PHYSIOLOGY & PATHOPHYSIOLOGY OF RAAS

1. DR.SANDEEP R SR , DEPT. OF CARDIOLOGY PHYSIOLOGY &PATHOPHYSIOLOGY OFRAAS CMC CALICUT

2. 1) HISTORY • 2) ANGIOTENSINOGEN • 3) RENIN • 4) ACE • 5)ANGIOTENSINS • 6)ANGIOTENSIN RECEPTORS • 7)LOCAL RAAS • 8)CARDIAC RAAS • 9)ALDOSTERONE • 9)PATHOPHYSIOLOGY OF RAAS

3. HISTORY • Robert Tigerstedt and Per Bergman from Sweden in their seminal 1898 report, Niere und Kreislauf, described the prolonged vasopressor effects of crude rabbit kidney extracts. • Tigerstedt named the unidentified active substance “renin” on the basis of its organ of origin. Steven A. Atlas, MDJTheRenin-AngiotensinAldosterone System: Pathophysiological Role and Pharmacologic Inhibition J Manag Care Pharm. 2007;13(8)(suppl S-b):S9-S20

4. Goldblatt and colleagues, published in 1934, that showed that renal ischemia induced by clamping of the renal artery could induce hypertension. • Ischemic kidney also released a heat-stable, short-lived pressor substance, in addition to renin. • This finding eventually led to the recognition that renin’spressor activity was indirect and resulted from its proteolytic action on a plasma substrate (eventually termed “angiotensinogen”) to liberate a direct-acting pressor peptide. • This peptide was initially termed “angiotonin” or “hypertensin” by Page & colleagues(US) • Ultimately named “angiotensin” by Braun-Menendez & colleagues(ARGENTINA) Steven A. Atlas, MDJTheRenin-AngiotensinAldosterone System: Pathophysiological Role and Pharmacologic Inhibition J Manag Care Pharm. 2007;13(8)(suppl S-b):S9-S20

5. In the early 1950s, during attempts at purification, Skeggs & colleagues discovered that this peptide existed in 2 forms, eventually termed Ang I and II. In later work, they demonstrated that Ang I was cleaved by a contaminating plasma enzyme,termed “angiotensin-converting enzyme,” to generate the active pressor peptide Ang II. • Laragh, Genest, Davis, Ganong, and their colleagues, culminated in the discovery that Ang II also stimulated the release of the adrenal cortical hormone aldosterone Steven A. Atlas, MDJTheRenin-AngiotensinAldosterone System: Pathophysiological Role and Pharmacologic Inhibition J Manag Care Pharm. 2007;13(8)(suppl S-b):S9-S20

6. ANGIOTENSINOGEN • The primary source of systemic circulating angiotensinogen is the liver particularly the pericentral zone of the hepatic lobules. • It is coded by a single gene, composed of five exons and four introns, that spans approximately13 kb of genomic sequence on chromosome 1 (1q42-q43). • Also detected in many other tissues, including kidney, brain, heart, vascular, adrenal gland, ovary, placenta, and adipose tissue • Rise in response to glucocorticoids, estrogens and other sex steroids, thyroid hormone, inflammatory cytokines (e.g., interleukin-1 and tumor necrosis factor), and Ang II. Ron D, Brasier AR, Habener JF. Angiotensinogen gene-inducible enhancer-binding protein 1, a member of a new family of large nuclear proteins that recognize nuclear factor kappa B-binding sites through a zinc finger motif. Mol Cell Biol. 1991;11:2887-2895.

7. RENIN • Glycoprotein (Mw 37,326) • Renin gene located in CHR.1 • Synthesized from preprorenin • Mature active renin is an aspartyl protease secreted by juxtaglomerular cells Richard E. Gilbert, David S. Game, and Andrew Advani:BRENNERS TEXTBOOK OF NEPHROLOGY;2010;12;384-385

8. JUXTAGLOMERULAR APPARATUS • The Juxtaglomerular Apparatus consists of: • The juxtaglomerular cells (2) The macula densa (3) The lacis cells or agranular cells

9. RENIN -SYNTHESIS PREPRORENIN( 406 aminoacid) PRORENIN (383 aminoacid) RENIN (340 aminoacid) Renin is also synthesized In brain, adrenal gland, ovary, visceral adipose tissue, heart and vasculature Half life of renin is 80 mt Only action of renin is conversion of angiotensinogen to angiotensin I

10. REGULATION OF RENIN • Active renin secretion is regulated principally by 4 interdependent factors: • (1) A renal baroreceptor mechanism in the afferent arteriole that senses changes in renal perfusion pressure, • (2) Changes in delivery of NaCl to the macula densa cells of the distal tubule • (3) Sympathetic nerve stimulation via beta-1 adrenergic receptors • (4) Negative feedback by a direct action of Ang II on the JG cells. HAYO CASTROP, KLAUS HO ¨ CHERL, ARMIN KURTZ, FRANK SCHWEDA, VLADIMIR TODOROV, AND CHARLOTTE WAGNER; Physiology of Kidney Renin: Physiol Rev 90: 607–673, 2010;

