桦木酸在酸性微环境中对PC-3细胞生物学行为及破骨细胞分化的作用

Study on the effect of betulinic acid on the biological behavior and osteoclast differentiation of PC-3 cells in acidic microenvironment

目的研究桦木酸在酸性微环境中对PC-3细胞增殖、迁移、黏附、成球、干细胞特性及破骨细胞分化的影响。方法使用pH 7.4和pH 6.5的培养基培养PC-3细胞,然后加入浓度为0、1、5、10、15、20、30、40μmol/L的桦木酸继续培养,24 h后采用CCK-8法和Transwell检测桦木酸对PC-3细胞增殖与迁移的影响,通过黏附试验、成球试验检测BA对PC-3细胞干细胞特性的变化,通过破骨细胞分化实验检测桦木酸对破骨细胞分化的影响,利用qPCR和Western blot检测PC-3细胞中CD31和CD44蛋白的表达。结果CCK-8结果表明与中性对照组(pH 7.4)相比,酸性微环境(pH 6.5)能促进PC-3细胞增殖,而桦木酸呈浓度依赖性抑制PC-3细胞在酸性微环境中的增殖(P<0.05)。此外,酸性微环境促进PC-3细胞的迁移、黏附、瘤体形成及CD31和CD44表达,在此基础上加入桦木酸后,PC-3细胞的迁移能力、黏附、瘤体形成、干细胞特征和破骨细胞分化明显下调(P<0.05)。结论桦木酸可抑制酸性微环境中PC-3细胞的增殖、迁移、干细胞特性及破骨细胞分化。

Objective To investigate the effects of Betulinic acid(BA)on proliferation,migration,adherence properties,tumorsphere formation,stem cell property and osteoclast differentiation in human prostate cancer PC-3 cells in acidic microenvironment.Methods Culturing PC-3 cells in pH 7.4 and pH 6.5 medium.Effects of BA on proliferation and migration of PC-3 cells were assessed with the CCK-8 assay and Transwell assays,respectively.Effects of BA on stem cell characteristics of PC-3 cells were evaluated by cell-matrix adhesion assay.The osteoclastogenesis activity of BA in RAW264.7 cells was examined by osteoclast differentiation assay.The express levels of CD31 and CD44 in PC-3 cells were measured by qPCR and Western blot.Results The result of CCK-8 test showed that acidic medium significantly enhanced cell viability of PC-3 cells than neutral medium,whereas BA suppressed the proliferation of PC-3 cells in a concentration-dependent under acidic microenvironment(P<0.05).In addition,the acidic medium also significantly enhanced cell migration,adherence,tumor sphere formation and expressed CD31,CD44 of PC-3 cells than neutral medium.After that,BA was added and the migration,adherence,tumor sphere formation,stem cell property and osteoclast differentiation of PC-3 cells were significantly inhibited(P<0.05).Conclusion Betulinic acid significantly inhibited the proliferation,migration,adherence,tumor sphere formation,stem cell property and osteoclast differentiation of PC-3 cells under acidic microenvironment.