肝细胞生长因子和机械生长因子对肛提肌成肌细胞增殖最佳作用浓度的研究

Study on the optimal concentration of hepatocyte growth factor and mechano growth factor on the proliferation of levator ani myoblasts

目的筛选肝细胞生长因子(hepatocyte growth factor,HGF)和机械生长因子(mechano growth factor,MGF)促进妊娠期压力性尿失禁(stress urinary incontinence,SUI)患者肛提肌成肌细胞增殖的最佳浓度,并检测最佳作用浓度下成肌细胞骨架蛋白Laminin、Dystrophin的表达,初步探索HGF、MGF对SUI患者治疗作用的分子生物学机制。方法以妊娠期SUI患者第3代肛提肌成肌细胞为研究对象,HGF、MGF分别设8个浓度组:50、100、150、200、400、600、800、1000 ng/mL,以SUI组(0 ng/mL)为空白对照,各组分别干预72 h后用四甲基偶氮唑盐(methyl thiazolyl tetrazolium,MTT)法测定细胞存活率,筛选出最佳作用浓度,并用反转录聚合酶链反应(reverse transcription polymerase chain reaction,RT-PCR)法检测肛提肌成肌细胞Laminin、Dystrophin的表达。结果与SUI组相比,HGF组、MGF组肛提肌成肌细胞存活率均增加,差异比较有统计学意义(P<0.05);与其他浓度相比,当HGF、MGF浓度为150 ng/mL时,肛提肌成肌细胞存活率最高,差异比较有统计学意义(P<0.05);与正常对照组相比,SUI组肛提肌成肌细胞Laminin、Dystrophin mRNA相对表达量降低,差异比较有统计学意义(P<0.05);与SUI组相比,HGF组(150 ng/mL)、MGF组(150 ng/mL)肛提肌成肌细胞Laminin、Dystrophin mRNA相对表达量增加,差异比较有统计学意义(P<0.05)。结论HGF、MGF可促进体外培养的人肛提肌成肌细胞增殖,并且最佳作用浓度均为150 ng/mL;SUI患者存在肛提肌成肌细胞Laminin、Dystrophin mRNA相对表达量的降低;浓度为150 ng/mL HGF、MGF可促进肛提肌成肌细胞Laminin、Dystrophin的表达。

Objective To screen the optimal concentration of hepatocyte growth factor(HGF)and mechano growth factor(MGF)for promoting the proliferation of levator ani myoblasts in patients with stress urinary incontinence(SUI)during pregnancy,and to detect the expression of skeleton proteins Laminin and Dystrophin in myoblasts at the optimal concentration,so as to preliminarily explore the molecular biological mechanism of the therapeutic effects of HGF and MGF on SUI patients.Methods Taking the third generation levator ani myoblasts of SUI patients during pregnancy as the research object,HGF and MGF were divided into eight concentration groups:50 ng/mL,100 ng/mL,150 ng/mL,200 ng/mL,400 ng/mL,600 ng/mL,800 ng/mL and 1000 ng/mL,SUI group(0 ng/mL)as the blank control.Each group was treated for 72 h,and the cell viability was determined by methyl thiazolyl tetrazolium(MTT)assay,and the optimal concentration was selected.The expression of Laminin and Dystrophin in levator ani myoblasts was detected by reverse transcription polymerase chain reaction(RT-PCR).Results Compared with SUI group(0 ng/mL),the survival rate of levator ani myoblasts in HGF group and MGF group increased,and the difference was statistically significant(P<0.05).Compared with other concentrations,when the concentration of HGF and MGF was 150 ng/mL,the survival rate of levator ani myoblasts was the highest,and the difference was statistically significant(P<0.05).Compared with the normal control group,the relative expression of Laminin and Dystrophin mRNA in the levator ani myoblasts of the SUI group was decreased,and the difference was statistically significant(P<0.05).Compared with the SUI group,the relative expression of Laminin and Dystrophin mRNA in the levator ani myoblasts of HGF group(150 ng/mL)and MGF group(150 ng/mL)was increased,and the difference was statistically significant(P<0.05).Conclusion HGF and MGF can promote the proliferation of human levator ani myoblasts cultured in vitro,and the optimal concentration is 150 ng/mL.The relative expression of Laminin and Dystrophin mRNA in levator ani myoblasts was decreased in SUI patients.The concentration of HGF and MGF was 150 ng/mL can promote the expression of Laminin and Dystrophin in levator ani myoblasts..