Optimizing myocardial cell protection with xanthine derivative KMUP-3 potentiates autophagy through the PI3K/Akt/eNOS axis.

BACKGROUND: Autophagy can have either beneficial or detrimental effects on various heart diseases. Pharmacological interventions improve cardiac function, which is correlated with enhanced autophagy. To assess whether a xanthine derivative (KMUP-3) treatment coincides with enhanced autophagy while also providing cardio-protection, we investigated the hypothesis that KMUP-3 treatment activation of autophagy through PI3K/Akt/eNOS signalling offered cardioprotective properties. METHODS: The pro-autophagic effect of KMUP-3 was performed in a neonatal rat model targeting cardiac fibroblasts and cardiomyocytes, and by assessing the impact of KMUP-3 treatment on cardiotoxicity, we used antimycin A-induced cardiomyocytes. RESULTS: As determined by transmission electron microscopy observation, KMUP-3 enhanced autophagosome formation in cardiac fibroblasts. Furthermore, KMUP-3 significantly increased the expressions of autophagy-related proteins, LC3 and Beclin-1, both in a time- and dose-dependent manner; moreover, the pro-autophagy and nitric oxide enhancement effects of KMUP-3 were abolished by inhibitors targeting eNOS and PI3K in cardiac fibroblasts and cardiomyocytes. Notably, KMUP-3 ameliorated cytotoxic effects induced by antimycin A, demonstrating its protective autophagic response. CONCLUSION: These findings enable the core pathway of PI3K/Akt/eNOS axis in KMUP-3-enhanced autophagy activation and suggest its principal role in safeguarding against cardiotoxicity.

KMUP-1, a GPCR Modulator, Attenuates Triglyceride Accumulation Involved MAPKs/Akt/PPARγ and PKA/PKG/HSL Signaling in 3T3-L1 Preadipocytes.

Xanthine-based KMUP-1 was shown to inhibit phosphodiesterases (PDEs) and modulate G-protein coupled receptors (GPCRs) to lower hyperlipidemia and body weight. This study further investigated whether KMUP-1 affects adipogenesis and lipolysis in 3T3-L1 preadipocytes. KMUP-1 (1⁻40 µM) concentration-dependently attenuated Oil Red O (ORO) staining and decreased triglyceride (TG) accumulation, indicating adipogenesis inhibition in 3T3-L1 cells. In contrast, the ß-agonist ractopamine increased ORO staining and TG accumulation and adipogenesis. KMUP-1 (1⁻40 µM) also reduced MAPKs/Akt/PPARγ expression, PPARγ1/PPARγ2 mRNA, and p-ERK immunoreactivity at the adipogenesis stage, but enhanced hormone sensitive lipase (HSL) immunoreactivity at the lipolysis stage. Addition of protein kinase A (PKA) or protein kinase G (PKG) antagonist (KT5720 or KT5728) to adipocytes did not affect HSL immunoreactivity. However, KMUP-1 did increase HSL immunoreactivity and the effect was reduced by PKA or PKG antagonist. Simvastatin, theophylline, caffeine, and sildenafil, like KMUP-1, also enhanced HSL immunoreactivity. Phosphorylated HSL (p-HSL) was enhanced by KMUP-1, indicating increased lipolysis in mature 3T3-L1 adipocytes. Decreases of MAPKs/Akt/PPARγ during adipogenesis contributed to inhibition of adipocyte differentiation, and increases of PKA/PKG at lipolysis contributed to HSL activation and TG hydrolysis. Taken together, the data suggest that KMUP-1 can inhibit hyperadiposity in 3T3-L1 adipocytes.

Xanthine-based KMUP-1 improves HDL via PPARγ/SR-B1, LDL via LDLRs, and HSL via PKA/PKG for hepatic fat loss.

The phosphodiesterase inhibitor (PDEI)/eNOS enhancer KMUP-1, targeting G-protein coupled receptors (GPCRs), improves dyslipidemia. We compared its lipid-lowering effects with simvastatin and explored hormone-sensitive lipase (HSL) translocation in hepatic fat loss. KMUP-1 HCl (1, 2.5, and 5 mg/kg/day) and simvastatin (5 mg/kg/day) were administered in C57BL/6J male mice fed a high-fat diet (HFD) by gavage for 8 weeks. KMUP-1 inhibited HFD-induced plasma/liver TG, total cholesterol, and LDL; increased HDL/3-hydroxy-3-methylglutaryl-CoA reductase (HMGR)/Rho kinase II (ROCK II)/PPARγ/ABCA1; and decreased liver and body weight. KMUP-1 HCl in drinking water (2.5 mg/200 ml tap water) for 1-14 or 8-14 weeks decreased HFD-induced liver and body weight and scavenger receptor class B type I expression and increased protein kinase A (PKA)/PKG/LDLRs/HSL expression and immunoreactivity. In HepG2 cells incubated with serum or exogenous mevalonate, KMUP-1 (10(-7)∼10(-5) M) reversed HMGR expression by feedback regulation, colocalized expression of ABCA1/apolipoprotein A-I/LXRα/PPARγ, and reduced exogenous geranylgeranyl pyrophosphate/farnesyl pyrophosphate (FPP)-induced RhoA/ROCK II expression. A guanosine 3',5'-cyclic monophosphate (cGMP) antagonist reversed KMUP-1-induced ROCK II reduction, indicating cGMP/eNOS involvement. KMUP-1 inceased PKG and LDLRs surrounded by LDL and restored oxidized LDL-induced PKA expresion. Unlike simvastatin, KMUP-1 could not inhibit (14)C mevalonate formation. KMUP-1 could, but simvastatin could not, decrease ROCK II expression by exogenous FPP/CGPP. KMUP-1 improves HDL via PPARγ/LXRα/ABCA1/Apo-I expression and increases LDLRs/PKA/PKG/HSL expression and immunoreactivity, leading to TG hydrolysis to lower hepatic fat and body weight.

