antagonists
• There is a decrease in BMP expression in the dorsal neural tube at the time of melanoblast migration
Steel Factor
Early melanoblast development requires the presence of the cytokine SF (mast/SCF, c-Kit ligand) and its tyrosine kinase transmembrane receptor c-Kit
Stem Cell Factor
crucial role in permitting melanoblast survival and promoting proliferation, both initially in the dorsolateral migration pathway and later from the dermal mesenchyme to colonize the follicles and epidermis
Regulation of microphthalmia-associated transcription factor (Mitf) transcription by ____ Signaling
Wnt Signalling
When Wnt is not bound to Frizzled is inactive
Wnt binding activates Frizzled leading to GSK3β (enzyme glycogen synthase kinase 3β) dissociation from β-catenin, which then translocates to the nucleus where it binds with specific transcription factors (Tr factor to initiate Mitf transcription).
What is c-Kit?
a tyrosine kinase receptor involved in melanoblast expansion, survival and migration
Expressed by epidermal keratinocytes; as soon as c-Kit is expressed on melanoblasts, these cells begin their migration to their final destination
Endothelins: (ETs)
Involved in the late steps of melanoblast migration from the dermis to the basal layer of the epidermis.
ET1, ET2, ET3
ET3: Synthesized by ectodermal cells
EdnrB receptor interacts with ET3; important during melanoblast migration along the dorso-lateral pathway; proper expression is required for survival, proliferation, and/or migration of melanoblasts and normal formation of enteric nerves.
Defects in ET3 or EdnrB (Waardenburg Syndrome 4) results in prominent melanocytes loss
What cells synthesize ET3?
Synthesized by ectodermal cells - interact with endothelia receptor B on melanoblasts
EdnrB receptor
interacts with ET3; important during melanoblast migration along the dorso-lateral pathway; proper expression is required for survival, proliferation, and/or migration of melanoblasts and normal formation of enteric nerves.
Defects in ET3 or EdnrB result in prominent melanocytes loss
Hepatocyte Growth Factor: (HGF/Scatter Factor)
Involved in the late steps of melanocyte migration from the dermis into the epidermis
Ligand for the transmembrane tyrosine kinase receptor Met.
Also regulates cadherin expression in melanocytes (down regulates E-cadherin)
The cytoplasmic domains for cadherins in melanocytes bind to what structure?
In melanocytes, their cytoplasmic domains bind to β-catenin
E-cadherins where before entry into the epidermis?
induced in melanoblasts before their entry into the epidermis; later when the melanoblasts migrate into the hair follicle, expression is muted, and the cells begin to express P-cadherin
Role in suppressing melanocytes proliferation in the epidermis
Cadherin expression by melanocytes matches the cadherin expression of surrounding cells
Melanosome
unique membrane-bound organelle in which melanin biosynthesis takes place; thought to represent a modified lysosome as it contains enzymes and other proteins
- Proteins found in both lysosomes and melanosomes: liposomal-associated membrane protines (LAMPs), acid phosphatase
- Like a lysosome, melanosome can endocytose receptors that are targeted for degradation
Where are the genes located for proper melanosome function?
located on the pink-eye dilution and mottled loci
What are the two TYPES of melanosomes?
Can be divided into eumelanosomes and pheomelanosomes,
Eumelanosomes: large (0.9 x 0.3 μm), elliptical in shape, contain a highly structured fibrillar glycoprotein required for eumelanin synthesis.
Pheomelanosomes: smaller (0.7 μm in diameter), spherical in shape, their glycoprotein is loose and disorganized
What are the four stages of melanosome maturation?
Stage I melanosomes: (premelanosomes) develop from endoplasmic reticulum; have an amorphous matrix and display internal vesicles that form as a result of membrane invagination; already contain glycoprotein Pmel17 (gp100) but requires further processing to become a component of the final fibrillar matrix
Stage II melanosomes: Eumelanosomes at this stage have organized structured fibrillar matrix, but no active melanin synthesis. They contain tyrosinase In pheomelanosomes, melanin synthesis already takes place
Stage III melanosomes: deposition of melanin on the fibrillar matrix is detected within eumelanosomes
Stage IV melanosomes: fully melanized and their internal matrix is masked by melanin deposits
Stage I melanosomes
(premelanosomes) develop from endoplasmic reticulum; have an amorphous matrix and display internal vesicles that form as a result of membrane invagination; already contain glycoprotein Pmel17 (gp100) but requires further processing to become a component of the final fibrillar matrix
Stage II melanosomes
Eumelanosomes at this stage have organized structured fibrillar matrix, but no active melanin synthesis. They contain tyrosinase In pheomelanosomes, melanin synthesis already takes place
Stage III melanosomes
deposition of melanin on the fibrillar matrix is detected within eumelanosomes
Stage IV melanosomes
fully melanized and their internal matrix is masked by melanin deposits
Where are melanogenic proteins produced in the cell?
In general: structural proteins involved in melanogenesis are thought to originate from the RER and enzymatic proteins are through to originate from the Golgi.
What process is required for proper association of tyrosinoase with melanosomes and its activity?
*****Glycosylation steps are required for proper association of tyrosinase with melanosomes as well as for its activity
- Mature tyrosinenase is folded in the ER, the trafficked/sorted in the Gogli apparatus & then into endosomes & finally melanosomes
Tyrosinase mutations
- Found in Oculocutaneous albinism type I
- Complete inability to synthesize melanin in skin, hair and eyes
- Melanocytes are present
TRP-2
****also known as DOPAchrome tautomerase
Synthesized in the ER; undergoes several maturation steps in the Golgi and trans-Gogli network
Eventually localizes in the melanosome & spans its membrane
W/n the melanosome, it complexes with tyrosinase and TRP-1
***During melanin synthesis, TRP-2 converts DOPAchrome to the carboxylated derivative DHICA
Requires zinc for proper function
Protein Kinase C-β
involved in regulating tyrosinase activity
Phosphorlyates serine residues on the cytoplasmic domains of tyrosinase, thus activating it
In melanocytes, activated PKC-β is associated with melanosomes
Pmel17
Glycoprotein that plays a critical role in fibril matrix formation within eumelanosomes
Transcription is induced by α-MSH through Mitf
Synthesized in the ER, glycosylated and cleaved then transported to stage I melanosomes to form a fibrillar structure that is the backbone of eumelanosome matrix, contributes to melanosome ellipsoid shape, and promotes melanin polymerization
MART-1/Melan A
membrane-associated protein present in stage I and II melanosomes
Forms a complex with Pmel17; affects the expression, stability, trafficking, and processing of Pmel17 within the melanosomes
P Protein
(pink-eyed dilution); a transmembrane protein responsible for maintaining acidic environment within the melanosomes
SLC24A5
melanosome-associated cation exchanger; mutations in slc24α5 in zebrafish lead to hypopigmentation
Lysosomal-Associated Membrane Proteins
linked to melanosome membranes and/or matrix; thought to protect melanosomal integrity by acting as scavengers of free radicals that are produced during melanin biosynthesis
Microphthalmia-Associated Transcription Factor
***Regulatory Protein***
Mitf
Termed the master gene for melanocyte survival
- Key factor regulating the transcription of the major melanogenic proteins (tyrosinase, TRP-1, TRP-2, PKC-β, MART-1)
Mitf Role in Melanocyte Proliferation and Survival
Promotes melanocyte survival by upregulating the expression of a major anti-apoptotic protein, BCL2
- It is frequently overexpressed or amplified in melanomas
**Mutations in MITF are found in Waardenburg syndrome type 2
Melanocortin 1 Receptor
Comprise a family of 5 related receptors; each with 7 transmembrane domains that belong to the G-protein coupled receptor superfamily
MC3R and MC4R: found in the CNS, absent in melanocytes
MC2R: expressed in adrenal cortex
MC5R: expressed in peripheral adipocytes
MC1R
expressed in a number of cells, including endothelial cells, fibroblasts, keratinocytes; but HIGHEST expression is found in melanocytes
ACTH &α-MSH activate MC1R which leads to G protein-dependent activation of adenylate cyclase increased intracellular cAMP level, induced Mitf transcription and upregulating the level of melanogenic proteins (tyrosinase)
Polymorphisms with MC1R
largely responsible for different skin/hair color among humans
Agouti gene encodes an antagonist for MC1R
_____ gene encodes an antagonist for MC1R
Agouti
α-MSH is an antagonist for which cytokine?
IL-1
What is the Raper-Mason pathway?
pathway for the biosynthesis of melanin. The bifunctional enzyme, tyrosinase (A) converts tyrosine and dopa to dopaquinone. Tyrosinase is assumed to be the sole enzyme associated with this pathway and the rest of the reactions (B) are presumed to be of nonenzymatic origin
**inhibition of this reaction blocks melanin synthesis
Melanosome Transfer
The melanosome is transferred from the origin in melanocyte perikaryon to the tips of the dendrites
Takes place on microtubules, arranged parallel to the long axis of the dendrite
Describe the melanosome exocytosis pathway
involves fusion of the melansomal membrane with the melanocyte plasma membrane, melanosome release into the intercellular space, and phagocytosis by surrounding keratinocytes.
Melanosome Cytophagocytosis
indicating the phagocytosis of a live cell or a portion of it; the keratinocyte cytophagocytose the tip of a melanocyte dendrite, which then fuses with lysosomes inside the keratinocyte, which is transported to a supranuclear location where the phagolysosome membranes break up, releasing the melanosomes.
Melanosome Fusion of plasma membranes
Fusion of keratinocyte and melanocyte plasma membranes creates a space through which melanosomes are transferred from the melanocyte to the keratinocyte. There is evidence of filopodia (slender, filiform, pointed cytoplasmic projections at the tip of melanocyte dendrites) that adhere and fuse with keratinocyte plasma membranes.
What is protease-activated receptor-2 (PAR-2)?
- Controls melanosome ingestion and phagocytosis by keratinocytes
- Exerts a regulatory role in skin pigmentation
- UV induced activity and expression
- Inhibition of PAR-2 willprevents UVB-induced tanning
Melanosomes are degraded by what enzymes?
Undergo degradation by lysosomal acid hydrolase enzymes (released from keratinocytes) and are extruded into the stratum corneum
Melanin is discarded; amino acids, lipids, and carbohydrates may be reabsorbed below the cornified layer and reutilized in epidermal metabolism
List melanogenic inhibitors
Sphingolipids: a class of membrane-associated lipids that act as signal transducers; decrease melanocgenesis (in part by enhancing Mitf degradation)
BMP-4: downregulates tyrosinase expression
TNFa
IL-1
IL-6
What is the function of melanin?
Provide protection against UV-induced DNA damage by absorbing & scattering UV radiation (280 – 400 nm)
- Radiation is converted into heat (less toxic form of energy)
What is the tanning response? immediate tanning? Delayed tanning?
UV irradiation induces tanning: facultative skin color; Increase above baseline or constitutive skin pigmentation that provides protection against future UV irradiation
Occurs within 5 to 10 minutes of exposure and fades within minutes to days
Represent melansomal relocation from the perikaryon to the melanocyte dendrites
Occurs within 3 to 4 days after exposure (both UVB (280-340) & UVA (320-400))
Increase in epidermal melanocytes, melanocyte dendricity & melanosome transfer to keratinocytes with greater melanization of individual melanosomes
Total amount of epidermal melanin in increased
What are the direct effects of UV Irradiation?
