Collecting Duct Renin Research Articles

Collecting Duct Pro(Renin) Receptor Contributes to Unilateral Ureteral Obstruction-Induced Kidney Injury via Activation of the Intrarenal RAS.

Although the concept of the intrarenal renin-angiotensin system (RAS) in renal disease is well-described in the literature, the precise pathogenic role and mechanism of this local system have not been directly assessed in the absence of confounding influence from the systemic RAS. The present study used novel mouse models of collecting duct (CD)-specific deletion of (pro)renin receptor (PRR) or renin together with pharmacological inhibition of soluble PRR production to unravel the precise contribution of the intrarenal RAS to renal injury induced by unilateral ureteral obstruction. We examined the impact of CD-specific deletion of PRR, CD-specific deletion of renin, and S1P (site-1 protease) inhibitor PF429242 treatment on renal fibrosis and inflammation and the indices of the intrarenal RAS in a mouse model of unilateral ureteral obstruction. After 3 days of unilateral ureteral obstruction, the indices of the intrarenal RAS including the renal medullary renin content, activity and mRNA expression, and Ang (angiotensin) II content in obstructed kidneys of floxed mice were all increased. That effect was reversed with CD-specific deletion of PRR, CD-specific deletion of renin, and PF429242 treatment, accompanied by consistent improvement in renal fibrosis and inflammation. On the other hand, renal cortical renin levels were unaffected by unilateral ureteral obstruction, irrespective of the genotype. Similar results were obtained via pharmacological inhibition of S1P, the key protease for the generation of soluble PRR. Our results reveal that PRR-dependent/soluble PRR-dependent activation of CD renin represents a key determinant of the intrarenal RAS and, thus, obstruction-induced renal inflammation and fibrosis.

Abstract P376: Collecting Duct Renin And Its Receptor Contributes To Unilateral Ureteral Obstruction-induced Kidney Injury Via Activation Of The Intrarenal Ras

The collecting duct (CD) is well known for its role in fine-tuning urinary excretion of electrolytes and water. Emerging evidence suggests a potential role played by the CD in immunity and renal injury but the underlying mechanism largely remains elusive. Within the CD, (pro)renin receptor (PRR) via site-1 protease (S1P)-derived soluble PRR interacts with renin controls the intrarenal renin-angiotensin system (RAS) to regulate Na+ and water balance and blood pressure. The present study attempted to employ mouse models of CD-specific deletion of PRR and renin (CD PRR KO and CD renin KO, respectively) and a specific S1P inhibitor PF-429242 (PF) to explore the pathogenic role of this local system in renal injury induced by unilateral ureteral obstruction (UUO). Male 3-4-month-old CD PRR KO and CD renin KO, and their respective floxed controls were subjected to UUO surgery or sham operation for 3 days, followed by analysis of renal fibrosis, inflammation, and indices of the intrarenal RAS. Male 3-4-month-old C57BL/6 mice were subjected to the same surgical procedures and infused with vehicle or PF at 20 mg/kg/day via minipump for 3 days, followed by analysis of the same parameters. 3-day UUO in floxed mice induced renal abundance of fibronectin and α-SMA by 8.6 folds and 5.8 folds, respectively, as assessed by immunoblotting analysis. qRT-PCR detected similar magnitude of increases in mRNA expression of fibronectin, α-SMA, as well as collagen III, collagen I, and TGF-β in the obstructed kidneys. Similar results were obtained by Trichrome Masson staining. In parallel, the obstructed kidney exhibited 4-5-fold increases in renal medullary renin mRNA, renin activity, and active renin content contrasting to unaltered renin levels in the renal cortex. CD-specific deletion of PRR and renin induced comparable attenuation of various indices of renal fibrosis, inflammation, renal medullary renin levels so did PF treatment. UUO induced a 2-fold increase in plasma sPRR, which was completely blocked by PF treatment. Taken together, our results demonstrated that CD PRR/sPRR-dependent activation of renin represents a key determinant of the intrarenal RAS and thus obstruction-induced renal inflammation and fibrosis.

