Handle Region Peptide Research Articles

Cardiovascular effect of preeclampsia upon offspring development: Are (Pro) renin-renin receptor ((P)RR) and gender related?

Preeclampsia (PE) is a complication of pregnancy that might increase progeny risk of cardiovascular and metabolic problems, mainly in males. Renin angiotensin aldosterone system is known to be involved. (Pro) renin/renin receptor ((P)RR) has been shown to participate in cardiovascular pathology. The aim of this work was to evaluate (P)RR expression and function upon cardiovascular and renal tissues from PE dams' offspring. We used offspring from normal pregnant and preeclamptic rats, evaluating body, heart, aorta and kidney weight, length, and blood pressure along 3 months after birth. Subsets of animals received handle region peptide (HRP) (0.2 mg/Kg, sc). Another group received vehicle. Animals were sacrificed at first, second, and third months of age, tissues were extracted and processed for immunoblot to detect (P)RR, PLZF, β-catenin, DVL-1, and PKCα. (P)RR and PLZF were also measured by RT-PCR. We found that offspring developed hypertension. Male descendants remained hypertensive throughout the whole experiment. Female animals tended to recover at second month and returned to normal blood pressure at third month. HRP treatment diminished hypertension in both male and female animals. Morphological evaluations showed changes in heart, aorta, and kidney weight, and HRP reverted this effect. Finally, we found that (P)RR, PLZF, and canonical WNT transduction pathway molecules were stimulated by PE, and HRP treatment abolished this increase. These findings suggest that PE can induce hypertension in offspring, and (P)RR seems to play an important role through the canonical WNT pathway and that gender seems to influence this response.

(Pro)renin receptor inhibition attenuated liver steatosis, inflammation, and fibrosis in mice with steatohepatitis.

The (Pro)renin receptor (PRR) is reportedly involved in hepatic lipid metabolism and hepatocyte PRR knockdown protects mice against hepatosteatosis. However, the impact of PRR inhibition on liver inflammation and fibrosis in nonalcoholic steatohepatitis (NASH) remains unclear. Herein, C57BL/6 mice were fed a normal chow diet or fast food diet (FFD) for 24 weeks. Lentivirus-mediated PRR short hairpin RNA (shRNA) or handle region peptide (HRP), a PRR blocker, was administered for PRR inhibition. Mouse primary hepatocytes were cultured with palmitic acid, prorenin, siRNA-targeted PRR, and HRP. In FFD-fed mice, PRR inhibition via lentivirus-mediated PRR knockdown or HRP significantly attenuated liver steatosis, inflammation, and fibrosis. Mechanistically, PRR knockdown or HRP decreased hepatic acetyl-CoA carboxylase (ACC) abundance and upregulated peroxisome proliferator-activated receptor-alpha (PPARα). HRP treatment also decreased hepatic PRR expression. In addition, intrahepatic oxidative stress, apoptosis and inflammatory cell recruitment were ameliorated by PRR knockdown or HRP treatment, along with suppression of proinflammatory cytokine expression. PRR inhibition downregulated the hepatic expression of profibrotic factors, as well as TGF-β1/SMAD3 pathway. In primary mouse hepatocytes, PRR knockdown with siRNA or HRP downregulated cellular ACC and increased PPARα expression. In conclusion, our findings revealed that PRR inhibition attenuated hepatic steatosis, inflammation, and fibrosis in mice with NASH. Accordingly, targeting PRR signaling may serve as a potential treatment for NASH.

Effect of HRP Upon the Expression and Intracellular Signaling of PRR in Cardiovascular and Renal Systems in Preeclamptic Rats Offspring

