Potential of Renin-Angiotensin-Aldosterone System Modulations in Diabetic Kidney Disease: Old Players to New Hope!

Due to a tragic increase in the incidences of diabetes globally, diabetic kidney disease (DKD) has emerged as one of the leading causes of end-stage renal diseases (ESRD). Hyperglycaemia-mediated overactivation of the renin-angiotensin-aldosterone system (RAAS) is key to the development and progression of DKD. Consequently, RAAS inhibition by angiotensin-converting enzyme inhibitors (ACEi) or angiotensin receptor blockers (ARBs) is the first-line therapy for the clinical management of DKD. However, numerous clinical and preclinical evidences suggested that RAAS inhibition can only halt the progression of the DKD to a certain extent, and they are inadequate to cure DKD completely. Recent studies have improved understanding of the complexity of the RAAS. It consists of two counter-regulatory arms, the deleterious pressor arm (ACE/angiotensin II/AT1 receptor axis) and the beneficial depressor arm (ACE2/angiotensin-(1-7)/Mas receptor axis). These advances have paved the way for the development of new therapies targeting the RAAS for better treatment of DKD. In this review, we aimed to summarise the involvement of the depressor arm of the RAAS in DKD. Moreover, in modern drug discovery and development, an advance approach is the bispecific therapeutics, targeting two independent signalling pathways. Here, we discuss available reports of these bispecific drugs involving the RAAS as well as propose potential treatments based on neurohormonal balance as credible therapeutic strategies for DKD.

Restoring Angiotensin Type 2 Receptor Function Reverses PFOS-Induced Vascular Hyper-Reactivity and Hypertension in Pregnancy.

Perfluorooctane sulfonic acid (PFOS) exposure during pregnancy induces hypertension with decreased vasodilatory angiotensin type-2 receptor (AT2R) expression and impaired vascular reactivity and fetal weights. We hypothesized that AT2R activation restores the AT1R/AT2R balance and reverses gestational hypertension by improving vascular mechanisms. Pregnant Sprague-Dawley rats were exposed to PFOS through drinking water (50 µg/mL) from gestation day (GD) 4-20. Controls received drinking water with no detectable PFOS. Control and PFOS-exposed rats were treated with AT2R agonist Compound 21 (C21; 0.3 mg/kg/day, SC) from GD 15-20. In PFOS dams, blood pressure was higher, blood flow in the uterine artery was reduced, and C21 reversed these to control levels. C21 mitigated the heightened contraction response to Ang II and enhanced endothelium-dependent vasorelaxation in uterine arteries of PFOS dams. The observed vascular effects of C21 were correlated with reduced AT1R levels and increased AT2R and eNOS protein levels. C21 also increased plasma bradykinin production in PFOS dams and attenuated the fetoplacental growth restriction. These data suggest that C21 improves the PFOS-induced maternal vascular dysfunction and blood flow to the fetoplacental unit, providing preclinical evidence to support that AT2R activation may be an important target for preventing or treating PFOS-induced adverse maternal and fetal outcomes.

Investigational Drugs for the Treatment of Postherpetic Neuralgia: Systematic Review of Randomized Controlled Trials.

