TLR2配体肽聚糖对骨髓间充质干细胞增殖和细胞周期的影响

Effects of Peptidoglycan on Proliferation and Cell Cycle of Human Bone Marrow-derived Mesenchymal Stem Cells

本研究探讨不同剂量的Toll样受体2(TLR2)的配体肽聚糖(peptidoglycan,PGN)在体外对人骨髓间充质干细胞(BM-MSC)增殖和细胞周期的影响。采用Ficoll淋巴细胞分离液、密度梯度离心法分离BM-MSC,体外扩增,并通过流式细胞术检测其表面标志以鉴定纯度;将分离培养的BM-MSC按一定浓度接种于96孔板中,以1、10、20μg/ml的PGN与BM-MSC共培养为实验组,不加PGN的BM-MSC为对照组;培养第1至第7天,MTT法检测BM-MSC的增殖情况;以上述浓度的PGN与BM-MSC共培养72小时后,流式细胞术检测BM-MSC的细胞周期。结果表明,BM-MSC与PGN共培养后,BM-MSC的增殖指数明显增加,呈时间、剂量依赖性,以10μg/mlPGN促BM-MSC增殖作用最明显。与对照组比较,PGN干预组G0/G1期细胞比例降低,S期和G2/M期细胞比例增加,以10μg/ml组最明显。结论:在培养条件下PGN可以促进更多的BM-MSC进入DNA合成期,从而有更多的细胞分裂,产生大量的增殖细胞,并且这种影响呈时间、剂量依赖性。

This study was aimed to explore the effects of peptidoglycan （PGN） on proliferation and cell cycle of human bone marrow mesenchymal stem cells （MSCs）. MSCs were isolated from human bone marrow by density gradient centrifugation. The purity of MSCs with the spindle fibroblastic morphology was identified by microphotography and the phenotypes were detected by flow cytometry （FCM）. MSCs incubated with different doses of PGN （ 1,10,20 μg/ml） were used as test groups, and those incubated without PGN were regarded as control group. The isolated and cultured MSCs were inoculated into 96-well plates according to a certain concentration. Cell cycle was measured by flow cytometry after incubated with PGN for 72 hours. The results showed that the cell proliferation index was significantly increased in dose and time dependent manners after MSCs was incubated with PGN. Its effects on the proliferation of MSCs were highest in 10 μg/ml group. Compared with the control group, PGN could significantly decrease proportion of MSCs in G0/G1 phase and increase them in S and G2/M phases（p 〈0.05 ）. It is concluded that PGN can promote more MSCs to enter the DNA synthesis phase and proliferate many much MSCs in dose and time dependent manners.