Natriuretic Peptide receptor-A mRNA Research Articles

Localization of natriuretic peptide receptors A, B, and C in healthy and diseased mouse kidneys

The natriuretic peptides (NPs) ANP (atrial natriuretic peptide) and BNP (B-type natriuretic peptide) mediate their widespread effects by activating the natriuretic peptide receptor-A (NPR-A), while C-type natriuretic peptide (CNP) acts via natriuretic peptide receptor-B (NPR-B). NPs are removed from the circulation by internalization via the natriuretic peptide clearance receptor natriuretic peptide receptor-C (NPR-C). In addition to their well-known functions, for instance on blood pressure, all three NPs confer significant cardioprotection and renoprotection. Since neither the NP-mediated renal functions nor the renal target cells of renoprotection are completely understood, we performed systematic localization studies of NP receptors using in situ hybridization (RNAscope) in mouse kidneys. NPR-A mRNA is highly expressed in glomeruli (mainly podocytes), renal arterioles, endothelial cells of peritubular capillaries, and PDGFR-receptor β positive (PDGFR-β) interstitial cells. No NPR-A mRNA was detected by RNAscope in the tubular system. In contrast, NPR-B expression is highest in proximal tubules. NPR-C is located in glomeruli (mainly podocytes), in endothelial cells and PDGFR-β positive cells. To test for a possible regulation of NPRs in kidney diseases, their distribution was studied in adenine nephropathy. Signal intensity of NPR-A and NPR-B mRNA was reduced while their spatial distribution was unaltered compared with healthy kidneys. In contrast, NPR-C mRNA signal was markedly enhanced in cell clusters of myofibroblasts in fibrotic areas of adenine kidneys. In conclusion, the primary renal targets of ANP and BNP are glomerular, vascular, and interstitial cells but not the tubular compartment, while the CNP receptor NPR-B is highly expressed in proximal tubules. Further studies are needed to clarify the function and interplay of this specific receptor expression pattern.

Angiotensin II represses Npr1 expression and receptor function by recruitment of transcription factors CREB and HSF-4a and activation of HDACs

The two vasoactive hormones, angiotensin II (ANG II; vasoconstrictive) and atrial natriuretic peptide (ANP; vasodilatory) antagonize the biological actions of each other. ANP acting through natriuretic peptide receptor-A (NPRA) lowers blood pressure and blood volume. We tested hypothesis that ANG II plays critical roles in the transcriptional repression of Npr1 (encoding NPRA) and receptor function. ANG II significantly decreased NPRA mRNA and protein levels and cGMP accumulation in cultured mesangial cells and attenuated ANP-mediated relaxation of aortic rings ex vivo. The transcription factors, cAMP-response element-binding protein (CREB) and heat-shock factor-4a (HSF-4a) facilitated the ANG II-mediated repressive effects on Npr1 transcription. Tyrosine kinase (TK) inhibitor, genistein and phosphatidylinositol 3-kinase (PI-3K) inhibitor, wortmannin reversed the ANG II-dependent repression of Npr1 transcription and receptor function. ANG II enhanced the activities of Class I histone deacetylases (HDACs 1/2), thereby decreased histone acetylation of H3K9/14ac and H4K8ac. The repressive effect of ANG II on Npr1 transcription and receptor signaling seems to be transduced by TK and PI-3K pathways and modulated by CREB, HSF-4a, HDACs, and modified histones. The current findings suggest that ANG II-mediated repressive mechanisms of Npr1 transcription and receptor function may provide new molecular targets for treatment and prevention of hypertension and cardiovascular diseases.

Expression and Regulation of Brain Natriuretic Peptide and Natriuretic Peptide Receptor A (NPR-A) in L6-S1 Dorsal Root Ganglia in a Rat Model of Chronic Nonbacterial Prostatitis.

