糖尿病肾脏疾病患者转化生长因子调节元件结合蛋白β1对胆固醇1活化和功能的调控作用研究
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摘要
目的探讨转化生长因子β1(TGF-β1)在DKD发病中的作用机制。方法 TGF-β1(2 ng/ml)和/或高糖(HG,24. 4 mmol/L)处理肾小球系膜细胞(GMC),Western blot检测胆固醇调节元件结合蛋白1(SREBP-1)和脂肪酸合成酶(FAS),油红O确定GMC中脂滴形成;SD大鼠左肾灌注携带TGF-β1基因腺病毒载体(AdTGF-β1)和空载体(AdLacZ)。处死后取左肾,免疫组织化学检测SREBP-1和p-Smad3蛋白表达。结果与正常细胞对照组(Con)比较,TGF-β1作用于GMC后,mSREBP-1蛋白表达增加,且呈时间依赖性[(1. 06±0. 55)vs(1. 50±0. 44)vs(1. 59±0. 68)],[(1. 23±0. 52)vs(1. 34±0. 65)];TGF-β1+HG作用也增加了mSREBP-1的蛋白表达。与Con组比较,TGF-β1处理增加了GMC FAS的蛋白表达[(1. 18±0. 04)vs(1. 38±0. 11)vs(1. 40±0. 13),P<0. 05],油红O证实GMC出现脂滴,呈红染颗粒状;大鼠肾脏灌注AdTGF-β1免疫组织化学法发现,TGF-β1能明显增加SREBP-1和p-Smad3的蛋白表达。结论 TGF-β1是参与DKD的重要致病因子,其作用机制可能与诱导SREBP-1活化,引起GMC细胞内脂质沉积密切相关。TGF-β1可能为DKD的预防和治疗提供新方法。
ObjectiveTo explore the mechanism of TGF-β1 during the process of diabetic kidney diseases(DKD).MethodsMesangial cells(GMC)were treated with TGF-β1(2 ng/ml)and/or high glucose(HG,24. 4 mmol/L). SREBP-1 and FAS were detected by western blot. The formation of lipid droplets were tested by Oil Red O staining. The left kidney of SD rat was perfused with adenovirus carrying TGF- β1 gene(AdTGF-β1)and empty vector(AdLacZ). Then the rat was sacrificed and the left kidney was removed. The expression of SREBP-1 and p-Smad3 protein were detected by IHC.ResultsThe ex-pression of mSREBP-1 was significantly increased in TGF-β1 treated GMC as compared with control group(Con group),and the effect was time dependent[(1. 06±0. 55)vs(1. 50±0. 44)vs(1. 59±0. 68)],[(1. 23±0. 52)vs(1. 34±0. 65)]. The expression of mSREBP-1 was also significantly increased by TGF-β1+HG in GMC. TGF-β1 treatment significantly increased the expression of FAS in GMC[(1. 18±0. 04)vs(1. 38±0. 11)vs(1. 40±0. 13),P<0. 05]. Red granular lipid droplets could be seen in GMC cells by oil red O staining. TGF-β1 treatment could increase the expression of SREBP-1 and p-Smad3 protein in rat kidney perfusion with adenovirus vector bring TGF-β1 gene.ConclusionTGF-β1 is an important patho-genic factor involved in the occurrence of DKD. The mechanism partly correlated with TGF-β1 inducing acti-vation of SREBP-1 resulting in deposition of lipid in GMC. Thus,TGF-β1 may be used as a new interven-tion target in clinical treatment of DKD.
ObjectiveTo explore the mechanism of TGF-β1 during the process of diabetic kidney diseases(DKD).MethodsMesangial cells(GMC)were treated with TGF-β1(2 ng/ml)and/or high glucose(HG,24. 4 mmol/L). SREBP-1 and FAS were detected by western blot. The formation of lipid droplets were tested by Oil Red O staining. The left kidney of SD rat was perfused with adenovirus carrying TGF- β1 gene(AdTGF-β1)and empty vector(AdLacZ). Then the rat was sacrificed and the left kidney was removed. The expression of SREBP-1 and p-Smad3 protein were detected by IHC.ResultsThe ex-pression of mSREBP-1 was significantly increased in TGF-β1 treated GMC as compared with control group(Con group),and the effect was time dependent[(1. 06±0. 55)vs(1. 50±0. 44)vs(1. 59±0. 68)],[(1. 23±0. 52)vs(1. 34±0. 65)]. The expression of mSREBP-1 was also significantly increased by TGF-β1+HG in GMC. TGF-β1 treatment significantly increased the expression of FAS in GMC[(1. 18±0. 04)vs(1. 38±0. 11)vs(1. 40±0. 13),P<0. 05]. Red granular lipid droplets could be seen in GMC cells by oil red O staining. TGF-β1 treatment could increase the expression of SREBP-1 and p-Smad3 protein in rat kidney perfusion with adenovirus vector bring TGF-β1 gene.ConclusionTGF-β1 is an important patho-genic factor involved in the occurrence of DKD. The mechanism partly correlated with TGF-β1 inducing acti-vation of SREBP-1 resulting in deposition of lipid in GMC. Thus,TGF-β1 may be used as a new interven-tion target in clinical treatment of DKD.