Triggers several biologic reactions though interaction with cellular chromophores that absorb photons
Most UVA effects - results of oxidative damage mediated through UVA absorption by cellular chromophores like melanin precursors that act as photosensitizers
UVB irradiation - directly absorbed by cellular DNA, leading to the formation of DNA lesions (mainly cyclobutane dimmers and pyrimidine pyrimidone photoproducts)
p53 protein is activated; plasma membrane lipids release DAG which then activates PKC-β to stimulate melanogenesis by activating tyrosinase
How does the Maltese dilution effect coat color in cats?
Maltese dilution: non-uniform distribution of very large, irregularly sized and shaped melanin granules.
an autosomal recessive trait of cats that dilutes black cats to blue, and orange cats to cream
Describe the Horses Coat Color
Dominant W allele: horse will lack pigment from birth but have blue or brown eyes
C allele: causes pigment dilution
G allele: horses are fully colored at birth but subsequently acquire increasing number of white hairs with age; skin and eyes remain pigmented
Ex: Lippizaner, Camarque
Leukoderma in horses
Common in horses
May be localized or multifocal; temporary or permanent
Often seen as a complication of onchocerciasis, dourine, herpes coital examthema, lupus erythematosus, pressure sores, ear papillomas, ventral midline dermatitis, regressing viral papillomatosis, or freezing and burns (chemical, thermal, radiation)
Reported to follow contact with phenols and rubber (many rubbers contain ***monobenzyl ether of hydroquinone*** which inhibits melanogenesis)
_____ may cause hyperpigmentation in dogs, attributed to iron deposition. Change in humans may be seen with parenteral/topical application of silver, gold, or mercury.
Minocycline
Color dilution and cerebellar degeneration in Rhodesian Ridgebacks
Age: Birth
Breed: Rhodesian Ridgebacks
Cause: Autosomal recessive
Clinical Signs: Pups born with dilute color in coat, neurologic abnormalities by 2 weeks of age. Coat color is bluish. Pups are ataxic, difficulty crawling and nursing – open eyes late
Diagnosis: Purkinje cell degeneration and cutaneous abnormalities typical of color dilution alopecia
What is piebaldism?
Genetically determined white spotting. Dominant trait. Melanocytes are absent or incompletely differentiated in affected sites.
Graying
Age: Mature dogs
Breed:German Shepherds, Irish setters, Labrador retrievers, Goldens more likely to develop gray muzzles and chin at younger age
Cause: Thought to result from reduction in melanocyte replication through senescence. Appears to be age and genetically related since certain breeds are more prone
Clinical Signs: Graying of hair
Tyrosinase Deficiency
Breed: Chow chows.
Cause: Deficiency of tyrosinase.
Clinical Signs: Normal bluish/black tongue turns pink. Portions of hair shafts turn white. Buccal mucosa may rapidly depigment.
Diagnosis: Confirm with skin biopsy
Histopath: Tyrosine added to histo prep. Incubated. Melanin measured after tissue staining.
Tx: No effective tx, however, melanin reappears in 2-4 months spontaneously.
Chromatophore
Pigment-containing and light-reflecting cells found in amphibians, fish, reptiles, crustaceans, and cephalopods
Largely responsible for generating skin and eye colour in cold-blooded animals
Generated in the neural crest during embryonic development
Grouped into subclasses based on their colour (more properly “hue”) under white light:
• xanthophores (yellow)
• erythrophores (red)
• iridophores (reflective / iridescent)
• leucophores (white)
• melanophores (black/brown)
• cyanophores (blue
Cyanophores
A cell in the skin of frogs, chameleons, etc, in which pigment is concentrated or dispersed, causing the animal to change color.
Merkel Cell - What are the markers and stains? Function?
Markers and Stains: Cytokeratin-20
Functions:
1) Slow Adapting Mechanoreceptor innervated with type I
2) Control sweat production, hair follicle and blood flow
3) Keratinocyte proliferation
Cutaneous vasculature function
Skin metabolism
Temperature regulation
Defense system against invaders:
Leukocyte trafficking
Wound healing
Angiogenesis
Nutritional supply of the skin – for epidermis, sweat glands, sebaceous glands, hair
Vasculature arrangement in dog model
3 intercommunicating plexuses of arteries and veins (humans only have 2)
- Deep plexus
- Middle plexus
- Superficial plexus
Deep plexus
Found at the interface of the dermis and subcutis
Branches from this plexus descend into the subcutis and ascend to supply the lower portions of the hair follicles and the epitrichial sweat glands, feed the middle plexus
Middle plexus
Lies at the level of the sebaceous glands gives off branches to the arrector pili muscles
Ascending and descending braches supply the middle portions of the hair follicles and the sebaceous glands
Ascending branches to feed the superficial plexus
Superficial plexus
Just below the basement membrane
Contains capillary loops that supply the epidermis and upper portion of the hair follicles
Arterioles
Consist of endothelial cells surrounded by basement membrane & two layers of smooth muscle cells
Function: as part of the resistance vessels in skin
Arterial and venous capillaries
Lack surrounding smooth muscle cells
Postcapillary venules
The majority of the superficial dermal vessels
Most physiologically reactive segment of the microcirculation
Site where inflammatory cells migrate from the vascular space into tissues
Site where endothelial cells develop gaps that result in increase vascular permeability during inflammation
Immue complexes are typically deposited here
Blood vessel endothelial cells are characterized by?
- Ultrastructurally by a peripheral basement membrane and intracytoplasmic Weibel-Palade bodies (rod-shaped tubular structures enveloped in a continuous single membrane)
- Storage for Factor VIII, P-selectin, histamine
- By their possession of factor VIII (vonWillebrand) antigen, plasminogen activators, & prostaglandins
- By CD31 (platelet endothelial cell adhesion molecule [PECAM])
- By being phagocytic (esp in the presence of carbon, mycobacteria or fibrin)
Blood vessel endothelial cells key function?
key regulators of leukocyte trafficking
Selectins are upregulated on what structures during inflammation?
(E- and P-selectin) and members of the immunoglobulin superfamily (ICAM-1, vascular cell adhesion molecule-1 [VCAM-1]) are upregulated on endothelial cells with inflammatory stimulation
What two inflammatory cytokines induce E-selectin
induced by IL-1 and TNFα
P-selectin
Stored in Weibel Palade bodies
Specific for leukocyte recruitment to the skin
Mediate rolling, adhesion, and transmigration of leukocytes from the blood stream
ICAM-1/ICAM-2
expressed on skin microvessel endothelial cells
ICAM-1 is upregulated by IL-1, TNFα, and IFNγ
Both serve as the binding site to leukocyte LFA-1
General adhesion molecules of lymphocytes
VCAM-1 (Vascular cell adhesion molecule)
Not normally present on endothelial cells
General adhesion molecule for lymphocytes
Induced by TNFα or LPS
Glomus: Hoyer-Grosser’s organ
- A special arteriovenous shunt located within the deep dermis
- Consist of an arterial segment (Sucquet-Hoyer canal) and a venous segment
- Rich cholinergic nerve supply
Pericytes
- Aligned parallel to blood vessels on the dermal side
- Vary from fusiform to clublike in appearance
- Contractile cells, containing actin-like and myosin-like filaments
- Important in regulating capillary flow
- Larger # of pericytes surround the endothelial cells in post capillary venules
- Pericytes and endothelial cells make numerous contacts over their entire surface
Function of pericytes?
- Important in regulating capillary blood flow
- Essential for microvessel stability
- Participate in the control of angiogenesis
- Major contractile cells responsible for producing the endothelial cel gaps in the postcapillary venular segment where the events of inflammation occur
- Endothelial cell gaps develop both, at the interendothelial cell junction and through the cell itself
- Gap formation is associated with agents such as histamine, bradykinin
Veil cells
- Surround all dermal microvessels, entirely external to blood vessel walls
- Flat, adventitial, fibroblast-like cells
- Stain for factor XIIIa - component of the dermal dendrocyte system
- Negative for 1 or B cell markers and CD1a negative
- Demarcates the vessel from the surrounding dermis
- Perivascular mast cells are usually present in the space between the vascular wall and the surrounding veil cells
Innervation of the cutaneous blood vessels
Under dual vasomotor control
Separate nervous signals for vasoconstriction and vasodilatation
Under normothermic conditions, skin blood flow averages approximately 5% of cardiac output
Reflex vasoconstriction (in response to cold and baroreflexes) is mediated through what fibers?
adrenergic sympathetic fibers distributed widely over most of the skin (epinephrine/norepinephrine)
In most of the skin vasodilatation occurring during heat exposure depends on _____ signals that cause blood vessels to dilate = active vasodilatation
sympathetic nervous signals
o Occurs in almost all skin outside the acral regions
o Vasodilatation occurs near the onset of sweating & therefore is not sensible to small temperature changes
o The neurotransmitter is unknown however, there is evidence that active vasodilatation is mediated by release of acetylcholine.
Sympathetic nervous system receptors
β-receptors: relax smooth muscle cells causing vasodilation
α-receptors: constrict smooth muscle cells causing vasoconstriction
Mediators of smooth muscle tone:
Catecholamines: activate both α & β receptor; Because α receptors predominate, vasoconstriction occurs
Vasodilators:
Histamine
Bradykinin
Kallikrein
Carbon dioxide
Lactate
NO
Substance P
Skin temperature affects heat loss responses also through direct actions on the effectors themselves
Specialized organelles of endothelial cells
Weibel-Palade bodies (WPB)
Fenestrae
Caveloar system
Weibel-Palade bodies (WPB):
Elongated, membrane-bound organelles that display longitudinal striations
Distributed randomly throughout the cytoplasm in vascular endothelial cells
Contain von Willebrand factor (vWF, factor VIII-related antigen), P-selectin (CD62P), lysosomal-membrane-associated glycoprotein 3 (LAMP-3 or CD63), 1,3-fucosyltransferase VI, and IL-8
Endothelial junctions
Neighboring endothelial cells connect to each other by forming interdigitating protrusions connected by junctions
Vary depending on the site of the vascular tree – allow for varying degrees of permeability for proteins and cells
Intercellular tight (occluding) junctions:
Adherens junctions
Gap junctions
Complexus adherens or syndesmos
Intercellular tight (occluding) junctions
Consist of continuous belt-like meshwork of six anastomosing junctional strands, which prevent the transport of macromolecules
Virtually absent in postcapillary venules
What are gap junction proteins?
connexins
Molecular markers of endothelial cells
vWF, CD31 (PECAM-1),ICAM-2,CD141 (thrombomodulin)
vWF
most widely used marker, which may be detected in WPB or within Golgi apparatus or endoplasmic reticulum; also found in megakaryocytes and platelets
CD31 (PECAM-1)
found on endothelium, platelets & circulating leukocytes
Nearly all endothelium secretes and binds CD143 (angiotensin-converting enzyme) to the cell surface
Most endothelial cells express anticoagulant protein CD141 (thrombomodulin)
ICAM-2
May be the single best marker for endothelium mainly as it is not generally found on other cell types
Endothelial Role in Leukocyte trafficking
Initial tethering, rolling, and firm adhesion
Initial tethering: mediated by selectins
L-selectin (CD62L), E-selectin (CD62E), and P-selectin (CD62P)
P-selectin: endothelial cells
L-selectin: leukocytes
E-selectin: expressed by endothelial cells; induced by TNF-α or IL-1β
Adhesion of leukocytes to endothelium is mediated by leukocyte integrins
β2 integrins (CD18): LFA-1 & Mac-1; induced by PAF – Bind to ICAM-1
Emigration/Diapedesis
Lymph vessel location
o Lie in the superficial dermis and surround the adnexae, usually not seen above the middle dermis
o Arise from capillary networks ; drain into a subcutaneous lymphatic plexus
Lymph vessel distinguishing features from capillaries
Have wider and more angular lumina; have flatter and more attenuated endothelial cells
Have no pericytes
Contain little to no blood in normal situations
Lymph vessel function
o Regulate pressure of the interstitial fluid by resorption of fluid released from vessels
o Clear the tissues of cells, proteins, lipids, bacteria, and degraded substances including materials that have penetrated the skin
o Link the skin and regional lymph nodes
o Lymph flow within the skin depends on movements of the tissue caused by arterial pulsations, muscle contractions and body movement; backflow is prevented by bicuspid-like valves within the vessels
Intermediate filaments
Intermediate filaments connect the nuclear membrane to hemidesmosomes on the keratinocytes membrane. These filaments are responsible for maintaining the integrity and structure of basal epidermal cells.
