Extracellular matrix and fluids, compartements and solutes

Question 1

What is the extracellular matrix? What cells deposit it?

Answer 1

Network of macromolecules - minaly proteins and carbs filling spaces between cells

Deposited by fibroblast cells consisting of both fibrillar and non-fibrillar components

Question 2

What are the key functions of the ECM?

Answer 2

Provides physical support

Determines mechanical and physiochemical properties of the tissue

Influences the growth, adhesion and differentiation status of the cells and the tissue with which it interacts

Essential for development, tissue function and organogenesis

Question 3

What are the three types of connective tissues?

Answer 3

Collagens
Type I, II, III (fibrillar)
Type IV (basement membrane

Multi adhesive glycoproteins
fibronectin, fibrinogen
laminins

Proteoglycans
Aggrecan, versican, decorin
perlecan - basement membrane

Question 4

Explain the structure of type I collagen v type II and II collagen

Answer 4

→ Each collagen molecules has three alpha chains forming a triple helix
→ Type I collagen has chains from two different genes
→ Types II and III have one chain

Every third position in the alpha chains is.glycine

Question 5

Describe simply how an single collagen α chain forms a collagen fibre

Answer 5

synthesis of pro-α chain → One α chain → hydroxylation and glycosylation → three α chains (self assembly) → Secretion from cell and cleavage of propeptides → collagen fibril (self assembly) → collagen fibre (crosslinking)

Question 6

What is the purpose of cross-links in collagen?

Answer 6

Provides tensile strength and stability

Both lysine and hydroxy-lysine residues involved

• Crosslinks are tissue specific and change with age

Question 7

What is the role of lysine and proline hydroxylation in collagen formation?
What is required?

Answer 7

Contribute to interchain hydrogen bond formation

Prolyl and lysyl hydroxylases require Fe2+ and vitamin C - so deficiency leads to underhydroxylated collagens with consequences for tissue stability (scurvy)

Question 8

What is ehlers-danlos syndrome?

Answer 8

Group of inherited connective tissue disorders whose symptoms include stretchy skin and loose joints

several disorders arise due to mutations in collagen which negatively effect

• collagen production
• collagen structure
• collagen processing

Question 9

What type of collagens are IX and XII and what do they do?

Answer 9

Fibril-associated collagens

associate with fibrillar collages and regulate organisation of collagen fibrils

Question 10

What type of collagen is type IV?

Answer 10

Network framing collagen - present in all basement membranes (molecular constitution varies from tissue to tissue)

Question 11

What are basememt membranes/basal lamina ?

Answer 11

Flexible thin mats of extracellular matrix underlying epithelial sheets and tubes
surround muscle, peripheral nerevs and fat cells

highly specialised extracellular matrices containing distinct repertoire of collagens, glycoproteins and proteoglycans.

Question 12

How is basement membrane affected in diabetic nephropathy?

Answer 12

Diabetic nephropathy - an accumulation of extracellular matrix leading to a thickened glomerular basement membrane, this restricts renal filtration and lead to renal failure

Question 13

How is glomerular basement membrane affected in alports syndrome?

Answer 13

mutations in collagen IV result in an abnormally split and laminated GBM which is associated with progressive loss of kidney function and hearing loss

Question 14

What is the role of elastic fibres in the ecm?

How is extent of their purpose limited

Answer 14

for elasticity of tissues ie as skin, blood vessels and lungs

Collagen and elastic fibres can be interwoven to limit the extent of stretching
Elastic fibres consist of a core made up pf protein elastin and microfibrils which are rich in protein fibrillin

Question 15

What is protein fibrillin? What does a mutation in this lead to/

Answer 15

The protein in myofibrils around elastic fibres that determines integrity of the elastic fibres

Mutations in fibrillin-1 lead to marfans syndrome leading to disease manifestations in skeletel, ocular and cardiovascular systems - individuals predisposed to aortic ruptures

Question 16

What is elastin?