Introduction Preeclampsia (PE) is a condition that affects both, the mother and the child, conditioning in both a greater susceptibility to develop cardiovascular diseases in future stages. It has been studied that prorenin renin receptor (PRR) has a broad participation in the cardiovascular system of PE products, it seems to play an important role in the development of future conditions, such as arterial hypertension, involving the renin angiotensin aldosterone system (RAS). RAS hyperactivity has been implicated as a cardiovascular risk factor in patients who come from a pregnancy complicated by preeclampsia. Mechanisms mediating fetal programming of adult arterial hypertension are numerous, including vascular dysfunction and reduction of the density of the arterioles and capillaries. Handle Region Peptide (HRP) is a decoy peptide that can inhibit the binding of prorrenin to the PRR receptor. HRP mimics the loop region and therefore competitively binds to the receptor, thus preventing receptor-mediated activation of prorenin and reducing its activity in tissue. Objective To evaluate the HRP blockade upon the PRR receptor and its transduction pathways: WNT and MAPK in cardiovascular and renal tissue in offspring from a pre-eclampsia model. Material and methods Male and female offspring from PE and healthy pregnant wistar rats, with 30 and 120 days of age, where set up in groups with or without HRP treatment (0.2 mg/Kg, subcutaneously for 7 days before sacrifice). We measured blood pressure by tail-cuff method. At the end of the treatment the expression of PRR, DVL1, beta-catenin, PKC-alpha were measured by immunoblot and RT-PCR in cardiovascular and renal tissue. Results Offspring from PE rats showed sustained high blood pressure, both male and female at 30 and 120 days. HRP treatment reverted partially this increase. All PE-derived groups showed an increase in PRR expression in aorta, heart and kidney. WNT and MAPK pathways also seemed to be activated in PE-derived animals, as shown by β-catenin, DVL-1 and PKCa increased expression. These findings are important because WNT signaling pathway is associated with kidney diseases and cardiac hypertrophy. Besides, this work shows that PRR participates in high blood pressure development in offspring derived from PE pregnancies.

Antiproliferative Effects of Monoclonal Antibodies against (Pro)Renin Receptor in Pancreatic Ductal Adenocarcinoma

We previously reported that silencing of the PRR gene, which encodes the (pro)renin receptor [(P)RR], significantly reduced Wnt/β-catenin-dependent development of pancreatic ductal adenocarcinoma (PDAC). Here, we examined the effects of a panel of blocking mAbs directed against the (P)RR extracellular domain on proliferation of the human PDAC cell lines PK-1 and PANC-1 in vitro and in vivo We observed that four rat anti-(P)RR mAbs induced accumulation of cells in the G0-G1-phase of the cell cycle and significantly reduced proliferation in vitro concomitant with an attenuation of Wnt/β-catenin signaling. Systemic administration of the anti-(P)RR mAbs to nude mice bearing subcutaneous PK-1 xenografts significantly decreased tumor expression of active β-catenin and the proliferation marker Ki-67, and reduced tumor growth. In contrast, treatment with the handle region peptide of (pro)renin did not inhibit tumor growth in vitro or in vivo, indicating that the effects of the anti-(P)RR mAbs were independent of the renin-angiotensin system. These data indicate that mAbs against human (P)RR can suppress PDAC cell proliferation by hindering activation of the Wnt/β-catenin signaling pathway. Thus, mAb-mediated (P)RR blockade could be an attractive therapeutic strategy for PDAC.

Abstract P2015: Antiproliferative Effects of Monoclonal Antibodies Against (Pro)Renin Receptor in Pancreatic Ductal Adenocarcinoma

We previously reportedthat silencing of the PRR gene, which encodes the(pro)renin receptor ((P)RR) significantly reduced Wnt/β-catenin-dependent development of pancreatic ductal adenocarcinoma (PDAC). Here, we examined the effects of a panel of blocking monoclonal antibodies (mAbs) directed against the (P)RR extracellular domain on proliferation of the human PDAC cell lines PK-1 and PANC-1 in vitro andin vivo. We observed that four rat anti-(P)RR mAbs induced accumulation of cells in the G0/G1 phase of the cell cycle and significantly reduced proliferation in vitroconcomitant with a significant reduction in the expression of active β-catenin and cyclin D1. Systemic administration of the anti-(P)RR mAbs to nude mice bearing subcutaneous PK-1 xenografts significantly decreased tumor expression of active β-catenin and the proliferation marker Ki-67 and reduced tumor growth. In contreffects ast, treatment with the handle region peptide of (pro)renin did not inhibit tumor growth in vitro orin vivo, indicating that the effects of the anti-(P)RR mAbs was independent of the renin-angiotensin system. These data indicatethat mAbs against human (P)RR can suppress PDAC cells proliferation by hindering activation of the Wnt/β-catenin signaling pathway. Thus, mAb-mediated (P)RR blockade could be an attractive therapeutic strategy for PDAC.

