毒胡萝卜素诱导大鼠冠状动脉平滑肌细胞内质网应激模型的建立

发布时间：2018-03-23 05:32

本文选题：冠状动脉平滑肌细胞　切入点：组织块贴壁法　出处：《中国病理生理杂志》2017年01期 　论文类型：期刊论文

【摘要】：目的:探讨SD大鼠冠状动脉平滑肌细胞(coronary artery smooth muscle cells,CASMCs)原代培养方法,并建立CASMCs内质网应激(endoplasmic reticulum stress,ERS)模型。方法:用组织块贴壁法培养CASMCs,光学显微镜观察细胞形态。免疫荧光技术检测CASMCs的标志分子α-SMA和SM-MHC的表达。采用Western blot检测ERS发生的标志物BiP和CHOP的蛋白表达水平。结果:冠状动脉组织块贴壁培养6 d后,细胞从组织块边缘爬出,呈长梭型,9~10 d细胞生长汇合后表现出平滑肌细胞典型的"峰-谷"状。免疫荧光技术鉴定结果显示,α-SMA和SM-MHC表达呈阳性。不同浓度(0.5、1和2μmol/L)的毒胡萝卜素(thapsigargin,TG)处理CASMCs 24 h后,Western blot结果显示,TG(1和2μmol/L)处理组的BiP和CHOP蛋白表达与对照组相比增加,差异具有统计学意义。与对照组相比较,1μmol/L TG处理CASMCs 24和48 h后BiP和CHOP蛋白表达显著性增加。结论:采用组织块贴壁法可以成功培养CASMCs。利用1μmol/L TG诱导24 h可建立CASMCs的ERS模型。
[Abstract]:Objective: to investigate the primary culture method of coronary artery smooth muscle cells (CASMC) in SD rats. The endoplasmic reticulum stress stress (CASMCs) model was established. Methods: the cells were cultured with tissue attachment method, the morphology of cells was observed by optical microscope, the expression of 伪 -SMA and SM-MHC, the markers of CASMCs were detected by immunofluorescence technique, and ERS was detected by Western blot. Results: the tissue mass of coronary artery was cultured for 6 days. The cells crawled out from the edge of the tissue mass, After 10 days of growth and confluence, the cells showed typical "peak-valley" shape of smooth muscle cells. The expression of 伪 -SMA and SM-MHC was positive by immunofluorescence technique. The CASMCs 24 was treated with different concentrations of carotene (0.5 渭 mol / L) and 2 渭 mol / L (TG2 渭 mol / L). After h, the expression of BiP and CHOP protein in TGG 1 and 2 渭 mol / L treated group was increased compared with the control group, the results of Western blot showed that the expression of BiP and CHOP in the treated group was higher than that in the control group. Compared with the control group, the expression of BiP and CHOP protein was significantly increased after treated with 1 渭 mol/L TG for 24 and 48 h. Conclusion: the expression of BiP and CHOP protein can be successfully cultured by tissue mass adherence method. ERS model of CASMCs can be established by 1 渭 mol/L TG for 24 h.
【作者单位】：汕头大学医学院;广东省人民医院医学研究部广东省医学科学院;
【基金】：国家自然科学基金资助项目(No.81273516;No.81470440)
【分类号】：R54

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