Obestatin ameliorates water retention in chronic heart failure by downregulating renal aquaporin 2 through GPR39, V2R and PPARG signaling.

AIMS: Obestatin regulates water metabolism by inhibiting arginine vasopressin (AVP) release and upregulated obestatin has been detected in patients with chronic heart failure (CHF). However, the significance of obestatin in CHF, particularly with regard to water retention and aquaporin 2 (AQP2) expression, remains unknown. MAIN METHODS: Using a CHF rat model, the effects of 2-week exogenous obestatin administration were evaluated. Expression of AQP2 was evaluated by immunoblotting, immunohistochemical staining, and quantitative real-time PCR (qPCR) in CHF rat model and mouse inner medullary collecting duct (mIMCD) 3 cell line. Moreover, the influence of obestatin on the genetic transcription profile in mIMCD3 cells was evaluated by microarray, and the potential regulatory mechanisms of obestatin on AQP2 were evaluated by RNA silencing of vasopressin receptor 2 (V2R), peroxisome proliferator-activated receptor gamma (PPARG), and G protein-coupled receptor 39 (GPR39). KEY FINDINGS: Obestatin increased urinary output and improved expression of CHF biomarker without significantly altering cardiac function, plasma electrolyte concentrations, or the plasma AVP concentration. AQP2 expression was significantly reduced. The results of microarray analyses and qPCR indicated that mRNA levels of Aqp2, Pparg, and V2r were significantly decreased. Inhibition of V2r and Pparg mRNA further reduced the expression of AQP2, while the inhibitory efficacy of obestatin on AQP2 was significantly offset after Gpr39 knockdown. SIGNIFICANCE: Long-term treatment with obestatin improves water retention in CHF by increasing urinary output through downregulation of AQP2 expression in renal IMCD cells. These effects may be at least partially mediated by regulation of GPR39, V2R and PPARG signaling.

Obestatin Downregulating Aquaporin 2 Plasma Membrane Distribution Through a Short-Term Regulatory Effect.

BACKGROUND: Previous studies have found that obestatin significantly inhibited water drinking and reduced the arginine vasopressin levels in the brain to decrease renal water reabsorption. However, obestatin is unable to cross the blood-brain barrier. Its effect on the body's kidney water metabolism in peripheral remains unknown. MATERIALS AND METHODS: Expression and subcellular distribution of aquaporin 2 (AQP2) were detected by immunoblotting and immunofluorescence in mouse inner medullary collecting duct-3 (mIMCD-3) cells and congestive heart failure model rats. Moreover, expression of phosphorylated AQP2 (P-AQP2; Ser256) in mIMCD-3 cells was evaluated by immunoblotting. RESULTS: After a 30-minute treatment with obestatin in mIMCD-3 cells and congestive heart failure model rats, the AQP2 plasma membrane distribution decreased, while AQP2 protein level, P-AQP2 (Ser256) protein level and phosphorylation ratio of AQP2 showed no significant change. CONCLUSIONS: These findings suggest that obestatin has a short-term regulatory effect on the AQP2 plasma membrane distribution. In addition, obestatin decreases the APQ2 plasma membrane distribution probably by promoting the endocytosis of AQP2.

Increased expression of microRNA-378a-5p in acute ethanol exposure of rat cardiomyocytes.

Alcohol abuse is a risk factor for a distinct form of congestive heart failure, known as alcoholic cardiomyopathy (ACM). Here, we investigate how microRNAs may participate in the induction of cardiomyocyte apoptosis associated with ethanol exposure in vitro. Increasing the concentrations of ethanol to primary rat cardiomyocytes resulted in elevated apoptosis assessed by annexin V and propidium iodide staining, and reduced expression of an enzyme for alcohol detoxification aldehyde dehydrogenase 2 (ALDH2). These ethanol effects were accompanied by a substantial elevation of miR-378a-5p. Driving miR-378a-5p overexpression in cardiomyocytes decreased ALDH2. The specific interaction of miR-378a-5p with the 3'UTR of ALDH2 was examined by luciferase reporter assays, and we found that miR-378a-5p activity depends on a complementary base pairing at the 3'-UTR region of ALDH2 mRNA. Finally, ethanol-induced apoptosis in cardiomyocytes was attenuated in the presence of anti-miR378a-5p. Collectively, these data implicate a likely involvement of miR-378a-5p in the stimulation of cardiomyocyte apoptosis through ALDH2 gene suppression, which might play a potential role in the pathogenesis of ACM.

Adeno-associated virus-mediated osteoprotegerin gene transfer protects against joint destruction in a collagen-induced arthritis rat model.

OBJECTIVE: To evaluate the in vivo joint protection effect of recombinant adeno-associated virus-mediated gene transfer of human osteoprotegerin (rAAV-hOPG). METHODS: Collagen-induced arthritis (CIA) rat model was established. CIA rats were randomly divided into three groups: CIA control group (PBS), rAAV-EGFP (enhanced green fluorescent protein) group and rAAV-hOPG (100 µL/d) group, which received corresponding intra-articular injection treatment. The thickness of the palms and soles, arthritis index, radiological score, pathological score, bone damage factor and protein expression of inflammatory factors were measured and compared with normal control group rats. RESULTS: Positive fluorescence of frozen section confirmed that rAAV-hOPG was efficiently transduced into the synovial tissues of test rats. In rAAV-hOPG group compared with CIA control group, the radiological score was 30.18% lower (P<0.05); the expression of OPG protein was 93.41% higher (P<0.05); the expression of matrix metalloproteinase-3 (MMP-3) protein was 35.38% lower (P<0.05); however, the expression of IL-1ß was not significant; the scores of pannus and inflammation in rAAV-hOPG group have no significant difference. CONCLUSION: These results suggest that adeno-associated virus-mediated transfer of human osteoprotegerin is effectively transducted into the synovial tissues of CIA model, and protects against articular cartilage and bone destruction, but has no obvious efficiency on inflammation. The results also demonstrate that gene transfer using rAAV-hOPG may be a feasible and effective therapeutic candidate to treat or prevent joint destruction in inflammatory arthritis.