红景天苷上调HIF-1α减轻高糖诱导的大鼠肾小球内皮细胞损伤
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摘要
目的:观察体外不同浓度葡萄糖对大鼠肾小球内皮细胞(rRGECs)表达低氧诱导因子1α(HIF-1α)、血管内皮生长因子A(VEGFA)及血管内皮钙黏素(VE-cadherin)的影响,探讨红景天苷减轻高糖诱导rRGECs损伤的作用及可能相关机制。方法:体外培养rRGECs,分为正常糖组、高糖(20、30和50 mmol/L)组、高渗组及红景天苷+高糖组。采用MTT法检测rRGECs的活力;RT-qPCR法检测rRGECs内HIF-1α、VEGFA及VE-cadherin的mRNA表达;Western blot法检测rRGECs内HIF-1α蛋白的表达。结果:与正常糖组相比,培养24 h后,高糖(20mmol/L)组rRGECs HIF-1α的mRNA及蛋白表达均上调(P <0.05);培养120 h后,高糖组HIF-1αmRNA表达下调(P <0.05)。与正常糖组相比,培养24 h和120 h后,高糖组rRGECs内VE-cadherin的mRNA表达下调(P <0.05)。与正常糖组相比,培养24 h后,高糖组rRGECs内VEGFA的mRNA表达上调(P <0.05);培养120 h后,高糖组rRGECs内VEGFA的mRNA表达下调(P <0.05)。与正常糖组相比,培养24 h和120 h后,高渗组rRGECs内HIF-1α、VE-cadherin及VEGFA的mRNA表达无变化。与高糖组相比,培养24 h后,红景天苷(50μmol/L)组rRGECs的活力增加(P <0.01),且细胞内HIF-1α和VE-cadherin的mRNA及HIF-1α蛋白表达均上调(P <0.05)。结论:体外高糖培养能影响rRGECs表达HIF-1α,可能与细胞活力、葡萄糖的浓度、作用时间及HIF/VEGF通路有关。红景天苷能减轻高糖诱导的rRGECs损伤,其机制可能与增加rRGECs内HIF-1α的表达有关。
AIM:To observe the expression of hypoxia-inducible factor-1α(HIF-1α),vascular endothelial growth factor A(VEGFA)and vascular endothelial cadherin(VE-cadherin)in cultured rat renal glomerular endothelial cells(rRGECs)exposed to glucose at different concentrations in vitro,and to verify the hypothesis that salidroside attenuates high glucose(HG)-induced injury of rRGECs by boosting HIF-1αlevel.METHODS:rRGECs were divided into 4group:normal glucose(NG)group,HG groups,hypertonic group and salidroside+HG group.The viability of rRGECs was measured by MTT assay.The mRNA expression of VEGFA,VE-cadherin and HIF-1αwas detected by RT-qPCR.The protein expression of HIF-1αwas determined by Western blot.RESULTS:Compared with NG group,the mRNA and protein expression of HIF-1αwas increased when the rRGECs were treated with glucose at concentration of 20 mmol/L for 24h(P<0.01).Compared with NG group,the mRNA expression of HIF-1αwas decreased in HG groups for 120 h(P<0.05).Compared with NG group,the mRNA expression of VE-cadherin was significantly down-regulated in HG groups for24 h or 120 h(P<0.05).Compared with NG group,the mRNA expression of VEGFA was increased in HG groups at 24h(P<0.05),while the mRNA expression of VEGFA was decreased at 120 h(P<0.05).Compared with NG group,no statistical difference in the mRNA expression levels of HIF-1α,VE-cadherin and VEGFA in DM group was observed.Compared with HG group,salidroside promoted the viability of rRGECs(P<0.01),and up-regulated the mRNA expression of HIF-1αand VE-cadherin,and the protein expression of HIF-1α(P<0.05).CONCLUSION:High glucose regulates HIF-1αexpression in rRGECs in connection with cell viability,the concentration of glucose,the culture time and HIF/VEGF signaling.Salidroside promotes rRGEC growth against high glucose-induced cell apoptosis via up-regulating the expression of HIF-1α.
