Cardiovascular System and the Kidney

• The following notes are from the March 2024 lecture
• Prof Steve Safrany 341 and Dermot Cox from the School of Pharmacy and Biomolecular Sciences presented

Learning Outcomes

• Mechanisms of action are described for diuretics, ACE inhibitors, ARBs, and mineralocorticoid receptor antagonists
• Mechanisms of action covered for natriuretic peptides and neprylisin inhibitors
• The role of immune modulators in kidney disease is described
• The compositions and uses of intravenous fluids are compared

Role of Sodium

• Increase in plasma volume is, in part, due to an increase in osmotic pressure
• Sodium increase can cause an increase in resistance, leading to small resistance vessel and large elastic artery remodeling
• Endothelial dysfunction can be a reduction in NO production
• Increased sympathetic tone

RAAS (Renin-Angiotensin-Aldosterone System)

• Angiotensinogen is a 452 amino acid peptide synthesized in the liver
• Angiotensin I is the first 10 amino acids cleaved by renin
• Angiotensin I is the inactive product of renin
• ACE (angiotensin-converting enzyme) removes the terminal two amino acids from angiotensin I, producing Ang II (Asp-Arg-Val-Tyr-Ile-His-Pro-Phe)
• Ang II is further cleaved into bioactive peptides

Angiotensinogen Details

• Ang 1-7 is a vasodilator that increases NO and counterbalances Ang II
• COVID virus attaches to ACE2, reducing its function
• Des(Ang I) AGT makes up 98% of the protein and has no known function

Angiotensin II Receptors

• Ang II acts on angiotensin II receptors
• AT1 and AT2
• G-protein-coupled receptors are Gq-linked
• Ang II binding to AT1 is primarily responsible for vasoconstriction through increased NAd production and sodium reabsorption

Aldosterone

• Is a steroid hormone
• Increases Na⁺ reabsorption
• Increases BP

Diuretics

• Check Med1 and CVS hypertension lecture
• Kidneys filter ions from plasma, which brings water
• Some of the filtered ions are reabsorbed
• Diuretics increase urine production
• Loop, thiazide and potassium-sparing diuretics are key classes
• Carbonic anhydrase inhibitors, osmotic diuretics and vasopressin receptor antagonists are further classes

Thiazide Diuretics

• These diuretics are based on benzothiadiazine
• They block Na⁺ reabsorption, increasing water loss
• They block the Na/Cl channel in the proximal segment of the distal convoluted tubule
• Bendroflumethiazide is most common in the UK, and not used in the US
• Hydrochlorothiazide used more commonly in the US than the UK
• Thiazide-like diuretics are structurally different but have a similar MOA
• Indapamide, mainly in the UK, is seldom used in the US
• Chlorthalidone is another thiazide-like diuretic

Thiazide Diuretics - Adverse Effects

• Can cause hyponatremia and increase the amount of urine Na⁺ leading to compensation
• Can also increase Na⁺/Ca²⁺ exchange, causing hypercalcaemia
• Increasing secretion of K⁺ can cause hypokalaemia, metabolic alkalosis, and hyperglycaemia
• An increase in urate absorption results in hyperuricemia, leading to gout

Loop Diuretics

• Highly protein bound and transported into cells by OAT in the proximal convoluted tubule
• Excreted into the urine by multidrug resistance-associated protein 4 (MRAP4)
• Inhibit the Na⁺/K⁺/Cl⁻ co-transporter in the ascending limb of the Loop of Henle
• Blocks the re-absorption of Na⁺, K⁺, and Cl⁻
• Adverse effects include; renal dysfunction, hyponatraemia, hypokalaemia and hypochloraemia, metabolic alkalosis
• Furosemide, bumetanide, and torsemide are loop diuretics

Potassium-Sparing Diuretics

• Hypokalaemia is a common adverse effect with many diuretics
• Epithelial Na channel (ENaC) blockers, such as amiloride and triamterene, cause diuresis without hypokalaemia
• Mineralocorticoid receptor antagonists; spironolactone, eplerenone, and finerone also cause this
• ACE inhibitors and ARBs are potassium-sparing, but are not diuretics

Loss of Potassium

• Hypotension activates RAAS and aldosterone secretion which stimulates Na⁺ reabsorption in the collecting tubule
• Na+ reabsorption can increase K⁺ secretion and cause hypokalemia
• These diuretics prevent Na⁺ reabsorption in the collecting tubule, and prevent hypokalaemia which can cause hyperkalaemia
• It can be used with thiazide/loop diuretics to prevent hypokalaemia, but is not a very powerful diuretic by itself