Collecting duct renin regulates potassium homeostasis in mice.

The kaliuretic action of the renin-angiotensin-aldosterone system (RAAS) is well established as highlighted by hyperkalemia side effect of RAAS inhibitors but such action is usually ascribed to systemic RAAS. The present study addresses the involvement of intrarenal RAAS in K+ homeostasis with emphasis on locally generated renin within the collecting duct (CD). Wild-type (Floxed) and CD-specific deletion of renin (CD renin KO) mice were treated for 7 days with a high K+ (HK) diet to investigate the role of CD renin in kaliuresis regulation and further define the underlying mechanism with emphasis on analysis of intrarenal aldosterone biosynthesis. In floxed mice, renin levels were elevated in the renal medulla and urine following a 1-week HK diet, indicating activation of the intrarenal renin. CD renin KO mice had blunted HK-induced intrarenal renin response and developed impaired kaliuresis and elevated plasma K+ level (4.45 ± 0.14 vs. 3.89 ± 0.04 mM, p < 0.01). In parallel, HK-induced intrarenal aldosterone and CYP11B2 expression along with expression of renal outer medullary K+ channel (ROMK), calcium-activated potassium channel subunit alpha-1 (α-BK), α-Na+ -K+ -ATPase, and epithelial sodium channel (β-ENaC and cleaved-γ-ENaC) expression were all significantly blunted in CD renin KO mice in contrast to the unaltered responses of plasma aldosterone and adrenal CYP11B2. Taken together, these results support a kaliuretic action of CD renin during HK intake.

Revisiting the relationship between (Pro)Renin receptor and the intrarenal RAS: focus on the soluble receptor.

The (pro)renin receptor (PRR), also termed as ATPase H+ transporting accessory protein 2 (ATP6AP2), was originally cloned as a specific receptor for prorenin and renin [together called (pro)renin]. Given the wide tissue distribution of PRR, PRR was further postulated to act as a regulator of tissue renin. However, assigning a physiological role of PRR within the renin-angiotensin system (RAS) has been challenging largely due to its pleotropic functions in regulation of embryogenesis, autophagy, and H+ transport. The current review will summarize recent advances in understanding the roles of sPPR within the intrarenal RAS as well as those outside this local system. Site-1 protease (S1P) is a predominant source of sPPR at least in the kidney. So far most of the known physiological functions of PRR including renal handling of electrolytes and fluid and blood pressure are mediated by sPRR. In particular, sPRR serves as a positive regulator of collecting duct renin to activate the intrarenal RAS during water deprivation or angiotensin-II (AngII) infusion. However, PRR/sPRR can act in renin-independent manner under other circumstances. S1P-derived sPRR has emerged as a key regulator of kidney function and blood pressure and its relationship with the intrarenal RAS depends on the physiological context.

α-Ketoglutarate Upregulates Collecting Duct (Pro)renin Receptor Expression, Tubular Angiotensin II Formation, and Na+ Reabsorption During High Glucose Conditions.

Diabetes mellitus (DM) causes high glucose (HG) levels in the plasma and urine. The (pro)renin receptor (PRR) is a key regulator of renal Na+ handling. PRR is expressed in intercalated (IC) cells of the collecting duct (CD) and binds renin to promote angiotensin (Ang) II formation, thereby contributing to Na+ reabsorption. In DM, the Kreb's cycle is in a state of suppression in most tissues. However, in the CD, expression of glucose transporters is augmented, boosting the Kreb's cycle and consequently causing α-ketoglutarate (αKG) accumulation. The αKG receptor 1 (OXGR1) is a Gq-coupled receptor expressed on the apical membrane of IC cells of the CD. We hypothesize that HG causes αKG secretion and activation of OXGR1, which increases PRR expression in CD cells. This effect then promotes intratubular AngII formation and Na+ reabsorption. To test this hypothesis, streptozotocin (STZ)-induced diabetic mice were treated with or without montelukast (ML), an OXGR1 antagonist, for 6 days. STZ mice had higher urinary αKG and PRR expression along with augmented urinary AngII levels and Na+ retention. Treatment with ML prevented all these effects. Similarly, primary cultured inner medullary CD cells treated with HG showed increased PRR expression, while OXGR1 antagonist prevented this effect. αKG increases PRR expression, while treatments with ML, PKC inhibition, or intracellular Ca2+ depletion impair this effect. In silico analysis suggested that αKG binds to mouse OXGR1. These results indicate that HG conditions promote increased levels of intratubular αKG and OXGR1-dependent PRR upregulation, which impact AngII formation and Na+ reabsorption.

