kmup-1 and Disease-Models--Animal

To explore the potential mechanism of KMUP-1 in the vascular calcification of chronic renal failure (CRF) through mediating NO/cGMP/PKG pathway, and provide novel insights into the CRF treatment.. CRF rats were treated by KMUP-1 with/without L-NNA (a NOS inhibitor) and then performed by ELISA, alizarin red staining, Von Kossa staining, Masson's trichrome, Sirius red staining and CD3 immunohistochemical staining. Simultaneously, vascular smooth muscle cells (VSMCs) were collected from rats to confirm the effect of KMUP-1 on vascular calcification in vitro via NO/cGMP/PKG pathway. Besides, protein and mRNA expressions were determined via Western blotting and qRT-PCR, respectively.. CRF rats were elevated in 24-h urine protein, blood urea nitrogen (BUN), serum creatinine, Cys-C levels and inflammatory cytokines. Besides, CRF rats also showed increased calcium content and ALP level with up-regulated mRNA of osteogenic differentiation-related markers. Furthermore, the up-regulated expressions of eNOS and PKG, as well as down-regulated levels of NOx and cGMP were also found in CRF rats. However, renal failure and vascular calcification of CRF were improved significantly by KMUP-1 treatment via activation of NO/cGMP/PKG pathway. Moreover, KMUP-1 treatment attenuated calcified VSMCs, accompanied by the decreases in the calcified nodules, level of calcium and activity of ALP. In addition, either L-NNA treatment for CRF rats or the calcified VSMCs could antagonize the improving effect of KMUP-1.. KMUP-1 can improve the renal function and vascular calcification in CRF rats at least in part by activating NO/cGMP/PKG pathway.

Topics: Animals; Calcium; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Kidney Failure, Chronic; Male; Myocytes, Smooth Muscle; Nitric Oxide; Osteogenesis; Piperidines; Rats; Rats, Sprague-Dawley; Signal Transduction; Vascular Calcification; Xanthines

Pulmonary fibrosis (PF) is a progressing lung injury initiated by pulmonary inflammation (PI). Bleomycin (BLM) is the most common pathogenesis of PF through early PI and extensive extracellular matrix deposition. This study is aimed to determine whether NO-releasing KMUP-1 inhibits PI and PF, and if so, the benefits of KMUP-1S resulted from simvastatin (SIM)-bonding to KMUP-1.. C57BL/6 male mice were intra-tracheally administered BLM (4 U/kg) at day 0. KMUP-1 (1-5 mg/kg), KMUP-1S (2.5 mg/kg), SIM (5 mg/kg), Plus (KMUP-1 2.5 mg/kg + SIM 2.5 mg/kg), and clarithromycin (CAM, 10 mg/kg) were orally and daily administered for 7 and 28 days, respectively, to mice, sacrificed at day-7 and day-28 to isolate the lung tissues, for examining the inflammatory and fibrotic signaling and measuring the cell population and MMP-2/MMP-9 activity in broncholaveolar lavage fluid (BAL).. KMUP-1 and KUP-1S significantly decreased neutrophil counts in BAL fluid. Fibroblastic foci were histologically assessed by H&E and Masson's trichrome stain and treated with KMUP-1 and references. Lung tissues were determined the contents of collagen and the expressions of TGF-β, α-SMA, HMGB1, CTGF, eNOS, p-eNOS, RhoA, Smad3, p-Smad3, MMP-2 and MMP-9 by Western blotting analyses, respectively. These changes areregulated by NO/cGMP and inhibited by various treatments. KMUP-1 and KMUP-1S predominantly prevented HMGB1/MMP-2 expression at day-7 and reduced TGF-β/phosphorylated Smad3 and CTGF at day-28.. KMUP-1 and KMUP-S restore eNOS, inhibit iNOS/ROCKII/MMP-2/MMP-9, attenuate histologic collagen disposition and reduce BALF inflammatory cells, potentially useful for the treatment of BLM-lung PF.