AIM:To observe the expression of hypoxia-inducible factor-1α(HIF-1α),vascular endothelial growth factor A(VEGFA)and vascular endothelial cadherin(VE-cadherin)in cultured rat renal glomerular endothelial cells(rRGECs)exposed to glucose at different concentrations in vitro,and to verify the hypothesis that salidroside attenuates high glucose(HG)-induced injury of rRGECs by boosting HIF-1αlevel.METHODS:rRGECs were divided into 4group:normal glucose(NG)group,HG groups,hypertonic group and salidroside+HG group.The viability of rRGECs was measured by MTT assay.The mRNA expression of VEGFA,VE-cadherin and HIF-1αwas detected by RT-qPCR.The protein expression of HIF-1αwas determined by Western blot.RESULTS:Compared with NG group,the mRNA and protein expression of HIF-1αwas increased when the rRGECs were treated with glucose at concentration of 20 mmol/L for 24h(P<0.01).Compared with NG group,the mRNA expression of HIF-1αwas decreased in HG groups for 120 h(P<0.05).Compared with NG group,the mRNA expression of VE-cadherin was significantly down-regulated in HG groups for24 h or 120 h(P<0.05).Compared with NG group,the mRNA expression of VEGFA was increased in HG groups at 24h(P<0.05),while the mRNA expression of VEGFA was decreased at 120 h(P<0.05).Compared with NG group,no statistical difference in the mRNA expression levels of HIF-1α,VE-cadherin and VEGFA in DM group was observed.Compared with HG group,salidroside promoted the viability of rRGECs(P<0.01),and up-regulated the mRNA expression of HIF-1αand VE-cadherin,and the protein expression of HIF-1α(P<0.05).CONCLUSION:High glucose regulates HIF-1αexpression in rRGECs in connection with cell viability,the concentration of glucose,the culture time and HIF/VEGF signaling.Salidroside promotes rRGEC growth against high glucose-induced cell apoptosis via up-regulating the expression of HIF-1α.
引文
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[22]Zheng KY,Zhang ZX,Guo AJ,et al.Salidroside stimulates the accumulation of HIF-1alpha protein resulted in the induction of EPO expression:a signaling via blocking the degradation pathway in kidney and liver cells[J].Eur J Pharmacol,2012,679(1-3):34-39.
[23]Guo XQ,Qi L,Yang J,et al.Salidroside accelerates fracture healing through cell-autonomous and non-autonomous effects on osteoblasts[J].Cell Tissue Res,2017,367(2):197-211.
[24]Li QY,Wang HM,Wang ZQ,et al.Salidroside attenuates hypoxia-induced abnormal processing of amyloid precursor protein by decreasing BACE1 expression in SH-SY5Y cells[J].Neurosci Lett,2010,481(3):154-158.
[25]Li D,Fu Y,Zhang W,et al.Salidroside attenuates inflammatory responses by suppressing nuclear factor-κB and mitogen activated protein kinases activation in lipopolysaccharide-induced mastitis in mice[J].Inflamm Res,2013,62(1):9-15.
[2]Maezawa Y,Takemoto M,Yokote K.Cell biology of diabetic nephropathy:roles of endothelial cells,tubulointerstitial cells and podocytes[J].J Diabetes Investig,2015,6(1):3-15.
[3]Lee JW,Bae SH,Jeong JW,et al.Hypoxia-inducible factor(HIF-1)α:its protein stability and biological functions[J].Exp Mol Med,2004,36(1):1-12.
[4]Vordermark D,Kraft P,Katzer A,et al.Glucose requirement for hypoxic accumulation of hypoxia-inducible factor-1α(HIF-1α)[J].Cancer Lett,2005,230(1):122-133.
[5]Xiao H,Gu Z,Wang G,et al.The possible mechanisms underlying the impairment of HIF-1αpathway signaling in hyperglycemia and the beneficial effects of certain therapies[J].Int J Med Sci,2013,10(10):1412-1421.
[6]Schenck CA,Maeda HA.Tyrosine biosynthesis,metabolism,and catabolism in plants[J].Phytochemistry,2018,149:82-102.
