Cardiovascular and diabetes

Question 1

What is haemostasis?

Answer 1

A set of well-defined processes that accomplish two important
functions:
* Maintains blood in a fluid, clot-free state in normal
vessels.
* Facilitates rapid localised response to vessel injury to
stop blood loss by sealing the vessel wall (blood
clot/haemostatic plug)

Question 2

What is thrombosis?

Answer 2

• A corruption of haemostasis (pathological opposite)
• Unwanted/excess blood clotting (thrombus)
• Occurs if haemostasis is unregulated due to:
• Impaired inhibitory pathways
• If intensity of stimulus exceeds natural anti-coagulation
  processes

Question 3

What is embolus?

Answer 3

• Blood clot that travels through vessels

Question 4

What are the 4 stages of haemostasis?

Answer 4

1. Vasoconstriction
2. Platelet plug formation
3. Clot formation
4. Fibrinolysis

Question 5

Describe vasoconstriction in haemostasis?

Answer 5

• Direct effect of injury
• Proportionate to the degree of trauma
• Contraction of muscular walls of vessels/vascular spasm
• Minimisation of acute blood loss.

Question 6

Describe platelet plug formation in haemostasis?

Answer 6

• At the site of the disrupted endothelial lining the
  ECM/collagen becomes exposed to the blood
  components.
• ECM releases cytokines and inflammatory molecules
• Tethering of platelet by glycoprotein GP1b-V-IX receptor
  complex to the von Willibrand Factor on endothelial cells
• GP1b-V-IX also binds to P-Selectin
• Interactions between GPVI and collagen
• These steps initiate platelet aggregation cascades

Adhered platelets undergo specific changes:
1. Release cytoplasmic granules which contain ADP,
thromboxane A2, serotonin.
2. Change morphology (pseudopodia,  surface area)
3. Release cytokines.
4. GPllb/lla becomes activated: This occurs by Intact endothelial cells release prostacyclin
which bind receptors in platelets and induces
synthesis of cAMP, cAMP stabilises inactive GPIIb/IIIa and
stabilises granules containing platelet aggregation agents or Ca2+.
This causes the formation of a primary platelet plug

Question 7

Describe the coagulation cascade in haemostasis?

Answer 7

Initial development of a temporary platelet plug. Subject to further
cross linking and integration of Fibrin, more platelets and red blood
cells to produce a more stable thrombus/blood clot.

FIBRIN IS KEY TO THE COAGULATION CASCADE:
Fibrinogen is converted to fibrin.
Fibrin forms a mesh which captures more platelets and blood cells

Question 8

Describe the extrinsic pathway of the coagulation cascade?

Answer 8

• Also known as tissue factor pathway
• Induced by damage to surrounding tissues such as traumatic injury
• Step 1: Upon contact with blood plasma damaged cells release factor lll
  (thromboplastin/tissue factor)
• Step 2: Factor lll (thromboplastin/tissue factor) activates factor VII
  (proconvertin) complex with calcium forming an enzyme complex
• Step 3: This enzyme complex leads to activation of factor X (Stuart-Prower
  factor) which activates the common pathway

Question 9

Describe the intrinsic pathway of the coagulation cascade?

Answer 9

• Also known as contact activation pathway (Intrinsic because the factors are
  within the bloodstream)
• Prompted by damage to tissue from internal factors (arterial disease) Tissue factor
• Step 1: Initiated by factor XII (Hageman factor) which activates factor XI
  (anti-hemolytic factor C).
• Step 2: Factor XI then activates factor IX (antihemolytic factor b)
• Step 3: Factor VIII (antihemolytic factor a) then complexes with activated
  factor IX
• Step 4: the factor VIII/IX complex then activates factor X (Stuart-Prower
  factor) leading to the common pathway.