(Pro)renin Receptor Blockade Ameliorates Heart Failure Caused by Chronic Kidney Disease

The (pro)renin receptor [(P)RR)] is involved in the activation of local renin-angiotensin system and subsequent development of cardiovascular disease. We investigated the therapeutic effect of a (P)RR blocker, handle-region peptide (HRP), on chronic kidney disease (CKD)-associated heart failure. CKD was induced in C57BL/6J mice by means of five-sixths nephrectomy. Eight weeks later, cardiac dysfunction and cardiac dilatation with hypertension developed. Mice were then assigned to 1 of the 3 following groups: vehicle, low-dose (0.01 mg·kg-1·d-1) HRP, or high-dose (0.3 mg·kg-1·d-1) HRP for 4 weeks. High-dose HRP treatment reversed left ventricular dilation and significantly improved cardiac dysfunction with ameliorated hypertension compared with the vehicle. The hearts with high-dose HRP treatment showed significant attenuation of cardiac fibrosis, cardiomyocyte hypertrophy, macrophage infiltration, and oxidative DNA damage. This treatment decreased the myocardial expressions of angiotensin (Ang) II, Ang II type 1 receptor, transforming growth factor β1, extracellular matrix-related proteins, and lipid peroxidation. Autophagy was activated in the cardiomyocyte from nephrectomized mice, but HRP treatment had no effect on cardiomyocyte autophagy. This study indicates that (P)PR blockade is a beneficial strategy by suppressing cardiac fibrosis and hypertrophy to ameliorate heart failure caused by CKD.

Aortic constriction induces hypertension and cardiac hypertrophy via (pro)renin receptor activation and the PLC‑β3 signaling pathway.

The (pro)renin receptor [(P)RR] serves an important role in cardiovascular complications. However, the precise mechanisms of (P)RR in the heart remain obscure. The authors hypothesized that overexpression of (P)RR would be associated with activation of the relevant signal pathway which could lead to organ injury. The aim of the present study was to test the role of cardiac (P)RR and its potential signaling pathway components including phospholipaseC(PLC), protein kinaseC(PKC), extracellular signal‑regulated kinase(ERK)1/2 and Raf‑1 proto‑oncogene, serine/threonine kinase(Raf‑1). Hypertension and cardiac hypertrophy were induced by partial abdominal aortic ligation in Sprague‑Dawley rats. The expression levels of cardiac (P)RR, PLC‑β3, PKC, ERK1/2 and Raf‑1 were measured following administration of the handle region peptide (HRP) and PLC‑β3 inhibitor U73122. The expression of (P)RR and PLC‑β3 significantly increased in the left ventricle (P<0.05). Levels of PKC‑α, ERK1/2 and Raf‑1 in the heart rose significantly (P<0.05). HRP and U73122 significantly decreased the levels of cardiac (P)RR and PLC‑β3. Furthermore, levels of PKC‑α, ERK1/2 and Raf‑1 were also decreased (P<0.05). Cardiac parameters, blood pressure and plasma Angiotensin(Ang)I and AngII levels were altered significantly (P<0.05). The results demonstrated that hypertension induced by aortic restriction activated the (P)RR in the heart. This action led to hypertension and cardiac hypertrophy via the (P)RR‑PLC‑β3‑PKC‑ERK1/2‑Raf‑1 signaling pathway. These results provide a mechanism by which elevated (P)RR levels in hypertension may contribute to the development of cardiac remodeling.

Inhibition of (pro)renin Receptor Contributes to Renoprotective Effects of Angiotensin II Type 1 Receptor Blockade in Diabetic Nephropathy.