Potassium-Sparing Diuretics- MOA

• Block ENaC to prevent Na⁺ reabsorption from urine
• This prevents exchange of Na⁺ for K⁺ in collecting tubules, which prevents hypokalaemia
• Aldosterone is a steroid agonist on mineralocorticoid receptor
• This upregulates ENaC and Na⁺/K⁺ATPase

Carbonic Anhydrase

• This catalyses the interconversion of bicarbonate, carbon dioxide and water
• Meaning HCO3- + H+ ⇌ CO2 + H2O
• It is widely distributed
• In the kidney, it facilitates the reabsorption of Na+, H+ and H2O from the urine
• Inhibiting carbonic anhydrase leads to an increase in excretion of Na+, H2O and HCO3-
• Acetazolamide is a Carbonic Anhydrase inhibitor.
• Diuresis is weak
• These are beneficial in patients with oedema and metabolic alkalosis as the loss HCO3- causes an acidosis which corrects the alkalosis

Acetazolamide Effects

• In urine pH is low and H₂CO₃ turns into H₂O + CO₂
• H₂O and CO₂ go into cell at proximal tubule
• Raised pH is created and converts H₂O + CO₂ = H⁺ + HCO₃-
• NHE3, Na and H exchanger, kNBC1, Na and HCO₃ transporter
• In the Urine of AZM patients, H₂CO₃ becomes H₂O + CO₂
• This means HCO₃ is trapped in urine and that H⁺ + HCO₃‾ not raised in the cell
• NHE3 cannot reuptake Sodium
• kNBC1 cannot transport Na/HCO₃ into blood

Osmotic Diuretics

• Mannitol is a poorly absorbed alcohol sugar
• This is given intravenously
• Retains water in the urine due to increased osmotic pressure
• Creates a water diuresis, and has little impact on sodium
• Initially act to increase blood volume

Sodium-Glucose-Linked Transport Inhibitors

• Found in the kidney
• Secondary active transporters pump out 3 Na+ in exchange for 2 K+
• Glucose is then imported into cells
• SGLT-2 recovers glucose from the urine, allowing increased glucose in urine which acts as an osmotic diuretic

Vasopressin / Anti-Diuretic Hormone

• ADH/vasopressin is a nonapeptide synthesised in the hypothalamus
• Acts on V receptors
• V1 are Gq-linked receptors on blood vessels increasing vascular tone
• V2 receptors are Gs-linked and found in the nephron
• Cause increased surface expression of aquaporin 2 (AQP2)
• AQP2 is a water channel that leads to increase in water absorption

Vasopressin Receptor Antagonists

• Vasopressin receptor antagonists are "vaptans"
• Tolvaptan is V2 selective
• Conivaptan is non-selective
• Increased water loss and plasma Na+ concentration
• Conivaptan may decrease blood pressure
• Vaptans should not be used for more than 30 days or in patients with liver disease due to hepatotoxic effects

Natriuretic Peptides

• These are a family of peptides that increase Na+ excretion
• Atrial (ANP), Brain (BNP) and C-type (CNP) natriuretic peptide types exist
• They bind to specific receptors like NPR-A, -B & -C
• Neprilysin metabolises them , inhibiting this leads to effects
• Sacubitril is used in conjunction with valsartan as an ARNI (angiotensin receptor neprilysin inhibitor)
• This increases Na+ and H2O excretion
• Used in heart failure

Effects of ARNIs

• Neprilysin breaks down NPs & AngII
• Inhibitors will protect them
• Adding ARB can prevent red outcome

Clinical use of diuretics

• Treats hypertension, which occurs when increasing Sodium excretion reduces blood pressure
• Primarily thiazide diuretics, with Loop diuretics being second inline
• Treats heart failure associated with oedema
• Use Loop blockers + Thiazides + Potassium-sparing to supplement other diuretics
• Treats glaucoma - carbonic anhydrase inhibitors
• Treats raised intracranial pressure with osmotic diuretics