Question 10

Describe the common pathway of the coagulation cascade

Answer 10

Intrinsic and extrinsic pathways converge on one common
pathway. This process involves formation of a fibrin mesh
* Step 1: Activated factor X with factor V (co-factor) convert
prothrombin to thrombin
* Step 2: Thrombin converts Factor l (Fibrinogen) to Fibrin.
* Step 3: The transglutaminase, Factor XIII (fibrin-stabilising factor) is
also activated by Thrombin. Activated factor XIII covalently cross-links
fibrin to form a mesh-like structure, further stabilising the clot.

Question 11

Describe fibrinolysis in haemostasis?

Answer 11

• Plasminogen produced in liver and binds to
  fibrin.
• Tissue plasminogen activator (t-PA) converts
  plasminogen to plasmin
• Plasmin degrades blood plasma proteins
• Facilitates the degradation of blood clot.

Question 12

What are the 2 ways that coagulation can be regulated?

Answer 12

1. Thrombin (factor lla) circulates as the inactive zymogen Pro-thrombin (Factor ll)
   * Requires activation→Thrombin
   * Intrinsic feedback mechanisms control balance between activation and inhibition
   * Coagulation cascade is an amplification cascade-concentrations of molecules at each subsequent step
   is exponentially increased e.g. FXII (30 μg/mL) compared to Fibrinogen (3 mg/mL)
2. Control of Thrombin activity
   A) Thrombomodulin
   B) Circulating thrombin inhibitors

Question 13

Describe how thrombomodulin regulates blood clots?

Answer 13

• Thrombomodulin is an integral membrane protein.
• Found on surface of endothelial cells
• Cofactor for Thrombin
• Converts thrombin to an anti-coagulant enzyme from a
  procoagulant enzyme via direct interactions.
  Step 1: Thrombomodulin binds with Thrombin
  Step 2: Thrombin activation of protein C proceeds slowly but after
  formation of complex with TM there is a 1000-fold increase
  in the activation of Protein C to Activated Protein C (APC)
  Step 3: APC is a potent anticoagulant which degrades Factors Va
  and VIIIa limiting coagulation
  Deficiency in Protein C -> prone to venous thrombus

Question 14

Name 4 circulating thrombin inhibitors

Answer 14

1. Anti-thrombin III
   * Accounts for 75% anti-thrombin activity
   * Also inhibits Factors lX, Xa, Xla and Xlla
2. Heparin
   * Binds to cationic site of Anti-thrombin lll
   * Induces a conformational change and promotes
   inhibitory action of Anti-thrombin lll
3. α-Macroglobulin
4. α1-antitrypsin

Question 15

Describe the pathology of deep venous thrombosis and how it can be treated?

Answer 15

• DVT blood clot (thrombus) in one or more of the deep veins in the body, usually in the leg.
• Symptoms: pain, swelling of site but can present with no symptoms.
• Risk factors: Age, obesity, prolonged sedentary periods, injury, surgery, birth control pills,
  smoking.
• Blood clots in small vessels can break and travel, particularly to the lungs and cause
  pulmonary embolism

Treatment goals:
* Prevent growth of the clot
* Prevent the clot from breaking and relocating
* Reduce chances of another DVT

Treatments:
* Blood thinners (don’t break up clots but prevent growth)
e.g. IV administration of Heparin
* Clot busters: thrombolytics (only if severe)
* Filters
* Compression stockings

Question 16

Describe 2 treatments that reduce the formation of blood clots?

Answer 16

Aspirin
* Platelet aggregation inhibitor
* Thromboxane A2 promotes aggregation of
platelets
* Aspirin inhibits COX-1 which reduces
production of thromboxane A2 and hence
platelet aggregation.

