左归丸含药血清通过JNK信号通路诱导MC3T3成骨细胞分化的研究  

Effect of serum containing Zuogui Pills on differentiation of MC3T3 osteoblast via JNK pathway

作　　者：刘立萍[1] 任艳玲[1] 李然[1] 王智民[2] 蒿长英[1] 宋囡[1] 

机构地区：[1]辽宁中医药大学基础医学院,辽宁沈阳110847 [2]辽宁中医药大学第一临床学院,辽宁沈阳110847

出　　处：《中成药》2012年第8期1433-1437,共5页Chinese Traditional Patent Medicine

基　　金：国家自然基金项目(30873226);教育部高等学校博士学科点专项科研基金(20102133110001);辽宁省高等学校优秀人才支持计划资助(LR201025)

摘　　要：目的探讨JNK信号通路对左归丸(熟地、菟丝子、川牛膝、龟甲胶、鹿甲胶、山药、山茱萸、枸杞子)含药血清干预MC3T3成骨细胞分化的影响。方法选用JNK特异抑制剂SP600125,制备大鼠含药血清,实验分为空白对照组、SP600125组、左归丸组、左归丸加SP600125组、倍美力组、倍美力加SP600125组。孵育48 h后,采用PNPP法检测碱性磷酸酶(ALP)活性,采用Western blot法分析ALP蛋白表达水平;孵育7 d,采用改良钙钴染色法检测ALP活性;孵育14 d,采用茜素红染色法测定矿化沉积情况。结果与空白对照组比较,左归丸组显著促进MC3T3成骨细胞分化,明显上调ALP蛋白表达水平(P<0.01);与左归丸组比较,左归丸加SP600125组孵育48 h对ALP活性及蛋白影响不显著,但显著对抗左归丸含药血清对孵育7 d的ALP活性和孵育14 d的矿化结节的促进作用(P<0.01)。结论左归丸含药血清可能部分通过JNK信号通路干预MC3T3成骨细胞分化,在分化末期尤其依赖JNK通路。AIM To investigate the role of c-Juk N-terminal kinase （JNK） pathway in osteoblast MC3T3 differentiation stimulated by serum containing Zuogui Pills （Rehmanniae Radix praeparata, Cuscutae Semen, Cyathulae Radix, Testudinis Carapacis et Plastr Colla, Cervi Comus Colla, Dioscoreae Rhizoma, Corni Fructus, Lycii Fructus）. METHODS Sixty rats were used to prepare the rat serum containing Zuogui Pills. Blood cells were preineubated in the absense or presence of the specific inhibitors of JNK SP6001250, and were assigned to six groups, including control, SP600125, Zuogui Pills, Zuogui Pills plus SP600125, premarin and premarin plus SP600125 group. The test for alkaline phosphatase （ALP） activity and protein expression was carried out with PNPP and Western blot method, respectively, placed in incubator for 48 h. The ALP test was measured by modified calcium-combat staining method on the seventh day and alizarin red staining method was applied to the detection of the mineralization deposition. RESULTS As compared with the control group, Zuogui Pills group markedly promoted the differentiation of MC3T3 osteoblast and upregulated the expression of ALP. As compared with the Zuogui Pills group, in Zuogui Pills plus SP600125 in 48 h incubation, ALP activity and protein expression were unaffected, but opposed ALP activity in the seventh day incubation and mineralized deposition on the fourteenth day incubation with serum containing Zougui Pills. CONCLUSION Serum containing Zuogui Pills in part intervenes differentiation of MC3T3 osteoblast through JNK pathway, especially in telophase.

关 键 词：左归丸 含药血清 MC3T3细胞 碱性磷酸酶活性 矿化沉积 JNK信号通路 

分 类 号：R285.5[医药卫生—中药学]