Potassium Intake Prevents the Induction of the Renin-Angiotensin System and Increases Medullary ACE2 and COX-2 in the Kidneys of Angiotensin II-Dependent Hypertensive Rats.

In angiotensin II (Ang II)-dependent hypertensive rats there is an increased expression of proximal tubule angiotensinogen (AGT), collecting duct renin and angiotensin converting enzyme (ACE), which contributes to intratubular Ang II formation. Ang II acts on Ang II type 1 receptors promoting sodium retention and vasoconstriction. However concurrently, the ACE2-Ang-(1–7) axis and the expression of kallikrein and medullary prostaglandins counteract the effects of Ang II, promoting natriuresis and vasodilation. Human studies demonstrate that dietary potassium (K+) intake lowers blood pressure. In this report we evaluate the expression of AGT, ACE, medullary prorenin/renin, ACE2, kallikrein and cyclooxygenase-2 (COX-2) in Ang II-infused rats fed with high K+ diet (2%) for 14 days. Dietary K+ enhances diuresis in non-infused and in Ang II-infused rats. The rise in systolic blood pressure in Ang II-infused rats was attenuated by dietary K+. Ang II-infused rats showed increased renal protein levels of AGT, ACE and medullary prorenin and renin. This effect was attenuated in the Ang II + K+ group. Ang II infusion decreased ACE2 compared to the control group; however, K+ diet prevented this effect in the renal medulla. Furthermore, medullary COX-2 was dramatically induced by K+ diet in non-infused and in Ang II infused rats. Dietary K+ greatly increased kallikrein immunostaining in normotensive rats and in Ang II-hypertensive rats. These results indicate that a high K+ diet attenuates Ang II-dependent hypertension by preventing the induction of ACE, AGT and collecting duct renin and by enhancing medullary COX-2 and ACE2 protein expression in the kidney.

Abstract P109: Collecting Duct Renin and Aldosterone Regulate Potassium Homeostasis

The kalliuric action of the renin-angiotensin-aldosterone system (RAAS) is well established as highlighted by hyperkaliemia side effect of RAAS inhibitors but such action is usually ascribed to systemic RAAS. The present study attempted to address the involvement of intrarenal RAAS in K + homeostasis with emphasis on locally generated renin and aldosterone (Aldo) within the collecting duct (CD). In normal C57BL/6 mice, a 1-wk high K + (HK) intake (5% KCl in diet) induced parallel increases in renal prorenin (cortex: 298.8%; medulla: 323.8%) assessed by immunoblotting, and urinary prorenin/renin content (URC) (2.5-fold) assessed by ELISA, and urinary renin activity (URA) (1.5-fold), contrasting to suppressed plasma prorenin/renin concentration (by 25.6%) and renin activity (by 61.6%). Following 1-wk HK loading, mice lacking renin in the CD (CD renin KO) had decreased urinary K + excretion (by 20.2%) and elevated plasma K + level (KO+HK: 4.35 ± 0.14 vs. Floxed+HK: 3.89 ± 0.04 mM, P &lt;0.01), accompanied with a reduction of URC (by 40%), URA (by 58.5%), urinary free and total Aldo excretion (by 50.6% and 36.6%, respectively), and kidney cortical and medullary Aldo (by 33.5% and 49.9%, respectively), without affecting plasma Aldo or renin levels. HK upregulated renal protein expression of Aldo synthase CYP11B2 (cortical: 133.4%, medullary: 221.5%), renal outer medullary K + channel (ROMK) (146.4%), calcium-activated potassium channel subunit alpha-1 (α-BK) (160.9%), α-Na + -K + -ATPase (155.8%), β-ENaC (165.1%), and cleaved-γ-ENaC (1800%), all of which were significantly blunted in CD renin KO mice (by 25-75%). We have developed an inducible renal tubule-wide CYP11B2 KO using the Pax8/LC1 transgenes (termed as RT CYP11B2 KO). While the homozygous deletion is lethal, the heterozygous mice exhibited normal development and were subjected to 3-day HK. K + -loaded RT CYP11B2 KO mice had decreased urinary K + excretion (by 30.5%) and hyperkalemia (KO+HK: 4.63 ± 0.05 vs . Floxed+HK: 3.98 ± 0.09 mM, P &lt;0.01), accompanied with a reduction of urinary free and total Aldo excretion (by 20.4% and 34.4%, respectively) without affecting plasma Aldo levels. Taken together, these results support a local action of CD renin and Aldo in regulation of K + homeostasis.

