CAJ | 학술논문

目的：通过观察补肾活血方含药血清对人成骨细胞的增殖分化的影响，研究补肾活血方对 p38 MAPK 和 PI3K/ Akt 信号转导通路的活性相互关联及影响，以探讨补肾活血方临床防治骨质疏松症的作用机制。方法将人成骨细胞株分为空白组（Control 组）、补肾活血方含药血清组（ Z组）、含药血清+ p38 MAPK 通路抑制剂 SB203580组（ SB 组），含药血清+ PI3K/ Akt 通路抑制剂LY294002组（LY 组）、含药血清+ SB203580+ LY294002组（SB + LY 组），通过四甲基偶氮唑盐比色法（MTT 法）检测成骨细胞的增殖情况，生化方法分析检测成骨细胞碱性磷酸酶（ ALP）的活性，采用Western blot 法检测 p38 MAPK 通路磷酸化 p38（p - p38）和 PI3K/ Akt 通路磷酸化 AKT（pAKT）的表达水平。结果补肾活血方含药血清能促进成骨细胞的增殖，使成骨细胞的分化增强，与 Control 组比较差异均有统计学意义（P <0.01）；阻断 p38 MAPK 通路后，对成骨细胞的增殖无明显抑制（ P >0.05），而阻断 PI3K/ Akt 通路后细胞增殖下降（P <0.01）；两条通路对成骨细胞的分化功能均有显著抑制作用（P <0.01）；Western blot 结果显示，补肾活血方含药血清能刺激 p - p38和 pAkt 表达，阻断p38 MAPK 和 PI3K/ Akt 信号转导通路后，p - p38含量和 pAkt 明显减少（P <0.01）；补肾活血方激活的两条通路之间存在一定的关联性，可互相影响。结论补肾活血方通过 PI3K/ AKT 信号通路调控成骨细胞的增殖，通过同时激活 p38 MAPK 和 PI3K/ AKT 两条信号通路介导成骨细胞的分化，这可能是补肾活血方临床治疗 OP 的作用机制之一。
목적：통과관찰보신활혈방함약혈청대인성골세포적증식분화적영향，연구보신활혈방대 p38 MAPK 화 PI3K/ Akt 신호전도통로적활성상호관련급영향，이탐토보신활혈방림상방치골질소송증적작용궤제。방법장인성골세포주분위공백조（Control 조）、보신활혈방함약혈청조（ Z조）、함약혈청+ p38 MAPK 통로억제제 SB203580조（ SB 조），함약혈청+ PI3K/ Akt 통로억제제LY294002조（LY 조）、함약혈청+ SB203580+ LY294002조（SB + LY 조），통과사갑기우담서염비색법（MTT 법）검측성골세포적증식정황，생화방법분석검측성골세포감성린산매（ ALP）적활성，채용Western blot 법검측 p38 MAPK 통로린산화 p38（p - p38）화 PI3K/ Akt 통로린산화 AKT（pAKT）적표체수평。결과보신활혈방함약혈청능촉진성골세포적증식，사성골세포적분화증강，여 Control 조비교차이균유통계학의의（P <0.01）；조단 p38 MAPK 통로후，대성골세포적증식무명현억제（ P >0.05），이조단 PI3K/ Akt 통로후세포증식하강（P <0.01）；량조통로대성골세포적분화공능균유현저억제작용（P <0.01）；Western blot 결과현시，보신활혈방함약혈청능자격 p - p38화 pAkt 표체，조단p38 MAPK 화 PI3K/ Akt 신호전도통로후，p - p38함량화 pAkt 명현감소（P <0.01）；보신활혈방격활적량조통로지간존재일정적관련성，가호상영향。결론보신활혈방통과 PI3K/ AKT 신호통로조공성골세포적증식，통과동시격활 p38 MAPK 화 PI3K/ AKT 량조신호통로개도성골세포적분화，저가능시보신활혈방림상치료 OP 적작용궤제지일。
Objective To observe the impact of the drug - contained serum of bushen huoxue for-mula on the proliferation and differentiation of human osteoblasts and study the correlation and impact of the formula on p38MAPK and PI3K/ Akt signal pathway so as to explore the action mechanism of the formula in the clinical treatment of osteoporosis. Methods The human osteoblasts were divided into a control group,a formula drug - contained serum group(Z group),a drug - contained serum + p38MAPK pathway inhibitor SB203580 group( SB group),a formula drug - contained serum + PI3K/ Akt pathway inhibitor LY294002 group(LY group)and a formula drug - contained serum + SB203580 + LY294002 group(SB + LY group). MTT was adopted to detect the proliferation of osteoblasts and the biochemical analysis was to detect osteo-blast ALP activity and Western blot method to determine p - p38 and pAKT expression. Results The drug -contained serum of bushen huoxue formula promoted osteoblast proliferation and enhanced the differentiation, presenting the significant difference compared with the control group( P < 0. 01). After the blockage of p38MAPK pathway,the proliferation of osteoblasts was not apparently inhibited(P > 0. 05),but the cell pro-liferation was reduced(P < 0. 01). Both of the pathways presented the significant inhibitory effect of osteo-blast differentiation(P < 0. 01). Western blot result showed that the formula drug - contained serum stimula-ted p - p38 and pAkt expression. After the blockage of p38MAPT and PI3K/ Akt signal pathway,p - p38 and pAkt expressions were apparently reduced(P < 0. 01). The two pathways activated by bushen huoxue formula existed a certain correlation and affected interactively. Conclusion Bushen huoxue formula regulates osteo-blast proliferation via regulating PI3K/ AKT,simultaneously,mediated osteoblast differentiation via activating p38MAPK and PI3K/ AKT pathways. All of these probably are the mechanisms of buhen huoxues formula for the clinical treatment of osteoporosis.