The pharmacological treatment of postherpetic neuralgia (PHN) is unsatisfactory, and there is a clinical need for new approaches. Several drugs under advanced clinical development are addressed in this review. A systematic literature search was conducted in three electronic databases (Medline, Web of Science, Scopus) and in the ClinicalTrials.gov register from 1 January 2016 to 1 June 2023 to identify Phase II, III and IV clinical trials evaluating drugs for the treatment of PHN. A total of 18 clinical trials were selected evaluating 15 molecules with pharmacological actions on nine different molecular targets: Angiotensin Type 2 Receptor (AT2R) antagonism (olodanrigan), Voltage-Gated Calcium Channel (VGCC) α2δ subunit inhibition (crisugabalin, mirogabalin and pregabalin), Voltage-Gated Sodium Channel (VGSC) blockade (funapide and lidocaine), Cyclooxygenase-1 (COX-1) inhibition (TRK-700), Adaptor-Associated Kinase 1 (AAK1) inhibition (LX9211), Lanthionine Synthetase C-Like Protein (LANCL) activation (LAT8881), N-Methyl-D-Aspartate (NMDA) receptor antagonism (esketamine), mu opioid receptor agonism (tramadol, oxycodone and hydromorphone) and Nerve Growth Factor (NGF) inhibition (fulranumab). In brief, there are several drugs in advanced clinical development for treating PHN with some of them reporting promising results. AT2R antagonism, AAK1 inhibition, LANCL activation and NGF inhibition are considered first-in-class analgesics. Hopefully, these trials will result in a better clinical management of PHN.

POLYMORPHISM OF ACE AND AT2R1 GENES AS A GENETIC BACKGROUND FOR DIFFERENT TYPES OF ENCEPHALOPATHIES.

OBJECTIVE: The aim: To study the prevalence of ACE I/D and AT2R1 A1166C gene polymorphisms in patients with CTE, SVD, AIE, and PIE and to assess the influence of the presence of a particular genotype of the studied genes on the occurrence and/or progression of encephalopathies. PATIENTS AND METHODS: Materials and methods: A total of 96 patients with encephalopathies of various genesis (chronic traumatic encephalopathy (CTE) n=26; chronic alcohol-induced encephalopathy (AIE) n=26; microvascular ischemic disease of the brain (or cerebral small vessel disease, (SVD)) n=18; post-infectious encephalopathy (PIE) n=26) were involved in the study. The molecular genetic study was performed in the molecular genetics laboratory of the State Institution «Reference Center for Molecular Diagnostics of the Ministry of Health of Ukraine¼, Kyiv. Statistical processing of the results was performed using the STATISTICA 10.0 program. RESULTS: Results: In patients with various types of encephalopathies, probable changes in the frequency distribution of genotypes of polymorphic variants I/D of the ACE gene were established (11.11% vs. 33.33% - carriers of the I/I genotype, 27.78% vs. 50.00% - carriers of the I/D genotype and 61.11% vs. 16.67% - carriers of the D/D genotype) and A1166C of the AT2R1 gene (22.22% vs. 66.67% - carriers of the A/A genotype, 50.00% vs. 25.00% - carriers A/C genotype, 27.78% versus 8.33% - carriers of the C/C genotype) compared to individuals of the control group only in patients with SVD. The presence of the D allele and the D/D genotype of the ACE gene is associated with a statistically significant increase in the risk of SVD development and progression (respectively, 4.2 times (95% CI (1.39-12.72)) and 7.9 (95% CI ( 1.31-47.05)) times). A similar trend was established for the carrier of the C allele of the A1166C polymorphic variant of the AT2R1 gene in patients with SVD: a 4.3-fold increase in the risk of development and progression (95% CI (1.30-13.86). In addition, there is a probable dependence between carrier genotype A/C of the AT2R1 gene and increased risk of PIE and AIE by 4.8 and 5.7 times, respectively. CONCLUSION: Conclusions: Therefore, results suggest the reasonability to include the I/D of the ACE gene polymorphism investigation in the genetic panel of encephalopathies.

CGP42112: the full AT2 receptor agonist and its role in the renin-angiotensin-aldosterone system: no longer misunderstood.

For years, the AT2R-selective ligand CGP42112 has been erroneously characterized as a partial agonist, partly due to its ability to also interact with the AT1R at high concentrations. As late as 2009, it was still being characterized as an antagonist as well. In this perspective/opinion piece, we try to resolve the ambiguity that surrounds the efficacy of this compound by extensively reviewing the literature, tracing its beginnings to 1989, showing that CGP42112 has never been convincingly shown to be a partial agonist or an antagonist at the AT2R. While CGP42112 is now routinely characterized as an AT2R agonist, regrettably, there is a paucity of studies that can validate its efficacy as a full agonist at the AT2R, leaving the door open for continuing speculation regarding the extent of its efficacy. Hopefully, the information presented in this perspective/opinion piece will firmly establish CGP42112 as a full agonist at the AT2R such that it can once again be used as a tool to study the AT2R.

