Activation of SUR2B/Kir6.1-type K ATP channels protects glomerular endothelial,mesangial and tubular epithelial cells against oleic acid renal damage

Cumulative evidence suggests that renal vascular endothelial injury play an important role in initiating and extending tubular epithelial injury and contribute to the development of ischemic acute renal failure.Our previous studies have demonstrated that iptakalim's endothelium protection is related to activation of SUR2B/Kir6.1 subtype of ATP sensitive potassium channel(K ATP) in the endothelium.It has been reported that SUR2B/Kir6.1 channels are widely distributed in the tubular epithelium,glomerular mesangium,and the endothelium and the smooth muscle of blood vessels.Herein,we hypothesized that activating renal K ATP channels with iptakalim might have directly neroprotective effects.In this study,glomerular endothelial,mesangial and tubular epithelial cells which are the main cell types to form nephron were exposed to oleic acid(OA) at various concentrations for 24 h.0.25 μl/ml OA could cause cellular damage of glomerular endothelium and mesangium,while 1.25μl/ml OA could lead to the injury of three types of renal cells.It was observed that pretreatment with iptakalim at concentrations of 0.1,1,10 or 100 μmol/L prevented cellular damage of glomerular endothelium and tubular epithelium,whereas iptakalim from 1 to 100 μmol/L prevented the injury of mesangial cells.Our data showed iptakalim significantly increased survived cell rates in a concentration-dependent manner,significantly antagonized by glibenclamide,a K ATP blocker.Iptakalim played a protective role in the main cell types of kidney,which was consistent with natakalim,a highly selective SUR2B/Kir6.1 channel opener.Iptakalim exerted protective effects through activating SUR2B/Kir6.1 channels,suggesting a new strategy for renal injury by its endothelial and renal cell protection.

Synergisms of cardiovascular effects between iptakalim and amlodipine, hydrochlorothiazide or propranolol in anesthetized rats

The primary object of this fundamental research was to survey the synergistic cardiovascular effects of iptakalim, a novel ATP–sensitive potassium channel(KATP) opener, and clinical first-line antihypertensive drugs, such as calcium antagonists, thiazide diuretics and β receptor blockers by a 2×2 factorial-design experiment. It would provide a theoretical basis for the development of new combined antihypertensive therapy program after iptakalim is applied to the clinic. Amlodipine besylate, hydrochlorothiazide and propranolol were chosen as clinical first-line antihypertensive drugs. Blood pressure, heart rate(HR) and cardiac functions were observed in anesthetized normal rats by an eightchannel physiological recorder. The results showed that iptakalim monotherapy in a low dose could produce significant antihypertensive effect. There was no interaction between iptakalim and amlodipine on the maximal changes of systolic blood pressure(SBP), diastolic blood pressure(DBP), mean arterial blood pressure(MABP), the left ventricular systolic pressure(LVSP), and the left ventricular end-diastolic pressure(LVEDP)(P>0.05). However, the effects of combination iptakalim/amlodipine on the maximal changes of SBP, DBP, MABP, LVSP and LVEDP were more obvious than those of iptakalim or amlodipine monotherapy. And there was strong positive interaction between iptakalim and amlodipine on the maximal changes of HR(P>0.05). According to the maximal changes of DBP, MABP, LVSP and LVEDP(P<0.05) of combination iptakalim with hydrochlorothiazide, there was strong positive interaction between them. But there was no interaction between iptakalim and hydrochlorothiazide on the maximal drop of SBP and HR(P>0.05). According to the maximal drops of DBP, MABP of combination iptakalim with propranolol, there was strong positive interaction between them(P<0.05). But there was no interaction between iptakalim and propranolol on the maximal changes of SBP, LVSP,LVEDP and HR(P>0.05). In conclusion, it was the first time to study the effects of amlodipine, hydrochlorothiazide or propranolol, which had different mechanisms of action from iptakalim, on cardiovascular effects of iptakalim in anesthetized normal rats. This study proved that the combination of iptakalim with hydrochlorothiazide or propranolol respectively had significant synergism on lowering blood pressure, while the combination of iptakalim/amlodipine had additive action on lowering blood pressure. Meanwhile the antihypertensive effect was explicit, stable and long-lasting. Iptakalim thus appears suitable for the clinical treatment of hypertensive people who need two or more kinds of antihypertensive agents.