BackgroundThe study aimed to investigate the expression of brain natriuretic peptide (BNP) and natriuretic peptide receptor A (NPR-A) in L6–S1 dorsal root ganglia (DRG) in a rat model of chronic nonbacterial prostatitis (CNP).Material/MethodsOne hundred specific pathogen-free (SPF) male Sprague–Dawley rats were randomly divided into a control group (N=50) and a study group (N=50). The control group underwent prostatic injection of 0.1 ml of normal saline on days 3, 7, 10, 14, and 28. The study group, or rat model of CNP, underwent prostatic injection of 0.1 ml of complete Freund’s adjuvant on days 3, 7, 10, 14, and 28. At the end of the study, the rats were euthanized, and the prostate tissues and L6–S1 DRG were removed. Histology was performed on the prostate tissue from the rats in the study group and control group. Real-time fluorescence-based quantitative polymerase chain reaction (PCR) and Western blot were used to study the expression of BNP and NPR-A mRNA and protein in the DRG from the rats in the study group and control group.ResultsIn the rat model of CNP, the expression of BNP and NPR-A were significantly increased in L6–S1 DRG compared with the controls.ConclusionsIn a rat model of CNP, the increased expression of BNP and NPR-A in L6–S1 DRG may have a role in pain signaling pathways associated with chronic prostatitis.

Association of fat mass profile with natriuretic peptide receptor alpha in subcutaneous adipose tissue of medication-free healthy men:A cross-sectional study.

Background: Atrial natriuretic peptide increases lipolysis in human adipocytes by binding to natriuretic peptide receptor-A (NPRA). The aim of the current study was to examine the associations of NPRA mRNA of subcutaneous adipose tissue with fat mass, fat-free mass, body mass index (BMI) and arterial blood pressure in medication-free healthy men. Method: Thirty-two volunteers [age (years): 36.06±7.36, BMI: 27.60±4.63 (kg/m 2)] underwent assessments of body height/weight, % fat mass, fat-free mass (kg), blood pressure, and a subcutaneous adipose tissue biopsy via a surgical technique. Results: We found that NPRA mRNA was negatively associated with % fat mass (r=-0.40, R 2=0.16, p=0.03) and BMI (r=-0.45, R 2=0.20, p=0.01). Cohen's f 2 effect size analyses showed a small effect size between NPRA mRNA and BMI ( f 2 =0.25). One-way analysis of variance with Bonferroni post-hoc tests showed a tendency for mean differences of NPRA mRNA across BMI categories (p=0.06). This was confirmed by Cohen's d effect size analyses revealing a large effect size of NPRA mRNA between obese individuals (BMI≥30 kg/m 2) and either normal weight (BMI=19-25 kg/m 2; d=0.94) or overweight (BMI=25-30 kg/m 2; d=1.12) individuals. Conclusions: NPRA mRNA is negatively associated with % fat mass and BMI in medication-free healthy men, suggesting a possible role of NPRA in the control of fat mass accumulation.

Association of fat mass profile with natriuretic peptide receptor alpha in subcutaneous adipose tissue of medication-free healthy men: A cross-sectional study

Background: Atrial natriuretic peptide increases lipolysis in human adipocytes by binding to natriuretic peptide receptor-A (NPRA). The aim of the current study was to examine the associations of NPRA mRNA of subcutaneous adipose tissue with fat mass, fat-free mass, body mass index (BMI) and arterial blood pressure in medication-free healthy men. Method: Thirty-two volunteers [age (years): 36.06±7.36, BMI: 27.60±4.63 (kg/m 2)] underwent assessments of body height/weight, % fat mass, fat-free mass (kg), blood pressure, and a subcutaneous adipose tissue biopsy via a surgical technique. Results: We found that NPRA mRNA was negatively associated with % fat mass (r=-0.40, R 2=0.16, p=0.03) and BMI (r=-0.45, R 2=0.20, p=0.01). Cohen’s f 2 effect size analyses showed a small effect size between NPRA mRNA and BMI ( f 2=0.25). One-way analysis of variance with Bonferroni post-hoc tests showed a tendency for mean differences of NPRA mRNA across BMI categories (p=0.06). This was confirmed by Cohen’s d effect size analyses revealing a large effect size of NPRA mRNA between obese individuals (BMI≥30 kg/m 2) and either normal weight (BMI=19-25 kg/m 2; d=0.94) or overweight (BMI=25-30 kg/m 2; d=1.12) individuals. Conclusions: NPRA mRNA is negatively associated with % fat mass and BMI in medication-free healthy men, suggesting a possible role of NPRA in the control of fat mass accumulation.