Topics: Animals; Bleomycin; Blotting, Western; Bronchoalveolar Lavage Fluid; Clarithromycin; Disease Models, Animal; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Piperidines; Pneumonia; Pulmonary Fibrosis; Signal Transduction; Simvastatin; Time Factors; Xanthines

To investigate whether 7-[2-[4-(2-chlorophenyl) piperazinyl] ethyl]-1,3-di-methylxanthine (KMUP-1) inhibits the effects of testosterone on the development of benign prostatic hyperplasia and sensitizes prostate contraction.. A benign prostatic hyperplasia animal model was established by subcutaneous injections of testosterone (3 mg/kg/day, s.c.) for 4 weeks in adult male Sprague-Dawley rats. Animals were divided into six groups: control, testosterone, testosterone with KMUP-1 (2.5, 5 mg/kg/day), sildenafil (5 mg/kg/day) or doxazosin (5 mg/kg/day). After 4 weeks, the animals were killed, and prostate tissues were prepared for isometric tension measurement and western blotting analysis. KMUP-1, Y27632, zaprinast, doxazosin or tamsulosin were used at various concentrations to determine the contractility sensitized by phenylephrine (10 μmol/L).. KMUP-1 inhibited testosterone-induced phosphorylation of extracellular signal-regulated phosphorylated protein kinase and mitogen-activated protein kinase kinase and Rho kinase-II activation. Sildenafil and doxazosin significantly decreased benign prostatic hyperplasia-induced mitogen-activated protein kinase kinase and Rho kinase-II activation. The decreased expressions of soluble guanylate cyclase α1 was reversed by KMUP-1, doxazosin and sildenafil. Soluble guanylate cyclase β1 and protein kinase G were increased by KMUP-1, doxazosin, and sildenafil in the testosterone-treated benign prostatic hyperplasia group. Phosphodiesterase-5A was increased by testosterone and inhibited by KMUP-1 (5 mg/kg/day) or sildenafil (5 mg/kg/day). KMUP-1 inhibited phenylephrine-sensitized prostate contraction of rats treated with testosterone.. Mitogen-activated protein kinase 1/extracellular regulated protein kinases kinase, soluble guanylate cyclase/cyclic guanosine monophosphate, protein kinase/protein kinase G and Rho kinase-II are related to prostate smooth muscle tone and proliferation induced by testosterone. KMUP-1 inhibits testosterone-induced prostate hyper-contractility and mitogen-activated protein kinase 1/extracellular regulated protein kinases kinase-phosphorylation, and it inactivates Rho kinase-II by cyclic guanosine monophosphate, protein kinase and α1A-adenergic blockade. Thus, KMUP-1 might be a beneficial pharmacotherapy for benign prostatic hyperplasia.

Topics: Animals; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Guanylate Cyclase; Male; MAP Kinase Signaling System; Piperidines; Prostatic Hyperplasia; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; rho-Associated Kinases; Soluble Guanylyl Cyclase; Xanthines

This study investigates whether endothelin-1 (ET-1) mediates monocrotaline (MCT)-induced pulmonary artery hypertension (PAH) and right ventricular hypertrophy (RVH), and if so, whether the G-protein coupled receptor antagonist KMUP-1 (7-{2-[4-(2-chlorobenzene)piperazinyl]ethyl}-1,3-dimethylxanthine) inhibits ET-1-mediated PA constriction and the aforementioned pathological changes. In a chronic rat model, intraperitoneal MCT (60 mg/kg) induced PAH and increased PA medial wall thickening and RV/left ventricle + septum weight ratio on Day 21 after MCT injection. Treatment with sublingual KMUP-1 (2.5 mg/kg/day) for 21 days prevented these changes and restored vascular endothelial nitric oxide synthase (eNOS) immunohistochemical staining of lung tissues. Western blotting analysis demonstrated that KMUP-1 enhanced eNOS, soluble guanylate cyclase, and protein kinase G levels, and reduced ET-1 expression and inactivated Rho kinase II (ROCKII) in MCT-treated lung tissue over long-term administration. In MCT-treated rats, KMUP-1 decreased plasma ET-1 on Day 21. KMUP-1 (3.6 mg/kg) maximally appeared at 0.25 hours in the plasma and declined to basal levels within 24 hours after sublingual administration. In isolated PA of MCT-treated rats, compared with control and pretreatment with l-NG-nitroarginine methyl ester (100 μM), KMUP-1 (0.1-100 μM) inhibited ET-1 (0.01 μM)-induced vasoconstriction. Endothelium-denuded PA sustained higher contractility in the presence of KMUP-1. In a 24-hour culture of smooth muscle cells (i.e., PA smooth muscle cells or PASMCs), KMUP-1 (0.1-10 μM) inhibited RhoA- and ET-1-induced RhoA activation. KMUP-1 prevented MCT-induced PAH, PA wall thickening, and RVH by enhancing eNOS and suppressing ET-1/ROCKII expression. In vitro, KMUP-1 inhibited ET-1-induced PA constriction and ET-1-dependent/independent RhoA activation of PASMCs. In summary, KMUP-1 attenuates ET-1-induced/ET-1-mediated PA constriction, and could thus aid in the treatment of PAH caused by MCT.