Angiotensin II type-2-receptor stimulation ameliorates focal and segmental glomerulosclerosis in mice.

Podocyte damage and loss are the early event in the development of focal segmental glomerulosclerosis (FSGS). Podocytes express angiotensin II type-2-receptor (AT2R), which may play a key role in maintaining kidney integrity and function. Here, we examined the effects of AT2R deletion and AT2R agonist compound 21 (C21) on the evolution of FSGS. FSGS was induced by adriamycin (ADR) injection in both male wild-type (WT) and AT2R knockout (KO) mice. C21 was administered to WT-FSGS mice either one day before or 7 days after ADR (Pre-C21 or Post-C21), using two doses of C21 at either 0.3 (low dose, LD) or 1.0 (high dose, HD) mg/kg/day. ADR-induced FSGS was more severe in AT2RKO mice compared with WT-FSGS mice, and included profound podocyte loss, glomerular fibrosis, and albuminuria. Glomerular cathepsin L expression increased more in AT2RKO-FSGS than in WT-FSGS mice. C21 treatment ameliorated podocyte injury, most significantly in the Pre C21-HD group, and inhibited glomerular cathepsin L expression. In vitro, Agtr2 knock-down in mouse podocyte cell line given ADR confirmed the in vivo data. Mechanistically, C21 inhibited cathepsin L expression, which protected synaptopodin from destruction and stabilized actin cytoskeleton. C21 also prevented podocyte apoptosis. In conclusion, AT2R activation by C21 ameliorated ADR-induced podocyte injury in mice by the inhibition of glomerular cathepsin L leading to the maintenance of podocyte integrity and prevention of podocyte apoptosis.

2-Alkyl substituted benzimidazoles as a new class of selective AT2 receptor ligands.

Ligands comprising a benzimidazole rather than the imidazole ring that is common in AT2R ligands e.g. in the AT2R agonist C21, can provide both high affinity and receptor selectivity. In particular, compounds encompassing benzimidazoles, substituted in the 2-position with small bulky groups such as an isopropyl (Ki = 4.0 nM) or a tert-butyl (Ki = 5.3 nM) or alternatively a thiazole heterocycle (Ki = 5.1 nM) demonstrate high affinity and AT2R selectivity. An n-butyl chain, as found in the AT1R selective sartans, makes the ligand less receptor selective. The isobutyl group on the biaryl scaffold present in most AT2R selective ligands reported so far was originally derived from the nonselective potent AT1R/AT2R ligand L-162,313. Notably, in all ligands discussed herein, the isobutyl group was substituted by an n-propyl group and ligands with high affinity to AT2R were provided and in addition the majority of them demonstrate a favorable AT2R/AT1R selectivity. The introduction of fluoro atoms in various positions had no pronounced effect on the affinity data. Ligands with a thiazole or a tert-butyl group attached to the 2-position and with a terminal trifluoromethyl butoxycarbonyl sidechain exhibited a similar stability as C21 in human liver microsomes, while other ligands examined were less stable in the microsome assay.

Angiotensin II AT2 receptor ligands with phenylthiazole scaffolds.

