葛根素促进体外培养大鼠颅骨成骨细胞增殖和分化过程中对cAMP/PKA信号通路的影响  

Effects of Puerarin in Promoting Proliferation and Differentiation of Rats' Osteoblast by Culture in Vitro on cAMP/PKA signal Pathway

作　　者：邓银栓[1] 樊宇[2] 惠升明 杨小杰[1] 董成辉 

机构地区：[1]兰州军区兰州总医院运动医学科 [2]兰州军区机关门诊部 [3]68202部队医院

出　　处：《解放军医药杂志》2017年第7期5-9,共5页Medical & Pharmaceutical Journal of Chinese People’s Liberation Army

基　　金：军区医药卫生课题(CLZ14JA05)

摘　　要：目的研究葛根素对体外培养大鼠颅骨成骨细胞(rats osteoblast,ROBs)增殖和分化及cAMP/PKA信号通路的影响。方法体外分离培养ROBs,观察不同浓度(1×10^(-4)~1×10^(-8)mol/L)葛根素对ROBs增殖和碱性磷酸酶(alkaline phosphatase,ALP)活性的影响;分析葛根素对骨形态发生蛋白(Bone morphogenetic protein,BMP-2)、Ⅰ型胶原(ColⅠ)、成骨特异性转录因子-2(Runx-2)和Oxterix mRNA表达水平的影响;观察1×10^(-6)mol/L葛根素对BMP-2、Col-Ⅰ、Runx-2、Oxterix、PKA和磷酸化PKA(p-PKA)蛋白表达水平,测定细胞中c AMP的变化。结果细胞增殖结果显示1×10^(-7)mol/L ROBs细胞浓度高于对照组,1×10^(-4)mol/L组ROBs细胞浓度低于对照组(P<0.01)。1×10^(-5)、1×10^(-6)、1×10^(-7)mol/L组ROBs ALP活性均高于对照组(P<0.05,P<0.01)。1×10^(-6)mol/L葛根素处理ROBs Runx-2、Oxterix、BMP-2、ColⅠmRNA和蛋白及p-PKA、c AMP表达水平均高于对照组(P<0.05,P<0.01)。结论 1×10^(-6)mol/L葛根素为促进ROBs增殖和分化的最适浓度,其可促进体外培养ROBs分化过程中c AMP/PKA信号通路的激活。Objective To investigate effects of Puerarin on promoting proliferation and differentiation of rats＇os- teoblasts （ROBs） by culture in vitro on cAMP/PKA signal pathway. Methods ROBs were isolated and cultured in vitro, and effects of different concentrations Puerarin （1 × 10^-4 mol· L^-1- × 10^-8 mol·L^-1） on proliferation and alka- line phosphatase （ALP） activity of ROBs were observed. Effects of Puerarin on bone morphogenetic protein-2 （BPM-2）, type I collagen （Col I ） , osteogenie specific transfer factor-2 （Runx-2） and Oxterix mRNA expression levels were analyzed. Effects of Puerarin （1×10^-6 mol/L） on BPM-2, collagen-l, Runx-2, Oxterix, PKA and phosphorylation-PKA （p-PKA） protein expression levels were observed, and cAMP changes was detected. Results The result of cell prolifer- ation showed that cell concentration of ROBs by 1 × 10^-7 mol/L Puerarin was significantly increased, but the concentration of ROBs by 1 x 10^-4 mol/L was significantly decreased compared with those in control groups （P 〈 0.01 ）. Values of ROBs ALP activity in 1 × 10^-5, 1 × 10^-6 and 1 × 10^-7 mol/L groups were significantly increased than those in control group （P 〈0.05, P 〈0.01 ）. ROBs Runx-2, Oxterix, BPM-2, collagen-1 mRNA and protein expression levels, and p- PKA and cAMP expression levels in 1×10^-6 mol/L Puerarin group were significantly increased compared with those in control group （ P 〈 0.05, P 〈 0.01 ）. Conclusion The 1 × 10^-6 mol/L Puerarin is the best concentration for promoting ROBs proliferation and differentiation, and it may promote activation of cAMP/PKA cell signal pathway during ROBs differentiation by culture in vitro.

关 键 词：葛根素 成骨细胞 颅骨 细胞增殖 细胞分化 大鼠 Wistar 

分 类 号：R332[医药卫生—人体生理学] R681[医药卫生—基础医学]