Cellular Patholgy Flashcards
What are the 4 types of primary tissue?
- Connective
- Epithelial
- Muscle
- Nervous
What is histology?
Study of tissues of the body an how these tissues are arranged to constitute organs
Involves all aspects of tissue biology to correlate the tissue structure with its function
What is a tissue?
Tissues are groups of similar cells that come together to perform a common function
What is cytology?
Involves looking at individual cells/ clusters of cells
What are the 6 steps of tissue preparation?
- Fixation
- Dehydration
- Clearing
- Embedding
- Sectioning
- Staining
Describe fixation
Critical step in preservation of sections
Tissues are pressed as close to the natural state as possible for examination
Describe dehydration
Series of alcohol solutions of ascending concentrations
Describe clearing
Clear the tissue to allow paraffin wax to embed into the tissue
Describe embedding
For light microscopes- paraffin
Electron- epoxy resin
Describe sectioning
Light- 5-15um thick
Electron- ultra thin section using a diamond knife (0.05-0.09um)
What is H&E staining?
Haematoxylin and Eosin
Haemotoxylin to react with anionic components
Eosin to react with cat ionic components
What is PAS staining?
Periodic Acid Schaffer
For CHO’S and carb rich molecules
Stains them magenta/ pink
Why does Mallory’s Trichrome stain do?
Stains collagen blue
What does Masson’s Trichrome stain do?
Differentiates smooth muscle (stains red) from tissue collagen
(blu/green)
What is the basis of immunohistochemistry?
IHC is the specificity of a reaction between an antigen and antibody
How does autoradiography work?
- Radioactive compound binds to the tissues
- Tissue is placed next to film
3.reaction produced by radioactivity is quantified and localised
What is the epithelium?
Structure that lines and covers all pars of the body and separate the organs in the body, it has no direct supply and is supported by connective tissue
What’s are the 2 types of epithelium?
Proper-covers the whole body and organs
Glandular- secretes hormones, digestive juices and sweat
List the function of the epithelial tissues
- Protection
- Sensory
- Secretions
- Absorption
- Excretion
How are the different types of epithelial tissue classified?
- Cell shape at surface
- No of cell layers
- Transitional epithelium
- Pseudostratified epithelium
What are the 4 types of epithelia?
Squamous
Cuboidal
Columnar
Pseudostratified
Describe simple squamous epithelium and their function
One layer of smooth surface flat cells attached to the basement membrane where cell width is larger than the height
Function- Exchange barrier and lubrication
What is the typical location of simple squamous epithelium?
Linguini of the heart
Body cavities
Bowman’s capsule
Describe stratified squamous epithelium and their function
Several layers of cells (deepest layer of cells only), squamous being the superficial layer and they can be keratinised/ non-keratinised
Function- Protection and barrier to dehydration + infection
Where can stratified squamous cells be found?
Esophogous
Vgaina
Epidermis (keratinised)
Describe simple cuboidal epithelium and their function
One layer of cuboidal cells in which the cell height= width= depth attached to the basement membrane
Nucleic are typically spherical and apical surfaces may have microvilli
Function- absorption and secretion barrier
Where are simple cuboidal epithelia found?
Surface of the ovaries
Kidney tubulars
Thyroid follicles
Describe stratified cuboidal epithelia and their function
2-3 layers of cells where the basal layer is attached to the membrane
Function- barrier and conduit (channel)
Where are stratified cuboidal epithelia generally found?
Sweat glands and ducts
Exocrine glands
Anorectal junction
Describe simple columnar epithelium and their function
Layer of columns cell attatched to absent of membrane, where the cell height is larger than the width
The nuclei are typically oval and the surface my be specialised depending on location
Function- Absorption and secretion
Where are simple columnar epithelial found?
Stomach
Intestines
Gallbladder
Describe stratified columnar cells and their function
2/3 layers of cells in the superficial layers
Not a common type of epithelium
Function- barrier and conduit (channel)
Where are stratified columnar epithelial cells found?
