摘要
目的:体外分离培养SD大鼠胸主动脉外膜成纤维细胞亚群,并对其增殖特性和血管功能进行初步分析。方法:克隆环法进行血管外膜成纤维细胞的单克隆培养,RT-PCR鉴定细胞亚群纯度,MTT检测血管外膜成纤维细胞亚群增殖能力。荧光定量PCR检测血管紧张素Ⅱ(Ang Ⅱ)及其受体拮抗剂氯沙坦(losartan)和PD-123319对前内皮素原-1(preproET-1)mRNA表达的影响。结果:克隆环法获得血管外膜成纤维细胞2个亚群:圆形细胞亚群和纺锤形细胞亚群。以β-肌动蛋白(β-actin)为内参照,经RT-PCR检测,结果显示无von Wille-brand因子、CD8、结蛋白和肌球蛋白重链的表达,是纯细胞系。2个细胞亚群的增殖情况有所不同,圆形细胞在第2～4 d进入指数分裂期;纺锤形细胞于第3～5 d进入指数分裂期。Ang Ⅱ在0～1 000 nmol/L范围内对纺锤形细胞preproET-1 mRNA的表达无显著影响,而呈浓度依赖性地增加圆形细胞preproET-1 mRNA的表达(P<0.01);losartan阻断了Ang II诱导的圆形细胞preproET-1 mRNA的表达,但对纺锤形细胞preproET-1 mRNA的表达无明显影响。结论:SD大鼠血管外膜成纤维细胞有2个细胞亚群:圆形细胞和纺锤形细胞,其生长特性略有不同。AngII显著促进圆形细胞preproET-1 mRNA的表达,提示这2个细胞亚群可能在血管重建和修复过程中发挥不同作用。
AIM： To observe in vitro monoclonal culture,proliferation characteristics and vascular functions of SD rat thoracic aorta adventitial fibroblasts.METHODS： Vascular adventitial fibroblasts were individually expanded using cloning rings.Phenotypic characteristics of the cells were identified by RT-PCR.The growth ability of the cells was determined by methyl thiazolyl tetrazolium（MTT） assay.Fluorescent quantitative PCR was used to detect the mRNA expression of preproendothelin-1（preproET-1） induced by angiotensin II（Ang II） and treated with Ang II receptor antagonist losartan and PD-123319.RESULTS： Spindle cells and round cells were isolated using cloning rings.The amplified PCR products of von Willebrand factor,CD8,desmin and myosin heavy chain was not expressed in both cells.For spindle cells,the exponential phase of growth was from the 2nd day to the 4th day after passage.For round cells,the exponential phase of growth was from the 3rd day to the 5th day after passage.The concentration dependence（0~1 000 nmol/L） of Ang II was observed to significantly increase the expression of preproET-1 mRNA in the round cells（P0.01）,and Ang II had no significant effect on the mRNA expression of preproET-1 in the spindle cells.Losartan blocked the Ang II-induced mRNA expression of preproET-1 in the round cells,and had no significant effect on the mRNA expression of preproET-1 in the spindle cells.CONCLUSION： Rat vascular adventitial fibroblasts can be isolated using cloning rings,and 2 different phenotypic cells are obtained.Ang II significantly increases the mRNA expression of preproET-1 in the round cell subpopulation,suggesting that the 2 subpopulations may play different roles in vascular remodeling and reparative process.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2012年第1期47-52,共6页
Chinese Journal of Pathophysiology
基金
河北省自然科学基金资助项目(No.C2009001037)