Aims: Renal renin-angiotensin system (RAS) plays a pivotal role in the development of diabetic nephropathy (DN). Angiotensin II (Ang II) type 1 receptor (AT1R) blockade elevates (pro)renin, which may bind to (pro)renin receptor (PRR) and exert receptor-mediated, angiotensin-independent profibrotic effects. We therefore investigated whether PRR activation leads to the limited anti-fibrotic effects of AT1R blockade on DN, and whether PRR inhibition might ameliorate progression of DN.Methods: To address the issue, the expression of RAS components was tested in different stages of streptozotocin (STZ)-induced diabetic rats (6, 12, and 24 weeks) and 6-week AT1R blockade (losartan) treated diabetic rats. Using the blocker for PRR, the handle region peptide (HRP) of prorenin, the effects of PRR on high glucose or Ang II-induced proliferative and profibrotic actions were evaluated by measurement of cell proliferation, matrix metalloproteinase-2 (MMP-2) activity, activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and transforming growth factor-β1 (TGF-β1) expression in rat mesangial cells (MCs).Results: PRR was downregulated in the kidneys of different stages of diabetic rats (6, 12, and 24 weeks). Moreover, 6-week losartan treatment further suppressed PRR expression via upregulating AT2R, and ameliorated diabetic renal injury. HRP inhibited high glucose and Ang II-induced proliferative and profibrotic effects in MCs through suppressing TGF-β1 expression and activating MMP-2. Meanwhile, HRP enhanced losartan's anti-fibrotic effects through further inhibiting phosphorylation of ERK1/2 and TGF-β1 expression. Moreover, the inhibitive effect of HRP on Ang II-induced TGF-β1 expression depended on the regulation of PRR expression by AT2R.Conclusions: Our findings suggest that inhibition of PRR contributes to renoprotection against diabetic nephropathy by AT1R blockade.

Shenfuqiangxin Capsule inhibits apoptosis through mitogen-activated protein kinase signal pathway in rats with cardio-renal syndrome induced by infrarenal aortic-clamping

OBJECTIVETo investigate the effect of Shenfuqiangxin capsule and the underlying mechanism on cardio-renal syndrome (CRS) in rats induced by infrarenal aortic-clamping after renal ischaemia. METHODSMale Wistar rats underwent infrarenal aortic-clamping after renal ischaemia or sham operation. The surviving CRS rats were divided randomly into three groups: CRS group (CRS + 10 mL · kg−1 · d−1 pure water by gavage), SFQX group (CRS +13.2 g crude drug · kg−1 · d−1 Shenfuqiangxin by gavage), and handle region peptide (HRP) group (CRS + 10mg/kg HRP by vein). Sham operation rats were given 10 mL · kg−1 · d−1 pure water. Treatments were given 8 weeks after surgery, which lasted for 4 weeks. The rats were detected for heart structure and function by transthoracic echocardiography. PPR mRNA was detected by Reverse Transcription Polymerase Chain Reaction (RT-PCR). To determine whether the mitogen-activated protein kinase (MAPK) signal pathway is included in the heart and kidney protective function of Shenfuqiangxin capsule, MAPK related proteins such as posophorylated C-Jun amino terminal kinase (p-JNK), posophorylated extracellular signal-regulated kinase ½ (p-ERK1/2), posophorylated p38 (p-p38) were examined by Western Blot. Apoptosis in heart and kidney tissues were detected by dUTP Nick End Labeling staining. RESULTSShenfuqiangxin capsule alleviated myocardial apoptosis and inhibited PRR mRNA expression and p-JNK, p-ERK1/2, p-p38 proteins expression in CRS rats. CONCLUSIONAll the results suggest that Shenfuqiangxin capsule improves the injured heart and kidney function maybe through inhibition of MAPK response pathway.

OS 15-01 ASSOCIATION OF CARDIAC (P) RR AND PLC-b3 EXPRESSION IN RATS WITH AORTIC CONSTRICTION

Objective: Recently, some findings support the (pro) renin-(Pro) renin receptor (P) RR interaction at exceptionally high (pro) renin levels in vitro. The aim of this study was to investigate the roles of cardiac (P) RR and its downward signals on mean arterial pressure (MAP) in rats with abdominal aortic constriction, following treatment of handle region peptide (HRP) and phospholipase C-β3 (PLC-b3) inhibitor, U73122. Design and Method: Seventy-five SD rats were divided into 5 groups (n=15 each group) as following: sham operated (SO), rats with the aortic ligation (AL), AL rats were given HRP (4 μg kg−1 d−1, SC), AL rats given U73122 (40 μg kg−1 d−1, SC) and AL rats given HRP+U73122. MAP was recorded using a tail-cuff method. After 4 weeks of treatment, levels of (P) RR, PLC-b3, PKC-a and ERK1/2 in the heart were examined by RT-PCR and western blot. Results: The levels of (P) RR and PLC-b3 significantly increased in the left ventricle in hypertensive rats (P < 0.01), respectively. MAP rose markedly (P < 0.01). HRP decreased the level of (P) RR and U73122 suppressed PLC-b3 expression. Treatment of both HRP and U73122 significantly decreased levels of PKC-a, ERK1/2 and Raf-1 in the heart (P < 0.001). Meanwhile, MAP was decreased after treatment. Conclusions: This study demonstrates that (P) RR inhibitor, HRP and PLC-b3 inhibitor, U73122 decreased levels of (P) RR, PLC-b3, PKC-a, ERK1/2 and Raf-1 in the heart. Meanwhile, administration of both reagents lowered MAP. These findings indicate that cardiac (P) RR may activate PLC-b3, PKC, ERK1/2 and Raf-1 signals and lead to hypertension. Acknowledgement: This study was supported by the Natural Science Foundation of China (No. 81270336).