Exocrine glands
Anorectal junction
Describe transitional epithelia and their function
Staratifed epithelium that line the urinary tract
Surface cells are often dome shaped (aka umbrella cells)
Function- Barrier and dispensable property
Where are transitional epithelium cells found?
Renal calyces
Ureter
Bladder
Describe psuedostratifed epithelium and their function
Layer of cells that vary in shape, size and height
Most are columnar but some are short basal
Pseudo= false
Functions- Secretion, conduit and absorption
Where are psuedostratified cells found?
Trachea
Bronchi
Male reproductive tract
How are glandular epithelium classified?
Exocrine- Secrete ONTO a surface: Merocrine, Aprocrine and holocrine
Endocrine- Secrete products INTO connective tissues that can then enter the bloodstream: Paracrine and Autocrine
What is the classification of exocrine glands based on?
- Number of cells- uni/ multi cellular
- Type of secretion- serous, mucus or mixed
How are multicellular cells sub-classified?
- Shape of secretary cells- tubular, alveolar, tube alveolar
- Presence/ absence of branching duct- simple= unbranched, compound= branched duct
Describe serous gland secretion
Watery, nuclei within the cells are rounded or oval in shape
Peri nuclear cytoplasm is basophilic due to extensive rough endoplasmic reticulum
Describe mucous gland secretion
Viscous / slimy
The nuclei within the cells are flat and compressed to the base of the cell
PAS +ve due to anionic oligosaccharides
Describe mixed gland secretion
Contain both mucus and serous secretory components
E.g Submandibular and Tubuloacinar gland
Describe the process of H&E staining?
- Take the section to water
- Flood with Haemotoxylin
- Wash the slides in water
- Remove excess dye by adding 1% alcohol then wash in water again
- “Blue” in ammonium water then wash in water again
- Cover in Eosin for 2-3 mins then rinse in water
- Dehydrate section
- Clear in ‘Cirtroclear’ and mount on coverslip in DPX
Describe the process of taking a section to water
- Stand slide in Citrocear for 5 mins
- Remove and rinse in absolute alcohol for 1 min
- Rinse in 95% alcohol for 1 min
- Rinse in 70% alcohol for 1 min
- Wash the slide in water
- Proceed to staining
Describe the process of dehydrating a section
- Wash the slide in water
- Rinse by dipping in 70% alcohol x5
- Rinse by dipping in 95% alcohol x5
- Rinse by dipping in 100% alcohol x5
- When ready to mount on slide immerse in Citroclear
What are the aims in preparing a tissue for microscopy?
- Identification of abnormalities
- Confirm clinical suspicions
- Cornmeal other lab data
List the different types of tissue preperation
Smear/ Imprints- Exfoliated cells
Histology- Touch or squash prep/ slice and H&E stain
Microbiology- Colony samples and Gram staining
Biochemistry- Centrifuge deposits + crystal films
Haematology- Blood smears, Leishman stain sections
What are the main causes of damage of specimens and what can be done to prevent it?
- Metabolic change
- Osmotic swelling/ shrinkage
- Evaporation
- Putrefaction- bacteria mould
- Autopsies
Prevention= Preservative
What is the effect of fixation on protein?
Stabilisation of proteins
What are the 2 different types of fixative?
Coagulant- removal of water leading to coagulation and denaturation of proteins (visible change)
Cross linking- form chemical bonds between proteins in tissue (no visible change)
List the 9 factors involved in fixation
- Buffers
- Temp
- Penetration
- Volume changes
- Osmolality
- Substances added
- Detergents
- Concentration
- Duration - rate of penetration
What is secondary fixation?
Use of secondary agent
What effect does formaldehyde have on cells?
Non coagulating
Adds glycol groups to proteins cross linking hem to form methylene bridges
Causes slight swelling, keeps tissue soft
Retains natural colour
Potential carcinogen
What effect does glutaraldehyde have on cells?
Binds to protein amino groups crosslinking them (like formalin)
Non coagulating.