Composition: Keratin-5 + Keratin-14
Inner plaque components
plectin + BPAG1-e
Outer plaque components
Integrin alpha-6/beta-4 + cytoplasmic domain (aminal terminus) of collagen XVII (BPAG2, BP180).
Anchoring filaments
Anchoring filaments connect hemidesmosomes to anchoring fibrils throughout the lamina lucida. The lamina lucida is thought to be a fixation artefact and probably does not exist in vivo.
Composition: intact and shed extracellular domain (carboxyl terminus) of collagen XVII (BPAG2, BP180).
What are focal contacts?
Focal contacts are inter-hemidesmosomal adhesion structures that are mostly functional during wound healing in which epidermal migration occurs.
Composition (main components): integrins alpha-2/beta-1, alpha-3/beta-1, alpha- 5/beta-1; laminin isoforms (3A32, 3A11); collagen XXIII; nidogens, fibulins.
In the skin, integrin alpha-6/beta-4 connects intermediate filaments, via hemidesmosomes, to what structure?
laminin-5 (Lm3A32)
Both alpha-2/beta-1, alpha-3/beta-1 and alpha-5/beta-1 integrins are present at non-hemidesmosomal focal contacts during _______ migration. In this case, they will link the laminin-5 (Lm3A32)/-6(Lm3A11) complex to the actin microfilament network.
basal keratinocyte migration
Large deletions in ____ have also been found to underlie JEB in Charolais calves and in Churra sheep.
ITGB4
Epidermal stratum germinativum differentiates into hair germs (primary epitheial germs) that give rise to what three structures?
hair follicles, sebaceous glands and epitrichial glands
Hair germs initially consist of an area of crowding of deeply basophilic cells in the basal layer of the epidermis. Subsequently, the areas of crowding become buds known what structure that protrudes into the dermis?
epidermal placodes
Beneath each epidermal placode lies a group of mesenchymal cells, referred to as what structures, from which the dermal hair papilla is later formed?
dermal condensate
The epidermal placode develops into three epithelial cylinders. The innermost cylinder forms the hair shft. The outermost cylinder forms what structure?
Outer root shealth
As the hair peg lengthens and develops into a hair follicle and hair, three buges appear. The lowest (deepest) of the bulges develops into the attachement for what structure?
arrector pili muscle
As the hair peg lengthens and develops into a hair follicle and hair, three buges appear. The middle buldge develops into the attachement for what structure?
sebaceous glands
As the hair peg lengthens and develops into a hair follicle and hair, three bulges appear. The uppermost bulge develops into the attachment for what structure?
epitrichial sweat glands
*** All these structures develop on the cranial side of the prinary hair follicle***
(secondary hairs develop on the caudal side)
The skin is an active site for ______ metabolism.
fatty acid
Signals controlling hair follicle induction, development, regression and reactivation have not been identified. however, what growth factors or their receptors have been localized to hair follciles and surrounding mesenchyme?
EGF, TGFb, TGFb2, neutotrophin 3
An interplay between class 1 major histocompatibility complex (MHC) expression and what two other factors are involved in the regulation of hair growth, especially during the catagen phase?
chondroitin proteoglycans and activated macrophages
The hair follicle is a source and target of ______, neutropharmacologic manipulations alter hair cycling.
neurotrophins
The trophic effects of cutaneous nerve on follicular growth are exerted via regulation of what four factors?
vascular tone
neuropeptides stimulation of receptors on follicular keratinocytes
dermal papilla fibroblasts
modulation of macrophages and mast cells
Hair growth responds premoninantly to ____; and to a lesser extent_____.
photoperiod
ambient temperature
** sinus hairs are not subjected to seasonal shedding**
The hair cycle and haircoat are also affected by hormonal changes. in general anagen is initated and advanced by thyroid hormone and growth hormone. Conversely, excessive amounts of ____ or ____ inhibit anagen and suppress hair growth rate.
Estrogens and glucocorticoids
The molecular components of the desmosome belong to what three major gene families?
plakins (desmoplakin), armadillo proteins (plakoglobin and plakophilin) and demosomal cadherens (desmoglein and desmocollin)
Linkage of the keratin intermediate filament (cytokeratin) network to the hemidesmosome and the basal keratinocyte plasma membrane involves several components including what 4 structures?
plaque proteins, bullous pemphigoid antigen 1 (BPAG1, BP230) and plectin, the transmembrane proteins a6b4 integrin and BPAGII (collagen XVII) and laminin 332A (laminin 5)
Release of profilagggrin from keratohyalin granules is followed by its _____ dependent cleavage into filaggrin monomers that function to aggregate, pack and align keratin filaments.
Calcium-dependent
Keratohyalin granules are composed of what products?
profillagrin, keratin filaments and loricin
What ultrastructural feature characterizes granular cells at the cell margins?
clustered lamellar granules
In rodents, what two morphologic forms of keratohyalin granules occur?
P-F granule is irregularly shaped and contains profilaggrin, whereas the L-granule is smaller, rounded and contains loricrin.
What layer is a fully keratinized, compact, thin layer of dead cells? This layer is anuclear, homogeneous and hyaline like, it contains regratile droplets and a semifluid substance called _____.
stratum lucidum
eleidin
How does the stratum lucidum differ from the stratum corneum histochemically?
Differs from the stratum corneum by being rich in protein bound lipids. The stratum lucidum is best developed in the foot pads and less developed in the nasal planum - absent from all other layers of the skin.
AKA stratum conjuctum
The termically differentiated corneocyte has a highly apecialized structure that forms under its plasma membrane, __________.
the cornified envelope (CE)
The CE develops under the plasma membrane of stratified epidermal cells, cells of the inner root sheath and medulla of the hair follicle and the cuticle of the claw.
Why is the corneocyte not contain a true cell membrane - what component is it missing?
CE contains no phopholipids
CE formation is associated with increased activity of what enzyme (calcium dependent) that catalyze cross-linking of soluable and particulate protein precursors into large insoluable polymers.
Transglutaminase
What are the known physiologic systems that regulate that are important to carry out the biologically processes of the skin (differentiation, immune response, proliferation, inflammation)?
- Protein Kinase C/Phospholipase C Second messenger system
- Receptor-linked Tyrosinekinase system
- Calcium/calmodulin second messenger system
- cAMP dependent protein kinase system
There appears to be a host of hormones and enzymes that can induce and increase the acivity of what enzyme during epidermal proliferation?
Ornithine decarboxylase - this enzyme is essential for biosynthesis of polyamines (putrescine, spermidine and spermine - known to be important for normal keratinization.
What are the four distinct cellular events in the process of cornification?
- keratinization (synthesis of the principal fibrous proteins of the keratinocyte)
- keratohyalin synthesis (incuding the histidine rich protein filaggrin)
- formation of the highly cross-linked insoluable stratum corneum cornified cell envelope (including the structural protein involucrin)
- Generation of neutral lipid enriched intercellular domains, resulting from the secretion of distincitive cellular lamellar granules.
Lamellar granules are synthesized primarily where?
lamellar granules are primarily synthesized in the stratum spinosum and displaced to the apex of the periphery of the cell as it reaches the stratum granulosum. They fuse with the plasma membrane and secrete their contents (phospoholipid, ceramides, FFA, hydrolytic enzymes and sterols).
What labeling technique has been shown that the turnover (cell renewal) time fot he viable epidermis of the dog is approximately 22 days?
Tritated thymidine
Clipping = 15 days
Cocker spaniels = 7 days
The use of what technique provides useful model for in vitro studies of epidermal kinetics, pathogenesis of various dermatological disease and cutaneous effected of arios pharmacologic agents?
cultured canine keratinocytes
***express antigens associated with PV, PF, and deposit laminin V and type IV collagen***
Synthesis of epidermal lamellar bodies is synthesized where in the epidermis?
Stratum spinosum
List the contents in lamellar vesicles?
- polar lipids
- glucosylcermadies
- sphingomyelin
- free sterols
- phospholipids
- hydrolytic enzymes
In response to increase in what the concentration in the stratum granulosum, will cause the lamellar bodies to fuse with the plasma membrane and secrete their contents into the intercellular spaces?
calcium
Ceramides are _____ linked fatty acids containing long-chan amino ______ (sphingoid base)
amide-linked
alcohol
_______ are the most important lipid component for lamellar arrangement in the stratum corneum and in barrier function.
Ceramides
_____ are importnat because they are incorporated into the ceramides.
PUFAs
Linoleic acid, an essential fatty acid for dogs and cats, in found in cermamides ____ , ___ and ___.
Ceramides 1, 4, 9
Which two ceramides bind to involucrin and other proteins of the CE, forming a scaffold for the binding of other lipids into an intercellular lipid bilayer located parallel to the corneocytes in the stratum corneum?
ceramides 1 and 2
PUFAs have important roles in the skin as precursors of _____.
Eicosanoids
PUFA fatty acids are liberated from phopholipids by ____ and are subsequently metabolized by ____ and ____ enzymes into prostagladines and leukotrienes.
PLA2
Cyclooxygenase, lipoxygenase
Skin surface lipids can be studied qualitatively by various extraction methods or quantitatively using ______________.
the lipometer or sebumeter.
Melanocytes, the second type of cell found in the ___ layer of the epidermis, are also found in the outer root shealth and hair matric of the hair folluicles, in the ducts of sebaceous and sweat glands and to a lesser extent in the superficial dermis.
basal layer of the epidermis
Melanocytes are derived from where? and mirgrate inot the epidermis in early fetal life.
neural crest
They are also found int he eyes (retinal pigment epithelium), cochlea (stria vascularis) and meniges.
Melanocytes have long cytoplasmic extensions (dendrites) that weave among keratinocytes and trasnfer pigment containing ______ to the keratinocytes.
melanosomes
*** In general - there is 1 melanocyte per 10 to 20 keratinocytes in the basal cell layer of the canine skin = epidermal melanin unit***
Although melanin absorbs light over a broad spectrum, including UVA and UVB, it is not partiularly efficient absorber of ____.
UVL
What is the different between UVA and UVB light?
Two types of UV light are proven to contribute to the risk for skin cancer: Ultraviolet A (UVA) has a longer wavelength, and is associated with skin aging. Ultraviolet B (UVB) has a shorter wavelength and is associated with skin burning.
