控制血糖对糖尿病肾脏疾病大鼠肾脏组织Wnt4/β-catenin信号通路及肾脏纤维化的影响
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摘要
目的观察降糖治疗对DKD大鼠肾脏组织Wnt4/βcatenin信号通路相关分子表达的影响,探讨降糖治疗延缓DKD肾纤维化的可能机制。方法 24只SD雄性大鼠,分为正常对照组(NC)、DKD组和胰岛素降糖组(INS),每组各8只。微量末梢血检测HbA1c,生化法测血糖、BUN、24 h UAlb。HE、Masson染色观察肾脏病理改变,免疫组织化学检测Wnt4和β连环蛋白(βcatenin)在肾脏组织的表达。Western blot检测Wnt4、βcatenin、磷酸化糖原合成酶激酶3β(pGSK3β)、糖原合成酶激酶3β(GSK3β)、α平滑肌肌动蛋白(αSMA)、上皮型钙黏附蛋白(Ecadherin)、I型胶原(CollagenⅠ)蛋白在大鼠肾脏组织中的表达。RTPCR检测Wnt4、βcatenin和CollagenⅠmRNA表达。结果与NC比较,DKD组Wnt4 mRNA和蛋白表达升高[(0. 27±0. 04)vs(1. 28±0. 42),(0. 03±0. 01)vs(0. 09±0. 03),P<0. 05],pGSK3β、βcatenin和αSMA蛋白表达升高[(0. 13±0. 02)vs(0. 92±0. 35),(0. 11±0. 03)vs(0. 47±0. 12),(0. 02±0. 002)vs(0. 20±0. 04),P<0. 05],CollagenⅠ沉积增加[(0. 01±0. 01)vs(0. 20±0. 07),P<0. 05]。与DKD组比较,INS组Wnt4 mRNA和蛋白表达降低[(1. 28±0. 42)vs(0. 78±0. 12),(0. 09±0. 03)vs(0. 05±0. 01),P<0. 05],pGSK3β、βcatenin和αSMA蛋白表达降低[(0. 92±0. 35)vs(0. 28±0. 02),(0. 47±0. 12)vs(0. 16±0. 03),(0. 20±0. 04)vs(0. 08±0. 01),P<0. 05],CollagenⅠ沉积减少[(0. 20±0. 07)vs(0. 03±0. 01),P<0. 05]。结论控制血糖可抑制Wnt4/βcatenin信号通路的活化,减少ECM沉积,延缓糖尿病大鼠肾纤维化的发展。
ObjectiveTo investigate the influence of blood glucose control on Wnt4/β-catenin sig-naling pathway in rats with diabetic kidney disease(DKD),and to explore the possible mechanism of blood glucose control in delaying DKD renal fibrosis.MethodsA total of 24 male Sprague-Dawley(SD)rats were randomly divided into control group(NC group,n=8),DKD group(n=8)and insulin - treated groups(INS group,n=8). Trace peripheral blood was used to test HbA1 c. Blood glucose,serum urea ni-trogen and 24 h UAlb were measured with biochemical methods. The pathological changes of renal tissuewere observed with HE and Mason staining. Immunohistochemical analysis was used to test the expression of Wnt4 and β - catenin in renal tissue. Western blot was used to detect the protein expression of Wnt4,β-catenin,p-GSK-3β,GSK-3β,E-cadherin,α-SMA,Collagen Ⅰ in renal tissues. The mRNA expression of Wnt4,β-catenin and α-SMA in renal tissue were detected by RT-PCR.ResultsThe protein and m RNA expression of Wnt4 significantly increased[(0. 27±0. 04)vs(2. 01±0. 62),(0. 03±0. 01)vs(0. 09±0. 03),P<0. 05],the protein expression of p-GSK-3β,β-catenin,α-SMA significantly increased[(0. 13±0. 02)vs(0. 92±0. 35),(0. 11±0. 03)vs(0. 47±0. 12),(0. 02±0. 00)vs(0. 20±0. 04),P<0. 05],and Collagen Ⅰ significantly increased[(0. 01±0. 01)vs(0. 20±0. 07),P<0. 05]in DKD group than in NC group. The mRNA and protein levels of Wnt4 were lower[(1. 28±0. 42)vs(0. 78±0. 12),(0. 09±0. 03)vs(0. 05 ± 0. 01),P<0. 05],the expression of p - GSK - 3β,β - catenin,and α - SMA were lower[(0. 92±0. 35)vs(0. 28±0. 02),(0. 47±0. 12)vs(0. 16±0. 03),(0. 20±0. 04)vs(0. 08±0. 01),P<0. 05],and Collagen Ⅰ protein was lower in INS group than in DKD group[(0. 20±0. 07)vs(0. 03±0. 01),P<0. 05].ConclusionControlling blood glucose could inhibit the abnormal activation of the Wnt4/β -catenin signaling pathway,decrease the deposition of extracellular matrix,and delay the develop-ment of renal fibrosis in diabetic rats.