Nephron prorenin receptor deficiency alters renal medullary endothelin-1 and endothelin receptor expression.

The endothelin (ET) and prorenin/renin/prorenin receptor (PRR) systems have opposing physiological effects on collecting duct (CD) salt and water reabsorption. It is unknown if the CD ET and renin/PRR systems interact, hence we examined the effects of deleting CD renin or nephron PRR on CD ET system components. PRR knockout (KO) mice were polyuric and had markedly increased urinary ET-1 and inner medullary CD (IMCD) ET-1 mRNA. PRR KO mice had greatly increased IMCD ETA receptor mRNA and protein, while ETB mRNA and protein were decreased. Water loaded wild-type mice with similar polyuria as PRR KO mice had modestly increased urinary ET-1 excretion and inner medullary ET-1 mRNA, while inner medullary ETA and ETB mRNA or protein expression were unaffected. In contrast to PRR KO, CD prorenin/renin KO did not alter ET system components. Taken together, these results suggest that the nephron PRR is involved in regulating CD ET system expression, but this effect may be independent of CD-derived renin.

PGE2 upregulates renin through E-prostanoid receptor 1 via PKC/cAMP/CREB pathway in M-1 cells.

During the early phase of ANG II-dependent hypertension, tubular PGE2 is increased. Renin synthesis and secretion in the collecting duct (CD) are upregulated by ANG II, contributing to further intratubular ANG II formation. However, what happens first and whether the triggering mechanism is independent of tubular ANG II remain unknown. PGE2 stimulates renin synthesis in juxtaglomerular cells via E-prostanoid (EP) receptors through the cAMP/cAMP-responsive element-binding (CREB) pathway. EP receptors are also expressed in the CD. Here, we tested the hypothesis that renin is upregulated by PGE2 in CD cells. The M-1 CD cell line expressed EP1, EP3, and EP4 but not EP2. Dose-response experiments, in the presence of ANG II type 1 receptor blockade with candesartan, demonstrated that 10-6 M PGE2 maximally increases renin mRNA (approximately 4-fold) and prorenin/renin protein levels (approximately 2-fold). This response was prevented by micromolar doses of SC-19220 (EP1 antagonist), attenuated by the EP4 antagonist, L-161982, and exacerbated by the highly selective EP3 antagonist, L-798106 (~10-fold increase). To evaluate further the signaling pathway involved, we used the PKC inhibitor calphostin C and transfections with PKCα dominant negative. Both strategies blunted the PGE2-induced increases in cAMP levels, CREB phosphorylation, and augmentation of renin. Knockdown of the EP1 receptor and CREB also prevented renin upregulation. These results indicate that PGE2 increases CD renin expression through the EP1 receptor via the PKC/cAMP/CREB pathway. Therefore, we conclude that during the early stages of ANG II-dependent hypertension, there is augmentation of PGE2 that stimulates renin in the CD, resulting in increased tubular ANG II formation and further stimulation of renin.

Role of Collecting Duct Renin in the Pathogenesis of Hypertension.

