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目的:探讨片仔癀对人骨肉瘤耐药株MG63/ADM增殖、凋亡的影响及可能的作用机制。方法:采用大剂量间隙干预建立骨肉瘤阿霉素(ADM)耐药细胞MG63/ADM,MTT检测其耐药性;不同浓度片仔癀(0、0.4、0.8、1.2mg/mL)作用MG63/ADM细胞后,MTT检测细胞增殖抑制率;Hoechst 33342染色检测细胞凋亡;Western Blot检测PTEN、Akt、p-Akt、PARP、Cleaved-PARP蛋白表达情况。结果:MG63/ADM细胞对阿霉素明显耐药,IC50为(1.98±0.58)μg/mL,耐药指数为11.47,同时对紫杉醇(PTX)、顺铂(DDP)耐药;片仔癀能明显抑制MG63/ADM的增殖,呈时间和剂量依赖性(P<0.01),Hoechst 33342染色显示片仔癀作用48h后细胞出现细胞核染色质浓聚,凋亡小体等细胞凋亡形态的表现;Wester Blot显示片仔癀作用后PTEN蛋白及Cleaved-PARP表达明显上调,p-Akt蛋白的表达降低,呈浓度依赖性,与对照组比较差异有统计学意义(P<0.05)。结论:片仔癀在体外可明显抑制MG63/ADM细胞的增殖,其机制可能与通过上调PI3K/Akt通路中PTEN的表达,抑制Akt蛋白的磷酸化,从而促进PARP蛋白的裂解,发挥诱导细胞凋亡的作用。
Objective: To investigate the effects of Pien Tze Huang on proliferation and apoptosis of human osteosarcoma drug-resistant MG63 / ADM cells and its possible mechanism. Methods: MG63 / ADM cells were treated with high dose of Gadolinium and MTT assay was used to detect the drug resistance of MG63 / ADM cells. MG63 / ADM cells were treated with different concentrations of Pien Tze Huang (0,0.4,0.8,1.2mg / mL) The inhibitory rate of cell proliferation was detected by MTT assay. Apoptosis was detected by Hoechst 33342 staining. The protein expressions of PTEN, Akt, p-Akt, PARP and Cleaved-PARP were detected by Western Blot. Results: MG63 / ADM cells were resistant to doxorubicin, the IC50 was (1.98 ± 0.58) μg / mL and the drug resistance index was 11.47. Meanwhile, the drug resistance to paclitaxel (PTX) and cisplatin The proliferation of MG63 / ADM cells in a time and dose-dependent manner (P <0.01). Hoechst 33342 staining showed the appearance of apoptotic bodies such as chromatin condensation and apoptotic bodies in the cells after 48 hours of Pien Tze Huang treatment. Wester Blot showed that Pien Tze Huang After treatment, the expression of PTEN and Cleaved-PARP were significantly up-regulated, while the expression of p-Akt was decreased in a concentration-dependent manner. The difference was significant compared with the control group (P <0.05). CONCLUSION: Pien Tze Huang can significantly inhibit the proliferation of MG63 / ADM cells in vitro. The mechanism may be through up-regulating the expression of PTEN in PI3K / Akt pathway, inhibiting the phosphorylation of Akt protein, promoting the cleavage of PARP protein and inducing cell apoptosis effect.