补肾益精中药复方对内皮间质转化的影响
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摘要
目的:探讨补肾益精中药复方对转化生长因子-β1(transforming growth factor-β1,TGF-β1)诱导的内皮间质转化的作用及机制。方法:TGF-β1诱导人脐静脉细胞(human umbilical vein endothelial cell,HUVEC)构建内皮间质转化模型,分别用对照血清、补肾益精复方含药血清、黄芪甲苷、Fli1 siRNA+含药血清干预模型,显微镜检测HUVEC细胞内皮间质转化过程,实时荧光定量聚合酶链式反应(quantitative polymerase chain reaction,QPCR)法检测各组VE-cadherin、FSP1、Fli1的表达。结果:①经TGF-β1诱导后,细胞形态呈现纤维化改变,VE-cadherin表达降低,FSP1表达升高,提示内皮间质转化模型诱导成功;同时Fli1在内皮间质转化模型中为低表达。②采用Fli1 siRNA干扰设计Fli1 siRNA序列,显示Fli1 siRNA-1明显抑制Fli1 mRNA表达,同时VE-cadherin表达明显下降,FSP1表达明显上升。③黄芪甲苷组、补肾益精复方含药血清组中Fli1、VE-cadherin的表达较内皮间质转化模型组升高,FSP1表达降低,提示血管内皮间质得到改善;Fli1siRNA+含药血清组较含药血清组Fli1下降、VE-cadherin下降、FSP1上升,其改善不如单独含药血清组明显。结论:补肾益精法中药复方可改善硬皮病血管内皮间质转化,其机制可能与上调Fli1有关。
Objective:To investigate the effect and mechanism of Bushen Yijing Compound Recipe on transforming growth factor-β1(TGF-β1)-induced endothelial mesenchymal transition.Methods:TGF-β1 induced human umbilical vein endothelial cells(HUVEC) to construct endothelial mesenchymal transition model,using control serum,Bushen Yijing compound serum,astragaloside IV,Fli1 siRNA+ serum containing Chinese medicine intervention model,and the endothelial mesenchymal transition process of HUVEC cells was detected by microscopy.The expression of VE-cadherin,FSP1 and Fli1 in each group was detected by real-time quantitative polymerase chain reaction(QPCR).Results:①After TGF-β1 induction,cell morphology showed fibrotic changes,VE-cadherin expression decreased,and FSP-1 expression increased,suggesting that the endothelial mesenchymal transition model was successfully induced;while Fli1 was low in the endothelial stroma transformation model.expression.②Fli1 siRNA sequence was used to design Fli1 siRNA sequence,which showed that Fli1 siRNA-1 significantly inhibited Fli1 mRNA expression,while VE-cadherin expression decreased significantly and FSP1 expression increased significantly.③The expression of Fli1 and VE-cadherin in the serum containing scutellariae and tonifying spleen were higher than those in the model of endothelial mesenchymal transition,and the expression of FSP-1 was decreased,suggesting that vascular endothelium was improved.Compared with that of the drug-containing serum group,the Fli1 in siRNA+ serum containing Chinese medicine group decreased,the VE-cadherin decreased,and the FSP-1 increased.The improvement was not as significant as that of the serum-containing serum alone group.Conclusion:Bushen Yijing method can improve the vascular endothelial endothelium transition in scleroderma,and its mechanism may be related to up-regulation of Fli1.