The presence of renin production by the principal cells of the collecting duct has opened new perspectives for the regulation of intrarenal angiotensin II (Ang II). Angiotensinogen (AGT) and angiotensin-converting enzyme (ACE) are present in the tubular fluid coming from the proximal tubule and collecting duct. All the components needed for Ang II formation are present along the nephron, and much is known about the mechanisms regulating renin in juxtaglomerular cells (JG); however, those in the collecting duct remain unclear. Ang II suppresses renin via protein kinase C (PKC) and calcium (Ca2+) in JG cells, but in the principal cells, Ang II increases renin synthesis and release through a pathophysiological mechanism that increases further intratubular Ang II de novo formation to enhance distal Na + reabsorption. Transgenic mice overexpressing renin in the collecting duct demonstrate the role of collecting duct renin in the development of hypertension. The story became even more interesting after the discovery of a specific receptor for renin and prorenin: the prorenin receptor ((P)RR), which enhances renin activity and fully activates prorenin. The interactions between (P)RR and prorenin/renin may further increase intratubular Ang II levels. In addition to Ang II, other mechanisms have been described in the regulation of renin in the collecting duct, including vasopressin (AVP), bradykinin (BK), and prostaglandins. Current active investigations are aimed at elucidating the mechanisms regulating renin in the distal nephron segments and understand its role in the pathogenesis of hypertension.

Collecting Duct Renin Does Not Mediate DOCA-Salt Hypertension or Renal Injury

Collecting duct (CD)-derived renin is involved in the hypertensive response to chronic angiotensin-II (Ang-II) administration. However, whether CD renin is involved in Ang-II independent hypertension is currently unknown. To begin to examine this, 12 week old male and female CD-specific renin knock out (KO) mice and their littermate controls were subjected to uni-nephrectomy followed by 2 weeks of deoxycorticosterone acetate (DOCA) infusion combined with a high salt diet. Radiotelemetric blood pressure (BP) was similar between KO and control mice at baseline; BP increased in both groups to a similar degree throughout the 2 weeks of DOCA-salt treatment. Urinary albumin excretion and plasma blood urea nitrogen were comparable between the two groups after DOCA-salt treatment. Fibrosis as assessed by Masson’s Trichrome stain/Sirius Red stain and collagen-1 mRNA expression was similar between control and KO mice. Compared to baseline, DOCA-salt treatment decreased plasma renin concentration (PRC), urinary renin excretion and medullary renin mRNA expression in both floxed and CD renin KO mice with no detectable differences between the two groups. Further, in primary culture of rat inner medullary CD, aldosterone treatment did not change renin activity or total renin content. Taken together, these data suggest that CD derived renin does not play a role in DOCA-salt hypertension.

Collecting duct (pro)renin receptor targets ENaC to mediate angiotensin II-induced hypertension.

The (pro)renin receptor (PRR) is abundantly expressed in the collecting duct (CD) and the expression is further induced by angiotensin II (ANG II). The present study was conducted to investigate the role of CD PRR during ANG II-induced hypertension and to further explore the underlying mechanism. Radiotelemetry demonstrated that a 1-wk ANG II infusion gradually and significantly induced hypertensive response in floxed mice and this response was significantly attenuated in mice lacking PRR in the CD (termed CD PRR KO). ANG II infusion in floxed mice increased urinary renin activity and selectively induced renal medullary α-epithelial sodium channel (α-ENaC) mRNA and protein expression, all of which were blunted in the null mice. In cultured mpkCCD cells grown in Transwells, transepithelial Na+ transport as measured by using a volt-ohmmeter was transiently stimulated by acute ANG II treatment, which was abolished by a PRR antagonist, PRO20. In a chronic setting, ANG II treatment induced α-ENaC mRNA expression in mpkCCD cells, which was similarly blocked by PRO20. Chronic intramedullary infusion of an ENaC inhibitor amiloride in rats significantly attenuated ANG II-induced hypertension. Overall, the present study suggests that CD PRR contributes to ANG II-induced hypertension at least partially via activation of renal medullary ENaC.

