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目的:观察复方接骨片含药血清对BMP-2沉默的骨髓间充质干细胞体外增殖及分化的影响。方法:设计三对BMP-2 si RNA,利用脂质体介导的si RNA转染骨髓间充质干细胞,PCR及Western blot法筛选活性;将复方接骨片含药血清加入BMP-2沉默的骨髓间充质干细胞中,MTT法及碱性磷酸酶法检测细胞增殖和分化情况。结果:三对si RNA中,si RNA3在抑制BMP-2的m RNA和蛋白表达的效果最好,抑制率可达61.6%和50.9%;正常骨髓间充质干细胞加入含药血清,48 h后可明显提高增殖效率(P<0.05)及碱性磷酸酶活性(P<0.01);骨髓间充质干细胞转染BMP-2 si RNA3,48 h后碱性磷酸酶活性显著降低(P<0.01),72 h后细胞活性显著降低(P<0.01),而加入含药血清干预,可显著拮抗上述抑制作用(P<0.01)。结论:复方接骨片含药血清能有效拮抗BMP-2 si RNA对骨髓间充质干细胞体外增殖及分化的抑制作用。
OBJECTIVE: To observe the effects of Fufangganggu tablet serum on the proliferation and differentiation of bone marrow mesenchymal stem cells (BMP-2) silenced in vitro. Methods: Three pairs of BMP-2 si RNAs were designed and transfected into bone marrow-derived mesenchymal stem cells by liposome-mediated siRNA. The activity of BMP-2 was determined by PCR and Western blot. Mesenchymal stem cells, MTT assay and alkaline phosphatase assay cell proliferation and differentiation. Results: Among the three pairs of si RNAs, si RNA3 had the best inhibitory effect on the expression of m RNA and protein of BMP-2, with the inhibitory rates up to 61.6% and 50.9%, respectively. The normal MSCs were added to the drug-containing serum for 48 h (P <0.05) and alkaline phosphatase activity (P <0.01). After transfection of BMP-2 si RNA3 by bone marrow mesenchymal stem cells for 48 h, alkaline phosphatase activity was significantly decreased (P <0.01) (P <0.01). However, the cell viability was significantly reduced after 72 h intervention (P <0.01). CONCLUSION: Fufangjiegu tablet serum can effectively inhibit the inhibitory effect of BMP-2 si RNA on the proliferation and differentiation of bone marrow mesenchymal stem cells in vitro.