Skin color is also influenced by ____ found in epidermal cells; red from oxygenated hemoglobin and blue from unoxygenated blood.
yellow carotenoids
Pheomelanins differ from eumelanins by containing a high proportion of ____.
sulfur
Despite different properties of the different melanins, they all arise from a common metabolic pathway in which _____ is the key intermediate.
dopaquinone
Melanin synthesis takes place exclusively within the ____ and ont he specialized organelle, the ______.
melanocyte
melanosome
The specific enzyme, ____, catalyzes conersion of tyrosine to dopa.
Tyrosinase
*** rate limiting enzyme in the melanin othway. It is a CU containing enzyme found exclusively in melanocytes, and is thus a good epcific marker for these cells***
What are the catalytic activities of tyrosinase?
The most critical is its tyrosine hydrozylase activity - converting tyrosine to dopa
It is also able to use dopa or 5,6-dihyroxyindole (DHI) as substrates for oxiudase activities.
***mutations in the tyrosine structural gene are responsible for several types of albinism**
Once dopa is formed, it can spontaneosly auto-oxidize into ____ without tyrosinase (although its slower) and continue through the melanin pathway to dopachrome, 5,6-dihydroyindole-2-carboxylic acid, DHI and indole-5,6-quinone.
dopaquinone
Classifications of Melanin: Eumelanin
Dark brown-black
alkali-insoluable
no sulfur content
Polymer of 5,6-dihydroxyindole and 5,6-dihydroxyindole - carboxylic acid
Ellipsoidal (melanosome shape)
Lamellae or fillaments (melanosome structure)
Classifications of melanin: Pheomelanin
Yellow-reddish brown
alkali-soluable
high sulfur content
Polymers of benzothiazine units derived from cyteinyldopa
Spherical (melanosome shape)
Microvesicles/microgranules (melanosome structure)
What enzymes converts dopachrome to DHICA and what metal is required for reaction?
TYRP2 - requires presence of zinc
The determination to produce eumelanins or pheomelanins is predominatly under ____ control.
genetic
If sulfhydryl groups are present, _____ is produced. It has been proposed that the “switching’ of melanin synthesis is mainly controlled by the levels of ______, with high levels producing eumelanins and low levels producing pheomelanins.
pheomelanins
tyrosinase
Melanosomes originate from the _____, where the tyrosinase enzyme is formed.
golgi apparatus
Transfer of melanin involes the endoytosis of the dendrite tips of the incorporated stage ___ melanosomes by the adjacent keratinocytes.
IV melanosomes
Melanocytes eject melanosomes into keratinocytes by a unique biological transfer process called _____.
cytocrinia
Steps in melanogenesis
- Melanoblast migration from neutral crest
- Melanoblast differentiation into melanochtes in skin
- Melanosome gene activation
- Synthesis of tyrosinease and melanogenic proteins
- Posttranslational processivng and glycosylation of tyosinases and transport
- fusion of vesicles to form. lanosomes
- Control of activity of tyrosinase and tyrosine-related proteins
- Melanin biosynthesis
- Melanosome transfer to keratinocyte
- Melanosome degradation
- Melanin removal with loss of cornified cells
A pivitol transcription factor in expression and differentiation of many enzymes involved in melanin synthesis is ________.
microphthalmia transcription factor (MITF)
One major determinant of melanogensis is the _____, a G-protein coupled receptor that regulates pigment phenotype.
melanocortin 1 receptor (MC1R)
Agonists of MCR1 include _______ and ______.
alpha-melanocte-stimulating hormone (MSH) and corticotropin.
_____ gene encodes an antagonist of MCR1.
Agouti
What is a tyrosinase kinase receptor involved in melanobast expansion, survival, and mirgration. Mutations in this have been identified in piebaldism and have leads to progression of lemanocytes toward metastatic melanoma phenotypes.
c-Kit
Melanocytes secrete what cytokine?
IL-8
What is a neutoimmunomodulatory and antiinflammatory peptide that is synthesized and released by keratinocytes, Langerhan cells, fibroblasts, endothelial cells, and melanocytes themselves?
a-MSH
a-MSH can modulated keratinocyte proliferation and differentiation, as well as endothelial cell and fibroblast cytokine and collagenase production.
It also downregulates the production of proinflammatory cytokines and accessory molecules on antigen-presenting cells.
It is an antagonist of IL-1.
Folliculr melanocytes undergo cyclic modifaction in coordination with the hair cycle. Melanocytes are located where in the air follicle during anagen? Melanocytes undergo apoptosis during late _____.
proximal hair bulb
catagen
Argentaffin stain
This stain relies on the abiity of melanin to reduce silver fron a cilver solution.
ex. Fontana Masson and Gomori methaniamine silver
These agents also stain neurosecretory granules and formalin pigment
Argyrophil stains
Similar to argentaffin stains but use an external silver reducer to produce elemantal silver.
example of argyrophil stain is Grimelius stain.
Argyophil stains also stain nerves, reticulum and elastic fibers.
Postassium permanganate can be used to ____ melanocytes to faciliate the study of internal organelles.
bleach
Merkel cell
Dendritic epidermal clear cells confined to the basal cell layer, or just below, and ocur predominatly in the tylotrich pads and hair follicle epithelium.
These specialized cells function as SLOW-ADAPTING TYPE 1 mechanorecptors.
Merkel cells contain a large cytoplasiic vauole that displaced the cell nucleus dorsally and are usually parallel to the skin surface.
Merkel cells may have other functions such as influencing cutaneous blood flow and sweat production (releasing. vasoactive intestinal peptide, coordinaing kerainotcyte proliferation and maintaining/stimulating stem cell population in hair follicle.
What are the IHC markers for merkle cells?
K8, K18,K19 and K20.
THe BMZ can be divided into the following 4 components:
- the basal cell plasma membrane
- the lamina lucida
- the lamina densa
- sublamina densa area (lamin fibroreticularis)
The first three componnets appear to be primarily of the epidermal origin. The epidermal basement membrane is composed of a wide variety of glycoproteins and other macromolecues.
What are the insoluable fibers of the dermis?
collagen and elastin
What are the major soluable macromolecules of the dermis?
proteoglycans and hyaluronan
What accounts for most of the tensile strength and elasticity of the skin; it is involved in regulation of cell growth, proliferation, adhesion, migration, and differentiation, and it modulates wound healing and the structure and function of the epidermis?
dermis
Why is the dermis of scrotal skin unique?
It contains numerous smooth muscle bundles
Most of the dermal extracellylar matrix (fibers and ground substance) is synthesized by what cell?
fibroblasts; which respod to a variety of stimuli such as growth factos elaborated by keraintocytes, infammatory cells and fibroblasts themselves.
What three collagen types predominate in the dermis?
Collagens I, II and V
Dermal fibers consist of what three structures?
Formed by fibroblasts and are collagenous, reticular and elastic
What fibers have GREAT tensile strength and are the largest and most numerous fibers. They are thick bands composed of multiple protein fibrils and are differentially stained by Masson Trichome.
Collagenous fibers
What two ‘unusual’ amino acids are measured in urine as indices of collagen turnover?
hydroxylysine and 4-hydroxyproline
What fibers are fine, branching structures that closely approximate collagen with age. They can be detected best with special silver stains?
Reticulin
What fibers are composed of single fine branches that possess great elasticity and accound for about 4% of the dermal extracellular matrix? They are well visualized by Verhoeff and van Gieson elastin stains.
Elastic fibers
The two components of elastic fibers are amorphous elastin, which contains what two unique cross-linked amino acids along microfibrils which contain ____ and _____?
desmosine and isodesmosine = where all the strength comes from
**fibrillin and type IV collagen**
What collagen type predominates the dermis? Describe it.
Type I collagen (87%), Type III and Type V predominate
Type I is it is triple helix structure composed of three polypeptide a chains coiled around each other much like a rope. This confirmation five the molecule a rodlike shape. Each a chain is composed of approximately 1000 aa with GLYCINE accounting for 1/3 - aa are arranged in repeated triplets.
The remarkable strenth of elastic fibers is dervied from the unusual cross-linking of desmosine and isodesmosine. What is the precurser to elastin?
tropoelastin
Collagen III are composed of how many alpha chains?
composed of three identical a chains with high content of hydroxyproline and glycine that form broad collagen fibers in the dermis and also in the gastrointestinal/vascular connective tissues and fetal skin.
Type V collagen
forms large fibrils and is found in the dermis in association with type I and III collagen
Type VI Collagen
form triple helical microfibrils arranged in a network to stabilize the large type I collagen molecules in the dermis.
Collagen III and collagen V are also concentrated around what structure?
blood vessels
Type IV collagen (lamina densa) and type V (lamina lucida) collagen are found in _____, and type VII collagen is found in the ______ of the BMZ.
BMZ
anchoring fibrils
What two collgen types are considered fibril-associated collagens with interrupted triple helices (FACITS)?
XII and XIV
What factors will cause collagen abnormalities?
Genetic defects, vit C deficiency, iron, copper and from B-aminoproprionitril poisoning.
Collagen synthesis is stimulated by what factors?
ascorbic acid, TGFb, IL-1, IGF1 (somatomedin C), IGF-2, superoxide generating systems and bleomycin.
Collagen synthesis is inhibited by what factors?
GC, retinoids, vit D, parathormone, prostaglandin E2, IFNy, D-penicilliamine and minoxidil.
Collagenases are members of the _____ gene family
MMP
What is a major source of collagenase under normal conidtions of remodeling?
Dermal fibroblasts
Under certain conditions, keratincytes, neutrophils, eosinopihils and macrophages can release a variety of proteolytic enzymes and collagenases.
The superficial dermis contains fine, loosely arranged collagen fibers that are irregularly distributed and a network of fine elastin fibers. The deep dermis contains what arrangement of collagen and elastin?
The deep dermis containsthick densely arranged collagen fibers that tend to parallel the skin surface, and elastin fibers that are thicker and less numerous than in the superficial dermis.
Elastic fibers have extensive cross-linkages owing to the presence of desmosine and isodemosine, which form colvaent cross-links resistant to ______.
degradation
In the superficial dermis, elastic fibers known as ______ fibers are organized in an arcade like arrangment. From these fibers, still thinner eslatic fibers called _____ fibers ascend almost vertically to terminate at the dermoepidermal juinction and anchor to the basement membrane.
elaunin
oxytalan
The _____ present in neutrophils and eosinophils are the most potent, and they readily degrade elastic fibers in disease states.
elastases (serine proteases)
What is the affected protein in Ehlers Danlos syndrome?
alpha chains of types I, III or V collgen; procollagen-lysine; lysyl hydroxylase; procollagen N peptidase; tenascin x; xylosylprotein 4-b-galactosyl-transferase
What is the affected proteins in Cutis laxa syndrome?
elastin, ATP-dependent copper transporter; fibulin 4; fibulin 5
What is the affected protein in EB?
a1 chains of types VII and XVII collagen
The ground substance is a viscoelastic gel-sol of fibroblast origin composed of _____that usually linked in vivo to ______.
GAGs
proteoglycans
The major proteoglycans and GAGs include ____ and _____.
hyaluronate and heparin in the epidermis synthesized by keratinocytes
heparin sulfate and chondroitin 6 sulfate in the BM
What are fibronectins?
glycoproteins capable of muliple interactions with cell surfaces and other matrix components.
They are produced by keratinocytes, fibroblasts, endothelial cells and histiocytes.
Moderate cell to cell interactions and cell adhesions to the substrate, and they modulate microvascular integrity, vascular permeability, basement membrane assembly, and wound healing.