Lack of collecting duct renin does not protect against DOCA‐salt hypertension and renal injury

Collecting duct (CD)‐derived renin is involved in the hypertensive response to chronic angiotensin‐II (Ang‐II) administration. However, whether CD renin is involved in Ang‐II independent hypertension is currently unknown. To begin to examine this, 12 week old male and female CD‐specific renin knock out (KO) mice and their littermate controls were subjected to uninephrectomy followed by 2 weeks of deoxycorticosterone acetate (DOCA) infusion combined with a high salt diet. Radiotelemetric blood pressure (BP) was similar between KO and control mice at baseline; BP increased in both groups to a similar degree throughout the 2 weeks of DOCA‐salt. Urinary albumin excretion and plasma blood urea nitrogen were comparable between the two groups after DOCA‐salt treatment. Further, renal fibrosis as assessed by Masson's Trichrome stain and Sirius Red stain were similar between control and KO mice. Taken together, these data suggest that CD derived renin does not play a role in DOCA‐salt hypertension. Further studies are needed to examine the role of CD renin in other Ang‐II independent hypertensive models.

Role of the Collecting Duct Renin Angiotensin System in Regulation of Blood Pressure and Renal Function.

Recent evidence suggests that the renal tubular renin angiotensin system regulates urinary Na(+) and water excretion and blood pressure. Three key components of the tubular renin angiotensin system, namely renin, prorenin receptor, and angiotensin-II type 1 receptor, are localized to the collecting duct. This system may modulate collecting duct Na(+) and water reabsorption via angiotensin-II-dependent and angiotensin-II-independent pathways. Further, the system may be of greatest relevance in hypertensive states and particularly those characterized by high circulating angiotensin-II. In this review, we summarize the current knowledge on the synthesis, regulation, and function of collecting duct-derived renin angiotensin system components and examine recent developments with regard to regulation of blood pressure and renal fluid and Na(+) excretion.

Sequential activation of the intrarenal renin-angiotensin system in the progression of hypertensive nephropathy in Goldblatt rats.

The intrarenal renin-angiotensin system (RAS) has an important role in generating and maintaining hypertension in two-kidney, one-clip (2K1C) rats. This study evaluated how various intrarenal RAS components contributed to hypertension not only in the maintenance period (5w; 5 wk after operation) but also earlier (2w; 2 wk after operation). We inserted a 2.5-mm clip into the left renal artery of Sprague-Dawley rats and euthanized them at 2w and 5w following the operation. Systolic blood pressure increased within 1 wk after the operation, and left ventricular hypertrophy occurred in 2K1C rats. At 2w, juxtaglomerular apparatus (JGA) and collecting duct (CD) renin increased in clipped kidney (CK) of 2K1C rats. The tubular angiotensin I-converting enzyme (ACE) was not changed, but peritubular ACE2 decreased in nonclipped kidney (NCK) and CK of 2K1C rats. At 5w, ACE and CD renin were enhanced, and ACE2 was still lessened in both kidneys of 2K1C rats. However, plasma renin activity (PRA) was not different from that in sham rats. In proximal tubules of CK, the ANG II type 1 receptor (AT1R) was not suppressed, but the Mas receptor (MasR) was reduced; thus the AT1R/MasR ratio was elevated. Although hypoxic change in CK could not be excluded, the JGA renin of CK and CD renin in both kidneys was highly expressed independent of time. Peritubular ACE2 changed in the earlier period, and uninhibited AT1R in proximal tubules of CK was presented in the maintenance period. In 2K1C rats, attenuated ACE2 seems to contribute to initiating hypertension while upregulated ACE in combination with unsuppressed AT1R may have a key role in maintaining hypertension.

PKC-α-dependent augmentation of cAMP and CREB phosphorylation mediates the angiotensin II stimulation of renin in the collecting duct.