Objective:To investigate the effect and mechanism of Bushen Yijing Compound Recipe on transforming growth factor-β1(TGF-β1)-induced endothelial mesenchymal transition.Methods:TGF-β1 induced human umbilical vein endothelial cells(HUVEC) to construct endothelial mesenchymal transition model,using control serum,Bushen Yijing compound serum,astragaloside IV,Fli1 siRNA+ serum containing Chinese medicine intervention model,and the endothelial mesenchymal transition process of HUVEC cells was detected by microscopy.The expression of VE-cadherin,FSP1 and Fli1 in each group was detected by real-time quantitative polymerase chain reaction(QPCR).Results:①After TGF-β1 induction,cell morphology showed fibrotic changes,VE-cadherin expression decreased,and FSP-1 expression increased,suggesting that the endothelial mesenchymal transition model was successfully induced;while Fli1 was low in the endothelial stroma transformation model.expression.②Fli1 siRNA sequence was used to design Fli1 siRNA sequence,which showed that Fli1 siRNA-1 significantly inhibited Fli1 mRNA expression,while VE-cadherin expression decreased significantly and FSP1 expression increased significantly.③The expression of Fli1 and VE-cadherin in the serum containing scutellariae and tonifying spleen were higher than those in the model of endothelial mesenchymal transition,and the expression of FSP-1 was decreased,suggesting that vascular endothelium was improved.Compared with that of the drug-containing serum group,the Fli1 in siRNA+ serum containing Chinese medicine group decreased,the VE-cadherin decreased,and the FSP-1 increased.The improvement was not as significant as that of the serum-containing serum alone group.Conclusion:Bushen Yijing method can improve the vascular endothelial endothelium transition in scleroderma,and its mechanism may be related to up-regulation of Fli1.
引文
[1]BLACK C M.Scleroderma--clinical aspects[J].J Intern Med,1993,234(2):115-118.
[2]郭刚.硬皮病诊断及分类探讨[J].内科理论与实践,2011,6(5):341-343.
[3]MANETTI M,ROMANO E,ROSA I,et al.Endothelial-to-mesenchymal transition contributes to endothelial dysfunction and dermal fibrosis in systemic sclerosis[J].Ann Rheum Dis,2017,76(5):924-934.
[4]王瑞风,杜宏.内皮间质转化在糖尿病大血管病变中的作用[J].中华临床医师杂志(电子版),2014,8(19):3508-3511.
[5]MENDOZA F A,PIERA-VELAZQUEZ S,FARBER J L,et al.Endothelial Cells Expressing Endothelial and Mesenchymal Cell Gene Products in Lung Tissue From Patients With Systemic Sclerosis-Associated Interstitial Lung Disease[J].Arthritis Rheumatol,2016,68(1):210-217.
[6]张丽霞,周竹,王倩,等.Cthrc1及TGF-β1/Smad信号通路在硬皮病成纤维细胞中的表达及其意义[J].中国皮肤性病学杂志,2015,28(4):345-348.
[7]邓铁涛.肺脾肾相关辨治硬皮病[J].中国中医药现代远程教育,2004,2(6):15-16.
[8]QI Q,MAO Y,YI J,et al.Anti-fibrotic effects of Astragaloside IV in systemic sclerosis[J].Cell Physiol Biochem,2014,34(6):2105-2116.
[9]KUBO M,CZUWARA-LADYKOWSKA J,MOUSSA O,et al.Persistent down-regulation of Fli1,a suppressor of collagen transcription,in fibrotic scleroderma skin[J].Am J Pathol,2003,163(2):571-581.
[10]万欢.丹参酮ⅡA对高糖环境下大鼠心肌微血管内皮细胞间质转分化的影响[D].南昌:南昌大学,2013.
[11]ARCINIEGAS E,FRID M G,DOUGLAS I S,et al.Perspectives on endothelial-to-mesenchymal transition:potential contribution to vascular remodeling in chronic pulmonary hypertension[J].Am J Physiol Lung Cell Mol Physiol,2007,293(1):L1-L8.
[12]JIMENEZ S A,PIERA-VELAZQUEZ S.Endothelial to mesenchymal transition (EndoMT) in the pathogenesis of Systemic Sclerosis-associated pulmonary fibrosis and pulmonary arterial hypertension.Myth or reality[J]?Matrix Biol,2016,51(1):26-36.
[13]COOLEY B C,NEVADO J,MELLAD J,et al.TGF-beta signaling mediates endothelial-to-mesenchymal transition (EndMT) during vein graft remodeling[J].Sci Transl Med,2014,6(2):227-234.
[14]TROJANOWSKA M.Noncanonical transforming growth factor beta signaling in scleroderma fibrosis[J].Curr Opin Rheumatol,2009,21(6):623-629.
[15]田永贞.基于补肾益精法复方的黄芪甲苷对硬皮纤维化的作用及机制研究[D].广州:广州中医药大学,2016.
[16]邱路萍,田永贞,张佳林,等.补肾益精法对系统性硬皮病血管内皮间质转化的影响[J].上海中医药大学学报,2017,31(1):63-68.