Present in lamina Lucida and lamina dense of BMZ
large glycoprotein that is prominently expressed at epithelial-mesenchymal interfaces. It plays a significant role in epithelial morphogenesis and proliferation and wound healing?
Tenasin
Mucin
In shar peis, large amounts of mucin (mainly HA) are found throughout the dermis, which could be confused with mucinosis in other breeds. Shar Peis have a high level of serum HA than dogs of thother breeds, and increased HA Synthase 2 mRNA transcription by dermal fibroblasts from Shar-Pei dogs.
The hair follicle is divided into what three parts?
medulla, cortex and cuticle
Medulla - the innermost region of the hair, is composed of longitudinal rows of cuboidal cells. The cells are solid near the hair root, the rest of the hair shaft contains air and glycogen vacuoles.
Cortex - the middle layer, consists of completely cornified spindle shaped cells whose long axis is paraellel to the hair shaft. These cells contain the pigment that give hair its color. This part also contributes most to the mechanical propers of the hair fibers.
The cuticle is the outermost region of the hair, and consists of flat, cornified, ancuclear cells arranged like ‘slate on a roof’
Compare secondary hairs and primary hairs
Secondary hairs have a narrower medulla and a more prominent cuticle than primary hairs, and lanugo hairs have NO medulla.
Cats and dogs have a _______ follicle arrangment.
compound hair follicle
In general, the cluster consists of 2 to 5 large primary hairs surrounded by groups of smaller secondary hairs. one of the primary hairs is the largest (central primary hair) and the remaining primary hairs are smaller (lateral primary hairs. Each primary hair has a sebaceus and apocrine gland and an arrector pili muslce.
Secondary hairs may be accompanied only by a sebaceous gland. The primary hairs generally emerge indenpendently through seperate pores; the secondary hairs emerge through a common pore.
List the FIVE major components of the hair follicle
- dermal hair papilla
- hair matrix
- hair itself
- inner root sheath
- outer root sheath
The pluripotent cells of the ____ give rise to the hair and the inner root sheath. The outer root sheath represents a downward extension of the _______.
hair matrix
epidermis
The anagen hair follicle is divided into these three anatomic segments:
- the infundibulum (aka pilosebaceous region - the upper portion which consists of the segment from the entrance of the sebaceous duct (where trichelemmal keratinization begins) to the skin surface.
- isthmus (the middle portions, which consists of the segment between the entrace of the sebaceous duct and the attachment of the arrector pili muslce. This is where the trichohyaline granule containing cellular portion of the inner root sheath begins.
- The ingerior segement (the lowest portion, which extends from the attachemnt of the arrector pili muscle to the dermal papilla).
Where does the trichohyaline granule containing cellular portion of the inner root sheath begins?
isthmus
What constitutes that permanent portion of the hair follicle/ what segement is transitory?
The infundibulum and isthmus make up the permanent portion of the hair follicle.
The inferior segment is transitory.
***dogs have no anatomic structure analogous to the human hair follicular buldge**
Studies in dogs suggest that the stem cells are distributed through the follucular infundibulum and isthmus rather than being concentrated in the bulsge area, as they are through to be in humans.
The inner root sheath is composed of what three concentric layers?
(going from in to out)
- inner root sheath cuticle - a flattened single layer of overlaping cells that point towards the hair bulb and interblock the the hair cuticle)
- the Huxley layer ( one to three nucleared cells thick)
- Henle layer (a single layer of non-nucleated cells
These layers contain eosinophiic cytoplasmic granules called ‘trichohyalin granules’
Describe trichohyalin granules
Eosinophilic cytoplasmic granules
Trichohyalin is a major protein component of these granules, which are morphologic hallmarks of the IRS and medually cells of the hair follicle.
Trichohyalin functions as a keratin-assocaited protein that promotes the lateral alignment and aggregation of parallel buncles of intermediate filaments in inner root sheath cells.
Expression of trichohyalin is closely associated with expression of filaggrin - the major keratohyalin granule protein.
What is the prime function of the IRS?
The prime function of the IRS is to mold the hair within it, which it accomplishes by hardening in advance of the hair.
** The telogen hair folliucle contains no inner root sheath nor inferior segment**
The amino acid ______ occurs in high concentrations in the hair and trichohyalin granules AND has been used as a marker for hair follicle differentiation.
citrulline
Describe the outer root sheath
The outer root sheath is thickest near the epidermis and granually decreases in thickness toward the hair bulb.
In its lower portion (from the isthmus otf the hair follicle downward), the ORS is covered by the inner root sheath. It does not undergo keratinixation , its cell shave a clear, vacuolated cytoplasm (glycogen).
The the middle portion of the hair follicle (isthmus), the ORS is no longer covered by the inner root sheath, and it undergoes trichilemmal keratinization
The the middle portion of the hair follicle (isthmus), the ORS is no longer covered by the inner root sheath, and it undergoes __________.
trichilemmal keratinization
What is the innermost layer of the outer root sheath?
The innermost cell layer of the ORS is a speacial single-cell layer, located just outside the Henle layer, whse pattern of cell idffereniation and keratinization is ultrastructually and immunohistochemically different from that of the min layer of the outer sheath. The cells are disctinctly flattened and lack lamellar bodies.
What are the the two prominent structures that surround the outer root sheath?
- BMZ (‘glassy membrane”)
- fibrous root sheath.
**perifollicular mineralization of the BMZ has been described in health toy poodles and Bedlington terriers***
The dermal hair papilla is continous with the ____ and is covered by a thin continuation of the ____.
dermal connective tissue, basement membrane
The inner root sheath and hair grow from a layer of plump _____ that cover the papilla.
plump nucleated epithelial cells.
These cells reguarly show mitosis and are called the hair matrix.
The morphology of the dermal papilla changes through the hair growth cycle, being ____ in volume in mature anagen and ____ in volume during telogen.
maximal, minimal
This is nostly a result of changes in the amount of extracellular matrix within the papilla. In the anagen hair follucle, dermal papilla volume is proportional to the volume of the hair. Just below the dermal papuilla is a fibroelastic cusion called Aaro-perkins corpuscle.
What structure is just below the dermal papilla is a fibroelastic cushion?
Aaro-perkins corpuscle.
What structure represents multiple corrugations of the inner root sheath that project into the pilar canal immediately below the sebaceous duct opening?
Follicular folds
** Believed to be artifact of fixation and processing, becuase they are not seen in unprocessed sections**
What are two specilized types of tactile hairs that are found in mammalian skin?
Sinus hairs and tylotrich hairs.
Sinus hairs (vibrissae, whiskers) are found on the muzzle, lip, eyelid, face and throat and on the palmar aspect of the carpus of cats.
Sinus hairs are characterized by an endothelium-lined blood sinus interposed between the external root sheath of the follucle and outer connective tissue capsule.
Describe the structure of sinus hairs?
They are characterized by an endothelium-lined blood sinus interposed between the external root sheath of the follucle and an out connective tissue capsule. The sinus is divded into a superior non-trabecular ring sinus and an inferior cavernous sinus.
A cusion like thickening of mesenchyme (sinus pad) projects into the annular sinus. The cavernous sinuses are covered by traeculae containing many nerve fibers.
Pancinian corpuscles are situated close ot the sinus hair follicles. Sinus hairs are thought to functions as slow-adapting mechanoreceptors.
What are Tylotrich hairs?
These hairs are scattered among ordinary body hairs, The hair follicles are larger than surrounding follicles and contain a single stout hair and an annular complex of neurovascular tissue that surround the follicle at the level of the sebaceous glands.
Tyrlotrich hairs are thought to function as rapid-adapting mechanoreceptors. Each tylotrich follicle is associated with a tylotrich pad.
Tylotrich pads are composed of a thickened and distinctive epidermis underlaid by a convex are of fine connective tissue that is highly vascularized and well innervated.
Unmyelinated nerve dibers end as flat plaques in assoacited with merkel cells, which serve as slow-adapting touch receptors.
What is the histologic appearance of the anagen hair follicle?
characterized by a well-developed spindle-shaped dermal papilla capped by the hair matrix to form the follicular bulb.
Hair matrix cells are often heavily melanized and show mitotic activity.
The anagen hair follicle extends into the deep dermis and subcutis.
Anagen has been divided into seven stages of growth.
What is the histologic appearance of the catagen hair follicle?
It is characterized by retraction toward the surface; a thickened, irregular, undulating BMZ; an increased number of apoptotic keratinocytes, a thickened BMZ between the hair matrix and the dermal papulla; a smaller bulb and an ovoid or round dermal papilla.
The catagen follucle epithelium is shortened in length and reduced in volume - shrinks from the position of the har bulb to just below the entry of the sebaceous duct - the volume reduction largely accompanied with apoptosis.
The TGFb pathway is involed in the induction of catagen, and caspases play essential roles in the execution of apoptosis.
What is the best morphologic feature of the catagen hair follicle?
Partial replacement of the inner root sheath by trichelemmal keratinization.
Additionally, melanogensis ceases, the proximal hair shaft is depigmented, the mitotic activity stops.
As the follicular epithelium is lost, the adventitial collagen and neurovasuclar network of the follicle condenses along the ract of the previous anagen follicle. This specialized adventitial struction the _____, is probably requed to prescisely direct the downward growth of the new follicular epithelium as the follicle reenters anagen.
follicular stile
What is the histologic appearance of the telogen hair follicle?
The telogen hair follicle is reduced to about one third of the former length and is characterized by the small dermal papilla that is separated from the matrix cells, no hair bulb, lack of melanin and mitotic activity, and absence of the inner root sheath and presence of a club hair.
Hair follicles show four morphologic patterns of keratinization
- infundibular (like epidermis - basketweave with keratohyaline granules)
- trichilemmal (a serrated, more closely packed eosinophilic keratin with scant or no keratohyaline granules)
- hair matrix or trichogenic - characterized by retention of nuclear outlines (ghost cells)
- inner root sheath and hair shaft medullary (compact and opaque keratin with blue-gray to eosinophilic color and red trichohyaline granules, as in the Henle and Huxley layers.
A hair plucked in anagn shows a larger expanded root that is moist and glistening, often pigmented and ____ at the end. A hair pluacked in telogen showes a ____ with little or no pigement.
square
tapered club root
What cells are present in the hair follicles outer root sheath and have been shown to migrate to the epidermis within 72 hours following ultraviolet radiation.
Langerhan cells; the hair follicle serves as a reservoir to repopulate the epidermis with Langerhan cells. This follicular reservois paradigm is also seen following thermal injury and in vitiligo, in which follicular keratinocytes and melanocytes replenish epidermal cell population.
What type of gland are simple or branched alveolar glands distirbued througout all haired skin in all mammals except whales and porpoises.
Sebaceous (holocrine) glands; these glands usually open through a duct inot the pulary canal in the infundibulum. Sebaceous glands are largest and most numeous near mucocutnaeous junctions, in the interdigital spaces, on the dorsal neck and rumo, the chin, and on the dorsal tail (supracaudal glad).
Sebaceous glands are NOT found in the food pad and nasal planum.
Sebaceous gland lobules are bordered by a BMZ on which ther sit a single layer of deeply basophilic basal cells called?
Reserve cells; these cells become progessively more lipidized and eventually disintegrate to form sebum toward the center of the lobule. The increase in lipid content can be detected by Oil Red staining.
Sebaceous glands express at least two different receptors involed in the uptake of circulating lipids; what are these two receptors?
FATP4 (fatty acid transporter 4) and LDL (low-density lipoprotein) receptor.