Association of fat mass profile with natriuretic peptide receptor alpha in subcutaneous adipose tissue of medication-free healthy men:A cross-sectional study.

Background: Atrial natriuretic peptide increases lipolysis in human adipocytes by binding to natriuretic peptide receptor-A (NPRA). The aim of the current study was to examine the associations of NPRA mRNA of subcutaneous adipose tissue with fat mass, fat-free mass, body mass index (BMI) and arterial blood pressure in medication-free healthy men. Method: Thirty-two volunteers [age (years): 36.06±7.36, BMI: 27.60±4.63 (kg/m 2)] underwent assessments of body height/weight, % fat mass, fat-free mass (kg), blood pressure, and a subcutaneous adipose tissue biopsy via a surgical technique. Results: We found that NPRA mRNA was negatively associated with % fat mass (r=-0.40, R 2=0.16, p=0.03) and BMI (r=-0.45, R 2=0.20, p=0.01). Cohen's f 2 effect size analyses showed a small effect size between NPRA mRNA and BMI ( f 2 =0.25). One-way analysis of variance with Bonferroni post-hoc tests showed a tendency for mean differences of NPRA mRNA across BMI categories (p=0.06). This was confirmed by Cohen's d effect size analyses revealing a large effect size of NPRA mRNA between obese individuals (BMI≥30 kg/m 2) and either normal weight (BMI=19-25 kg/m 2; d=0.94) or overweight (BMI=25-30 kg/m 2; d=1.12) individuals. Conclusions: NPRA mRNA is negatively associated with % fat mass and BMI in medication-free healthy men, suggesting a possible role of NPRA in the control of fat mass accumulation.

Localization of corin and atrial natriuretic peptide expression in human renal segments.

Atrial natriuretic peptide (ANP)-mediated natriuretic response is a well-established cardiac endocrine function. Corin is a transmembrane protease that activates ANP in the heart. Corin expression has been detected in non-cardiac tissues including the kidney. Here we examined corin, pro-ANP/ANP and natriuretic peptide receptor-A (NPR-A) expression in human renal segments. By immunostaining and in situ hybridization, we found similar corin, pro-ANP/ANP and NPR-A protein and mRNA expression in human renal segments. The expression was most abundant in the proximal convoluted tubules and the medullary connecting ducts. In the proximal tubules, corin protein was present in the apical membrane region underneath the brush border where the ANP-degrading protease neprilysin was abundant. These results suggest that corin-mediated pro-ANP activation may occur in renal segments and that locally produced ANP may act in an autocrine manner to regulate sodium and water reabsorption in situ Our results also point to the proximal convoluted tubules as a major site for local ANP action. Such a renal corin/ANP autocrine mechanism may differ from the cardiac corin/ANP endocrine mechanism in regulating sodium homoeostasis under physiological and pathological conditions.

Serum levels of ANP signaling in different degree of allergic asthmatics and its role in the pathogenesis of allergic asthma

To investigate the relationship between the severity of allergic asthma and the levels of atrial natriuretic peptide (ANP), and to analyze the potential role of ANP signaling in the pathogenesis of asthma. We recruited 96 subjects, including 23 healthy volunteers, 25 stable allergic asthmatics, 21 mild allergic asthmatics and 27 moderate allergic asthmatics, from the Affiliated Hospital of Guilin Medical University. ANP, IFN-γ and IL-4 levels in serum were detected by enzyme-linked immunosorbent assay (ELISA), and the mRNA and protein expressions of natriuretic peptide receptor A (NPRA), transcription factor T-bet and GATA3 were measured by RT-PCR and Western blot. The levels of ANP in serum and the expressions of NPRA mRNA and protein in the peripheral blood mononuclear cell (PBMC) from the mild asthma group or the moderate group were elevated compared with those in the stable asthma group or the mild group, respectively (P<0.05). Consistently, expressions of GATA3 and levels of IL-4 showed the same tendency (P<0.05). In addition, levels of ANP in serum were positively correlated with the severity of asthma, whereas negatively correlated with the ratio of T-bet/GATA3 and IFN-γ/IL-4 (r=-0.85, P<0.05; r=-0.88, P<0.05, respectively). Levels of ANP signaling in serum were significantly increased with the severity of allergic asthma, suggesting a close relation with the pathogenesis of asthma; ANP signaling may play a role in the pathogenesis of allergic asthma through inducing the Th2-type immune response.