ObjectiveTo investigate the influence of blood glucose control on Wnt4/β-catenin sig-naling pathway in rats with diabetic kidney disease(DKD),and to explore the possible mechanism of blood glucose control in delaying DKD renal fibrosis.MethodsA total of 24 male Sprague-Dawley(SD)rats were randomly divided into control group(NC group,n=8),DKD group(n=8)and insulin - treated groups(INS group,n=8). Trace peripheral blood was used to test HbA1 c. Blood glucose,serum urea ni-trogen and 24 h UAlb were measured with biochemical methods. The pathological changes of renal tissuewere observed with HE and Mason staining. Immunohistochemical analysis was used to test the expression of Wnt4 and β - catenin in renal tissue. Western blot was used to detect the protein expression of Wnt4,β-catenin,p-GSK-3β,GSK-3β,E-cadherin,α-SMA,Collagen Ⅰ in renal tissues. The mRNA expression of Wnt4,β-catenin and α-SMA in renal tissue were detected by RT-PCR.ResultsThe protein and m RNA expression of Wnt4 significantly increased[(0. 27±0. 04)vs(2. 01±0. 62),(0. 03±0. 01)vs(0. 09±0. 03),P<0. 05],the protein expression of p-GSK-3β,β-catenin,α-SMA significantly increased[(0. 13±0. 02)vs(0. 92±0. 35),(0. 11±0. 03)vs(0. 47±0. 12),(0. 02±0. 00)vs(0. 20±0. 04),P<0. 05],and Collagen Ⅰ significantly increased[(0. 01±0. 01)vs(0. 20±0. 07),P<0. 05]in DKD group than in NC group. The mRNA and protein levels of Wnt4 were lower[(1. 28±0. 42)vs(0. 78±0. 12),(0. 09±0. 03)vs(0. 05 ± 0. 01),P<0. 05],the expression of p - GSK - 3β,β - catenin,and α - SMA were lower[(0. 92±0. 35)vs(0. 28±0. 02),(0. 47±0. 12)vs(0. 16±0. 03),(0. 20±0. 04)vs(0. 08±0. 01),P<0. 05],and Collagen Ⅰ protein was lower in INS group than in DKD group[(0. 20±0. 07)vs(0. 03±0. 01),P<0. 05].ConclusionControlling blood glucose could inhibit the abnormal activation of the Wnt4/β -catenin signaling pathway,decrease the deposition of extracellular matrix,and delay the develop-ment of renal fibrosis in diabetic rats.
引文
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[13]Liu CL,Yan L,Cai KR,et al. Effects of soybean isoflavones on Wnt/β-catenin and the TGF-β1 signaling pathway in renal tissue of type 2 diabetic rats. J Biol Regul Homeost Agents,2018,32:455-464.
[14]李淑珍,何云.改善微循环为主配合控制血糖治疗36例早期糖尿病肾病的疗效观察.中国医药指南,2014,14:128-129.
[15]Vatankhah N,Jahangiri Y,Landry GJ,et al. Effect ofsystemic in-sulin treatment on diabetic wound healing. Wound Repair Regen,2017,25:288-291.
[16]Herman-Edelstein M,Scherzer P,Tobar A,et al. Alteredrenal lipid metabolism and renal lipid accumulation inhuman diabetic ne-phropathy. J Lipid Res,2014,55:561-572.
[2] Chang B,Chen W,Zhang Y,et al. Tripterygiumwilfordii mitigates hyperglycemia-induced upregulated Wnt/β-catenin expres-sion and kidney injury in diabetic rats. ExpTher Med,2018,15:3874-3882.
[3]石明隽,肖瑛,郭兵,等.β-环连蛋白在糖尿病大鼠肾小管中表达及意义.中国现代医学杂志,2011,21:556-563.
[4]石明隽,肖瑛,郭兵,等. Wnt5a在糖尿病大鼠肾脏组织中表达及意义.中国公共卫生,2011,27:330-332.
[5] Edeling M,Raqi G,Huang S,et al. Developmental signalling-pathways in renal fibrosis:the roles of Notch,Wnt and Hedge-hog. Nat Rev Nephrol,2016,12:426-439.
[6]于海英,孙中华,崔俊生,等.胰岛新生实验研究中1型糖尿病大鼠模型的构建.医学信息,2011,24:3118.
[7] Liu Y. Cellular and molecular mechanisms of renal fibrosis. Nat RevNephrol,2011,7:684-696.
[8] Wang Y,Zhou CJ,Liu Y. Wnt signaling in kidney development and disease. Prog Mol Biol Transl Sci,2018,153:181-207.
[9] Zhang HX,Luo WL,Sun YH,et al. Wnt/β-catenin signaling me-diated-UCH-L1 expression in podocytes of diabetic nephropathy.Int J Mol Sci,2016,17:1404.
[10]Lan X,Wen H,Aslam R,et al. Nicotine enhances mesangial cell proliferation and fibronectin production in high glucose milieu via activation of Wnt/β-catenin pathway. Bioscience Reports,2018,38:BSR20180100.
[11]Li Y,Rankin SA,Sinner D,et al. Sfrp5 coordinates foregut spec-ification and morphogenesis by antagonizing both canonical and noncanonical Wnt11 signaling. Genes Dev,2008,22:3050-3063.
[12]Satoh W,Matsuyama M,Takemura H,et al. Sfrp1,Sfrp2,and Sfrp5 regulate the Wnt/β-cate-nin and the planar cell polarity pathways during early trunk formation in mouse. Genesis,2008,46:92-103.
[13]Liu CL,Yan L,Cai KR,et al. Effects of soybean isoflavones on Wnt/β-catenin and the TGF-β1 signaling pathway in renal tissue of type 2 diabetic rats. J Biol Regul Homeost Agents,2018,32:455-464.
[14]李淑珍,何云.改善微循环为主配合控制血糖治疗36例早期糖尿病肾病的疗效观察.中国医药指南,2014,14:128-129.
[15]Vatankhah N,Jahangiri Y,Landry GJ,et al. Effect ofsystemic in-sulin treatment on diabetic wound healing. Wound Repair Regen,2017,25:288-291.
[16]Herman-Edelstein M,Scherzer P,Tobar A,et al. Alteredrenal lipid metabolism and renal lipid accumulation inhuman diabetic ne-phropathy. J Lipid Res,2014,55:561-572.