摘要
目的探讨降钙素基因相关肽（CGRP）体外定向诱导前软骨干细胞（PSCs）向成骨细胞分化的作崩及其机制。方法将分离培养的大鼠乳鼠PSCs分别置于含有不同浓度（包括0，1×10^-8mol／L，1×10^-9mol／L，1×10^-10nml／L）CGRP的DMEM／F12培养液中培养，观察细胞形态学变化。于细胞培养后不同时间点采用CCK-8榆洲细胞增殖情况；^将细胞随机分为实验组及对照组，实验组置于禽有CGRP（1×10μmol／L）的成骨培养基中培养，对照组则酋于不含CGRP的成骨培养基中培养。对各组细胞进行茜素红染色并检测碱性磷酸酶（AI．P）岔量；采用RT—pCR法检测各组细胞成骨相关基因[如Runt相关基因转录因子2（RUNX2）、骨桥蛋白（OPN）及骨钙蛋白（BGP）等]表达；采用Westernblot法检测B-catenin蛋白表达以评估CGRP对wut／β-catenin通路激活的影响。结果 与对旦H组（其培养基中无CGRP）比较，CGRP在体外能显著促进PSCs细胞ALP含量增加，矿化结节增大、增多，显著上调RUNX2、OPN、BGP等成骨相关基因及蛋门β—catenin表达，但对PSCs增殖无明显影响。结论CGRP可能通过激活Wnt／β-catenin通路促进PSCs向成骨细胞方向分化。
Objective To investigate the effect of ealcitonin gene-related peptide （CGRP） in inducing os- teogenic differentiation of rat precartilaginous stem cells in vitro and the underlying mechanisms. Methods Rat pre- cartilaginous stem cells （PSCs） were cultured in complete osteogenesis medium containing DMEM/F-12 medium and different e, oncentrations （0, 10 s, 10 9,10-10tool/L） of CGRP, the morphology changes of PSCs were observed. The proliferation of PSCs was examined at different time points by CCK-8. All tile PSCs were then randomly assigned to an experimental group and a control group. The PSCs in the experimental group were cultured in complete osteogenesis medium with 10^-10 mnl/L CGRP, while the control group cultured merely in complete osteogenesis medium was re- ceived no special intervention. Both groups were stained by Alizarin Red and the expression of alkaline phosphatase （ALP） was detected. The osteogenic genes （RUNX2,OPN and BGP） were measured by use of RT-PCR. The activa- tion of Wnt/β-catenin signaling pathway was tested by using Western blotting to evaluate the effect of CGRP. Results Compared to the control group （ the concentration of CGRP was 0 mol/L） , the concentration of ALP was significantly higher in the experimental group, calcium deposition was significantly more obvious, and the expression of the osteogenic genes such as RUNX2,OPN and BGP as well as the β-eatenin protein expression were up-regulated significantly. However, CGRP had no effect on cell proliferation. Conclusion CGRP activated Wnt/β-catenin sig- nal pathway and induced osteogenic differentiation of precartilaginous stem cells.
出处
《中华物理医学与康复杂志》
CAS
CSCD
北大核心
2016年第5期329-334,共6页
Chinese Journal of Physical Medicine and Rehabilitation
基金
国家自然科学基金面上项目（31070831）