[7]黄菲菲,李耀浙,张婷,等.红景天苷通过抑制氧化应激防治大鼠低氧性肺动脉高压[J].中国病理生理杂志,2018,34(3):500-506.
[8]Khanna K,Mishra KP,Ganju J,et al.Golden root:a wholesome treat of immunity[J].Biomed Pharmacother,2017,87:496-502.
[9]张华,周光群.红景天甙减轻大鼠酒精性肝损伤[J].中国病理生理杂志,2018,34(7):1311-1316.
[10]梁淼,张压西,熊玮.红景天联合氯沙坦对早期糖尿病肾病的临床疗效观察[J].中国中西医结合肾病杂志,2008,9(8):694-697.
[11]Gilbert RE.The endothelium in diabetic nephropathy[J].Curr Atheroscler Rep,2014,16(5):410.
[12]Gavard J.Endothelial permeability and VE-cadherin:a wacky comradeship[J].Cell Adh Migr,2014,8(2):158-164.
[13]Singh A,Friden V,Dasgupta I,et al.High glucose causes dysfunction of the human glomerular endothelial glycocalyx[J].Am J Physiol Renal Physiol,2011,300(1):F40-F48.
[14]Zhao L,Zhao J,Wang X,et al.Serum response factor induces endothelial-mesenchymal transition in glomerular endothelial cells to aggravate proteinuria in diabetic nephropathy[J].Physiol Genomics,2016,48(10):711-718.
[15]Miyata T,Suzuki N,van Ypersele DSC.Diabetic nephropathy:are there new and potentially promising therapies targeting oxygen biology?[J].Kidney Int,2013,84(4):693-702.
[16]Marfella R,D'Amico M,Di Filippo C,et al.Myocardial infarction in diabetic rats:role of hyperglycaemia on infarct size and early expression of hypoxia-inducible factor 1[J].Diabetologia,2002,45(8):1172-1181.
[17]Malhotra R,Tyson DG,Sone H,et al.Glucose uptake and adenoviral mediated GLUT1 infection decrease hypoxiainduced HIF-1alpha levels in cardiac myocytes[J].J Mol Cell Cardiol,2002,34(8):1063-1073.
[18]Nishikawa T,Araki E.Impact of mitochondrial ROS production in the pathogenesis of diabetes mellitus and its complications[J].Antioxid Redox Signal,2007,9(3):343-353.
[19]Klimova T,Chandel NS.Mitochondrial complex III regulates hypoxic activation of HIF[J].Cell Death Differ,2008,15(4):660-666.
[20]Kimura H,Weisz A,Kurashima Y,et al.Hypoxia response element of the human vascular endothelial growth factor gene mediates transcriptional regulation by nitric oxide:control of hypoxia-inducible factor-1 activity by nitric oxide[J].Blood,2000,95(1):189-197.
[21]Inoguchi T,Li P,Umeda F,et al.High glucose level and free fatty acid stimulate reactive oxygen species production through protein kinase C-dependent activation of NAD(P)Hoxidase in cultured vascular cells[J].Diabetes,2000,49(11):1939-1945.
[22]Zheng KY,Zhang ZX,Guo AJ,et al.Salidroside stimulates the accumulation of HIF-1alpha protein resulted in the induction of EPO expression:a signaling via blocking the degradation pathway in kidney and liver cells[J].Eur J Pharmacol,2012,679(1-3):34-39.
[23]Guo XQ,Qi L,Yang J,et al.Salidroside accelerates fracture healing through cell-autonomous and non-autonomous effects on osteoblasts[J].Cell Tissue Res,2017,367(2):197-211.
[24]Li QY,Wang HM,Wang ZQ,et al.Salidroside attenuates hypoxia-induced abnormal processing of amyloid precursor protein by decreasing BACE1 expression in SH-SY5Y cells[J].Neurosci Lett,2010,481(3):154-158.
[25]Li D,Fu Y,Zhang W,et al.Salidroside attenuates inflammatory responses by suppressing nuclear factor-κB and mitogen activated protein kinases activation in lipopolysaccharide-induced mastitis in mice[J].Inflamm Res,2013,62(1):9-15.