The syntheses and the AT1R and AT2R binding data of a series of new small molecule ligands are reported. These ligands comprise a phenylthiazole scaffold rather than the biphenyl or phenylthiophene scaffolds found in essentially all of the previously described ligands originating from the nonselective AT1R/AT2R ligand L-162,313 and the AT2R selective agonist C21, the latter now in Phase II/III clinical trials. A phenylthiazole rather than the phenylthiophene scaffold that is present in the AT2R selective agonist C21 and in the AT2R selective antagonist C38 had a deleterious effect on the affinity to AT2R. Nevertheless, a significant improvement could be accomplished by introduction of a small bulky alkyl group in the 2-position of the imidazole ring attached through a methylene group bridge to the phenylthiazole scaffold. Hence, a combination of a 2-tert-butyl or a 2-isopropyl group and a butoxycarbonyl furnished potent AT2R selective ligands. Furthermore, a high affinity ligand derived from L-162,313 and exhibiting a > 35 fold selectivity for AT1R was identified (10). The ligand 21 with the 2-tert-butyl group and âˆ¼ 35 fold selectivity for AT2R, demonstrated high stability in human, rat and mouse liver microsomes and a very attractive profile with regard to the inhibition of common drug-metabolizing CYP enzymes. Thus, very low levels of inhibition of CYP 3A (5%), 2D6 (12%), 2C8 (26%), 2C9 (23%) and 2B6 (24%) were observed with the 2-tert-butyl derivative comprising the methoxycarbonyl sulfonamide function, levels that are significantly lower than those obtained with C21 under the same experimental conditions.

Tripeptide IRW Protects MC3T3-E1 Cells against Ang II Stress in an AT2R Dependent Manner.

Multiple strategies including the use of bioactive peptides and other nutraceuticals are being adopted to maintain bone health. This study provides an improved and deeper understanding of the pharmacological effects that a bioactive peptide IRW (Ile-Arg-Trp) extends on bone health. Our results showed that IRW treatment protects osteoblasts against Ang II induced decline in cell proliferation and restores protein levels of collagen type I alpha 2 chain (COL1A2) and alkaline phosphatase (ALP) levels in MC3T3-E1 cells (p < 0.05). Apart from augmentation of these mineralization factors, the angiotensin II (Ang II) induced apoptotic stress in osteoblasts was mitigated by IRW as well. At the molecular level, IRW abolished the cytochrome-c release via modulation of pro-and anti-apoptotic genes in MC3T3-E1 cells (p < 0.05). Interestingly, IRW also increased cellular levels of cytoprotective local RAAS factors such as MasR, Ang (1−7), ACE2, and AT2R, and lowered the levels of Ang II effector receptor (AT1R). Further, our results indicated a lower content of inflammation and osteoclastogenesis biomarkers such as cyclooxygenase 2 (COX2), nuclear factor kappa B (NF-κB), and receptor activator of nuclear factor kappa-B ligand (RANKL) following IRW treatment in MC3T3-E1 cells (p < 0.05). The use of an antagonist-guided cell study indicated that IRW contributed to the process of cytoprotection and proliferation of osteoblasts via Runt-related transcription factor 2 (RUNX2) in face of Ang II stress in an AT2R dependent manner. The key findings of our study showed that IRW could potentially have a therapeutic role in the treatment and/or prevention of bone disorders.

AT2R deficiency in mice accelerates podocyte dysfunction in diabetic progeny in a sex-dependent manner.

AIMS/HYPOTHESIS: The angiotensin II receptor type 2 (AT2R) may be a potential therapeutic target for the treatment of hypertension and chronic kidney disease (CKD). The expression and function of AT2R in the vasculature and kidney appear sexually dimorphic. We hypothesised that Agtr2 knockout dams (AT2RKO) with gestational diabetes would program their offspring for subsequent hypertension and CKD in a sex-dependent manner. METHODS: Age- and sex-matched offspring of non-diabetic and diabetic dams of wild-type (WT) and AT2RKO mice were followed from 4 to 20 weeks of age and were monitored for development of hypertension and nephropathy; a mouse podocyte cell line (mPOD) was also studied. RESULTS: Body weight was progressively lower in female compared with male offspring throughout the lifespan. Female but not male offspring from diabetic AT2RKO dams developed insulin resistance. Compared with the offspring of non-diabetic dams, the progeny of diabetic dams had developed more hypertension and nephropathy (apparent glomerulosclerosis with podocyte loss) at 20 weeks of age; this programming was more pronounced in the offspring of AT2RKO diabetic dams, particularly female AT2RKO progeny. Female AT2RKO offspring had lower basal ACE2 glomerular expression, resulting in podocyte loss. The aberrant ACE2/ACE ratio was far more diminished in glomeruli of female progeny of diabetic AT2RKO dams than in male progeny. Knock-down of Agtr2 in mPODs confirmed the in vivo data. CONCLUSIONS/INTERPRETATION: AT2R deficiency accelerated kidney programming in female progeny of diabetic dams, possibly due to loss of protective effects of ACE2 expression in the kidney.