Modulation by desensitized nicotinic receptors on metabolism of DA in striatum derived from the hemiparkinsonian model

Objective: The purpose of this study was to investigate the effects of desensitized nicotinic receptors(n ACh Rs) on striatal dopaminergic system in the hemiparkinsonian rats treated with 6-hydroxydopamine(6-OHDA). Methods: We examined the effects of desensitized n ACh Rs on the levels of dopamine(DA) and its metabolites, m RNA expression of dopamine receptor D1,D2 and monoamine oxidase B(MAO-B) in the striatum of 6-OHDA-lesioned rats using high-performance liquid chromatography and reverse transcription-polymerase chain reaction. Results: The results showed that n ACh Rs desensitization following repeated nicotine stimulation could reverse significantly the decrease of striatal DA and its metabolites levels and the increase in DA turnover in lesioned side striatum of hemiparkinsonian rats. Dopamine D1 receptor m RNA expression increased significantly, whereas dopamine D2 receptor m RNA expression remained unchanged in lesioned side striatum of nicotine-treated rats compared to 6-OHDA-lesioned rats when n ACh Rs were desensitized. Meanwhile, nicotine-treated rats displayed a significant decrease in MAO-B m RNA expression in lesioned side striatum compared to 6-OHDA-lesioned rats after n ACh Rs desensitization. Conclusion: These results suggest that n ACh Rs desensitization could promote DA level, upregulate dopamine D1 receptor expression and downregulate MAO-B expression in striatum of hemiparkinsonian rats.

Stimulation of endothelial non-neuronal muscarinic receptor attenuates the progression of atherosclerosis via inhibiting endothelial cells activation

Objective To investigate the effects of non-neuronal muscarinic receptors(NNMR) stimulation on atherosclerosis and endothelial cells activation.Methods Atherosclerosis model was established in ApoE^(-/-)mice by a high fat diet for 7 weeks.During the experimental periods,animals were received a low(7 mg/kg/d) or a high(21 mg/kg/d) dose of arecoline by gavage.At the termination of the treatments,serum total cholesterol and NO levels were measured,and the aorta morphology was analyzed by hematoxylin and eosin staining.The gene expression of monocyte chemoattractant protein-1(MCP-1) and adhesion molecules in the thoracic aortas was determined by RT-PCR,and the MCP-1 protein expression and NFkB activity were detected by Western blot analysis.NO production,MCP-1 secretion in cultured rat aortic endothelial cells(RAECs),and monocyte-endothelium adhesion assay were also performed after arecoline treatments.Results Arecoline efficiently decreased atherosclerotic plaque areas,increased serum nitric oxide(NO) content,suppressed the mRNA and protein expression of MCP-1,and modulated the IκB-αdegradation and P65 phosphorylation in the aortae of ApoE^(-/-) mice.Furthermore,arecoline promoted NO production and suppressed MCP-1 secretion in cultured RAECs after ox-LDL exposure,and either atropine or NG-nitro-L-arginine methylester could abrogate these effects.Arecoline also significantly inhibited the adherence of U937 monocytes to the ox-LDL injured human umbilical vein endothelial cells,which could be abolished by atropine.Conclusion Our results indicate that arecoline attenuates the progression of atherosclerosis and inhibits endothelial cells activation and adherence by stimulating endothelial NNMR.These effects,at least in part,are due to its modulation on NF-kB activity.

Iptakalim,an ATP-sensitive potassium channel opener,confers neuroprotection against cerebral ischemia/reperfusion injury in rats by protecting neurovascular unit cells

Objective:To investigate the role of iptakalim,an ATP-sensitive potassium channel opener,in transient cerebral ischemia/reperfusion (I/R) injury and its involved mechanisms.Methods:Intraluminal occlusion of middle cerebral artery (MCAO) in a rat model was used to investigate the effect of iptakalim at different time points.Infarct volume was measured by staining with 2,3,5-triphenyltetrazolium chloride,and immunohistochemistry was used to evaluate the expressions of Bcl-2 and Bax.In vitro,neurovascular unit (NVU) cells,including rat primary cortical neurons,astrocytes,and cerebral microvascular endothelial cells,were cultured and underwent oxygen-glucose deprivation (OGD).The protective effect of iptakalim on NVU cells was investigated by cell viability and injury assessments,which were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and release of lactate dehydrogenase.Caspase-3,Bcl-2 and Bax mRNA expressions were evaluated by real-time polymerase chain reaction (PCR).Results:Administration of iptakalim 0 or 1 h after reperfusion significantly reduced infarct volumes,improved neurological scores,and attenuated brain edema after cerebral I/R injury.Iptakalim treatment (0 h after reperfusion) also reduced caspase-3 expression and increased the ratio of Bcl-2 to Bax by immunohistochemistry.Iptakalim inhibited OGD-induced cell death in cultured neurons and astrocytes,and lactate dehydrogenase release from cerebral microvascular endothelial cells.Iptakalim reduced mRNA expression of caspase-3 and increased the ratio of Bcl-2 to Bax in NVU cells.Conclusions:Iptakalim confers neuroprotection against cerebral I/R injury by protecting NVU cells via inhibiting of apoptosis.