***Sebaceous ducts are lined by squamous epithelium
Enzyme histochenical studies have indicated that all mammalian sebaceous glands contain what three enzymes?
succinate acid dehydrogenase, cytochrome oxidase and a few esterases.
Describe epitrichial (apocrine) glands?
Epitrichial sweat glads are generally coiled and saccular or tubular and are distributed throughout all haired skin. They are not present in foodpads or nasal planum. These glands are located below the sebaceous glands, and they usually open through a duct into the pilary canal in the infundibulum, above the sebaceous duct opening.
Epitrichial glands tend to be larger in areas where there are less hair follicles. They are largest and most numerous near mucocutaneous junctions, interdigital spaces and over the dorsal aspect of the neck.
In general, epitrichial glands do not appear to be innervated.
Enzyme shitochemical studies have demonstrated alkaline phosphatase and acid phophatase in epitrichial sweat gland secreotry epithelium.
IHC has shown acini and ducts are positive for cytokeratin and S-100 protein.
Atrichial (merocrine) sweat glads are found on in ______.
footpads
These glands are small and tightly coiled and they are located in the deep dermis and subcutis of the footpads. Secroty coils consist of a single layer of cuboidal ro columnar epithlial cells and as ingle layer of fusiform myoepithelial cells.
Enzyme histochemical studies have demonstrated cytochrome oxidase, succinate and other dehydrogenases, phophorylases and alkaline phosphatase.
These glands are richly supplied with cholinesterase-positive nerves.
Normal dogs with intact pelage have a critical temperature of? If coat is shaved off, it increases to?
14C (57F)
23C (77F)
Describe the circumanal gland
these glands are present at birth and develop from ducts that originate from the inner and outer perianal side of the extneral root sheath of hair follicles. These glands are also found in the skin of the prepuce and dorsal/ventral aspects of the tail.
These glands are positive for cytokeratin.
Tail gland
aka supracaudal gland, preen gland
is an oval shaped area located on the dorsal surface of the tail over the fifth to seventh coccygeal vertebrae, about 5cm distal to the anus.
This gland is consistently present in most dogs histologically; visible in only 5%.
Histologically, the canine tail gland is composed predominately of hepatoid cells that are identical to those in the hepatoid circumanal glands - dependent on testosterone to form the topographic-ethnologic unit.
The tail gland of the cat consists of numeous large sebaceous glands that run the entire length of the dorsal surface of the tail. Excess accumulation of glandular secretion in the area is called ‘stud tail’
Describe arrector pili muscles
These muscles are of msenchymal origin and consist of smooth muslce with intracellular and extracellular vacuoles. They are present in all haired skin and are largest on the dorsal neck and rump.
Arrector pili muscles originate in the superficial dermis and insert approximately perpendicular on the primary hair follicles.
These muslces receive cholinergic innervation and contact in response to epinephrine and norepeinephrine, producing piloerection.
Arrector pili mucles probably function int he termoregulation and in the emptying of sebaceous glands. They are also important in providing socail signals.
Cutaneous blood vessles are generally arranged in three intercommunicating plexuses of arteries and veins. Identify and describe these three plexuses?
deep plexus - is found at the interface of the dermis and subvutous. Branches from this plexus descend into the subcutis and ascend to supple the lower portiosn of the hair follicles and epitrichial sweat glands.
The ascending vessels continue upward to feed the middle plexus, whih lies at the level of the sebaceous glands. The middle plexus give off branches to the affector pili muscles, ascending and descending brances that supply the middle portions of the hair follicles and sebaceous glands, and ascending branches to feed the superficial plexus.
Capillary loops immediately below the epidermis emanate from the superficial plexus and supply the epidermis and upper portion of the hair folicles.
The majority of superficial dermal vessels are ______, which are the most physiologically reactive sements of the microcirculation and also the site where inflammatory cells migrate from the vascular space into tissues and endothelial cells develop gaps the result in increased vascular permeability.
postcapillary venules
Blood vessels endothelial cells are mesenchymal in origin and are characterized by what 4 factors?
- Ultrastucturally by a peripheral basement membrane and intracytoplasmic Weibel-Palade bodies (rod-shaped tubular structures enveloped in a continuous single membrnae)
- By their possession of factor VIII (von willebrane) antigen, plasminogen activators, and protaglandins.
- by CD31 (platelet endothelial cell adhesion molecule
- by being phagocytic; Endothelial cells are strateically located between intravascular and tissue compartments and serve as key regulators of leukocyte trafficking.
Angiogensis is controlled in part by what three factors?
mast cells (histamine), macrophages (TNFa) and TGFB
Pericytes
vary from fusiform to clublike in appeance, are aligned parallel to blood vessels on their dermal side. They are contractile cells containing actin-like and myosin-like filaments, and they are important in regulating capillary flow.
Veil cells
are flat, adventitial, fibroblast-like cells that surround all dermal microvessels. Although their exact function and nature are undetermined, the fact that they stain for factor XIIIa indicated that they are component of the dermal dendrocyte system.
Unlike pericytes, which are an integral component of the vascular wall and enmeshed in the mural basement membrane material, veil cells are entirely external to the wall. The weil cell dermarcates the vessel from the durround dermis. Perivascular mast cells are usually present in the space between the vascular wall and surround veil cells.
______ is an excellent noninvasive technique for meauring cutaneous microcirculation. The technique can be aplied to monitor inflammation caused by various drugs, chemicals and allergens related to blood flow.
Laser doppler flowmetry
Where do cutaneous lymphatics arise from?
Lymphatics arise from capillary networks that lie in the superficial dermis and surround the adnexa.
Vessels arising from these networks drain into a subcutanous lymphatic plexus.
Lymph vessels are not usually seen above the middle dermis in route in histologic preparation of normal skin.
What is the function of lymphatics in the skin?
Lymphatics control the movement of interstitial tissue fluid. The supply, permeation, and removal of tissue fluid are important for proper skin function.
Lymphatics are the drains that take away the debris and excess matter that results from daily wear and tear. They are essential channels for the return of protein and cells from the tissues to the blood stream.
How are lymph vessels distinguished from blood vessels?
- possessing wider and more angular lumina
- having a flatter and more attentuated endothelial cells
- having no pericytes
- containing no blood
The subcutis is about 90% _____ by weight?
triglyceride
What are the functions of the subcutis?
- energy reserve
- thermogenesis and insulation
- protective padding and support
- maintaining surfaec contours
- steroid reservois and as the site of steroid metabolism and estrogen production.
Basal cell keratinocytes express which keratins?
K1, K5, K6 and K14
CERs are sphingoid consisting of sphingoid bases that are amide-linked to what?
fatty acids
In the stratum spinosum CERs are synthesized by the de novo pathway and then converted to _____ and _____. These are then packed into lamellar granules.
glucosylceramides and sphingomyelin synthase
the SG-SC boundaries, glucosylceramides and sphingomyelins are converted back to CERs by _______ and ______ respectively.
B-glucocerebrosidase and sphingomyelinase respectively.
Ceramides in the SC initially are formed as fee extractable CERs. But some are subsequently degraded to protein bound CERs. The free-extractable and protein-bound CERs are cleaved to generates ________ by spingosine kinase. It is though that S1p has anti-inflammatory activity. S1p is irreversibly digested to degradation products by S1p lyase (SGPL1
sphingosine-1-phosphate
the amount of the anti-inflammatory product S1p in the skin and plasma is decreased in dogs with CAD and this change is probably associated with up-regulation of ____ gene transcription.
SGPL1
In response to ______, the keratohyalin granules are degranulated, their content is released to the cytoplasm and profilaggrin is broken down into filaggrin molecules at the SC-SG boundary
Increased intracellular calcium concentration
What is the major role of filaggrin monomers?
- The major role of FLG monomers in the SC is in the aggregation of keratin intermediate filaments into tight bundles, thereby leading to collapse of the cells, which then have a flattened appearance and providing mechanical strength to the cells.
- It is also though that pyrrolidone carboxylic a and urocanic acids, the degraded products of FLG monomers act as natural moisturizing factors in the upper SC.
- FLG appears to play a role in providing mechanical strength to corneocytes and hampering percutaneous entry of foreign materials through the SC.
- It is debatable whether the FLG degradation products (NFF) provide water holding capacity to the SC, and this needs further investigation.
In the stratum spinosum layer, kertinocytes begin to express K1/K10; characterisitic of terminal differentiation. K5 and K14 are still present in the cytoplasm. Additionally, dogs also express which other keratins?
K4, K15,K16
At the interface of the granular and cornified layers. the lamellar granules fuse with what structure at the apical surface of the cell and secrete lipid laden disks into the intercellular space?
plasma membrane
What is the key component of the corneocyte cornified cell envelope?
loricrin
what are the three types of filaments that form the cytoskeleton of all epithelial cells?
microfilaments, microtubules and intermediate filaments
Keratin intermediate filaments
Comprised of keratin proteins assembled in precise fashion. The keratin proteins have a tripartite structure, compsied of a central alpha-helical rod domain, an amino-termal head and a carboxy-termal tail.
The epidermal keratin termal domains exhibit structural motids known as “glucin loops” which are also seen in loricrin and corneodesmosin and are suggested to mediate intermolecular adheson by acting like velcro.
Describe KIF assembly
keratins are though to self-asseble into filaments. The assembly starts with type I and type II keratin molecules spontaneously forming coiled-coil heterodimers. The heterodimers align in antiparallel and staggered pattern to form ouble coiled-coil tetramers.
The tetramers are the building blocks of the kIP and assemble end to end to form protofilaments (2-3 nm diameter) and laterally into protofibrils (two protofilements) and then finally into intermediate filaments (4 protofibrils).
This process relies on lateral interactions between the staggered coiled-coils as well as head-to-head interactions between the same keratin proteins, and keratins are cross-liked by interchain disulfide bonds.
What are the associated proteins that contribute to the organization of the KIFs?
Filaggrin plays an important role in promoting the keratin bundling. Filaggrin is a histadine rich protein.
Profilaggrin, the precursor molecule for filaggrin and primary component of keratohyalin granuesl, consists of multiple filagggrin untis (4 in dogs). Between N and C termal domains.
How are natural moisturizing factors produced?
Profillagrin is cleaved into filaggrin at the transiion from the granular to cornified layers, and filaggrin binds the KIFs, facilitating the flattening of the cornified cells.
Fillagrin is ultimately degraded to a mixture of hygroscopic molecules, such as amino acids, urocaniic acid, pyrrolidone carboxylic acid, lactic acid, cirtrate and sugars.
These procuts of filaggrin degradation are categorized as the natural moisturizing factors = contibute to hydration, pH and barrier function, urocanic acid plays a role in photoprotetion.
The protease caspase-14 is required for filaggrin degradation.
Where does the CE start and what enzyme is required?
The formation of CE starts inside of the granular keratinocyte cell membrnae and involved transglutaminases, which are a family of calcium dependent enzymes that catalyze the formation of N (epsolon) - gamma glutyl-lysine isopeptide bonds.
Increases in extracellyklar calcium levels in the suprabasal layers leads to increased intracelllar calcium levels which, in turn, increases the TG activity. The isopeptide bonds are highly resistant to most common proteolytic enzymes, contributing to the strucutal strength of the stratum corneum.
As the CE formation begins, where are envoplakin, periplakin and involucrin localized?
Envoplakin and periplakin are expressed at the desmosomal plauqes, and involucrin, a highly soluable glutamin-rich protein, locailzied to the cell membrane.