N-(Methyloxycarbonyl)thiophene sulfonamides as high affinity AT2 receptor ligands.

A series of meta-substituted acetophenone derivatives, encompassing N-(alkyloxycarbonyl)thiophene sulfonamide fragments have been synthesized. Several selective AT2 receptor ligands were identified, among those a tert-butylimidazole derivative (20) with a Ki of 9.3 nM, that demonstrates a high stability in human liver microsomes (t½ = 62 min) and in human hepatocytes (t½ = 194 min). This methyloxycarbonylthiophene sulfonamide is a 20-fold more potent binder to the AT2 receptor and is considerably more stable in human liver microsomes, than a previously reported and broadly studied structurally related AT2R prototype antagonist 3 (C38). Ligand 20 acts as an AT2R agonist and caused an AT2R mediated concentration-dependent vasorelaxation of pre-contracted mouse aorta. Furthermore, in contrast to imidazole derivative C38, the tert-butylimidazole derivative 20 is a poor inhibitor of CYP3A4, CYP2D6 and CYP2C9. It is demonstrated herein that smaller alkyloxycarbonyl groups make the ligands in this series of AT2R selective compounds less prone to degradation and that a high AT2 receptor affinity can be retained after truncation of the alkyloxycarbonyl group. Binding modes of the most potent AT2R ligands were explored by docking calculations combined with molecular dynamics simulations.

Glucagon-like peptide-1 attenuates cardiac hypertrophy via the AngII/AT1R/ACE2 and AMPK/mTOR/p70S6K pathways.

Glucagon-like peptide-1 (GLP-1), a novel type of glucose-lowering agent, has been reported to exert cardioprotective effects. However, the cardioprotective mechanism of GLP-1 on spontaneous hypertension-induced cardiac hypertrophy has not been fully elucidated. In this study, we revealed that liraglutide or alogliptin treatment ameliorated spontaneous hypertension-induced cardiac hypertrophy, as evidenced by decreased levels of cardiac hypertrophic markers (atrial natriuretic peptide, brain natriuretic peptide, and ß-myosin heavy chain), as well as systolic blood pressure, diastolic blood pressure, mean arterial pressure, and histological changes. Both drugs significantly reduced the levels of angiotensin II (AngII) and AngII type 1 receptor (AT1R) and upregulated the levels of AngII type 2 receptor (AT2R) and angiotensin-converting enzyme 2 (ACE2), as indicated by a reduced AT1R/AT2R ratio. Simultaneously, treatment with liraglutide or alogliptin significantly increased GLP-1 receptor expression and adenosine monophosphate-activated protein kinase (AMPK) phosphorylation and downregulated the phosphorylation of mammalian target of rapamycin (mTOR), p70 ribosomal S6 protein kinase, and eukaryotic translation initiation factor 4E binding protein 1 in spontaneous hypertension rats. Furthermore, our data demonstrated that the AMPK inhibitor compound C or mTOR activator MHY1485 inhibited the anti-hypertrophic effect of GLP-1. In summary, our study suggests that liraglutide or alogliptin protects the heart against cardiac hypertrophy by regulating the expression of AngII/AT1R/ACE2 and activating the AMPK/mTOR pathway, and GLP-1 agonist can be used in the treatment of patients with cardiac hypertrophy.