Answer 16

• Unusual protein consisting of two types segments that alternate along the polypeptide chain: hydrophobic regions and a-helical regions rich in alanine and lysine
• Many lysine side chains are covalently cross-linked

Question 17

What is the role of multi-adhesive glycoproteins?

Answer 17

Determine modular architecture of extracellular matrix proteins
Many large modular proteins are multi-adhesive binding various matrix components and cell surface receptors

Question 18

What are laminins?

How do they interact in the ECM?

What mutations can arise in these?

Answer 18

Heterotrimeric proteins
made up of an α chain, a β chain and a γ chain, which form a large cross shaped molecules

Muti-adhesive proteins - interact with various cell surface receptors including integrins and dystroglycan

self-associate as part of the basement membrane matrix, but also interact with other matrix components like type IV collagen, nidogen and proteoglycans

Mutations in specific chains are associated with inherited diseases such as….
○ Congenital muscular dystrophy can rise from an absence of the alpha-2 chain in laminin 2 - Symptoms include hypotonia (abnormally decreases muscle tension), generalised weakness and deformities of the joints
○ epidermolysis bullosa

Question 19

What are fibronectins in the ECM? How are they derived

Answer 19

• fibronectins are multi-adhesive proteins glycoproteins, made up of large multidomain molecule linked together by disulphide bonds
• They either exist as an insoluble fibrillar matrix or as a soluble plasma protein
• They are derived from a single gene, with alternate splicing of mRNAs giving rise to the different types

Question 20

What do fibronectins interact with and what are they important for?

Answer 20

• Fibronectins interact with cell surface receptors and other matrix molecules to regulate cell adhesion and migration in a variety of processes, ie embryogenesis and tissue repair

form a continuum with actin cytoskeleton of many cell types.
→ Integrin receptors at the cell surface provide the linkage between the matrix and cytoskeleton

• They are important for wound healing and helping promote clotting (ALS2- haemostasis)

Question 21

What are proteoglycans?

Answer 21

core proteins to which glycosaminoglycan (GAG) chains are attached
GAGs (repeating disaccharide units) are sulfated or carboxylated so carry a high negative charge
this attracys cloud of cations - Na= included so water is sucked into the ECM

Question 22

What are the different proteoglycan families and give examples

Answer 22

→ Basement membrane proteoglycans ie perlecan

→ Aggregating proteoglycans (interact with hyaluronan) eg aggrecan

→ Small leucine-rich proteoglycans ie decorin

→ Cell surface proteoglycans ie syndecans 1-4

Question 23

What is hyaluronan (hyaluronic acid)?

Answer 23

Found in the extracellular matric of soft connective tissue
a carbohydrate chain with no core protein
Unsulfated and made of repeating disaccharides

Can undergo a high degree of polymerisation, creating enormous molecules thus can occupy a large volume

Its typically of high viscosity - vitreous humour of eye and synovial fluid of joints where it protects cartilaginous surface from damage

Question 24

What is aggrecan in the cartilage matrix suited for?

Answer 24

Resists compressive forces

Under compressive load water is given up, but regained once the load is reduced.

Question 25

What are two types of disorders of the ECM?

Answer 25

Osteoarthritis
Erosive disease resulting in excessive extracellular matrix degradation
• The cushioning properties of cartilage over the end of bones is lost, so with increasing age aggrecan is cleaved by aggrecanases and metalloproteinases.
• This results in a loss of aggrecan fragments to the synovial fluid

Fibrotic diseases
Arises as a result of excessive production of fibrous connective tissue

Question 26

What does a histopathologist do?

Answer 26

deals with tissues. She/he will examine sections, noting the architecture of the tissue and asking what it tells us about a particular condition.

works with 
Biopsies 
Resection specimens
Frozen specimens 
Post-mortems

Question 27

What does a cytopatholgist do?

Answer 27

deals with cells. They are often are the individuals responsible for taking the cells from the patient, preparing them for examination and then delivering their expert diagnosis on the cell sample.