Adipose tissue (P)RR regulates insulin sensitivity, fat mass and body weight

ObjectiveWe previously demonstrated that the handle-region peptide, a prorenin/renin receptor [(P)RR] blocker, reduces body weight and fat mass and may improve insulin sensitivity in high-fat fed mice. We hypothesized that knocking out the adipose tissue (P)RR gene would prevent weight gain and insulin resistance. MethodsAn adipose tissue-specific (P)RR knockout (KO) mouse was created by Cre-loxP technology using AP2-Cre recombinase mice. Because the (P)RR gene is located on the X chromosome, hemizygous males were complete KO and had a more pronounced phenotype on a normal diet (ND) diet compared to heterozygous KO females. Therefore, we challenged the female mice with a high-fat diet (HFD) to uncover certain phenotypes. Mice were maintained on either diet for 9 weeks. ResultsKO mice had lower body weights compared to wild-types (WT). Only hemizygous male KO mice presented with lower total fat mass, higher total lean mass as well as smaller adipocytes compared to WT mice. Although food intake was similar between genotypes, locomotor activity during the active period was increased in both male and female KO mice. Interestingly, only male KO mice had increased O2 consumption and CO2 production during the entire 24-hour period, suggesting an increased basal metabolic rate. Although glycemia during a glucose tolerance test was similar, KO males as well as HFD-fed females had lower plasma insulin and C-peptide levels compared to WT mice, suggesting improved insulin sensitivity. Remarkably, all KO animals exhibited higher circulating adiponectin levels, suggesting that this phenotype can occur even in the absence of a significant reduction in adipose tissue weight, as observed in females and, thus, may be a specific effect related to the (P)RR. Conclusions(P)RR may be an important therapeutic target for the treatment of obesity and its associated complications such as type 2 diabetes.

Prorenin/renin receptor blockade promotes a healthy fat distribution in obese mice.

Administration of the handle region peptide (HRP), a (pro)renin receptor blocker, decreases body weight gain and visceral adipose tissue (VAT) in high-fat/high-carbohydrate (HF/HC) diet-fed mice. The objective of this study was to elucidate potential mechanisms implicated in these observations. Mice were given a normal or a HF/HC diet along with saline or HRP for 10 weeks. In HF/HC-fed mice, HRP increased the expression of several enzymes implicated in lipogenesis and lipolysis in subcutaneous fat (SCF) while the expression of the enzyme implicated in the last step of lipogenesis decreased in VAT. A reduction was also observed in circulating free fatty acids in these animals which was accompanied by normalized adipocyte size in VAT and increased adipocyte size in SCF. ''Beiging'' is the evolution of a white adipose tissue toward a brown-like phenotype characterized by an increased mitochondrial density and small lipid droplets. HRP increased the expression of' "beiging" markers in SCF of HF/HC diet-fed mice. HRP treatment may favor healthy fat storage in SCF by activating a triglyceride/free fatty acid cycling and "beiging," which could explain the body weight and fat mass reduction.

The critical role of the central nervous system (pro)renin receptor in regulating systemic blood pressure.

The systemic renin-angiotensin system (RAS) has long been recognized as a critically important system in blood pressure (BP) regulation. However, extensive evidence has shown that a majority of RAS components are also present in many tissues and play indispensable roles in BP regulation. Here, we review evidence that RAS components, notably including the newly identified (pro)renin receptor (PRR), are present in the brain and are essential for the central regulation of BP. Binding of the PRR to its ligand, prorenin or renin, increases BP and promotes progression of cardiovascular diseases in an angiotensin II-dependent and -independent manner, establishing the PRR a promising antihypertensive drug target. We also review the existing PRR blockers, including handle region peptide and PRO20, and propose a rationale for blocking prorenin/PRR activation as a therapeutic approach that does not affect the actions of the PRR in vacuolar H(+)-ATPase and development. Finally, we summarize categories of currently available antihypertensive drugs and consider future perspectives.