Macrophage angiotensin II type 2 receptor triggers neuropathic pain.

Peripheral nerve damage initiates a complex series of structural and cellular processes that culminate in chronic neuropathic pain. The recent success of a type 2 angiotensin II (Ang II) receptor (AT2R) antagonist in a phase II clinical trial for the treatment of postherpetic neuralgia suggests angiotensin signaling is involved in neuropathic pain. However, transcriptome analysis indicates a lack of AT2R gene (Agtr2) expression in human and rodent sensory ganglia, raising questions regarding the tissue/cell target underlying the analgesic effect of AT2R antagonism. We show that selective antagonism of AT2R attenuates neuropathic but not inflammatory mechanical and cold pain hypersensitivity behaviors in mice. Agtr2-expressing macrophages (MΦs) constitute the predominant immune cells that infiltrate the site of nerve injury. Interestingly, neuropathic mechanical and cold pain hypersensitivity can be attenuated by chemogenetic depletion of peripheral MΦs and AT2R-null hematopoietic cell transplantation. Our study identifies AT2R on peripheral MΦs as a critical trigger for pain sensitization at the site of nerve injury, and therefore proposes a translatable peripheral mechanism underlying chronic neuropathic pain.

AT2R agonist NP-6A4 mitigates aortic stiffness and proteolytic activity in mouse model of aneurysm.

Clinical and experimental studies show that angiotensin II (AngII) promotes vascular pathology via activation of AngII type 1 receptors (AT1Rs). We recently reported that NP-6A4, a selective peptide agonist for AngII type 2 receptor (AT2R), exerts protective effects on human vascular cells subjected to serum starvation or doxorubicin exposure. In this study, we investigated whether NP-6A4-induced AT2R activation could mitigate AngII-induced abdominal aortic aneurism (AAA) using AngII-treated Apoe-/- mice. Male Apoe-/- mice were infused with AngII (1 µg/kg/min) by implanting osmotic pumps subcutaneously for 28 days. A subset of mice was pre-treated subcutaneously with NP-6A4 (2.5 mg/kg/day) or vehicle for 14 days prior to AngII, and treatments were continued for 28 days. NP-6A4 significantly reduced aortic stiffness of the abdominal aorta induced by AngII as determined by ultrasound functional analyses and histochemical analyses. NP-6A4 also increased nitric oxide bioavailability in aortic tissues and suppressed AngII-induced increases in monocyte chemotactic protein-1, osteopontin and proteolytic activity of the aorta. However, NP-6A4 did not affect maximal intraluminal aortic diameter or AAA incidences significantly. These data suggest that the effects of AT2R agonist on vascular pathologies are selective, affecting the aortic stiffness and proteolytic activity without affecting the size of AAA.

Does activation of the protective Renin-Angiotensin System have therapeutic potential in COVID-19?

Infection of lung cells by the corona virus results in a loss of the balance between, on the one hand, angiotensin II-mediated stimulation of the angiotensin II type 1 receptor and, on the other hand, stimulation of the angiotensin II type 2 receptor and/or the Mas receptor. The unbalanced enhanced stimulation of the angiotensin II type 1 receptor causes inflammation, edema and contributes to the pathogenesis of severe acute respiratory distress syndrome. Here we hypothesize that stable, receptor-specific agonists of the angiotensin II type 2 receptor and of the Mas receptor are molecular medicines to treat COVID-19 patients. These agonists have therapeutic potential in the acute disease but in addition may reduce COVID-19-associated long-term pulmonary dysfunction and overall end-organ damage of this disease.

Brain Angiotensin Type-1 and Type-2 Receptors in Physiological and Hypertensive Conditions: Focus on Neuroinflammation.