11. REGULATION OF RENIN SECRETION • 1.INTRARENAL MECHANISM • Renal baroreceptor mechanism • Renin secretion increases as the blood pressure falls below 90mmhg • The precise mechanism , how the pressure signal is transduced into renin release is still unknown, although postulated mediators include stretch-activated calcium channels, endothelins, and prostaglandins • JG cells are strongly electrically coupled to the neighboring cells of the afferent arteriole . • Interestingly, their resting membrane potential changes in situ from -60 to -80 mV in nonpressurizedarterioles to approximately -40 mV in pressurized arterioles • Since the depolarization of JG cells is accompanied by the suppression of renin release, the depolarization in response to an increase in perfusion pressure might directly or indirectly contribute to the known pressure-dependent inhibition of renin secretion HAYO CASTROP ET AL; Physiology of Kidney Renin: Physiol Rev 90: 607–673, 2010;

12. 2.NEURAL CONTROL • The JGA is endowed with a rich network of noradrenergic nerve endings and their β1 receptors • Stimulation of the renal sympathetic nerve activity leads to renin secretion that is independent of changes in renal blood flow, glomerular filtration rate (GFR), or Na+ resorption • Moreover, this effect can be blocked surgically by denervationand pharmacologically by the administration of β blockers

13. 3.TUBULAR CONTROL • Chronic diminution in luminal NaCl delivery to the macula densa is a potent stimulus for renin secretion • This mechanism is thought to account for the chronically high plasma renin activity (PRA) in subjects who adhere to a low-salt diet • The initial step of the MD-dependent control of renin secretion is the detection of the NaCl concentration in the tubular lumen by the MD cells. HAYO CASTROP ET AL; Physiology of Kidney Renin: Physiol Rev 90: 607–673, 2010;

14. INCREASED UPTAKE INTO MACULA DENSA THROUGH NA/K+/2CL CHANNEL DISTAL TUBULAR SODIUM ↑ MACULA DENSA Na+ & FLUID CONTENT↑ SWELLING OF MACULA DENSA & STRETCH STRETCH CAUSES ADENOSINE RELEASE A2 CAUSES ↓ RENIN SECRN NO SYNTHETASE ↓ DECREASES RENIN

15. Na+ UPTAKE IN MACULA DENSA ↓ • DISTAL TUBULAR SODIUM ↓ ACTIVN OF NO SYNTHETASE PG SYNTHESIS STIMULATE RENIN RELEASE

16. The detection done by Na-K-2Cl cotransporter in the apical membrane of tubular cells • Three mediators - prostanoids, NO, and adenosine/ATP Castrop et al ; Physiology of Kidney ReninPhysiol Rev 90: 607–673, 2010

17. REGULATION OF RENIN

18. FACTORS THAT DECREASE RENIN RELEASE ANG II VASOPRESSIN A N P IL6 5) TNF - ALPHA 6) ADENOSINE • FACTORS THAT INCREASE RENIN RELEASE • 1)Catecholamines • 2)Bradykinin • 3)Dopamine • 4)NO • 5)Prostaglandins Castrop et al ; Physiology of Kidney ReninPhysiol Rev 90: 607–673, 2010

19. MOLECULAR MECHANISM OF RENIN RELEASE • C-amp mediated - sympathetic, prostaglandin E2 & I2,dopamine • Calcium paradox - increase in intracellular Ca2+ decrease renin release-endothelin,vasopressin,Ang II • C-GMP Low consc. stimulates & High consc. Inhibits - NO Castrop et al ; Physiology of Kidney ReninPhysiol Rev 90: 607–673, 2010

20. PRORENIN • Traditionally, prorenin was considered the inactive precursor of renin • The Current studies implicate prorenin and renin as direct cardiac and renal toxins • Prorenin is inactive because a 43–amino acid hinge is closed and prevents it from binding to angiotensInogen • Prorenin & renin levels increased by ACE inhibitor , ARB , DRI Richard E. Gilbert, David S. Game, and Andrew Advani:BRENNERS TEXTBOOK OF NEPHROLOGY;2010;12;384-385

21. PRORENIN RECEPTOR • The kidneys convert inactive prorenin to active renin by enzymatic cleavage of this inhibitory hinge region • When circulating prorenin binds to a newly discovered (pro)renin receptor in the heart and kidneys, the hinge is opened (but not cleaved), and this nonenzymatic process fully activates prorenin Direct Renin Inhibition: Focus on Aliskiren James L. Pool, MD JMCP October 2007 Vol. 13, No. 8, S-b