Topics: Animals; Blood Pressure; Body Weight; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Endothelin-1; Guanylate Cyclase; Heart Rate; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; In Vitro Techniques; Male; Monocrotaline; Nitric Oxide Synthase Type III; Piperazines; Piperidines; Pulmonary Artery; Purines; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Receptors, G-Protein-Coupled; rho-Associated Kinases; rhoA GTP-Binding Protein; Signal Transduction; Sildenafil Citrate; Soluble Guanylyl Cyclase; Sulfonamides; Vasoconstriction; Xanthines

Axonal regeneration in peripheral nerves after injury is a complicated process. Numerous cytokines, growth factors, channels, kinases, and receptors are involved, and matrix metalloproteinase-9 (MMP-9) has been implicated in the pathogenesis subsequent to nerve injury. In this study, the effect of KMUP-1, an activator of large-conductance Ca(2+)-activated potassium channel, on functional recovery, myelinated axon growth, and immunoreactivity of MMP-9 was evaluated in rats subjected to sciatic nerve crush injury.. A total of 144 male Sprague-Dawley rats were divided into the following six groups (n = 24/group): group 1, sham-operated; group 2, sciatic nerve injury without treatment; group 3, injured and vehicle-treated; group 4, injured and treated with 1 mM KMUP-1 by topical application; group 5, injured and treated with 10 mM KMUP-1; group 6, injured and treated with 50 mM KMUP-1. Functional recovery was evaluated using walking track analysis at 1, 2, 3, and 4 weeks (n = 6/group at each time point) after injury. In addition, the number of myelinated axons and MMP-9 in the nerve was also examined.. Animals subjected to sciatic nerve crush injury had decreased motor function, a reduced number of myelinated axons, and increased MMP-9 in the nerve. Treatment with KMUP-1 concentration-dependently improved functional recovery, increased the number of myelinated axons, and decreased MMP-9.. These results suggest that KMUP-1 may be a novel agent for assisting peripheral nerve recovery after injury. The beneficial effect is probably due to known ability of the compound in activating the nitric oxide/cGMP/protein kinase G pathway.

Topics: Animals; Axons; Disease Models, Animal; Male; Matrix Metalloproteinase 9; Nerve Crush; Nerve Regeneration; Peripheral Nerve Injuries; Piperidines; Rats; Rats, Sprague-Dawley; Recovery of Function; Sciatic Nerve; Xanthines

To determine whether KMUP-1, a novel xanthine-based derivative, attenuates isoprenaline (ISO)-induced cardiac hypertrophy in rats, and if so, whether the anti-hypertrophic effect is mediated by the nitric oxide (NO) pathway.. In vivo, cardiac hypertrophy was induced by injection of ISO (5 mg.kg(-1).day(-1), s.c.) for 10 days in Wistar rats. In the treatment group, KMUP-1 was administered 1 h before ISO. After 10 days, effects of KMUP-1 on survival, cardiac hypertrophy and fibrosis, the NO/guanosine 3'5'-cyclic monophosphate (cGMP)/protein kinase G (PKG) and hypertrophy signalling pathways [calcineurin A and extracellular signal-regulated kinase (ERK)1/2] were examined. To investigate the role of nitric oxide synthase (NOS) in the effects of KMUP-1, a NOS inhibitor, N(omega)-nitro-L-arginine (L-NNA) was co-administered with KMUP-1. In vitro, anti-hypertrophic effects of KMUP-1 were studied in ISO-induced hypertrophic neonatal rat cardiomyocytes.. In vivo, KMUP-1 pretreatment attenuated the cardiac hypertrophy and fibrosis and improved the survival of ISO-treated rats. Plasma NOx (nitrite and nitrate) and cardiac endothelial NOS, cGMP and PKG were all increased by KMUP-1. The activation of hypertrophic signalling by calcineurin A and ERK1/2 in ISO-treated rats was also attenuated by KMUP-1. All these effects of KMUP-1 were inhibited by simultaneous administration of L-NNA. Similarly, in vitro, KMUP-1 attenuated hypertrophic responses and signalling induced by ISO in neonatal rat cardiomyocytes.. KMUP-1 attenuates the cardiac hypertrophy in rats induced by administration of ISO. These effects are mediated, at least in part, by NOS activation. This novel agent, which targets the NO/cGMP pathway, has a potential role in the prevention of cardiac hypertrophy.

Topics: Animals; Calcineurin; Cardiomegaly; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Drug Delivery Systems; Fibrosis; Isoproterenol; Male; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nitric Oxide; Nitric Oxide Synthase; Piperidines; Rats; Rats, Wistar; Signal Transduction; Survival Rate; Xanthines