Activation of the (pro)renin receptor in the paraventricular nucleus increases sympathetic outflow in anesthetized rats.

Previous studies have indicated that hyperactivity of brain prorenin receptors (PRR) is implicated in neurogenic hypertension. However, the role of brain PRR in regulating arterial blood pressure (ABP) is not well understood. Here, we test the hypothesis that PRR activation in the hypothalamic paraventricular nucleus (PVN) contributes to increased sympathetic nerve activity (SNA). In anaesthetized adult Sprague-Dawley (SD) rats, bilateral PVN microinjection of human prorenin (2 pmol/side) significantly increased splanchnic SNA (SSNA; 71 ± 15%, n = 7). Preinjection of either prorenin handle region peptide, the PRR binding blocker (PRRB), or tiron (2 nmol/side), the scavenger of reactive oxygen species (ROS), significantly attenuated the increase in SSNA (PRRB: 32 ± 5% vs. control, n = 6; tiron: 8 ± 10% vs. control, n = 5; P < 0.05) evoked by prorenin injection. We further investigated the effects of PRR activation on ROS production as well as downstream gene expression using cultured hypothalamus neurons from newborn SD rats. Incubation of brain neurons with human prorenin (100 nM) dramatically enhanced ROS production and induced a time-dependent increase in mRNA levels of inducible nitric oxide synthase (iNOS), NAPDH oxidase 2 subunit cybb, and FOS-like antigen 1 (fosl1), a marker for neuronal activation and a component of transcription factor activator protein-1 (AP-1). The maximum mRNA increase in these genes occurred 6 h following incubation (iNOS: 201-fold; cybb: 2 -fold; Ffosl1: 11-fold). The increases in iNOS and cybb mRNA were not attenuated by the AT1 receptor antagonist losartan but abolished by the AP-1 blocker curcumin. Our results suggest that PVN PRR activation induces sympathoexcitation possibly through stimulation of an ANG II-independent, ROS-AP-1-iNOS signaling pathway.

The Role of the (Pro)renin Receptor in Hypertensive Disease.

Tissue angiotensin generation depends on the uptake of circulating (kidney-derived) renin and/or its precursor prorenin (together denoted as (pro)renin). Since tissue renin levels are usually higher than expected based upon the amount of (renin-containing) blood in tissue, an active uptake mechanism has been proposed. The (pro)renin receptor ((P)RR), discovered in 2002, appeared a promising candidate, although its nanomolar affinity for renin/prorenin is many orders of magnitude above their levels in blood. This review discusses (P)RR-related research since its discovery. First, encouraging in vitro findings supported detrimental effects of (pro)renin-(P)RR interaction, even resulting in angiotensin-independent signaling. Moreover, the putative (P)RR blocker "handle region peptide" (HRP) yielded beneficial effects in various cardiovascular animal models. Then doubt arose whether such interaction truly occurs in vivo, and (P)RR deletion unexpectedly turned out to be lethal. Moreover, HRP results could not be confirmed. Finally, it was discovered that the (P)RR actually is a component of vacuolar-type H(+)-ATPase, a multisubunit protein found in virtually every cell type which is essential for vesicle trafficking, protein degradation, and coupled transport. Nevertheless, selective (P)RR blockade in the brain with the putative antagonist PRO20 (corresponding with the first 20 amino acids of prorenin's prosegment) reduced blood pressure in the deoxycorticosteroneacetate (DOCA)-salt model, and (P)RR gene single nucleotide polymorphisms associate with hypertension. To what degree this relates to (pro)renin remains uncertain. The concept of (P)RR blockade in hypertension, if pursued, requires rigorous testing of any newly designed antagonist, and may not hold promise given the early death of tissue-specific (P)RR knockout animals.

(Pro)renin receptor in skeletal muscle is involved in the development of insulin resistance associated with postinfarct heart failure in mice.