引文
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[15]Derynck R.Smaddependent and Smadindependent pathways in TGFbeta family signaling.Nature,2003,425:577584.
[16]Weng HB,Han WK,Xiong YW,et al.Taxus chinensis ameliorates diabetic nephropathy through downregulating TGFβ1/Smad pathway.Chin J Nat Med,2018,16:9096.
[17]LontchiYimagou E,Sobngwi E,Matsha TE,et al.Diabetes mellitus and inflammation.Curr Diab Rep,2013,13:435444.
[2]Xu Y,Wang L,He J,et al.Prevalence and control of diabetes in Chineseadults.JAMA,2013,310:948959.
[3]Chen G,Wang T,Uttarwar L,et al.SREBP1 is a novel mediator of TGFβ1 signaling in mesangial cells.J Mol Cell Biol,2014,6:516530.
[4]Castro NE,Kato M,Park JT,et al.Transforming growth factorβ1(TGFβ1)enhances expression of profibrotic genes through a novel signaling cascade and microRNAs in renal mesangial cells.J Biol Chem,2014,289:2900129013.
[5]Brown MS,Goldstein JL.The SREBP pathway:regulation of cholesterol metabolism by proteolysis of a membranebound transcription factor.Cell,1997,89:331340.
[6]Daemen S,Kutmon M,Evelo CT.A pathway approach to investigate the function and regulation of SREBPs.Genes Nutr,2013,8:289300.
[7]Ghayur A,Liu L,Kolb M,et al.Adenovirusmediated gene transfer of TGFbeta1 to the renal glomeruli leads to proteinuria.Am J Pathol,2012,180:940951.
[8]Uttarwar L,Peng F,Wu D,et al.HBEGF release mediates glucoseinduced activation of the epidermal growth factor receptor in mesangial cells.Am J Physiol Renal Physiol,2011,300:F921F931.
[9]Fujita T,Ogihara N,Kamura Y,et al.Interleukin18 contributes more closely to the progression of diabetic nephropathy than other diabetic complications.Acta Diabetol,2012,49:111117.
[10]Luan P,Zhuang J,Zou J,et al.NLRC5 deficiency ameliorates diabetic nephropathy through alleviating inflammation.Faseb J,2018,32:10701084.
[11]Hills CE,Squires PE.The role of TGFβand epithelial to mesenchymal transition in diabetic nephropathy.Cytokine Growth Factor Rev,2011,22:131139.
[12]Yuan N,Zhang HF,Wei Q,et al.Expression of CD4+CD25+Foxp3+regulatory t cells,interleukin 10 and transforming growth factorβin Newly diagnosed type 2 diabetic patients.Exp Clin Endocrinol Diabetes,2018,126:96101.
[13]Yue Y,Meng K,Pu Y,et al.Transforming growth factor beta(TGFβ)mediates cardiac fibrosis and induces diabetic cardiomyopathy.Diabetes Res Clin Pract,2017,133:124130.
[14]Xu Y,Niu Y,Gao Y,et al.Borapetoside E,a clerodane diterpenoid extracted from tinospora crispa,improves hyperglycemia and hyperlipidemia in highfatdietinduced type 2 diabetes mice.JNat Prod,2017,80:23192327.
[15]Derynck R.Smaddependent and Smadindependent pathways in TGFbeta family signaling.Nature,2003,425:577584.
[16]Weng HB,Han WK,Xiong YW,et al.Taxus chinensis ameliorates diabetic nephropathy through downregulating TGFβ1/Smad pathway.Chin J Nat Med,2018,16:9096.
[17]LontchiYimagou E,Sobngwi E,Matsha TE,et al.Diabetes mellitus and inflammation.Curr Diab Rep,2013,13:435444.