胰岛素样生长因子-1对糖尿病大鼠结肠平滑肌细胞表达干细胞因子的影响

摘要

背景:糖尿病胃肠动力障碍与Cajal间质细胞(ICC)数量和超微结构异常有关.前期实验发现胰岛素样生长因子-1(IGF-1)可诱导正常大鼠胃肠平滑肌细胞(SMC)表达干细胞因子(SCF),从而有利于ICC的生存.目的:探讨IGF-1对糖尿病大鼠结肠SMC表达SCF的影响及其信号转导通路.方法:以链脲霉素建立糖尿病大鼠模型.分离、培养正常和糖尿病大鼠结肠SMC,设不同浓度(0、50、100、150μg/L)、不同时间(0、8、16、24、48 h)IGF-1干预组和MEK抑制剂PD98059+IGF-1、PI3K抑制剂LY294002+IGF-1干预组,以RT-PCR和蛋白质印迹法检测SMC中的SCF表达.结果:糖尿病大鼠结肠SMC的生长速度较正常大鼠减慢.无血清培养条件下,正常和糖尿病结肠SMC中SCFmRNA和蛋白表达较低,IGF-1可诱导SCF表达增加,最大有效浓度为100μg/L,诱导高峰时间正常对照组为16 h,糖尿病组为24 h.经PD98059预处理的SMC,IGF-1诱导的SCF表达部分受抑,LY294002预处理对IGF-1的作用无明显影响.结论:0～100μg/L IGF-1在24 h内能以剂量和时间依赖性方式诱导糖尿病大鼠结肠SMC表达SCF,但效应弱于其对正常大鼠结肠SMC的作用,该作用的发挥可能部分依赖于ERKMAPK信号通路.%Background: Deficiency and ultrastructural changes of interstitial cells of Cajal (ICC) may be associated with the gastrointestinal dysmotility in diabetes mellitus (DM). Previous studies have demonstrated that insulin-like growth factor-1 (IGF-1) may induce the gastrointestinal smooth muscle cells (SMC) of normal rats to express stem cell factor (SCF), which is necessary for the development and phenotype maintenance of ICC. Aims: To investigate the effect of IGF-I on expression of SCF in colonic SMC from DM rats and its signal transduction pathway. Methods: DM rat model was established by streptozotocin. Colonic SMC isolated from normal and DM rats were cultured and then incubated with different concentrations (0, 50, 100 and 150 μg/L) of IGF-1 for different time durations (0. 8, 16, 24 and 48 hours), and some were pretreated with MEK inhibitor PD98059 or PI3K inhibitor LY294002 before IGF-I intervention. Expression of SCF in SMC was determined by RT-PCR and Western blotting. Results: Growth of colonic SMC from DM rats was slower than those from normal rats. Only low level of SCF (mRNA and protein) was expressed in both normal and DM colonic SMC cultured in serum-free medium. Increased expression of SCF was induced by IGF-I with the maximum effective concentration located at 100 μg/L, and the peak time of increment was 16 hours in normal control group and 24 hours in DM group. The IGF-l-induced SCF expression was partially inhibited by pretreatment of PD98059, whereas LY294002 had no such effect. Conclusions: Expression of SCF in colonic SMC from normal and DM rats can be induced by IGF-1 (0-100 μg/L for 0-24 hours) in a dose- and time-dependent manner. This effect is partially mediated by ERKMAPK pathway and is more prominent in normal rats.