22. RENIN/PRORENIN RECEPTOR • The existence of high-affinity cell surface receptors that bind both renin and prorenin in several tissues, including heart, brain placenta, and kidney but its significance still unknown • The binding of renin to its receptor resulted in a fivefold increase in the catalytic activity compared with renin in solution • The binding of pro-renin to the receptor increased its enzymatic activity from virtually zero to values comparable to those of active renin in solution • Activation of the (pro)renin receptor increases TGF-β production, leading to collagen deposition and fibrosis Direct Renin Inhibition: Focus on Aliskiren James L. Pool, MD JMCP October 2007 Vol. 13, No. 8, S-b

23. ACE • ACE is responsible for the cleavage of Ang I to form the octapeptideAng II • ACE cleaves bradykinin into inactive fragments • Human ACE in encoded by a single gene located on chromosome 17 • The majority (∼90%) of ACE activity in the body is found in tissues; the remaining 10% of ACE activity is found in a soluble (non–membrane bound) form in the interstitium of the heart and vessel wall • It is seen pulmonary vascular endothelium, endothelium of vasculature ,cell membrane of heart , kidneys & brain

24. ACE 2 • ACE2 represents a zinc metalloprotease with carboxypeptidase activity that shares 42% identity with the catalytic site of somatic ACE and can be shed from cells • ACE2 can convert ANGII to ANG 1-7 & ANG I to ANG 1-9 • Preferable physiological substrate for ACE2 seems to be ANG II • The expression of ACE2 is (in comparison with ACE) relatively restricted to cardiac blood vessels and tubular epithelia of the kidneys • ACE 2 cannot hydrolyze bradykinin and is not inhibited by ACE inhibitors • Actual function and significance is still unknown • ACE2 is the functional receptor for coronavirus associated with the acute respiratory syndrome, i.e., SARS

26. Steven A. Atlas, MDJTheRenin-AngiotensinAldosterone System: Pathophysiological Role and Pharmacologic Inhibition J Manag Care Pharm. 2007;13(8)(suppl S-b):S9-S20

27. ANGIOTENSIN SYNTHESIS

28. ANGIOTENSIN RECEPTORS • ANGIOTENSIN RECEPTORS • 1)AT1 • 2)AT2 • 3)AT3 • 4)AT4 • 5)Mas receptor

29. AT1 RECEPTOR • AT1 - G protein coupled receptor • Chromosome 3 • AT1A receptors are found predominantly in kidney, lung, liver and vascular smooth muscle • AT1B receptors are expressed mainly in the adrenal and anterior pituitary glands. Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

30. AT1 receptors are primarily found in the • 1)Brain-hypothalamus,NTS and ventrolateral medulla in the hindbrain,ant.pituitary • 2)Adrenals- the zonaglomerulosa of the adrenal cortex and chromaffin cells of the adrenal medulla • 3) Heart - in the conducting system , nerves of myocardium • 4)Vasculature-the aorta, pulmonary and mesenteric arteries, are present in high levels on smooth muscle cells and low levels in the adventitia • 5) Kidney – glomerularmesangial cells and renal interstitial cells Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

31. AT1 RECEPTOR • The predominant angiotensin receptor in the vasculature is the AT1 receptor • Although both the AT1 and AT2 receptor subtypes are present in human myocardium • Ratio of AT1 to AT2 receptors decreases in heart failure Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

32. ACTIONS OF AT1 RECEPTOR • 1) Blood vessels – vasoconstriction leading to an increase in peripheral vascular tone and systemic blood pressure Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

33. 2)Heart • Positive ionotropic and chronotropic effects of Ang II on cardiomyocyte • 1)INCREASED SYMPATHETIC ACTIVITY • 2)INCREASED CA2+ INFLUX • Ang II is also known to mediate cell growth and proliferation in cardiac myocytes and fibroblasts, as well as in vascular smooth muscle cells via TGF,PDGF etc Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

34. 3) Adrenal -Ang II stimulates the release of catecholamines from the adrenal medulla and aldosterone from the adrenal cortex • Trophic factor for zonaglomerusa • 4) Brain- thirst , salt appetite, central control of blood pressure, stimulation of pituitary hormone release and has effects on learning and memory Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

35. 5)RENAL • It increases salt reabsorbtion – direct & indirect • Direct effect Renal arterioles constriction Proximal Tubular Epithelial cells Peritubular capillary pressure Sodium reabsorbtion Fluid reabsorption From tubules

36. INDIRECT ACTION OF ANG II IN KIDNEY • ANGIOTENSIN II ADRENAL CORTEX STIMLN. SYMPATHETIC ACTIVATION ALDOSTERONE SECRETION MESANGIAL CONTRN GFR SALT & WATER RETENTION

