摘要
背景:降钙素是强有力的破骨细胞抑制剂,可直接、快速而广泛地抑制骨吸收,近年来有学者报道其有促进骨折愈合的功能,推测降钙素对成骨细胞也有直接的作用。目的:体外观察降钙素对大鼠成骨细胞的药理作用,分析其是否通过细胞外信号调节激酶信号通路产生。设计、时间及地点:分组对照观察,于2007-06/2008-05在南京医科大学药理学实验室完成。材料:选用新生SD大鼠,分离培养颅骨成骨细胞。方法:取第2代成骨细胞进行分组实验:0.01μg/L降钙素组、0.1μg/L降钙素组、1μg/L降钙素组分别加入不同剂量降钙素进行干预,U0126+1μg/L降钙素组提前30min加入100μmol/L的细胞外信号调节激酶抑制剂U0126,并设空白对照组。主要观察指标:通过四甲基偶氮唑盐比色法、对硝基苯磷酸盐法和钙结节茜素红染色法分别检测降钙素对成骨细胞的增殖、分化、矿化能力的影响;应用反转录-聚合酶链反应技术检测促骨形成关键基因核心结合因子a1mRNA的表达情况;应用WesternBlotting技术检测磷酸化细胞外信号调节激酶1/2蛋白表达情况。结果:加入0.01,0.1,1μg/L的降钙素干预后,成骨细胞的增殖、分化、矿化能力以及核心结合因子a1mRNA的表达均随降钙素剂量增加而增强,且0.1,1μg/L组与空白对照组比较差异具有显著意义(P<0.05～0.01);此外降钙素促进了磷酸化细胞外信号调节激酶1/2蛋白的表达(P<0.05)。U0126可阻断以上所有效应(P<0.05～0.01)。结论:降钙素可能通过细胞外信号调节激酶信号通路调控促骨形成关键基因核心结合因子a1mRNA的表达,进而促进成骨细胞的增殖、分化、矿化能力。
BACKGROUND： Calcitionin is a kind of drastic osteoclast inhibitor, which can inhibit bone resorption directly, quickly and extensively. Recent studies demonstrated that calcitionin can promote union of fracture, suppose that it has direct effect on osteoblast. OBJECTIVE： To observe the effect of calcitionin on calvarial osteoblasts in vitro, in addition, to analyze if it can regulate extracellular signal-regulated kinase （ERK）. DESIGN, TIME AND SETTING： The grouping controlled experiment was performed in Pharmacology Laboratory of NanJing Medical University from July 2007 to May 2008. MATERIALS： The calvarial osteoblast was cultured from new born Sprague Dawley rats. METHODS： The second generation osteoblasts were divided into groups according to different calcitionin concentrations （0.01 μg/L, 0.1μg/L, 1 μg/L）, and another 1 μg/L calcitionin was added with U0126, in addition, a blank control group was prepared. MAIN OUTCOME MEASURES： The ability of cell proliferation, differentiation and mineralization were evaluated by MTT, PNPP and mineralized nodes staining respectively. The expression of core binding factor al （cbfal） mRNA in different groups was measured by RT-PCR, and the phosphor-ERK1/2 expression of the calvarial osteoblasts was analyzed by Western Blotting. RESULTS： The ability of proliferation, differentiation and mineralization, as well as the expression of al mRNA showed a dose-dependent manner after treated with 0.01, 0.1, 1 μg/L calcitionin, which had a significant differences compared with the 0.01 μg/L, 0.1 μg/L and the blank control group （P〈 0.05-0.01）. More over, calcitionin facilitated the expression of the phospho-ERK1/2 （P〈 0.05）, however, all the effects above could be blocked by U0126 （P〈 0.05-0.01）. CONCLUSION： Calcitionin facilitates the ability of proliferation, differentiation and mineralization by regulating the cbfalmRNA expression, which may related to the signal pathway of ERK-MAPK.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2009年第2期233-237,共5页
Journal of Clinical Rehabilitative Tissue Engineering Research