KMUP-1 inhibits pulmonary artery proliferation by targeting serotonin receptors/transporter and NO synthase, inactivating RhoA and suppressing AKT/ERK phosphorylation.

KMUP-1 inhibits monocrotaline (MCT)-induced pulmonary artery (PA) proliferation by targeting serotonin (5-HT) receptors, inactivating RhoA and reducing phosphorylation of AKT/ERK. In MCT-treated rats, KMUP-1 f (5 mg/kg p.o.; 1mg/kg i.p.x 21 days) decreased proliferation (PCNA-positive) cells and 5-HTT-expression in lung and 5-HT levels in plasma. In isolated PA, KMUP-1 and simvastatin (0.1-100 µM) inhibited 5-HT (10 µM)-induced PA constriction. l-NAME-pretreatment reduced KMUP-1-induced relaxation. In pulmonary arterial smooth muscle cells (PASMCs), KMUP-1 (1-100 µM) and simvastatin (10 µM) inhibited 5-HT-induced cell migration and proliferation and KMUP-1 (1-100 µM) inhibited 5-HT-induced Ca²+ influx. Similar to Y27632, KMUP-1 (1-100 µM) inhibited 5-HT-induced RhoA/ROCK expression, while KMUP-1, Y27632 and simvastatin at 10 µM inhibited 5-HT-induced 5-HTT expression and KMUP-1 inhibited 5-HT-induced phosphorylation of AKT and ERK1/2 in PASMCs. In human pulmonary arterial endothelial cell (HPAEC), KMUP-1 (1-100 µM) increased the expression of eNOS and 5-HT(2B) and also at 10 µM augmented eNOS expression and production of nitric oxide (NO) in 5-HT-treated HPAEC. In radioligand binding, the IC50/K(i) values of KMUP-1 for 5-HT(2A), 5-HT(2B) and 5-HT(2C) receptors were 0.34/0.0971, 0.04/0.0254, and 0.408/0.214 µM respectively. In conclusion, KMUP-1 inhibits MCT-induced PA proliferation by binding to 5-HT(2A), 5-HT(2B) and 5-HT(2C) receptors, increasing endothelial eNOS/5-HT(2B) receptor expression and NO release and inhibiting 5-HTT/RhoA/ROCK expression and AKT/ERK phosphorylation. KMUP-1 is suggested to be useful in the treatment of 5-HT-induced pulmonary artery proliferation.

Association of amino-terminal propeptide of type III procollagen and acute myocardial rejection in male patients receiving heart transplantation.

BACKGROUND: The amino-terminal propeptides of type I and III procollagens (PINP and PIIINP) are markers reflecting the status of collagen turnover. We hypothesized that measurement of these serum procollagen propeptides could be used to non-invasively assess acute rejection in heart transplant recipients. METHODS: In heart transplant recipients, endomyocardial biopsy specimens taken at 6 and 12 months after surgery were used for study. PINP and PIIINP were measured postoperatively at 3, 6, and 12 months. RESULTS: A total of 20 male heart transplant patients and seven male control subjects were enrolled. Five patients showed rejection 6 months after transplantation (group 1), while 15 patients showed no rejection (group 2). In group 2 patients, serum PINP and PIIINP levels decreased significantly 6 months after transplantation. In contrast, elevation of serum PINP and PIIINP levels persisted in group 1 patients 6 months after transplantation. At 6 months after transplantation, group 1 patients had significantly higher PIIINP levels than group 2 patients (p=0.025) and controls (p=0.003). After immunosuppressive therapy, all group 1 patients were free of rejection 12 months after transplantation and serial serum PIIINP levels decreased significantly in these patients. CONCLUSIONS: Serum PIIINP levels represent a non-invasive method to reflect the occurrence and resolution of acute rejection.

The influence of estimated creatinine clearance on plasma homocysteine in hypertensive patients with normal serum creatinine.

OBJECTIVES: To examine the relation of estimated creatinine clearance (eCrCl) and plasma total homocysteine (tHcy) in hypertensive patients with a normal serum creatinine level. DESIGN AND METHODS: A total of 137 hypertensive patients (mean age 66.6 years, 69 men) with serum creatinine level 60 mL/min/1.73 m(2)). The CRI group was older (p<0.001), had higher tHcy (p<0.001), higher serum urea nitrogen (p<0.001), higher serum creatinine (p<0.001), lower eCrCl (p<0.001), and lower diastolic blood pressure (p=0.001). In univariate analysis, eCrCl had the strongest correlation with tHcy (r=-0.453, p<0.001). Significant correlations, ranging in decreasing order from r=-0.418, p<0.001 to r=-0.170, p=0.047, were also noted between tHcy and twelve other variables. In multivariate analysis, only eCrCl (p<0.001), usage of fibrate (p<0.001), serum level of vitamin B(12) (p=0.002), serum level of folic acid (p=0.009), and smoking (p=0.027) were independent predictors of tHcy. CONCLUSION: eCrCl is a strong independent predictor of tHcy in hypertensive patients with normal serum creatinine.