Modulation in Natriuretic Peptides System in Experimental Colitis in Rats.

Renin-angiotensin system is involved in the pathophysiology of colonic inflammation. However, there are a few reports about modulation of natriuretic peptide system. This study investigates whether a local atrial natriuretic peptide (ANP) system exists in rat colon and whether ANP plays a role in the regulation of colonic motility in experimental colitis rat model. Experimental colitis was induced by an intake of 5 % dextran sulfate sodium (DSS) dissolved in tap water for 7 days. After rats were killed, plasma hormone concentrations and mRNAs for natriuretic peptide system were measured. Functional analysis of colonic motility in response to ANP was performed using taenia coli. DSS-treated colon showed an increased necrosis with massive infiltration of inflammatory cells. The colonic natriuretic peptide receptor-A mRNA level and particulate guanylyl cyclase activity in response to ANP from colonic tissue membranes were higher, and the mRNA levels of ANP and natriuretic peptide receptor-B were lower in DSS-treated rats than in control rats. ANP decreased the frequency of basal motility in a dose-dependent manner but did not change the amplitude. The inhibitory responses of frequency of basal motility to ANP and 8-bromo-cGMP were enhanced in DSS-treated rat colon. In conclusion, augmentation of inhibitory effect on basal motility by ANP in experimental colitis may be due an increased expression of colonic natriuretic peptide receptor-A mRNA. These data suggest that local natriuretic peptide system is partly involved in the pathophysiology of experimental colitis.

Atrial natriuretic peptide ameliorates peritoneal fibrosis in rat peritonitis model

Atrial natriuretic peptide (ANP) was recently reported to ameliorate fibrosis in the heart and experimental renal diseases and vascular thickening after balloon injury. Peritoneal fibrosis is an important complication of long-term peritoneal dialysis, and peritonitis is a factor in its onset. In the present study, we investigated the effects of ANP in a rat peritonitis-induced peritoneal fibrosis model. As pretreatment, an osmotic pump containing vehicle (saline) or ANP (0.15 or 0.3 μg/min) was inserted through the carotid vein in male Sprague-Dawley rats. ANP or saline was continuously infused using the osmotic pump. Three days after administration of ANP or saline, rats underwent peritoneal scraping in a blind manner and were sacrificed on Day 14. The effects of ANP were evaluated based on peritoneal thickness, immunohistochemistry and real-time polymerase chain reaction. In each experiment, we evaluated messenger RNA (mRNA) expression of the ANP receptor natriuretic peptide receptor A (NPR-A) in the peritoneum after scraping. The effects of ANP were also studied in cultured peritoneal fibroblasts and mesothelial cells. We observed a significant increase in NPR-A mRNA in the peritoneum. Peritoneal thickness increased with time and peaked on Day 14, but ANP significantly reduced peritoneal thickness. Parameters such as number of macrophages and CD-31-positive vessels and expression of type III collagen/transforming growth factor-β/plasminogen activator inhibitor-1 (PAI-1)/connective tissue growth factor (CTGF) were significantly suppressed by ANP. In cultured peritoneal fibroblasts and mesothelial cells, ANP suppressed angiotensin II-induced upregulation of CTGF and PAI-1. Our results suggest that ANP is useful in preventing inflammation-induced peritoneal fibrosis.