Quick to reach complete fixation – 1-2 hours
Widely used in electron microscopy
Inactivates all enzymes and so can’t be used for enzyme histochemistry.
Depresses staining with acid dyes
Highly toxic
What effect does osmium tetroxide have on cells?
Powerful oxidising agent
Stains lipids black – used to stain membranes
Used for small tissue samples (e.g. electron microscopy)
Expensive (£180 000 per Kg).
Highly dangerous – can cause blindness.
List the reagent that could be used for embedding media
Wax
Gelatin
Agar
Cellodin
Resin
List 3 agents that could be used to freeze agents
CO2 gas
Dry ice
Liquid gas Nitrogen
Why do ice crystals form?
Caused by slow cooling
How is microwave tech used in cell pathology?
Used in fixation, processing, decertification, staining and immunochemistry
Speeds up all areas by providing a faster response
What are the aims of staining?
Visualise cell structure
To see variation in structure
See changes in chemical structure due to disease
What are the 5 steps/ principles of staining?
A. Direct attachment of dye to tissue
B. Indirect attatchment of dye to tissue- Using a mordant
C. Differentiations- using mordants, acids or oxidising agents
D. Trapping agents- prevent yeast escaping
E. Differential action of dye- different structures identified using different dyes
Which cell component reacts with Haemotoxylin?
Nucleus
-ve charges of Nucleic acids bind with basophilic dye
Which cell components react with Eosin?
RBC’s, cytoplasm, muscle collagen etc
All of the above are basic so bind with acidophilllic eosin
What are the principles of H&E staining?
Chemical attraction between the tissue and the dye
Charges on dye/ tissue are attracted to each other via Van der Waal’s forces, H- bonding etc
Tissue charges are found in protein, lipids etc
List 3 basic yes and their colour
Methyl green- Green
Methylene blue- blue
Toluidine blue- blue
List 3 acidic dyes an their colour
Acid fuschsin- Red
Aniliiine blue- blue
Eosin- Red
Describe a Trichrome stain
Similar to H&E
Stronger Haemotoxylin is used
3 acid dyes are used to replace eosin: picric acid, acid fuschsin and methyl blue
Above bind to different densities of tissue
What is PAS staining?
Periodic acid- Schniff
Histochemical method based on:
1. Oxidation of periodic acid
2. Schniff reagent- colourless
3. Aldehydes at site of reaction restores chromophore= purple magenta
Used for CHO’s and CHO-rich molecules
What stains are used on a liver biopsy and why?
PAS/ D-PAS- to look for glycogen disorders
Perls iron- For iron deficiency, excess iron and heamochromatosis= over consumption by the gut
How are specimens analyse using TEM?
Examination of sections on grids, “stained” by heavy metals (lead, uranyl salts)
How are specimens analyse using SEM?
Examination of solid particulate material showing topographical
Details via Freeze fracturing, Freeze etching, Replication (metal shadowing) (viruses, proteins)
List some factors to consider when staining tissue
Shrinkage due to paraffin heating leaves gaps
Looking at 2D image of 3D strucuture
Consider the plane of sectioning
How does the epithelium demonstrate polarity?
They cover body surfaces, lining cavities and forming glands
Polarity is related to function and morphology
List the apical surface specialisations of cells
Cillia- assist in movement of particles
Microvilli- Inc cel surface area to facilitate absorption + sec
Steriocilia- Inc apical surface area of cell
Keratinized and Nonkeratinized- layer of dad cell fillled with keratin forming protective layer
How are cilia modified to carry out their function?
Elongate, motile structures, core of microtubules
5-10 um in length
Two central microtubules are surrounded by by 9 peripheral pairs
How are microvilli modified to carry out their function?
Small finger- like projections
1-3um in length
Ac as a brush border
Composed of actin micro filaments
How are sereocilia modified to carry out their function?