We previously reported that insulin resistance was induced by the impairment of insulin signaling in the skeletal muscle from heart failure (HF) via NAD(P)H oxidase-dependent oxidative stress. (Pro)renin receptor [(P)RR] is involved in the activation of local renin-angiotensin system and subsequent oxidative stress. We thus examined whether (P)RR inhibitor, handle region peptide (HRP), could ameliorate insulin resistance in HF after myocardial infarction (MI) by improving oxidative stress and insulin signaling in the skeletal muscle. C57BL6J mice were divided into four groups: sham operated (Sham, n = 10), Sham treated with HRP (Sham+HRP, 0.1 mg·kg(-1)·day(-1), n = 10), MI operated (MI, n = 10), and MI treated with HRP (MI+HRP, 0.1 mg/kg/day, n = 10). After 4 wk, MI mice showed left ventricular dysfunction, which was not affected by HRP. (P)RR was upregulated in the skeletal muscle after MI (149% of sham, P < 0.05). The decrease in plasma glucose after insulin load was smaller in MI than in Sham (21 ± 2 vs. 44 ± 3%, P < 0.05), and was greater in MI+HRP (38 ± 2%, P < 0.05) than in MI. Insulin-stimulated serine phosphorylation of Akt and glucose transporter 4 translocation were decreased in the skeletal muscle from MI by 48 and 49% of Sham, both of which were ameliorated in MI+HRP. Superoxide production and NAD(P)H oxidase activities were increased in MI, which was inhibited in MI+HRP. HRP ameliorated insulin resistance associated with HF by improving insulin signaling via the inhibition of NAD(P)H oxidase-induced superoxide production in the skeletal muscle. The (P)RR pathway is involved in the development of insulin resistance, at least in part, via the impairment of insulin signaling in the skeletal muscle from HF.

Impact of the prorenin/renin receptor on the development of obesity and associated cardiometabolic risk factors.

Obesity is a worldwide epidemic and current treatments have limited success thus, novel therapies are warranted. Our objective was to determine whether the prorenin/renin receptor [(P)RR] is implicated in obesity. Mice received a normal or high-fat/high-carbohydrate diet with the handle region peptide (HRP), a (P)RR blocker, or saline for 10 weeks. Post-menopausal non-diabetic obese women were enrolled in the Complication Associated with Obesity Study and were classified as insulin-resistant (IRO) or -sensitive (ISO) using a hyperinsulinemic-euglycemic clamp. In mice, obesity increased the (P)RR by twofold in adipose tissue. Likewise, renin increased by at least twofold. The HRP reduced weight gain in obese mice by 20% associated to a 19% decrease in visceral fat. This was accompanied by a 48% decrease in leptin mRNA in fat and 33% decrease in circulating leptin. Inflammatory markers were also decreased by the HRP treatment. HRP normalized triglyceridemia and reduced insulinemia by 34% in obese mice. Interestingly, we observed a 33% increase in (P)RR mRNA in the fat of IRO women compared to ISO. This is the first report of a potential implication in obesity of the (P)RR which may be a novel therapeutic target.

Combined renin inhibition/(pro)renin receptor blockade in diabetic retinopathy--a study in transgenic (mREN2)27 rats.

Dysfunction of renin-angiotensin system (RAS) contributes to the pathogenesis of diabetic retinopathy (DR). Prorenin, the precursor of renin is highly elevated in ocular fluid of diabetic patients with proliferative retinopathy. Prorenin may exert local effects in the eye by binding to the so-called (pro)renin receptor ((P)RR). Here we investigated the combined effects of the renin inhibitor aliskiren and the putative (P)RR blocker handle-region peptide (HRP) on diabetic retinopathy in streptozotocin (STZ)-induced diabetic transgenic (mRen2)27 rats (a model with high plasma prorenin levels) as well as prorenin stimulated cytokine expression in cultured Müller cells. Adult (mRen2)27 rats were randomly divided into the following groups: (1) non-diabetic; (2) diabetic treated with vehicle; (3) diabetic treated with aliskiren (10 mg/kg per day); and (4) diabetic treated with aliskiren+HRP (1 mg/kg per day). Age-matched non-diabetic wildtype Sprague-Dawley rats were used as control. Drugs were administered by osmotic minipumps for three weeks. Transgenic (mRen2)27 rat retinas showed increased apoptotic cell death of both inner retinal neurons and photoreceptors, increased loss of capillaries, as well as increased expression of inflammatory cytokines. These pathological changes were further exacerbated by diabetes. Aliskiren treatment of diabetic (mRen2)27 rats prevented retinal gliosis, and reduced retinal apoptotic cell death, acellular capillaries and the expression of inflammatory cytokines. HRP on top of aliskiren did not provide additional protection. In cultured Müller cells, prorenin significantly increased the expression levels of IL-1α and TNF-α, and this was completely blocked by aliskiren or HRP, their combination, (P)RR siRNA and the AT1R blocker losartan, suggesting that these effects entirely depended on Ang II generation by (P)RR-bound prorenin. In conclusion, the lack of effect of HRP on top of aliskiren, and the Ang II-dependency of the ocular effects of prorenin in vitro, argue against the combined application of (P)RR blockade and renin inhibition in diabetic retinopathy.