PURPOSE OF REVIEW: To review recent data that suggest opposing effects of brain angiotensin type-1 (AT1R) and type-2 (AT2R) receptors on blood pressure (BP). Here, we discuss recent studies that suggest pro-hypertensive and pro-inflammatory actions of AT1R and anti-hypertensive and anti-inflammatory actions of AT2R. Further, we propose mechanisms for the interplay between brain angiotensin receptors and neuroinflammation in hypertension. RECENT FINDINGS: The renin-angiotensin system (RAS) plays an important role in regulating cardiovascular physiology. This includes brain AT1R and AT2R, both of which are expressed in or adjacent to brain regions that control BP. Activation of AT1R within those brain regions mediate increases in BP and cause neuroinflammation, which augments the BP increase in hypertension. The fact that AT1R and AT2R have opposing actions on BP suggests that AT1R and AT2R may have similar opposing actions on neuroinflammation. However, the mechanisms by which brain AT1R and AT2R mediate neuroinflammatory responses remain unclear. The interplay between brain angiotensin receptor subtypes and neuroinflammation exacerbates or protects against hypertension.

Recent Research Advances in Renin-Angiotensin-Aldosterone System Receptors.

PURPOSE OF REVIEW: The renin-angiotensin-aldosterone system (RAAS) plays important roles in regulating blood pressure and body fluid, which contributes to the pathophysiology of hypertension and cardiovascular/renal diseases. However, accumulating evidence has further revealed the complexity of this signal transduction system, including direct interactions with other receptors and proteins. This review focuses on recent research advances in RAAS with an emphasis on its receptors. RECENT FINDINGS: Both systemically and locally produced angiotensin II (Ang II) bind to Ang II type 1 receptor (AT1R) and elicit strong biological functions. Recent studies have shown that Ang II-induced activation of Ang II type 2 receptor (AT2R) elicits the opposite functions to those of AT1R. However, accumulating evidence has now expanded the components of RAAS, including (pro)renin receptor, angiotensin-converting enzyme 2, angiotensin 1-7, and Mas receptor. In addition, the signal transductions of AT1R and AT2R are regulated by not only Ang II but also its receptor-associated proteins such as AT1R-associated protein and AT2R-interacting protein. Recent studies have indicated that inappropriate activation of local mineralocorticoid receptor contributes to cardiovascular and renal tissue injuries through aldosterone-dependent and -independent mechanisms. Since the mechanisms of RAAS signal transduction still remain to be elucidated, further investigations are necessary to explore novel molecular mechanisms of the RAAS, which will provide alternative therapeutic agents other than existing RAAS blockers.

Seminal levels of angiotensin II and angiotensin II type 2 receptor expression on spermatozoa in varicocele patients: Relation to fertility status.

Several theories were proposed to explain the pathophysiology of varicocele-related infertility seen in some patients. Our aim was to study the levels of angiotensin II in semen and angiotensin II type 2 receptor expression on spermatozoa in varicocele patients in relation to their fertility status and to evaluate the influence of varicocelectomy on their levels in infertile varicocele patients. Thirty fertile and 30 infertile varicocele patients and 30 healthy controls were subjected to measurement of reproductive hormones, semen analysis, measurement of seminal angiotensin II and evaluation of angiotensin II type 2 receptor expression on spermatozoa. Infertile varicocele patients underwent varicocelectomy and were re-evaluated for the same parameters after the operation. Sperm concentration, morphology, progressive motility, seminal angiotensin II and angiotensin II type 2 receptor expression were significantly lower in infertile varicocele patients compared with the other groups. Post-operative values showed significant increase in the studied parameters compared with the pre-operative values but not to other two groups. A significant positive correlation between angiotensin II type 2 receptor expression and progressive motility was detected in all studied groups. In conclusion, dysregulation of angiotensin II and angiotensin II type 2 receptor in varicocele patients may be involved in varicocele-related infertility.

Angiotensin II Triggers Peripheral Macrophage-to-Sensory Neuron Redox Crosstalk to Elicit Pain.