In contrast to the negative feedback of angiotensin II (ANG II) on juxtaglomerular renin, ANG II stimulates renin in the principal cells of the collecting duct (CD) in rats and mice via ANG II type 1 (AT1R) receptor, independently of blood pressure. In vitro data indicate that CD renin is augmented by AT1R activation through protein kinase C (PKC), but the exact mechanisms are unknown. We hypothesize that ANG II stimulates CD renin synthesis through AT1R via PKC and the subsequent activation of cAMP/PKA/CREB pathway. In M-1 cells, ANG II increased cAMP, renin mRNA (3.5-fold), prorenin, and renin proteins, as well as renin activity in culture media (2-fold). These effects were prevented by PKC inhibition with calphostin C, PKC-α dominant negative, and by PKA inhibition. Forskolin-induced increases in cAMP and renin expression were prevented by calphostin C. PKC inhibition and Ca2+ depletion impaired ANG II-mediated CREB phosphorylation and upregulation of renin. Adenylate cyclase 6 (AC) siRNA remarkably attenuated the ANG II-dependent upregulation of renin mRNA. Physiological activation of AC with vasopressin increased renin expression in M-1 cells. The results suggest that the ANG II-dependent upregulation of renin in the CD depends on PKC-α, which allows the augmentation of cAMP production and activation of PKA/CREB pathway via AC6. This study defines the intracellular signaling pathway involved in the ANG II-mediated stimulation of renin in the CD. This is a novel mechanism responsible for the regulation of local renin-angiotensin system in the distal nephron.

Roles of collecting duct renin and (pro)renin receptor in hypertension: mini review.

In angiotensin (Ang)-II-dependent hypertension, collecting duct renin synthesis and secretion are stimulated despite suppression of juxtaglomerular (JG) renin. This effect is mediated by Ang II type 1 (AT1) receptor independent of blood pressure. Although the regulation of JG renin is known, the mechanisms by which renin is regulated in the collecting duct are not completely understood. The presence of renin activity in the collecting duct may provide a pathway for intratubular Ang II formation since angiotensinogen substrate and angiotensin converting enzyme are present in the distal nephron. The recently named new member of the renin-angiotensin system (RAS), the (pro)renin receptor [(P)RR], is able to bind renin and the inactive prorenin, thus enhancing renin activity and fully activating prorenin. We have demonstrated that renin and (P)RR are augmented in renal tissues from rats infused with Ang II and during sodium depletion, suggesting a physiological role in intrarenal RAS activation. Importantly, (P)RR activation also causes activation of intracellular pathways associated with increased cyclooxygenase 2 expression and induction of profibrotic genes. In addition, renin and (P)RR are upregulated by Ang II in collecting duct cells. Although the mechanisms involved in their regulation are still under study, they seem to be dependent on the intrarenal RAS activation. The complexities of the mechanisms of stimulation also depend on cyclooxygenase 2 and sodium depletion. Our data suggest that renin and (P)RR can interact to increase intratubular Ang II formation and the activation of profibrotic genes in renal collecting duct cells. Both pathways may have a critical role in the development of hypertension and renal disease.

Renin and the (pro)renin receptor in the renal collecting duct: Role in the pathogenesis of hypertension.

The intrarenal renin-angiotensin system (RAS) plays a critical role in the pathogenesis and progression of hypertension and kidney disease. In angiotensin (Ang) II-dependent hypertension, collecting duct renin synthesis and secretion are stimulated despite suppression of juxtaglomerular (JG) renin. This effect is mediated by the AngII type I receptor (AT1 R), independent of blood pressure. Although the regulation of JG renin has been extensively studied, the mechanisms by which renin is regulated in the collecting duct remain unclear. The augmentation of renin synthesis and activity in the collecting duct may provide a pathway for additional generation of intrarenal and intratubular AngII formation due to the presence of angiotensinogen substrate and angiotensin-converting enzyme in the nephron. The recently described (pro)renin receptor ((P)RR) binds renin or prorenin, enhancing renin activity and fully activating the biologically inactive prorenin peptide. Stimulation of (P)RR also activates intracellular pathways related to fibrosis. Renin and the (P)RR are augmented in renal tissues of AngII-dependent hypertensive rats. However, the functional contribution of the (P)RR to enhanced renin activity in the collecting duct and its contribution to the development of hypertension and kidney disease have not been well elucidated. This review focuses on recent evidence demonstrating the mechanism of renin regulation in the collecting ducts and its interaction with the (P)RR. The data suggest that renin-(P)RR interactions may induce stimulation of intracellular pathways associated with the development of hypertension and kidney disease.