Involucrin forms a scaffold for the attachement of other components fo CE.
TG1 and TG3 catalyze the formation of isopeptide bonds between lysine and glutamine residues cross-linking involucin to most other CE proteins.
Involucrin glytamyl residues and other cornified envelope proteins covalently bind to omega-hydroxycermides.
In the finished CE structure, involucrin is localized to the outer protion of the CE scaffold and connects the corneocytes to the extracellular lipid layer.
After locicrin is released from keratohyalin granules, where does it accumulate?
Loricrin, a cystein rich highly flexible protein WITH glycine loops (velcro), is the MAJOR structural component of the CE. It comprises over 70% of its mass, and is gound primarily in the inner portions of the CE.
After being released from keratohyalin granules, Loricrin accumulates at the desmosomes.
TG3 cross-links loricrin and small proline rich peptides together, and TH1 cross-links those oligmers ot the developing CE.
Multiple other proteins including filaggrin, cystatin A, cornifelin, SPRs, elafin, S100 proteins, late envelope proteins, demoplakin, and keratins, are also crosslinked to the developing scaffold to become part of the final CE.
What are the remaining internal structures of the keratinocyte once the CE firmation progresses and the organelles of the keratinocyte are degraded?
The KIFs are covalently linked to several CE proteins, including lorcrin, envoplakin and involucrin, through lysine residues on teh head domain of type II kerain chains. The CE eventually replaces the plasma membrnae. The mature corneocyte is flattened cells.
KIF+Fillagrin (inside cell)
Loricrin (inside)/Involucrin (outside)
Where do lamellar bodies originate from?
In the upper spinous and granular layers, lamellar granules (aka lamellar bodies) appear from within the Golgi apparatus. These granules contain phopholipids, glucosylceramides, sphinogmyelin and cholesterol.
Numerous enzymes and their inhibitors are also released from the lamellar bodies. These inlcude lipid enzymes and their inhibitors are also released from lamellar bodies which incude lipid hydrolases (b-glucocerebrosidase, acidic sphinomyelinase and secretory PLAs, and serine and cystein proteeases, kallikrins and cathepsins)
Lamellar garanules also contain cornedesmosin, steroid sulfatase and AMPs such as B-defensin 2 and cathelicidin LL-37.
Trafficking of lamellar granules from the trans-golgi network to the plasma membrane is facilitated by what complex?
CHEVI (C-homologues in Endosome-Vesicle Interaction Complex)
What complex drives the fusion of the vesicle membrane with the plasma membrane?
THe CHEVI complex is through to interact with the SNARE protein complex which drives the fusion of the two membranes.
What is the primary signal for inducing lamellar granule secretion?
The increase in extracellular calcium concentration in the granular layer appers to be the primary signal for inducing lamellar granule secretion. The secreted lipds are arranged into intercellular lamellae parallel to the cell sugace and metabolzied by the secreted enzymes from polar to nonpolar products.
Glucosylceramides are converted into cermides (b-glucocerebrosidase)
Spingomyelin is converted into ceramides (acidic spingomyelinase)
Phospholipids are converted into FFA and glycorol (phoholipases). The free fatty acids contributed to the acidication of the SC which is important for functioning of many enzymes.
Glycerol is hygroscopic and helps to keep the SC hydrated.
What are the major lipid classes in the SC?
ceramides, free fatty acids and cholesterol
Ceramides are categoized into how many different structural groups?
11; cermides are amide-linked fatty acids attached to spingoid bases, which are long-chain amino alcohols. Ceramides are categozied into structural groups based on the shpingoid base and the types of attached fatty acids.
The shingoid bases include phytoshingosine (P), singosine (S), 6 0 hydroxylspingosine (H), and dihydrospinogsine (D), a-hydroxy fatty acids (A) and nonhydroxy fatty acids (N).
The stuctural group yields a ceramide group name such as EOS representing a shingosine base with an attached esterified -omega-hydroxy fatty acids.
What are the three omega-hydroxyceramides that are unique to the stratum corneum and carry very long chain amide-linked fatty acids?
EOS (ceramide 1)
EOH (cermide 2)
EOP (cermide 9)
CLE (corneocyte lipid envelope)
The omega cermaides are covalently bonded to involucrin primarily as well as envoplakin and periplakin in the CE and interdigitate with the intercellular lipid lamellae. This layer of omega-hydroxycermides bonded to the CE is called the CLE (corneocyte lipid envelope).
The CLE forms a key linkin the stratum corneum barrier, and malformations in the CLE result in ichthyosis.
The omega-hydroxyceramides are known to play an important role in epidermal barrier function.
What is the key rate limiting enzyme in free fatty acid synthesis pathways?
acetyl-CoA carboxylase and fatty acid synthase
What is the role of fatty acids in the epidermis?
Fatty acids are long carbon chains with methyl group at one end. ,the epidermis contains free fatty acids as well as fatty acids bound to triglyceridesm phospholipids, glycosylceramides and cermides.
Epidermal free fatty acids range in chain length from C12 to C24
The epidermis synthesizes saturated and monounsaturated FA but not omega-6 polyunsaturated essential fatty acids (EFAs) such as linoleic acid and alpha-lineoleni acid, which must be supplied by diet.
Linoleic acid is contained in phopholipids glycosylcermides, ceramide 1, ceramide 4 and cermide 9.
Cholesterol is synthesized from ____ in the lower epidermis in a pathway in which ____ reductase is the rate limiting enzyme.
acetate
hydroxymethylglutaryl CoA (HMG CoA) reducatse is the rate limiting enzyme.
Some cholesterol is incorporated into the lamellar granules and secreted to the granule-cornified layer interface uncahgned. Some of the cholesterol is fulgurylated to cholesterol sulfate.
Cholesterol sulfate plays a role in inhibiting ____ proteases involed in desqumation.
serine
Desmosomes are composed of mulitple proteins and are divided into several regions based upon their electron micrographi appearance. Those three regions are?
the desmoglea - extracellular portion between adhered keratinocytes
outer dense plaque - within keratinocyte PM (bordering the membrane)
inner dense plaque - within keratinocyte PM (intertwined with KF)
The molecular components of desmosomes fall into what three major gene families and consist of multiple proteins?
Cadherins: transmembrane molecules that form a Ca-dependent adhesive interface between neighboring keratinocytes. Their extracellular portions comprise the desmoglea (desmogelin, desmocollin)
Armadillo protein damily: at the desmosomes, these proteins bind to the cytoplasmic protions of the cadherins.
- Plakoglobin - binds to desmoplakin and cadherins
- Plakophilin - binds to cadherin complexes to strength the plaque
Plakin protein family
- Desmoplakin - binds KIFs to plakoglobin
- Envoplakin and periplakin
What are corneodesmosomes?
At the transition from the granular to the cornidied layer, the morphology of the desmosomes changes as they are integrated into the CE. The cornified layer, these sites of keratinocyte cohesion are known as corneodesmosomes, and they play a key role in SC barrier function.
What are the three moelcular components of corneodesmosomes?
desmoglein1, desmocollin1 and corneodesmosin (CDSN)
What is the function of CDSN?
It is a glycoprotein with high serine and glycine content particularly at its termal regions. Like keratins and loricrin, CDSN forms strutural motids called ‘glycine loops’ which mediated reversible and adjustable intermoelcular adhesion by acting like Velcro.
CDSN is synthesized by granular keratinocytes and secreted from lamellar granules into the itnercellular space starting in the second layer of the SG. CDSN binds ot the desmoglea shortly before the transformation of desmosomes to corneodesmosomes.
What are tight junctions?
aka zonulae occludens, are cell to cell contacts that leave virtually no intercellular space betwen neighbnoring cells. Tight junctions provide a semipermeable size and ion specific ‘gste’ barrier that regulates the diffusion of ions and solutes through the interceullular spaces in simple and stratified epithelia = paracellular pathway
Tight junctions also perform a ‘fence function’ to maintain cell polarity, demarcating the apical and basolateral cell membrane regions, by restricting the diffusion of apical and basolateral cell membrane molecules within the cell membrane.
Epidermal tight junctions are composed of what two families of tranmembrane molecules?
Claudins and occludins (and intracellular zonula occludens (ZO) proteins)
Claudins 1, 4, and 7 are expressed in the epidermis.
What are junctional adhesion molecules (JAMs)
JAMS are an IgG-like family of transmembrane receptors that interact with proteins of the tight junctions and adherens junction to regular barrier function, cell migration and cell proliferation.
Many of the components of moelcules of tight juntions are present in the non-cornified epidermal layers, but functional tight juntions are located ONLY in what layer?
Stratum Granulosum
In the stratum corneum, tight junction-related structures surround corneodesmosomes at the lateral esges of corneocytes preventing the acces of proteases involved in cornodesmosome degradation.
What are adherens junctions?
These are found on the lateral and apical membranes of basal cells and on the entire cell surface of suprabasal cells.
Adherens junctions are cadhein-based junctions that libnk the actin cytoskeleton to the plasma membrane.
Transmembrane cadherins, E-cedherin, form the physical linkages between E-cadherin molecules on adjacent cells. They initiatio and maintain cell-cell adhesion, regulate the organization of the actin cytoskeleton, and establish a hub for cell signaling and regulatation of gene transcription.
Adherens junctions are also involved in regulating keratinocyte proliferation and differentiation and play a role in wound healing.
What are adheren junctions composed of?
AJ are composed of a cadherin-catenin complex. The cytoplasmic tail of E-cadherin binds to the cytoplasmic catenins, B-catenin and p120 catenin. B-catenin binds a-catenin, and a-catenin link the F-actin cytoskeleton to the junction
There appears to be a dynamic relationship between adherens junctions and the intracellular actin cytoskeleton. The extracellular cadherin domains bind a cahderin on the opposing cell surface in calcium dependent manner.
What enzymes are involved in desquamation?
The enzymes include kallireins (serine proteases), and cathepsins (cystein proteases).
KLKs are synthesizes as proenzymes and activated by themselves or other proteases in a complex ph-dependent proteolytic network. As ph decreases, inhibitors are less effective.
KLK 1,5,6,7 and 14 are involved in the degradation of corneodesmosomes.
Multiple enzyme inhibitor regulate this enzymatic degradation, including the KLK inhibitor - LEKTI (lymphepithelial Kazal-type-related inhibtor), SPINK6, elafin, serpins adn cholesterol sulfate.
What is the cysteine protease inhibitor is localized on cornodesmosomes following secretion by lamellar granules and inhibits cathepsin V (cleaves corneodesmosomes)
Cystatin M/E
What is the role of Ca gradient in keratinocyte differentiation?
Calcium ions and their concentration in the epidermis play essemtial roles in regulating keratinocyte differentiation, skin barrier formation and pearmability homeostasis. Ca concentrations are low in the basal and spinous layers with progressively increasing levels towards the stratum granuloms (stored in ER) and decreasing levels again in the stratum corneum
Tight junctions and the stratum corneum are both through to the crucial to generate and maintain gradeitn by preventing Ca diffusion from the SG to SC.
Ca regulate the transcription of all genes encoding keratinocyte differentiatrion-specific proteins, adn the gradient relects the relative concentration of Ca required for state specific protein expression in each layer.
The high level of Ca in teh SG regulates which important processes for cornification?
The high levels of calcium in the granulaer layers regulates lamellar granule secretion, TG activity and the cleavage of profillagrin to filaggrin.