Warfarin
* Coumarin anti-coagulant
* Factors II, VII, IX, X require Vitamin K as a co-factor for their
synthesis by the liver.
* These factors undergo a vitamin K-dependent post-translational
modification
* Glutamic acid residues are carboxylated to  Carboxy-glutamyl
residues.
*  Carboxy-glutamyl residues are required to bind Ca2+ which is
essential for interaction between platelets and co-factors.
* The carboxylases that catalyse the reaction are vitamin K-
dependent.
* Warfarin antagonises the co-factor function of vitamin K by
inhibiting vitamin K epoxide reductase

Question 17

Describe the pathology of an ischaemic stroke and how it can be treated?

Answer 17

An ischemic stroke occurs when a vessel supplying blood to the brain is obstructed. It accounts for about 87% of all strokes.

The main cause of ischemic stroke is atherosclerosis, or fatty deposits (plaque) that line the vessel walls.

Atheroma
* Intracranial vessels
* Extracranial vessels

Hypertension
* Risk factors for atheroma anywhere in intra and extracranial vessels
* Also causes changes in cerebral vessel walls

Treatments:
* Thrombolytic/Fibrinolyitc drugs activate
plasminogen.
* 3 major classes:
– tissue plasminogen activator (tPA)
– streptokinase (SK)
– urokinase (UK)
* Differ in their mechanisms that alter their
selectivity for fibrin clots.
* Alteplase (Activase®; rtPA) is a recombinant
form of human tPA.
* Approved for MI (FDA: 1987; EMA: 1988)
* Approved for Acute Ischemic Stroke
within 3 hours (FDA: 1996; EMA:2002)

Question 18

Describe 2 genetic disorders that cause thrombosis?

Answer 18

• Thrombophilia: characterized by an increased tendency for
  thrombosis.
• Individuals with thrombophilia are prone to developing venous
  thrombi, e.g. deep vein thrombosis (DVT) and pulmonary
  embolism (PE).
• Genetic causes include:
• Deficiencies or dysfunction of natural anticoagulants that
  normally help regulate clot formation and prevent
  excessive coagulation, e.g. Protein C, Protein S, or
  Antithrombin
• Mutations in coagulation factors, e.g. Factor V mutation

Factor V Leiden
* Defect in coagulation factor, Factor V
* Glutamine instead of arginine at amino acid 506 makes
Factor V resistant to inactivation by Activated Protein C
* The resistant Factor V (Leiden) remains active longer
→ increased thrombin generation
→ higher risk of abnormal clotting (thrombophilia)
Treatment of Factor V Leiden
* Traditional and newer anti-coagulants
* Future Possibilities: CRISPR-based gene editing to correct
the mutation are in preclinical development

Question 19

Describe the 2 types of haemophilia

Answer 19

Haemophilia A
* Inherited blood disorder in which there is a lifelong defect in the clotting mechanism of blood
* Loss of clotting Factor VIII
* Increased bleeding (usually internally)
* X-linked recessive trait therefore mostly affects XY individuals.
* Treatment with transfusion of plasma with FVIII concentrates/Recombinant FVIII now available (expensive)

Haemophilia B
* Due to deficiency of Factor IX
* Less common than Haemophilia A
* Also X-linked
* Treated by transfusion with F-IX (plasma concentrates or recombinant)

Question 20

Describe how gene therapy can be used to treat haemophilia B

Answer 20

Hemgenix (Etranacogene Dezaparvovec):
* Approved as the first gene therapy for hemophilia B (FDA: 2022, EMA:2023)
* Single dose of AAV virus containing functional copy of Factor IX gene delivered to the liver.
* Increased endogenous production of F-IX → reducing the need for regular F-IX infusions.
* Decreased number of bleeding episodes

Roctavian (Valoctocogene Roxaparvovec):
* Approved as the first gene therapy for adults with severe hemophilia A (FDA: 2023, EMA:2022)
* An adeno-associated virus vector delivers a functional copy of the Factor VIII gene, enabling patients
to produce Factor VIII themselves.
* Patients see a reduction in bleeding episodes

Question 21

How are lipids transported in the body?

