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丹参凝集素促进血管平滑肌细胞的增殖机制研究 预览

Mechanisms of promoting proliferation of vascular smooth muscle cells by Salvia miltiorrhiza lectin
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摘要 目的探究丹参凝集素蛋白(SMLP)对大鼠血管平滑肌细胞(VSMCs)增殖的影响机制。方法通过蛋白分离纯化技术从中药丹参中分离得到具有活性的凝集素蛋白,噻唑蓝(MTT)法检测SMLP对大鼠VSMCs的增殖作用,蛋白质印迹法(Westernblot)检测相关增殖基因的表达水平。构建pet-28a-sml原核表达载体,利用蛋白质亲和层析等技术纯化体外表达的SMLP,并通过红细胞凝集实验确认纯化的SMLP的活性。结果丹参中分离的SMLP能够促进大鼠VSMCs增殖,增殖相关基因蛋白激酶B(AKT)基因与哺乳动物雷帕霉素靶蛋白(mTOR)的磷酸化水平明显增加,而凋亡相关基因JNK1和Bax表达水平明显降低。成功构建了pet-28a-smlp原核表达载体,并在体外表达纯化了SMLP;红细胞凝集实验表明,体外表达纯化的SMLP具有活性。结论SMLP能够促进大鼠VSMCs的增殖;成功建立SMLP的体外表达纯化方法,为进一步研究其功能奠定了基础。 Objective To explore the mechanism of the effect of Salvia miltiorrhiza lectin(SMLP)on the proliferation of vascular smooth muscle cells(VSMCs)in rats.Methods The SMLP was isolated from Salvia miltiorrhiza by protein separation and purification technology.The MTT assay was performed to detect the proliferation of VSMCs in rats after SMLP treatment.The expression levels of proliferation-related genes were detected by Western blot.The prokaryotic expression vector of pet-28a-sml was constructed,and SMLP was purified by affinity chromatography.The activity of purified SMLP was tested by RBC agglutination experiment.Results The SMLP isolated from Salvia miltiorrhiza could promote the proliferation of VSMCs in rats,and the expression levels of proliferation-related genes(p-AKT and p-mTOR)were significantly increased,while the expression levels of apoptosis-related genes(JNK1 and Bax)were decreased obviously.The prokaryotic expression vector pet-28a-sml was successfully constructed,and the SMLP was purified in vitro.The RBC agglutination experiment showed that the purified SMLP protein was functional in vitro.Conclusion SMLP can promote the proliferation of VSMCs in rats.The expression and purification method of SMLP in vitro was established,which lays a foundation for further study on the function of the protein.
作者 沈艳花 牛成群 覃韦宁 李珊 武福云 SHEN Yanhua;NIU Chengqun;QIN Weining;LI Shan;WU Fuyun(College of Basic Medicine,Hubei University of Medicine,Shiyan,Hubei 442000,China)
出处 《重庆医学》 CAS 2019年第7期1099-1102,共4页 Chongqing medicine
基金 国家自然科学基金资助项目(81502637、81702639) 湖北省教育厅科学技术研究计划青年项目(Q20162105) 大学生创新创业训练计划项目(201810929034)。
关键词 植物凝集素类 血管平滑肌细胞 细胞增殖 plant lectins vascular smooth muscle cells cell proliferation
作者简介 沈艳花(1996-),在读硕士,主要从事中药药理方面的研究;通信作者:武福云,E-mail:wufuyun100@126.com。
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