[17]ASANO Y.Epigenetic suppression of Fli1,a potential predisposing factor in the pathogenesis of systemic sclerosis[J].Int J Biochem Cell Biol,2015,67(1):86-91.
[18]TANIGUCHI T,ASANO Y,AKAMATA K,et al.Fibrosis,vascular activation,and immune abnormalities resembling systemic sclerosis in bleomycin-treated Fli-1-haploinsufficient mice[J].Arthritis Rheumatol,2015,67(2):517-526.
[2]郭刚.硬皮病诊断及分类探讨[J].内科理论与实践,2011,6(5):341-343.
[3]MANETTI M,ROMANO E,ROSA I,et al.Endothelial-to-mesenchymal transition contributes to endothelial dysfunction and dermal fibrosis in systemic sclerosis[J].Ann Rheum Dis,2017,76(5):924-934.
[4]王瑞风,杜宏.内皮间质转化在糖尿病大血管病变中的作用[J].中华临床医师杂志(电子版),2014,8(19):3508-3511.
[5]MENDOZA F A,PIERA-VELAZQUEZ S,FARBER J L,et al.Endothelial Cells Expressing Endothelial and Mesenchymal Cell Gene Products in Lung Tissue From Patients With Systemic Sclerosis-Associated Interstitial Lung Disease[J].Arthritis Rheumatol,2016,68(1):210-217.
[6]张丽霞,周竹,王倩,等.Cthrc1及TGF-β1/Smad信号通路在硬皮病成纤维细胞中的表达及其意义[J].中国皮肤性病学杂志,2015,28(4):345-348.
[7]邓铁涛.肺脾肾相关辨治硬皮病[J].中国中医药现代远程教育,2004,2(6):15-16.
[8]QI Q,MAO Y,YI J,et al.Anti-fibrotic effects of Astragaloside IV in systemic sclerosis[J].Cell Physiol Biochem,2014,34(6):2105-2116.
[9]KUBO M,CZUWARA-LADYKOWSKA J,MOUSSA O,et al.Persistent down-regulation of Fli1,a suppressor of collagen transcription,in fibrotic scleroderma skin[J].Am J Pathol,2003,163(2):571-581.
[10]万欢.丹参酮ⅡA对高糖环境下大鼠心肌微血管内皮细胞间质转分化的影响[D].南昌:南昌大学,2013.
[11]ARCINIEGAS E,FRID M G,DOUGLAS I S,et al.Perspectives on endothelial-to-mesenchymal transition:potential contribution to vascular remodeling in chronic pulmonary hypertension[J].Am J Physiol Lung Cell Mol Physiol,2007,293(1):L1-L8.
[12]JIMENEZ S A,PIERA-VELAZQUEZ S.Endothelial to mesenchymal transition (EndoMT) in the pathogenesis of Systemic Sclerosis-associated pulmonary fibrosis and pulmonary arterial hypertension.Myth or reality[J]?Matrix Biol,2016,51(1):26-36.
[13]COOLEY B C,NEVADO J,MELLAD J,et al.TGF-beta signaling mediates endothelial-to-mesenchymal transition (EndMT) during vein graft remodeling[J].Sci Transl Med,2014,6(2):227-234.
[14]TROJANOWSKA M.Noncanonical transforming growth factor beta signaling in scleroderma fibrosis[J].Curr Opin Rheumatol,2009,21(6):623-629.
[15]田永贞.基于补肾益精法复方的黄芪甲苷对硬皮纤维化的作用及机制研究[D].广州:广州中医药大学,2016.
[16]邱路萍,田永贞,张佳林,等.补肾益精法对系统性硬皮病血管内皮间质转化的影响[J].上海中医药大学学报,2017,31(1):63-68.
[17]ASANO Y.Epigenetic suppression of Fli1,a potential predisposing factor in the pathogenesis of systemic sclerosis[J].Int J Biochem Cell Biol,2015,67(1):86-91.
[18]TANIGUCHI T,ASANO Y,AKAMATA K,et al.Fibrosis,vascular activation,and immune abnormalities resembling systemic sclerosis in bleomycin-treated Fli-1-haploinsufficient mice[J].Arthritis Rheumatol,2015,67(2):517-526.