Answer 21

Non-polar lipids (triacylglycerol, cholesteryl esters) are
associated with amphipathic lipids (phospholipids,
cholesterol) and proteins to form water-miscible
lipoproteins
This helps transport the lipids in a water soluble medium.

Question 22

What is the generalized structure and function of a plasma lipoprotein?

Answer 22

• Lipoproteins are composed of:
• a single layer of amphipathic phospholipid and cholesterol
  molecules.
• Apolipoproteins
• Non-polar lipid core consisting of triacylglycerols (triglycerides)
  and cholesteryl esters
• Solubilise lipids via lipid binding domains.
• Deliver lipids to cells for:
• Membrane synthesis (permeability and fluidity).
• Steroid hormone production (precursors).
• Energy metabolism
• Lipoproteins are taken up by receptors in the liver and other tissues.
• Regulate lipid metabolism through signalling domains.
• As they circulate through the bloodstream they can be extensively
  remodelled by interchange and removal of apolipoproteins and
  lipids.

Question 23

What are the 5 lipoprotein classes and what are they classified based on?

Answer 23

5 Lipoprotein classes:
* HDL (Highest density)
* LDL
* IDL
* VLDL
* Chylomicrons

Classified based on size, density, lipid
composition and apolipoproteins

Question 24

What are the 3 major protein pathways in lipoprotein metabolism?

Answer 24

• Exogenous: digestion, absorption, and
  packaging dietary lipids for secretion into
  lymphatics and into bloodstream.
• Endogenous: Biosynthesis and secretion of
  lipids and apolipoproteins by the liver and
  small intestine
• HDL: Mediates reverse cholesterol
  transport by transfer back to the liver for
  removal

Question 25

Describe exogenous fat metabolism

Answer 25

* Step 1: Dietary triglycerides hydrolysed into free fatty acids by gastric and pancreatic lipases * Step 1a: Cholesteryl esters are cleaved by cholesteryl esterase (mostly cholesterol exists as free sterol). * Step 2: Emulsification of fats by bile acids result in fine lipid droplets which are taken up by intestinal enterocytes (intestinal absorptive cells which line the inner surface of the small and large intestine). * Step 3: Upon entering the enterocytes the lipids make their way to the endoplasmic reticulum where they are resynthesized into triglycerides, cholesteryl esters and phospholipids. * Step 4: These reformed lipids are then packaged in Chylomicrons with the addition of apoB-48, apo-A1, and ApoA-IV. * Step 5: Loaded chylomicrons are then secreted into lymph. * Step 6: Chylomicrons then enter muscle and fat capillaries lose ApoA-I and ApoA-IV and acquire apolipoproteins CII (activator of lipoprotein lipase), apoCIII and apoE. * Step 7: The triglycerides within the chylomicrons are rapidly hydrolysed by lipoprotein lipase forming chylomicron remnants that are taken up by the liver via the LDL Receptor

Question 26

Describe the steps of endogenous fat metabolism

Answer 26

* Step 1: VLDL synthesized by liver used to export excess triglycerides . VLDL levels increase with high levels of free fatty acids (diet etc). * Step 2: VLDL contains apoprotein B-100. Also contains Apo C-II on surface which activates endothelial LipoProtein Lipase (LPL) to break down TGs into FFAs which are taken up by cells. * Step 3: LPL processing results in formation of IDL. IDL are cholesterol- rich VLDL and chylomicron remnants that are cleared by the liver or metabolized by hepatic lipase into LDL. LDLK retains Apo-B-100. * Step 4: LDLs are super-cholesterol rich molecules. Non-cleared LDL are taken up by either hepatic LDL receptors or scavenger cell LDL receptors (e.g. macrophages) leading to foam cell formation.