Question 28

Biopsy?

Answer 28

biopsies are small sections of tissues that are removed from the patient and typically placed in a formalin solution which preserves the tissues by cross-linking proteins

mbedded in paraffin wax to allow thin sections to be cut by an instrument known as a microtome

Question 29

What does the Zieh-Neelsen stain do?

Answer 29

stain acid-fast bacteria red, aiding in the diagnosis of tuberculosis infection

Question 30

Resected specimens?

Primary used for?

Answer 30

Def- Taken from tissue that has been removed as part of a surgical procedure and can be processed for biopsy

Primarily used to look at the stage of the disease ie cancer

can also be donated to biobanks and used to inform genomic studies of the disease process, with the input of other medical professionals

Question 31

Frozen sections?

Answer 31

Frozen sections are taken during surgical procedures and are examined by pathologists while the patient is being operated u​pon - rapid diagnosis in minutes

freshly taken tissue is frozen by a machine known as a cryostat, cut then mounted on glass slides and stained as for biopsies

Question 32

Fine needle aspirates?

Answer 32

• A fine needle can be used to get into a lesion and aspirate (suck out) the cells which can be analysed as a smear
• powerful technique as the needle can penetrate relatively inaccessible tissues ( thyroid nodule) and assess the suspect mass without the need for surgery

Question 33

What indicates endothelial cell tumours?

Answer 33

CD31 immunohistochemical staining of a skin biopsy.
Brown colouration indicates CD31 positivity

antibody recognising the endothelial cell marker CD31 on the surface of endothelial cells allows us to define this biopsy as being of an endothelial cell tumour

Question 34

What are some antibody conjugates and what do they do?

Answer 34

Enzymes - peroxidase, alkaline phosphatase

Fluorescent probes - allow rapid measurement of hte levels of mokecules in a sample

Magnetic beads - purification of cell types

Drugs; ie kadcyla, anti-HER2 antibody linked to emtansine

Question 35

What are the three uses of manufactures antibodies?

Answer 35

Blood group serology - blood transfusion

Immunoassays - detection of hormones and circulating antibodies/antigens

Immunodiagnosis - infectious diseases, AB levels and IgE hypersensitivity

Question 36

What precaution is taken to stop donated organ deterioration during transport?

Answer 36

rapid cooling to 4 degrees
Perfused with cold solutions via arterial supply

ie universit of wisconsin solution (UW)

• lack of Na+ and Cl- so no influx possible
• Presence of extracellular impermeant solutes (lactobionate ions, raffinose).
• Presence of a macromolecular colloid (starch)

Question 37

What is the colloid osmotic pressure?

Answer 37

Higher concentration of plasma proteins inside the capillary than outside generates an osmotic pressure known as the colloid osmotic pressure (COP)

Question 38

What is inflammatory oedema?

Answer 38

cardinal signs of inflammation

Infectious and inflammatory stimuli often results in oedema
inflammation can be observed around the sites of insect bites which has caused local blood vessels to become leaky. Swelling occurs because the rate of leakage from the vessels is greater than the rate at which the lymphatics can drain it.

Question 39

What is hydrostatic oedema?

Answer 39

have high blood pressure, which means increased hydrostatic pressure in vessels.
This pushes more fluid out of the vessels, and can lead to accumulation of interstitial fluid

Question 40

Why does a breast cancer survivor often have oedema in arms?

Answer 40

likely to have had axillary (armpit) lymph nodes removed as part of her diagnosis or treatment
This can remove the pathway of drainage from the upper limb on the affected side, resulting in the accumulation of fluid.

Question 41

What happens in elephantiasis?

Answer 41

parasitic worms can block lymphatic vessels, thereby preventing drainage of the lymph

Question 42

Features of marfans syndrome?

Answer 42

long slender limbs, long fingers, a predisposition to aortic ruptures and heart murmur

Question 43

Features of Ehler’s-Danlos syndrome?

Answer 43

Joint hypermobility and extremely elastic skin