Atrial Natriuretic Peptide Stimulates Rat Pancreatic Beta‐Cell Growth

Natriuretic peptide receptor‐A (NPR‐A) is a guanylyl cyclase‐linked receptor that generates cGMP in response to atrial natriuretic peptide (ANP). Since elevated ANP levels are associated with diabetes mellitus and hyperglycemia, ANP effects on cultured isolated rat pancreatic islets and insulinoma cells (INS‐1e) were determined. Islets and INS‐1e cells expressed NPR‐A mRNA and protein. To determine the chronic effects of ANP, islets were cultured at 5.5 mM glucose with ANP (1 μM) for 7 days, which increased [3H]thymidine incorporation to 200±36% of control (p&lt;0.05); similarly, ANP increased INS‐1e cell [3H]thymidine incorporation to 163±18% of control (p&lt;0.05). 8‐Bromo‐cGMP (0.5 mM) also stimulated islet [3H]thymidine incorporation to 207±15% of control (p&lt;0.001). The mTOR inhibitor rapamycin failed to inhibit ANP‐stimulated [3H]thymidine incorporation. In ANP‐treated INS‐1e cells, the phosphatidylinositol 3‐kinase (PI3K)/Akt/Foxo1a pathway was activated. Phospho‐Akt and phospho‐Foxo1a were increased by 387±77% and 170 ±12% of control (p&lt;0.05), respectively, and this response was blocked by the PI3K inhibitor LY294002. Cyclin D2 mRNA was upregulated in ANP treated islets and INS‐1e cells by 336±71% and 332±36% of control (p&lt;0.05), respectively. The evidence suggests that the mitogenic response stimulated by ANP treatment involves activation of the PI3K/Akt/Foxo1a/cyclin D2 pathway.

Ontogeny of Atrial Natriuretic Peptide and Its Receptor In the Lung: Effects on Perinatal Surfactant Release

During the transition at birth to air breathing, regulation of surfactant release from alveolar type II (ATII) cells is critical. Atrial natriuretic peptide (ANP) stimulates natriuretic peptide receptor-A (NPR-A) and increases intracellular cGMP. We examined the changes in ANP and NPR-A in respiratory epithelium during the perinatal period using immunohistochemistry and studied the effect of ANP on surfactant release from ATII cells isolated from fetal and newborn lambs. NPR-A mRNA was detected in the fetal lung by Northern Blot and RT-PCR. At 100 d gestation (term 145 d), ANP staining was absent and NPR-A staining was weak in cuboidal epithelial cells. ANP and NPR-A staining was prominent in ATII cells at 136 d gestation and was undetectable postnatally. ANP stimulated (maximal effect at 10(-10)M) surfactant release from both late gestation fetal and neonatal ATII cells. Protein kinase G inhibition significantly blocked this release. We conclude that ANP stimulates surfactant release in isolated perinatal ATII cells by a cGMP-dependent mechanism. ANP and NPR-A expression in ATII cells is greatest in late gestation and declines sharply postnatally. We speculate that increased activity of the ANP/NPR-A pathway in late gestation may prime the surfactant system, preparing the lung for air breathing.

Natriuretic peptide guanylyl cyclase receptors in the kidney of the Japanese eel, Anguilla japonica

Natriuretic peptides are linked to osmoregulation, cardiovascular and volume regulation in fishes. The peptides bind to two guanylyl-cyclase-linked receptors, natriuretic peptide receptor-A (NPR-A) and NPR-B, to elicit their effects. Atrial natriuretic peptide (ANP) binds principally to NPR-A, whereas C-type natriuretic peptide (CNP) binds to NPR-B. The teleost kidney has an important role in the maintenance of fluid and electrolyte balance; therefore, the location of NPR-A and NPR-B in the kidney could provide insights into the functions of natriuretic peptides. This study used homologous, affinity purified, polyclonal antibodies to NPR-A and NPR-B to determine their location in the kidney of the Japanese eel, Anguilla japonica. Kidneys from freshwater and seawater acclimated animals were fixed overnight in 4% paraformaldehyde before being paraffin-embedded and immunostained. NPR-A immunoreactivity was found on the apical membrane of proximal tubule 1 and the vascular endothelium including the glomerular capillaries. In contrast, NPR-B immunoreactivity was located on the smooth muscle of blood vessels including the glomerular afferent and efferent arterioles, and on smooth muscle tissue surrounding the collecting ducts. No difference in the distribution of NPR-A and NPR-B was observed between freshwater and seawater kidneys. Immunoreactivity was not observed in any tissue in which the antibodies had been preabsorbed. In addition, there was no difference in NPR-A and NPR-B mRNA expression between freshwater-acclimated and seawater-acclimated eels. These results suggest that, although utilizing the same second messenger system, ANP and CNP act on different targets within the kidney and presumably elicit different effects.

Differential responses of newborn pulmonary arteries and veins to atrial and C-type natriuretic peptides.