Question 27

Describe HDL metabolism

Answer 27

* Step 1: HDL is secreted by the liver or small intestine as a lipid poor, nascent particle containing ApoAI and small amounts of phospholipids and triglycerides. * Step 2: Unesterified cholesterol and phospholipids are transported out of cells via the ABCA1 transporter and loaded into nascent HDL. * Step 3: Lecithin-cholesterol acyl transferase (LCAT) esterifies free cholesterol present on nascent HDL with unsaturated fatty acids from the sn-2 position of lecithin to create small, spherical HDL particles. * Step 4: HDL transports cholesterol to the liver and steroidogenic tissues by three distinct pathways: * 1. Indirectly by transfer of cholesterol esters to VLDL and IDL which are then taken up by the LDL and LRP receptors. * 2. selective uptake via hepatic SR-B1 scavenger receptors * 3. ApoE-rich HDL uptake via ApoE and ApoA1 binding sites.

Question 28

Describe autosomal dominant familial hypercholestrolemia and how it can be treated

Answer 28

* Archetypal monogenic disorder leading to excess LDL. * Caused by one of over 600 known mutations in the LDL receptor (LDLR) gene. * Usually results in reduced LDL receptor numbers * More common within certain populations * French Canadian, Jewish populations of Lithuanian origin, South African Afrikaners) * Coronary artery disease often clinically apparent before 10 years of age. * Can be identified at birth due to elevated cholesterol in umbilical cord blood. * Drug therapy (Statins) is required to lower cholesterol levels by reducing cholesterol synthesis. * Potential gene therapy approach holds promise.

Question 29

Describe the pathology of familial defective apolipoprotein B

Answer 29

* As common as familial hypercholesterolaemia. * Results in elevated LDL-cholesterol due to impaired clearance * Caused by a mutation affecting the ApoB gene on chromosome 2 (renders ApoB unable to bind LDL-R). * Resulting amino acid substitution disrupts ApoB binding to LDL receptor and impairs LDL uptake. * Heterozygous mutation increases LDL levels by about 50% * Often indistinguishable from FH, requiring genetic testing for definitive diagnosis.

Question 30

What is the danger of low HDL cholesterol?

Answer 30

* Low HDL levels: <35mg/dL for men, <45 mg/dL for women. * HDL cholesterol level is under tight genetic control. * Low HDL Increases risk of coronary artery disease * Genetic causes of low HDL: * ApoA1 (apolipoprotein A1) mutation: Analphalipoproteinaemia * LCAT (Lecithin-cholesterol acyltransferase) mutation: Fish Eye Disease * ABCA1 (cholesterol efflux regulatory protein): Tangier disease

Question 31

Describe 2 drugs that affect lipid metabolism

Answer 31

Statins: * HMG-CoA reductase inhibitors * Treat individuals with elevated LDL-cholesterol * HMG-CoA reductase is the enzyme that controls the rate-limiting step for cholesterol biosynthesis in the liver and other tissues. * Statins block cholesterol synthesis, reduce hepatocyte cholesterol content, and increase the expression of the LDL receptor. * Unequivocally proven to reduce the incidence of all- cause mortality, stroke and re-vasculisation procedures Bile acid sequestrants: * Used to treat elevated LDL-cholesterol as an adjunct to statins. * Synthetic anion compounds that bind bile acids but not cholesterol. * Function in the intestinal lumen and prevent bile acid reabsorption. * Bile acid sequestrants bind bile acids in the intestine, reducing their reabsorption and increasing bile acid excretion. This lowers hepatic cholesterol, leading to increased LDL clearance from circulation.

Question 32

What are 2 drugs that can elevate lipid levels

Answer 32

Protease inhibitors * May stimulate triglyceride synthesis and upregulate gene expression of key enzymes in the lipid biosynthesis pathway Immunosuppressive agents * Corticosteroids can lead to elevated lipid levels. * E.g. Sirolimus (mTOR inhibitor) increases plasma cholesterol and triglyceride levels and exacerbates hyperlipidaemia.

Question 33

What is an atheroma

Answer 33

A reversible accumulation of degenerative tissue in the intima of the arterial wall