Intravenous Fluids - Fluid Resuscitation

• Fluids treat blood loss for hypovolemia
• Can Correct electrolyte imbalance, like hyponatremia & hypernatremia, and provide glucose
• Can use multiple fluids or 2 types
• Water with electrolytes come as crystalloids, whether Isotonic or hypotonic
• Solutions like saline, Ringer's, or Lactated Ringer's (Hartmann's)
• Colloids: A suspension of insoluble particles such as Albumin and Dextrans

Crystalloids

• Standard fluid is normal isotonic saline containing buffered 0.9% NaCl
• Actually, has higher NaCl concentration than plasma which can cause Hyperchloremic acidosis
• There is reduced NaCl and a buffer such as Buffered crystalloid (balanced solutions), Bicarbonate or Lactate and also containing lactate ringers
• No clear evidence supports normal saline vs balanced crystalloid
• To Note: Normal saline as initial resuscitation fluid and Balanced crystalloid as second line or if a large volume of fluid is to be administered

Colloids

• Albumin or Dextrans in suspension is a colloid
• Hyperoncotic albumin is more effective at volume expansion
• But is very expensive, and is not as beneficial when compared to crystalloid
• Starch has been shown to increase the risk of acute kidney injury which results not recommenced for managing hypovolemia

Renal Failure

• Renal failure requires a patient to undergo dialysis.
• Dialysis is not an ideal treatment option
• A kidney transplant from a compatible donor is the most effective treatment
• Graft rejection is the biggest challenge to transplants
• Graft rejection treated with immunosuppressants

Role of IL-2

• 15.5 kDa protein and is the Important stimulator of cell-mediated immune system which Acts on IL-2 receptor
• Is an important target in pharmacotherapy as IL-2 plays plays a role in renal graft rejection
• Decrease IL-2 levels or Block IL-2 receptors - Block IL-2 signalling to help treat

Calcineurin Inhibitors

Cyclosporine

• Is a cyclic undecapeptide from the Tolypocladium inflatum fungus
• Inhibits mixed lymphocyte reaction (MLR) but was not cytotoxic (1972)
• Inhibits antibody production against T-cell dependent antigens like sheep erythrocytes, but not T-cell independent antigens like LPS

Tacrolimus

• Is a macrolide antibiotic isolated from the Streptomyces tsukubaensis fungus
• Similar activity to cyclosporine but more potent (10-100x)
• Found to inhibit MLR in 1980's
• Structurally unrelated to cyclosporine

Calcineurin Inhibitors - Mechanism

• Tacrolimus & CsA inhibit gene transcription, thus inhibiting IL-2,3&4, GM-CSF, TNFa and IFNy
• Specific for pathways involving increase in intracellular Ca2+
• Specific for T-cell receptor

Immunophilins

• Both drugs bind to a family of intracellular receptors known as immunophilins (0.1-0.4% of cell protein)
• These are Prolyl-peptidyl cis-trans isomerases and rotamases involved in protein folding
• Cyclosporine binds to cyclophilin (CyP)
• FK506 binds to FK506-binding protein (FKBP)

Calcineurin

• Immunosuppressive activity is independent of rotamase activity
• Drug-immunophilin complexes bind to and inhibit calcineurin, which is a serine-threonine phosphatase
• Nuclear factor of activated T-cells (NF-AT) is a substrate for calcineurin which is a transcription factor that binds to the IL-2 promoter

Rapamycin (Sirolimus)

• Structural homolog of FK506
• Isolated from Streptomyces hygroscopicus.
• Works by a different mechanism
• Similar profile to FK506 - Binds to FKBP
• Pre-existing antibiotic shown to have immunosuppressant activity (tested in 1989)

Targets of Rapamycin

• Rapamycin-FKBP complex does not inhibit calcineurin or gene induction
• Rapamycin inhibits signal transduction by IL-2 receptor
• Rapamycin-FKBP binds to targets of rapamycin (mTOR 1) a phosphatidylinositol 3-kinase
• Leads to reduced expression of adhesion molecules including sphingosine-1-phosphate receptor and Causes cell cycle arrest at G1

Other Immune Suppressants

Mycophenolate Mofetil

• Converted to active metabolite, mycophenolic acid
• Inhibits IMPDH, the Inosine monophosphate dehydrogenase
• Essential for guanine nucleotide synthesis
• Immunosuppressive and anti-proliferative and Used in renal transplants as prophylaxis with calcineurin inhibitors

Basiliximab

• Is Anti-IL-2 monoclonal antibody which has approval for renal transplant
• This is Used in induction therapy