摘要
通过体外实验研究雷公藤甲素(triptolide,TP)对VEGFA,SDF-1,CXCR4通路的影响,探讨其改善强直性脊柱炎(AS)患者血小板活化的机制。外周血单核细胞(PBMC)用于实验。分为4组:正常组(NC)、模型组(MC)、雷公藤甲素组(TP)、AMD3100组。MTT法检测TP最佳浓度。ELISA法检测TNF-α,IL-1β,IL-4,IL-10,VEGFA,VEGFR的表达,实时荧光定量PCR(RT-qPCR)法检测SDF-1,CXCR4,VEGFA的表达,Western blot法检测SDF-1,CXCR4,VEGFA,VEGFR蛋白的表达,免疫荧光法检测CD62p,CD40L,PDGFA的表达水平。MTT结果显示中剂量TP在24 h时对细胞抑制作用最强。ELISA及PCR法结果显示TP可抑制IL-1β,TNF-α,VEGFA,VEGFR,SDF-1,CXCR4,VEGFA mRNA的表达;Western blot结果说明TP可抑制SDF-1,CXCR4,VEGFA,VEGFR蛋白的表达;免疫荧光结果说明TP可抑制CD62p,CD40L,PDGFA的表达。TP可能通过下调SDF-1,CXCR4,VEGFA,VEGFR mRNA,进而下调IL-1β,TNF-α,上调IL-4,IL-10细胞因子的表达,从而调节血小板活化。
The effect of triptolide(TP)on VEGFA,SDF-1,CXCR4 pathway were investigated in vitro to explore the mechanism in improving platelet activation in patients with ankylosing spondylitis(AS).Peripheral blood mononuclear cells(PBMC)were used for the experiment and divided into 4 groups:normal group(NC),model group(MC),triptolide group(TP),and AMD3100 group.The optimal concentration of TP was measured by the MTT method.The expressions of TNF-α,IL-1β,IL-4,IL-10,VEGFA and VEGFR were detected by ELISA.The expressions of SDF-1,CXCR4 and VEGFA were detected by real-time quantitative PCR(RT-qPCR).The expressions of SDF-1,CXCR4,VEGFA and VEGFR were detected by Western blot.The expression levels of CD62p,CD40L and PDGFA were detected by immunofluorescence.MTT results showed that medium-dose TP had the strongest inhibitory effect on cells at 24 h.The results of ELISA and PCR showed that TP inhibited mRNA expressions of IL-1β,TNF-α,VEGFA,VEGFR and SDF-1,CXCR4 and VEGFA.The results of Western blot indicated that TP inhibited SDF-1,CXCR4 and VEGFA,VEGFR protein expressions;immunofluorescence results indicate that TP can inhibit the expressions of CD62p,CD40L,PDGFA.TP may regulate platelet activation by down-regulating SDF-1,CXCR4,VEGFA and VEGFR mRNA expressions,thereby down-regulating IL-1βand TNF-αexpressions,and up-regulating the expressions of IL-4 and IL-10 cytokines.
作者
方妍妍
万磊
董文哲
文建庭
刘健
FANG Yan-yan;WAN Lei;DONG Wen-zhe;WEN Jian-ting;LIU Jian(Graduate School of Anhui University of Traditional Chinese Medicine,Hefei 230038,China;the First Affiliated Hospital of Anhui University of Traditional Chinese Medicine,Hefei 230031,China)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2019年第16期3520-3525,共6页
China Journal of Chinese Materia Medica
基金
安徽省科技攻关项目(1604A0802085)
2016年中央引导地方科技发展专项(财教[2016]1188)
现代中医内科应用基础与开发研究安徽省重点实验室项目(财教[2016]518)
2015年技术创新服务体系专项(财教[2015]1551)
中医药行业科研专项(201307001)
安徽高校自然科学研究重点项目(KJ2017A281)
