CAJ | 학술논문

目的：观察龟鹿二仙胶含药血清诱导大鼠骨髓基质干细胞(BMSCs)成骨分化的作用并探讨其机制。方法全骨髓贴壁法分离BMSCs，培养至F3代。 MTT法检测龟鹿二仙胶含药血清增殖作用并确定最佳浓度；F3细胞分为10%FBS组、10% KB组、10%GLEXJ组和Classical组，检测碱性磷酸酶(Alkaline phosphatase，ALP)比活性，茜素红染色方法观察对BMSCs成骨分化的作用，RT-PCR方法和Western blot检测Fzd-2、Axin-2基因和蛋白表达的变化。结果 MTT结果显示浓度为10%的龟鹿二仙胶含药血清能促进BMSCs增殖；10%GLEXJ组和Classical组ALP比活性高于其余两组，差异有统计学意义(P<0.05)；且随着时间的延长，增加更明显(P<0.05或P<0.01)，茜素红染色见橘红色钙结节着色；10%GLEXJ组Fzd-2 mRNA和蛋白表达水平显著高于其余各组，差异均有统计学意义(P<0.05或P<0.01)；10%GLEXJ组Axin-2 mRNA和蛋白表达水平显著低于10%FBS组和10%KB组，差异均有统计学意义(P<0.05或P<0.01)。结论龟鹿二仙胶含药血清能促进BMSCs增殖并诱导其向成骨细胞分化，其机制可能与增强Fzd-2表达、抑制Axin-2表达和激活Wnt/β-catenin通路密切相关。
목적：관찰구록이선효함약혈청유도대서골수기질간세포(BMSCs)성골분화적작용병탐토기궤제。방법전골수첩벽법분리BMSCs，배양지F3대。 MTT법검측구록이선효함약혈청증식작용병학정최가농도；F3세포분위10%FBS조、10% KB조、10%GLEXJ조화Classical조，검측감성린산매(Alkaline phosphatase，ALP)비활성，천소홍염색방법관찰대BMSCs성골분화적작용，RT-PCR방법화Western blot검측Fzd-2、Axin-2기인화단백표체적변화。결과 MTT결과현시농도위10%적구록이선효함약혈청능촉진BMSCs증식；10%GLEXJ조화Classical조ALP비활성고우기여량조，차이유통계학의의(P<0.05)；차수착시간적연장，증가경명현(P<0.05혹P<0.01)，천소홍염색견귤홍색개결절착색；10%GLEXJ조Fzd-2 mRNA화단백표체수평현저고우기여각조，차이균유통계학의의(P<0.05혹P<0.01)；10%GLEXJ조Axin-2 mRNA화단백표체수평현저저우10%FBS조화10%KB조，차이균유통계학의의(P<0.05혹P<0.01)。결론구록이선효함약혈청능촉진BMSCs증식병유도기향성골세포분화，기궤제가능여증강Fzd-2표체、억제Axin-2표체화격활Wnt/β-catenin통로밀절상관。
Objective To observe the effect of the Guilu Erxian Glue(GLEXJ) containing serum on the proliferation and the osteogenic differentiation of BMSCs, and try to reveal the mechanism. Methods The BMSCs were isolated by the adherent cell cytopheresis and were expanded to the F3 generation, MTT method was used to observe the effects of GLEXJ-containing serum on the proliferation of BMSCs. Then cells were separated to 4 groups, 10%FBS group, 10%KB group,10%GLEXJ group and Classical group. The ALP activity and the Alizarin Red staining were used to define the osteogenic differentiation of BMSCs. And the mRNA and protein expression of Fzd-2 and Axin-2 were detected by RT-PCR and Western blot method. Results The results of MTT showed that the 10% concentration of GLEXJ-contain-ing serum can stimulate the proliferation of BMSCs. The ALP activity of 10%GLEXJ and Classical group were higher than the other two groups(P<0.05), and the level was gradually increased in a time-dependent manner(P<0.05 or P<0.01); the calcium nodules of orange-red color were observed in the GLEXJ-containing serum group and the Classical group in ARS.The mRNA and protein expression of Fzd-2 in 10%GLEXJ group were higher than the other groups(P<0.05 or P < 0.01); the mRNA and protein expression of Axin-2 in 10%GLEXJ group were lower than the 10%FBS group and the 10%KB group(P< 0.05 or P<0.01). Conclusion The GLEXJ-containing serum can enhance the prolif-eration and the osteogenic differentiation of BMSCs, the possible mechanism is by stimulating the Fzd-2 ex-pression and inhibiting the Axin-2 expression which related with the activity of Wnt/β-catenin pathway.