38. AT2 RECEPTOR • The AT2 receptor is also a seven transmembrane domain receptor, encoded by a 363-amino-acid protein( MW 41 kDa) • Shares only 34% sequence identity with the AT 1 receptor • The AT2 receptor -highly expressed in foetus but rapidly declines at birth • AT2 receptors are present in brain, heart, adrenal medulla, kidney and reproductive tissues Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

39. Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

40. AT2 RECEPOR & ITS ACTIONS • Brain – cerebellum • Heart - fibroblasts in interstitial regions • Adrenal- adrenal medulla • Kidney- the AT2 receptor is localized to glomeruli, tubules and renal blood vessel • ACTIONS • Vasodilation • Antiproliferative • Apoptosis • Thirst Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

41. AT3 & AT4 Type 3 (AT3) receptors – Function unknown The type 4 (AT4) receptors - mainly mapped in brain & kidney • Thought to mediate the release of plasminogen activator inhibitor 1 by Ang II and by the N-terminal truncated peptides (Ang III and Ang IV) • Suggested a role in mediating cerebral and renal blood flow, memory retention and neuronal development Diem T. DINH*, et aI:Angiotensin receptors : distribution,signalling and function:Clinical Science (2001) 100, ;481–492

42. Mas RECEPTOR Mas receptor - Acted upon by ANG1-7 Causes • Vasodilatation • Natriuresis • Antiproliferation • Cardiac protection Castrop et al ; Physiology of Kidney ReninPhysiol Rev 90: 607–673, 2010

43. LOCAL RAS • Angiotensin II is found to be synthesized in the various tissues through ACE & non ACE pathways • Independent Ang II - heart, peripheral blood vessels, kidney, brain, adrenal glands, pituitary, adipose tissue, testes, ovaries, and skin. • COMPONENTS : • 1) Renin and Prorenin receptor • 2) Serine proteases, including several kallikrein-like enzymes (tonins), cathepsin G, and chymase are thought to contribute to Ang II • 3) Non-ACE pathways- chymase is the dominant Ang II-generating pathway in the human heart, coronary arteries, and atherosclerotic aorta in vitro • 4)ACE 2 ,Mas receptor,AT2 , AT4 The Renin-AngiotensinAldosterone System: Pathophysiological Role and Pharmacologic Inhibition Vol. 13, No. 8, S-b October 2007 JMCP

44. LOCAL RAS

45. CARDIAC RAAS 1)RENIN 2)RENIN RECEPTOR 3)ANGIOTENSINOGEN 4)CHYMASE 5)ACE 2 6)AT1& AT2 RECEPTOR 1 MARTIN PAUL et alPhysiology of Local Renin-Angiotensin Systems :Physiology of Local Renin-Angiotensin Systems Physiol Rev 86: 747–803, 2006

46. CARDIAC RAS • Predominant physiological role of the cardiac RAS appears to be the maintenance of an appropriate cellular milieu balancing stimuli inducing and inhibiting cell growth and proliferation as well as mediating adaptive responses to myocardial stress, for example, after myocyte stretch

48. EFFECTS OF CARDIAC RAAS • 1)Ionotropic Effect • Positive ionotropic and chronotropic effects of Ang II on cardiomyocyte 1)INCREASED SYMPATHETIC ACTIVITY 2)INCREASED CA2+ INFLUX • 2)Hypertrophy • Ang II is also known to mediate cell growth and proliferation in cardiac myocytes and fibroblasts, as well as in vascular smooth muscle cells via TGF,PDGF MARTIN PAUL et alPhysiology of Local Renin-Angiotensin Systems :Physiology of Local Renin-Angiotensin Systems Physiol Rev 86: 747–803, 2006

49. 3)Mechanical stretch- stretch can cause release of ANG II causing hypertrophy and remodelling • 4)Cardiac remodelling – stretch causing fibroblast activation through AT1 receptor. • Increased activity of the system has also been linked to changes in the electrical physiology that lead to arrhythmias both in the ventricle and atria • 5) Apoptosis – remodelling during M.I , DCMPY etc MARTIN PAUL et alPhysiology of Local Renin-Angiotensin Systems :Physiology of Local Renin-Angiotensin Systems Physiol Rev 86: 747–803, 2006

50. VASCULATURE RAS • 1) Vasoconstriction • 2) Oxygen free radicals causing endothelial dysfunction • The production of ROS by NAD(P)H oxidase in reponse to ANG II stimulation in endothelial and vascular smooth muscle cells activates signal pathways such as MAP kinases, tyrosine kinases may lead to inflammation , artherosclerosis, hypertrophy • 3)Angiogenesis MARTIN PAUL et alPhysiology of Local Renin-Angiotensin Systems :Physiology of Local Renin-Angiotensin Systems Physiol Rev 86: 747–803, 2006