학술논문
秋水仙碱对LPS诱导内皮间质转化的影响及机制 / Effect of colchicine on LPS induced endothelial mesenchymal transformation and its mechanism
Document Type
Academic Journal
Source
中国药理学通报 / Chinese Pharmacological Bulletin. 40(2):243-248
Subject
Language
Chinese
ISSN
1001-1978
Abstract
目的 研究秋水仙碱(colchicine,Col)对脂多糖(li-popolysaccharide,LPS)诱导的人脐静脉血管内皮细胞(hu-man umbilical vein vascular endothelial cells,HUVECs)内皮间质转化的影响及相关机制.方法 采用LPS处理HUVECs建立内皮间质转化(endothelial to mesenchymal transition,EndMT)模型.CCK-8法检测细胞增殖率,LDH实验检测细胞毒性,筛选最适药物浓度.将细胞分为正常对照组,正常对照+秋水仙碱(10 nmol·L-1)组,LPS(10 mg·L-1)模型组,LPS+秋水仙碱(10nmol·L-1)组.倒置显微镜观察细胞形态学变化情况,Transwell实验检测细胞迁移能力,小管形成实验检测小管形成能力,Western blot检测内皮标志物(CD31/VE-cadherin)及间质转化标志物(α-SMA/FSP-1)表达情况;使用NF-κB信号通路抑制剂处理,检测相关信号通路变化情况.结果 CCK-8及LDH实验显示,10 nmol·L-1的Col为最适药物浓度;LPS可诱导细胞形态发生变化,Col在一定程度可逆转HUVECs的形态学变化;Transwell实验显示,LPS处理组HUVECs迁移能力明显增强(P＜0.05),而Col可明显逆转这种现象(P＜0.05);小管形成实验显示,LPS处理可抑制HUVECs小管形成能力(P＜0.05),Col可改善LPS诱导的小管形成能力受损(P＜0.05);Western blot结果显示,Col与LPS共同孵育后,CD31及VE-cadherin表达水平相比于模型组明显增加(P＜0.05),α-SMA及FSP-1表达水平相比于模型组明显下降(P＜0.05);在LPS诱导内皮细胞EndMT过程中,Col可抑制NF-KB/Snail信号通路激活.结论 Col能有效抑制LPS诱导的EndMT,其机制与调控NF-KB/Snail信号通路有关.
Aim To investigate the effect of colchicine on lipopolysaccharide(LPS)induced endothelial to mesenchymal transition(EndMT)in human umbilical vein vascular endothelial cells(HUVECs)and its re-lated mechanisms.Methods The EndMT model was established by treating HUVECs with LPS.Cell prolif-eration rate was detected by CCK-8 assay,cytotoxicity was detected by LDH assay,and the optimal drug con-centration was screened.The cells were divided into the normal control group,the normal control+colchi-cine(10 nmol·L-1)group,the LPS(10 mg·L-1)model group,and the LPS+colchicine(10 nmol· L-1)group.The morphologic changes of the cells were observed under an inverted microscope,the cell migra-tion ability was detected by Transwell assay,and the ability of tube formation was analyzed by tube formation assay.The expression of endothelial markers(CD31/VE-cadherin)and mesenchymal cell markers(α-SMA/FSP-1)were detected by Western blot.NF-KB inhibitor was used to detect the changes in related sig-naling pathways.Results CCK-8 and LDH experi-ments showed that 10 nmol·L-1 colchicine was the optimal concentration.LPS could induce morphological changes in HUVECs,and colchicine could reverse morphological changes in HUVECs to a certain extent.Transwell experiment showed that the migration ability of HUVECs in the LPS treatment group was significant-ly enhanced(P＜0.05),and colchicine could signifi-cantly reverse this phenomenon(P＜0.05).Tube for-mation experiment showed that LPS decreased the en-dothelial tube formation ability of HUVECs(P＜0.05),while colchicine treatment markedly improved LPS-induced tube formation defects(P＜0.05).Western blot assay showed that after colchicine co-cul-tured with LPS,the expression levels of CD31 and VE-cadherin significantly increased compared with the model group(P＜0.05),while the expression levels of α-SMA and FSP-1 significantly decreased compared with the model group(P＜0.05).During the induc-tion of EndMT by LPS,colchicine could inhibit the ac-tivation of the NF-KB/Snail signaling pathway.Con-clusions Colchicine can effectively inhibit EndMT in-duced by LPS,and the mechanism may be related to the regulation of the NF-KB/Snail signaling pathway.
Aim To investigate the effect of colchicine on lipopolysaccharide(LPS)induced endothelial to mesenchymal transition(EndMT)in human umbilical vein vascular endothelial cells(HUVECs)and its re-lated mechanisms.Methods The EndMT model was established by treating HUVECs with LPS.Cell prolif-eration rate was detected by CCK-8 assay,cytotoxicity was detected by LDH assay,and the optimal drug con-centration was screened.The cells were divided into the normal control group,the normal control+colchi-cine(10 nmol·L-1)group,the LPS(10 mg·L-1)model group,and the LPS+colchicine(10 nmol· L-1)group.The morphologic changes of the cells were observed under an inverted microscope,the cell migra-tion ability was detected by Transwell assay,and the ability of tube formation was analyzed by tube formation assay.The expression of endothelial markers(CD31/VE-cadherin)and mesenchymal cell markers(α-SMA/FSP-1)were detected by Western blot.NF-KB inhibitor was used to detect the changes in related sig-naling pathways.Results CCK-8 and LDH experi-ments showed that 10 nmol·L-1 colchicine was the optimal concentration.LPS could induce morphological changes in HUVECs,and colchicine could reverse morphological changes in HUVECs to a certain extent.Transwell experiment showed that the migration ability of HUVECs in the LPS treatment group was significant-ly enhanced(P＜0.05),and colchicine could signifi-cantly reverse this phenomenon(P＜0.05).Tube for-mation experiment showed that LPS decreased the en-dothelial tube formation ability of HUVECs(P＜0.05),while colchicine treatment markedly improved LPS-induced tube formation defects(P＜0.05).Western blot assay showed that after colchicine co-cul-tured with LPS,the expression levels of CD31 and VE-cadherin significantly increased compared with the model group(P＜0.05),while the expression levels of α-SMA and FSP-1 significantly decreased compared with the model group(P＜0.05).During the induc-tion of EndMT by LPS,colchicine could inhibit the ac-tivation of the NF-KB/Snail signaling pathway.Con-clusions Colchicine can effectively inhibit EndMT in-duced by LPS,and the mechanism may be related to the regulation of the NF-KB/Snail signaling pathway.