Elongated microvillli to help in absorption
Up to 120 um in length
Core= actin micro filament
List the lateral specialisations of cells
- Occluding junctions- Tight
- Anchoring junctions- Adhering and Desmosomes
- Communicating- Gap
- Morphologic- Lateral cell surface folds
Describe occluding junctions and their function
Separate luminal space and connective tissue compartment
Seals intercellular space
2 transport pathways:
Paracellular and Trans cellular
Permeability of junction depends on the strands-numbers, complexity and presence of aqueous channels
What are adhering anchoring junctions?
Also act as a belt, but allow some space in between cells
Cadaherin attaches to actin filament of cytoskeleton
What are desmosomes anchoring junctions?
Act as intermediate filaments between cells
Made of protein
What are gap junctions?
Membrane of adjacent cells
2 half channels= 2 connexons
Each connexon is made up of x6 connexin proteins
Describe the morphological specialisations of the ell surface
The infoldings of cytoplasmic processed of adjoining cells
They inc lateral cell surface area, which gives cells an inc chance to carry out absorption
What are the 2 basal membrane cell junctions specialisations?
- Cel-to-extracellular matrix junctions- focal adhesion and hemidesmosomes
- Basal cell membrane in foldings
What is focal adhesion?
An anchoring complex- links cytoskeleton actin filament to extracellular matrix proteins in the basement membrane
Plays a role in signal transduction
What are Hemidesmosomes?
Half the component of a desmosome
Found in epithelia subject to abrasion and mechanical shearing forces
Anchoring complex:
Cell to Extracellular Matrix Junctions epithelium attachment to basement membrane
Anchor Epithelial cell to basement membrane
Describe the basal membrane infoldings
Inc surface area of basal domain
Allow for more transport proteins and channels to be present
Well developed in cells performing active transport
What is connective tissue?
Underlies/ supports other 3 basic tissues structurally and functionally
What are the 4 functions of connective tissue?
- Support
- Defense
- Repair
- Nutrition
Describe fibroblasts
Most common cell in connective tissue
Synthesise fibres and ground substance
Spindle- shaped, can be active or inactive
Myofibroblasts= wound healing
How is connective tissue classified?
Embryonic- mesenchyme and mucous
Proper- loose and dense
Specialised- cartilage bone etc
What is mesenchymal embryonic tissue?
Part of 3 embryonic germ layers (mesoderm)
Occupies space between developing organ and mucous connective tissue in the umbilical cord cord
What is mucous connective tissue?
It is “jelly”-like, few cells or fibers - mainly ground substance
Few fibers and/ or cells
Abundant ground substance
Describe loose connective tissue
Fill spaces between other tissues, loosely thin and relatively sparse collagen fibers
Abundant cells of various types
More cells than fibres
Abundant ground substance
Flexible, rich blood supply, not resistant to stress
Located beneath the epithelia surfaces
Surrounds glands and the smallest blood vessels
Describe dense irregular connective tissue
Abundant fibers, mostly collagen fibers, irregularly arranged
Few cells of single type, mostly fibroblasts
Little ground substance
More fibers than cells
Can withstand tension exerted in all directions
Provide structural strength
Describe dense regular connective tissue
More fibers than cells
Specific orientation of collagen fibers
Few cells aligned between fiber bundles
Little ground substance
What is argrophyllic connective tissue and where are they found?
Network of reticular fibbers in a loose ground substance
Support other cell types and play a role in body protection
Found in liver, lymph nosed and bone marrow
What are elastic connective tissues and where are they found?
Predominant fibers are the Elastic fibers - yellow color Extremely elastic - 5x more than rubber (150% their resting length)
Found in the wall of elastic arteries (aorta), parts of the trachea and bronchi, vocal ligament, suspensory ligament of the penis
Provide durability with stretch, recoil
What is white adipose tissue and where is it found?
In areolar connective tissue, but very sparse; closely packed with adipocytes, or fat cells
Nucleus is pushed to the side by large fat droplet
Found under the skin; around kidneys and eyeballs; in bones and within abdomen; in the breasts
Provides reserve food fuel; insulates against heat loss; supports and protects organs.