Deterioration of kidney function by the (pro)renin receptor blocker handle region peptide in aliskiren-treated diabetic transgenic (mRen2)27 rats

Dual renin-angiotensin system (RAS) blockade in diabetic nephropathy is no longer feasible because of the profit/side effect imbalance. (Pro)renin receptor [(P)RR] blockade with handle region peptide (HRP) has been reported to exert beneficial effects in various diabetic models in a RAS-independent manner. To what degree (P)RR blockade adds benefits on top of RAS blockade is still unknown. In the present study, we treated diabetic TGR(mREN2)27 rats, a well-established nephropathy model with high prorenin levels [allowing continuous (P)RR stimulation in vivo], with HRP on top of renin inhibition with aliskiren. Aliskiren alone lowered blood pressure and exerted renoprotective effects, as evidenced by reduced glomerulosclerosis, diuresis, proteinuria, albuminuria, and urinary aldosterone levels as well as diminished renal (P)RR and ANG II type 1 receptor expression. It also suppressed plasma and tissue RAS activity and suppressed cardiac atrial natriuretic peptide and brain natriuretic peptide expression. HRP, when given on top of aliskiren, did not alter the effects of renin inhibition on blood pressure, RAS activity, or aldosterone. However, it counteracted the beneficial effects of aliskiren in the kidney, induced hyperkalemia, and increased plasma plasminogen activator-inhibitor 1, renal cyclooxygenase-2, and cardiac collagen content. All these effects have been linked to (P)RR stimulation, suggesting that HRP might, in fact, act as a partial agonist. Therefore, the use of HRP on top of RAS blockade in diabetic nephropathy is not advisable.

Abstract 184: Deterioration of Kidney Function by the (Pro)renin Receptor Blocker Handle Region Peptide in Aliskiren-treated Diabetic Transgenic (mRen2)27 Rats

Objective Renal function in patients with diabetes mellitus (DM) might be influenced by the level of prorenin due to its binding to the (pro)renin receptor ((P)RR) and the subsequently occurring angiotensin production and/or the direct agonistic effects of prorenin mediated via this receptor. Indeed, in DM elevated prorenin levels are correlated with the development of microvascular complications such as nephropathy. Design and Methods In this study we evaluated renal function in diabetic TGR(mREN2)27 rats (a high prorenin hypertensive model), treated with vehicle, the renin inhibitor aliskiren, or aliskiren plus the putative (P)RR antagonist HRP for 3 weeks. 24-hour urine was collected, and blood and kidney were evaluated for renin-angiotensin system components and pathology. Results Increased diuresis and proteinuria due to DM were prevented by aliskiren, but not aliskiren+HRP. Aliskiren+HRP additionally decreased creatinine clearance, and increased the plasma levels of the profibrotic marker plasminogen-activator inhibitor-1. The increased natriuresis and renal collagen-1 expression in this model were unaffected by aliskiren±HRP. Aliskiren increased rat renin expression in the renal cortex. This was associated with a decline in (P)RR and AT 1 receptor mRNA expression, and these changes were unaffected by HRP. Glomerular volume and interlobar arterial lumen diameter modestly increased in the aliskiren+HRP group, and were unaffected by aliskiren alone. Conclusions HRP, when given on top of aliskiren in DM TGR(mREN2)27 rats worsens kidney damage and counteracts the beneficial effects of aliskiren. (P)RR blockade is therefore contraindicated in DM.