Atrial natriuretic peptide (ANP) and C-type natriuretic peptide (CNP) are important dilators of the pulmonary circulation during the perinatal period. We compared the responses of pulmonary arteries (PA) and veins (PV) of newborn lambs to these peptides. ANP caused a greater relaxation of PA than of PV, and CNP caused a greater relaxation of PV than of PA. RIA showed that ANP induced a greater increase in cGMP content of PA than CNP. In PV, ANP and CNP caused a similar moderate increase in cGMP content. Receptor binding study showed more specific binding sites for ANP than for CNP in PA and more for CNP than for ANP in PV. Relative quantitative RT-PCR for natriuretic peptide receptor A (NPR-A) and B (NPR-B) mRNAs show that, in PA, NPR-A mRNA is more prevalent than NPR-B mRNA, whereas, in PV, NPR-B mRNA is more prevalent than NPR-A mRNA. In conclusion, in the pulmonary circulation, arteries are the major site of action for ANP, and veins are the major site for CNP. Furthermore, the differences in receptor abundance and the involvement of a cGMP-independent mechanism may contribute to the heterogeneous effects of the natriuretic peptides in PA and PV of newborn lambs.

Pulmonary natriuretic peptide system during rat development.

Maturational changes in the rat lung natriuretic peptide system were studied postnatally in 1-, 4-, and 22-day-old rats. Lung atrial natriuretic factor (ANF) content increased significantly from day 1 to day 4 (712+/-188 vs. 1905+/-520 pg/mg protein; p<0.01) but decreased to 532+/-41 pg/mg protein, on day 22. These changes paralleled ANF messenger RNA (mRNA) detected by reverse transcribed polymerase chain reaction (RT-PCR). Rat pulmonary development also was associated with quantitative and qualitative alterations in ANF receptors. Competitive-binding radioreceptor assays of lung membranes with 125I-ANF and increasing concentrations of unlabeled ANF revealed that the natriuretic peptide receptor-binding sites (Bmax) progressively increased with age from 112 +/-21 fmol/mg protein at day 1 to 211+/-16 (p<0.02) and 326+/-62 fmol/mg protein (p<0.04) in 4- and 22-day-old rats, respectively. Autoradiographic studies of 125I-tyr(0)CNP binding to lung sections revealed that the levels of the natriuretic peptide receptor B (NPR-B) were undetectable. On the other hand, binding of 125I-ANF increased with age, and the higher binding at 4 days was mainly due to increased density of the clearance receptor-C (NPR-C), and at 22 days due to increased natriuretic peptide receptor-A (NPR-A). The increase in natriuretic peptide binding was confirmed at the level of synthesis, where RT-PCR revealed that NPR-A mRNA significantly increased (p<0.01) in 22-day-old rats. In conclusion, these studies demonstrate that the rat pulmonary natriuretic peptide system is altered during development. The altered synthesis of lung natriuretic peptides and their receptors may play a role in the postnatal adaptation of pulmonary circulation.

Ligand-dependent regulation of NPR-A gene expression in inner medullary collecting duct cells.

Atrial natriuretic peptide (ANP) interacts with high-affinity, guanylyl cyclase-linked receptors in the inner medullary collecting duct (IMCD), where it exerts important regulatory control over sodium handling. We sought to determine whether receptor activity in these cells would be modulated (downregulated) by prolonged exposure to ligand. A number of natriuretic peptides (ANP, brain natriuretic peptide, and urodilatin) were found to decrease ligand-dependent natriuretic peptide receptor A (NPR-A) activity in IMCD cells. This inhibition was in direct proportion to their capacity to increase basal cGMP levels in this cell population. The reduction in receptor activity was accompanied by a dose- and time-dependent reduction in NPR-A mRNA levels in these cells. The decrease in transcript levels arose, in part, from a reduction in NPR-A gene transcription. ANP reduced NPR-A gene promoter activity in a transiently transfected IMCD cell population. 8-Bromo-cGMP was also effective in inhibiting NPR-A mRNA levels and NPR-A promoter activity, suggesting that the second messenger (i.e., cGMP) rather than ANP, itself, is responsible for downregulation of NPR-A gene expression.