Abstract 475: Vasopressin Type 2 Receptor (v2r) Activation Increases Renin Expression In Renal Collecting Duct Cells.

Renin synthesis in juxtaglomerular cells (JGC) is mediated by intracellular cAMP accumulation and activation of PKA/CREB pathway. Recent evidence demonstrated that renin is expressed in renal collecting duct (CD) cells. Furthermore, it has been shown that CD renin is augmented in animal models of hypertension and kidney disease, despite the suppressed expression observed in JGC. Vasopressin activates V2R stimulating cAMP/PKA/CREB pathway and aquaporin-2 expression in apical plasma membrane of principal cells of the CD. We hypothesized that activation of V2R increases renin expression in mouse CD cell line M-1 through cAMP/PKA/CREB pathway. Desmopressin (ddAVP, 10-6 mol/L, 6 hrs), a specific V2R agonist, increased renin mRNA, prorenin protein levels in cell lysates and prorenin secretion to the culture media. To determine if this effect was related to PKA pathway, we used the PKA inhibitor H89. Co-treatment with ddAVP + H89 prevented the ddAVP-mediated increase in renin expression. To further confirm if the stimulation of renin synthesis in M-1 cells was mediated by cAMP accumulation, we raised intracellular cAMP levels using forskolin (10-7 mol/L). Forskolin treatment significantly increased renin mRNA and prorenin protein levels as compared to controls. Additionally, ddAVP increased phosphorylated CREB, while H89 blunted this effect. Finally, shRNA against CREB prevented the ddAVP-induced renin synthesis. We additionally confirmed the stimulatory effects of Ang II + ddAVP on renin synthesis in mpkccdc14 cell line, a mouse cortical line composed only by CD principal cells. Tolvaptan (V2R antagonist) reduced the additive effect of Ang II + ddAVP on renin expression. To achieve in vivo relevance we further measured renin mRNA levels in renal inner medullary tissues from mice subjected to 16 hours of water deprivation and controls. Mice water-deprived showed significantly greater renin mRNA levels in the renal inner medulla than controls. These results indicate that the activation of V2R stimulates renin mRNA synthesis and prorenin secretion in M-1 cells via cAMP/PKA/CREB pathway.

Collecting duct-specific knockout of renin attenuates angiotensin II-induced hypertension.

The physiological and pathophysiological significance of collecting duct (CD)-derived renin, particularly as it relates to blood pressure (BP) regulation, is unknown. To address this question, we generated CD-specific renin knockout (KO) mice and examined BP and renal salt and water excretion. Mice containing loxP-flanked exon 1 of the renin gene were crossed with mice transgenic for aquaporin-2-Cre recombinase to achieve CD-specific renin KO. Compared with controls, CD renin KO mice had 70% lower medullary renin mRNA and 90% lower renin mRNA in microdissected cortical CD. Urinary renin levels were significantly lower in KO mice (45% of control levels) while plasma renin concentration was significantly higher in KO mice (63% higher than controls) during normal-Na intake. While no observable differences were noted in BP between the two groups with varying Na intake, infusion of angiotensin II at 400 ng·kg(-1)·min(-1) resulted in an attenuated hypertensive response in the KO mice (mean arterial pressure 111 ± 4 mmHg in KO vs. 128 ± 3 mmHg in controls). Urinary renin excretion and epithelial Na(+) channel (ENaC) remained significantly lower in the KO mice following ANG II infusion compared with controls. Furthermore, membrane-associated ENaC protein levels were significantly lower in KO mice following ANG II infusion. These findings suggest that CD renin modulates BP in ANG II-infused hypertension and these effects are associated with changes in ENaC expression.