Where is the intracellular calcium stored to maintain gradient?
Calcium is stored in the endoplasmic reticulum. Keraintocytes express several types of calcium channels which permit Ca influx into keratinocytes from extracellular sources.
In the granular layer, the calcium-sensing receptor, a subfamily of G protein coupled receptors in the keratiocyte plasma membrane, senses the rise in extracellular Ca and activates the intraceullar signals that lead to the release of Ca from intraceullar ER stores and stimulates Ca infux through calcium channles. The results is an increase in extraceullar Ca which promotes differentiation fro granulate keratinocytes to corneocytes.
In the non-cornified epidermal layers, what cells procide a barrier to water loss from the interior?
tight junctions
The plasma membrane of basal keratinocytes provides localization and organizational cues largely due to what structures?
Integrins
B4 binds to collagen XVII (BP180 or BPAG2) via fibronectin domains ___ and ___. It also binds to plectin and BPAG1 (BP230) via fibronectin domains __ and __.
Fibronectin domains III and IV
Fibroectin domains I and II
Laminin 332 is one of the major epidermal laminins. It is Initially synthesized, assebled and secreted by kerainocytes in precursor form and processed by what extracellular enzyme?
Tolloid
Laminin 511
Produced by dermal microvascular endothelial cells
Abundant in the interfollicular epidermis and dermal blood cells
Contains multiple globular chains that bind to nidogen (collagen IV)
Laminin 311
Consists of a3, B1 and y1 chains
Regulate cellular functions with laminin 332
Share binding domain in a3 chain
What are the two isoforms of nidogen?
Two isoforms:
Nidogen -1: binds perlecan and collagen IV
Nidogen-2: binds collagen IV and I and perlecan, NOT filbulin
Fibulin
Able to bridge carious BMZ molecules (fibronectin, collagen IV, perlecan, and nidogen 1)
Thought to stabilzie moelcular organization of extracellular membrane structures
Where are retinal pigments cells derived from?
Optic cup of the forebrain
What keratins are expressed throughout all layers and in hyper proliferative disorders?
K6/K16
What are the keratins of the ‘upper spinous layer’?
K2/K11
Which fatty acids are keratolytic and fungistatic?
caprylic, propionic and undecylenic acid
What two anti-inflammatory compounds can be synthesized from linoleum acid and linolenic acid?
DGLA and EPA
**DGLA is the elongation product of γ-linolenic acid. Elongases add additional carbon molecules to existing chain.
Delta-desaturase deficiency in atopic humans cannot convert DGLA, linoleic and linolenic acid into what substance?
Is there is a delta-6-deficiency - Linoleic, linolenic and DGLA accumulate locally, it cannot be metabolized into arachidonic acid. DGLA completes with AA for cyclooxygense and lipoixygenase enzymes. This competitive inhibition it thought to be the mechanism of anti-inflammatory action of fatty acid therapy -which generally involves the modification of leukotriene and prostaglandin synthesis.
What are the effects of prostaglandins on the skin?
Alteration of vascular permeability
Potentiation of vasoactive substance such as histamine
Modulation of lymphocyte function
Potentiation of pain and pruritus
**Prostaglandins and Leukotrienes potentiate each other
What are the effects of Leukotriene on the skin?
Alter vascular permeability
Activate neutrophils
Modify lymphocyte function
Cause potent neutrophil and eosinophil chemotaxis
y-Linoleic acid is found in relatively high concentrations in primrose, borage and black current oils. It is elongated TO DGLA, which directs completes with AA as a substrate for cyclooxyrgenase and 15-lipoxygenase. The result of DGLA metabolism is the formation of what two products?
Prostaglandin E and 15-hydroxy-8,11,13-ETA
***EPA is supplied by using old water marine oils and competes as a substrate for cyclooxygenase and lipoxygenase.
EPA can also be supplied by the conversion of a-linoLENic acid into EPA and what other fatty acid?
DHA
An excellent source if linoLENic acid is chia sed and flaxseed
Metabolism of EPA by lipoxygenase enzymes results in the formation of what two products?
LTB5 and 15-hydroxyEPA.
These two products are thought to inhibit LTB4 - which is a potent pro-inflammatory mediator.
THe mechanism of how EFAs function I controlling pruritus includes inhibition of _____ and metabolism o metabolic byproducts of ____ metabolism
AA
Fatty Acid
List hormones 5 hormones that stimulate hair cycle
List 6 hormones that inhibit hair cycle
Stimulators:
Thyroid homrone
Corticotropin
Melatonin
Androgens
Growth Hormone
Inhibitors:
Cortisol
Estrogen
CRH
Corticotripin
Prolactin
Parathyroid-related hormone
List 10 Growth Factors that stimulate hair cycle
FGF-7
HGF
Insulin-like growth factor
Sonic Hedge Hog
Keratinocyte growth Factor
Wnt
B-catenin
TGF-a
Nerve growth factor
GDNF
List 9 Growth Factors that inhibit hair cycle
Epidermal Growth Factor
FGF2
FGF5
Brain-derived neurotrophin
Neutrotropin 3
Neurotropin 4
TGFa
TGFb1
TGFb2
Which cytokine stimulates hair cycle?
Substance P
Which cytokines inhibits hair cycle?
IL-1 and IL-6
Do retinoids and calcitriols stimulate or inhibit hair cycle?
Inhibit!
Acivin, Noggin, Folistatin, Cyclosporine, Monoxidil and Finasteride stimulate
What are the 3 trophic effects of cutaneous nerves on follicular growth?
Regulate Vascular Tone
Neuropeptide stimulating of receptors on follicular keratinocytes and dermal papilla
modulating of macrophages and mast cell activity
What cells of the hair bulb consists of mesenchymal cells and is considered to play a fundamental role in epithelial differentiation?
Dermal papilla cells
These are then PRIMARY target cells of the hair follicle that respond to hormones and mediate growth-stimulating signals to the follicular epithelial cells.
White cats with blue eyes often have what abnormality?
Cochlear degeneration and deafness
What gene in cats results with the gradual replacement of eumelanin with pheomelanin seen in Norweigain Forest Cats?
Amber gene
What gene controls the acromelanism enzyme that converts melanin precursors into melanin by a process of oxidation?
C, tyrosinase gene, TYR
Tuxedo cats white spotting is controlled by what gene?
white gloves gene (KIT)
Hemidesmosomes and ____ are located on the basal surface of basal cells and mediate adhesion to the underlying extracellular matrix whereas desmosomes and ______ mediate adhesion between keratinocytes in all epidermal layers.
focal adhesions
adherens junctions
P-Cadherin is present in which layer of the epidermis?
Basal
What are the transmembrane proteins for focal adhering versus adherent junctions?
Focal adherens - B1 integrins
Adherens junctions - P and E Cadherins
What are the plaque proteins associated with focal adhesions versus adherens junctions?
focal adhesions - Talin, Vinculin, a-actinin, paxillin and zykin
adherens junctions - plakoglobin-a and b-catenin; alpha- actin; vinculin
Profilaggrin is cleaved by several enzymes when it is converted to the functional fillagrin protein. The profilaggrin linker domain is cleaved by ______. Fillagrin is further degraded into natural moisturizing factors by what enzyme?
kallikrein-related peptidase 5KLK5
Caspase 14
What locations does the CE develop beneath the plasma membrane?
stratified epidermal cells
cells of the inner root sheath
medulla of the hair follicle
cuticle of the claw
What protein binds ceramides covalently to create the backbone for the attachment of intracellular lipids?
Involucrin
Transglutaminases are chiefly expressed in what layers of the epidermis?
Stratum granulosum
Stratum spinosum (upper)
Ceramides 1 and 2 bind to what protein in the CE, forming a scaffold for the binding of other lipids into an intercellular lipid bilayer parallel to corneocytes in the SC?
Involucrin
Where are melanocytes found in the hair follicle?
Outer root sheath
What are the 3 catalytic activities of tyrosinase?
Converting tyrosine to Dopa (rate limiting step)
Can use Dopa or 5,6-dihydroxyindole asa substrates for oxidase activities.
Once Dopa is formed, it can spontaneously autooxidize to _______ without tyrosinase (although at a slower rate) and continue through the melanin pathway to dopa chrome, 5,6 dihydroxyindole-2-carbxylic acid, DHI and indole-5,6-quinone
dopaquinone
What are three agonists of MC1R?
a-MSH
Corticotropin
Agouti gene (antagonist)
What is the major determinant of melanogenesis that regulates pigment phenotype?
melanocortin-1-receptor
Loss of what receptors expression is associated with progression of melanocytes toward metastatic melanoma phenotypes?
c-kit - a tyrosine kinase receptor involved in melanoblast expansion, survival and migration.
**mutations are involved in piebaldism
What is a neuroimmunomodulatory and anti-inflammatory peptide that is synthesized and released by keratinocytes, Langerhan cells, fibroblasts, endothelial cells and melanocytes?
a-MSH
In dogs, dermal dendrocytes are positive for what markers?
CD1, Factor XIIIa, MCH II and Thy-1 positive (CD90)
Mast cells are present where in the dermis and near what structures?
Mast cells are most abundant around superficial derma blood vessel and appendages.
Toludine blue and acid orcein Giemsa will help more easily recognize these cells.
The inner root sheath keratinizing and disintegrates when it reaches what layer of the hair follicle?
Isthmus
Atrichial sweating from the footpads of cats and dogs can be provoked by what stimuli? and less by what other stimuli?
Cholinergic
Adrenergic
**Atropine blocks glandular responses to these agents
What muslces receive cholinergic innervation and contact in response to epinephrine and norepeinephrine, producing piloerection.
Arrector pili muscles
The_________ are a gene family critical for normal embryologic development; they likely play a very important role in development of skin adenexa, pigment system, and stratified epithelium during embryogenesis.
Homeobox (hox) genes
The arterioles consist of endothelial cells surrounded by two layers of smooth muscle cells, whereas arterial and venous capillaries lack what layer?
smooth muscle cells
Selectins and members of the ________(ICAM1; vascular cell adhesion molecule, VCAM1) are upregualted on endothelial cels following inflammatory stimulation and mediate rolling, adhesion and transmigration of leukocytes from the blood stream.
immunoglobulin SUPERFAMILY
Angiogenesis is controlled by what 3 things?
mast cells (histamine), TNFa and TGFb
What is a signalling protein that promotes the growth of new blood vessels. It is part of the mechanism that restores the blood supply to cells and tissues when they are deprived of oxygenated blood due to compromised blood circulation.
What is Vascular endothelial growth factor (VEGF)
Arteriovenous anastomoses are normal connection between arteries and veins that allow arterial blood to enter the venous circulation without passing through what?
capillaries
Most common over extremities (legs and ears)
A “glomus” is a shunt in the deep dermis - the partial segment is called “sucquet-hoyer canal” from the arteriole.
Glomus cells are generally considered modified smooth muscle cells.
The motor innervation of skin is attributable to sympathetic fibers of the _____ nervous system.
autonomic
What do the somatic sensory system mediate?
touch, head, cold, pressure, vibration proprioception, pain and pruritus
What to the nerves of the autonomic motor system control?
They control cutaneous vascular resistance, pilomotor responses, regulate the secretory activities of glands.
Although the cutaneous nerve trunks are considered somatic sensory nerves, they carry what types of fibers?
myelinated post-ganglionic sympathetic fibers
What are hairless, knob-like innervated structure that are precent in the haired skin of cats and dogs.
tylotrich pads
