摘要
目的 建立一种全新的膀胱出口梗阻(BOO)体外细胞水平的生物力学模型,为BOO的研究提供一个更为稳定科学的实验平台。方法 应用负压发生装置对贴壁生长在BioFlex培养皿内的正常膀胱平滑肌细胞施加不同程度的周期性张力,使其硅胶底部分别发生10 %、2 0 %、3 0 %的形变,利用免疫细胞化学检测收缩表型标志物αactin ,RT PCR检测增殖指标PCNA的mRNA表达以及Westernblot检测增殖细胞核抗原(PCNA)、cdk2kinase蛋白表达,观察它们的动态变化,利用MTT法观察细胞增殖性的变化。结果 随着形变量的增大,免疫细胞化学发现αactin先代偿增加后失代偿降低;RT PCR和Westernblotting发现PCNA的mRNA表达量逐渐增加,PCNA、cdk2kinase的蛋白表达量逐渐增加,3 0 %形变组显著高于正常对照组(P <0 0 1) ;MTT实验发现与正常对照组比,细胞的增殖活性显著增加(P <0 0 1)。结论 ①对培养逼尿肌细胞施加周期性张力负荷作用的方法建立了一种全新的体外细胞水平的BOO模型;②随周期性张力增加,逼尿肌细胞由“收缩型”逐渐转化为“分泌型”。
Objective To establish a brand new model of bladder outflow obstruction (BOO) at single cell level in vitro to offer a more stable and scientific experimental base. Methods Recurrent mechanical stretch generated by vacuum facility was applied to cells attached to flexible membrane of special culture plate, which led to 10%, 20% and 30% elongation of them. Immunocytochemistry was used to analyze the expression of α-actin (sign of contractile phenotype) and RT-PCR was performed to detect the dynamic changes of mRNA expression of PCNA and Western blotting to protein expression of PCNA and cdk2 kinase. MTT assay was used to observe the changes of proliferation of cells. Results In the case of gradual elongation, the expression of α-actin rose at first and fell afterwards; the mRNA expression of PCNA and protein expression of PCNA and cdk2 kinase rose gradually. Changes in 30% elongation group were greater than the control (P<0.01). MTT assay also revealed the significant rising of proliferation (P<0.01). Conclusion A model of BOO at single cell level in vitro was established by means of performing recurrent mechanical stretch upon cultured smooth muscle cells. Smooth muscle cells were remodeled from 'contractile phenotype' to 'secretory phenotype' by the increasing stretch.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2005年第10期968-971,共4页
Journal of Third Military Medical University
基金
国家自然科学基金资助项目 ( 30 30 0 350 )~~
关键词
逼尿肌
梗阻
模型
张力
重塑
destructor
obstruction
model
stretch
remodeling