Injury, inflammation, and nerve damage initiate a wide variety of cellular and molecular processes that culminate in hyperexcitation of sensory nerves, which underlies chronic inflammatory and neuropathic pain. Using behavioral readouts of pain hypersensitivity induced by angiotensin II (Ang II) injection into mouse hindpaws, our study shows that activation of the type 2 Ang II receptor (AT2R) and the cell-damage-sensing ion channel TRPA1 are required for peripheral mechanical pain sensitization induced by Ang II in male and female mice. However, we show that AT2R is not expressed in mouse and human dorsal root ganglia (DRG) sensory neurons. Instead, expression/activation of AT2R on peripheral/skin macrophages (MΦs) constitutes a critical trigger of mouse and human DRG sensory neuron excitation. Ang II-induced peripheral mechanical pain hypersensitivity can be attenuated by chemogenetic depletion of peripheral MΦs. Furthermore, AT2R activation in MΦs triggers production of reactive oxygen/nitrogen species, which trans-activate TRPA1 on mouse and human DRG sensory neurons via cysteine modification of the channel. Our study thus identifies a translatable immune cell-to-sensory neuron signaling crosstalk underlying peripheral nociceptor sensitization. This form of cell-to-cell signaling represents a critical peripheral mechanism for chronic pain and thus identifies multiple druggable analgesic targets.SIGNIFICANCE STATEMENT Pain is a widespread health problem that is undermanaged by currently available analgesics. Findings from a recent clinical trial on a type II angiotensin II receptor (AT2R) antagonist showed effective analgesia for neuropathic pain. AT2R antagonists have been shown to reduce neuropathy-, inflammation- and bone cancer-associated pain in rodents. We report that activation of AT2R in macrophages (MΦs) that infiltrate the site of injury, but not in sensory neurons, triggers an intercellular redox communication with sensory neurons via activation of the cell damage/pain-sensing ion channel TRPA1. This MΦ-to-sensory neuron crosstalk results in peripheral pain sensitization. Our findings provide an evidence-based mechanism underlying the analgesic action of AT2R antagonists, which could accelerate the development of efficacious non-opioid analgesic drugs for multiple pain conditions.

Angiotensin receptor expression revealed by reporter mice and beneficial effects of AT2R agonist in retinal cells.

The renin-angiotensin system (RAS) plays a vital role in cardiovascular physiology and body homeostasis. In addition to circulating RAS, a local RAS exists in the retina. Dysfunction of local RAS, resulting in increased levels of Angiotensin II (Ang II) and activation of AT1R-mediated signaling pathways, contributes to tissue pathophysiology and end-organ damage. Activation of AT2R on other hand is known to counteract the effects of AT1R activation and produce anti-inflammatory and anti-oxidative effects. We examined the expression of angiotensin receptors in the retina by using transgenic dual reporter mice and by real-time RT-PCR. We further evaluated the effects of C21, a selective agonist of AT2R, in reducing Ang II, lipopolysaccharide (LPS) and hydrogen peroxide induced oxidative stress and inflammatory responses in cultured human ARPE-19 cells. We showed that both AT1Ra and AT2R positive cells are detected in different cell types of the eye, including the RPE/choroid complex, ciliary body/iris, and neural retina. AT1Ra is more abundantly expressed than AT2R in mouse retina, consistent with previous reports. In the neural retina, AT1Ra are also detected in photoreceptors whereas AT2R are mostly expressed in the inner retinal neurons and RGCs. In cultured human RPE cells, activation of AT2R with C21 significantly blocked Ang II, LPS and hydrogen peroxide -induced NF-κB activation and inflammatory cytokine expression; Ang II and hydrogen peroxide-induced reactive oxygen species (ROS) production and MG132-induced apoptosis, comparable to the effects of Angiotensin-(1-7) (Ang-(1-7)), another protective component of the RAS, although C21 is more potent in reducing some of the effects induced by Ang II, whereas Ang-(1-7) is more effective in reducing some of the LPS and hydrogen peroxide-induced effects. These results suggest that activation of AT2R may represent a new therapeutic approach for retinal diseases.