调衡方对Lewis肺癌细胞凋亡标志物bax及bcl-2表达的影响

Expression of Bax and Bcl-2 of Lewis lung cancer cells apoptosis markers induced by Tiaoheng prescription

目的探讨调衡方含药血清诱导Lewis肺癌细胞凋亡的作用及其分子机制。方法采用细胞培养技术,用不同浓度的含药血清(无特定病原体雌性SD大鼠32只随机分为4组,每组8只,灌服给药,制备调衡方大、中、小剂量含药血清,正常血清。又将细胞分为5组,调衡方大、中、小剂量含药血清组,中剂量含药血清+AG490干预组,正常血清组)处理Lewis肺癌细胞,用溴化四唑蓝(MTT)法检测药物对细胞增殖的影响;以流式细胞仪测定碘化丙啶染色细胞周期的变化;采用实时聚合酶链式反应(PCR)法检测bax和bcl-2的mRNA表达。结果调衡方含药血清处理Lewis肺癌细胞后,随着药物浓度增加,作用时间延长,细胞增殖速度减慢。流式细胞仪检测结果显示,调衡方含药血清使Lewis肺癌细胞阻滞于S期,随着药物浓度的增加,细胞凋亡逐渐增加,中、大剂与中剂量+AG490组尤为明显(P<0.01)。调衡方含药血清还能够促进细胞中bax mRNA表达,减少bcl-2 mRNA表达(P<0.05,P<0.01),并呈浓度依赖性。结论调衡方含药血清能降低Lewis肺癌细胞内bcl-2含量及提高bax的表达,使bax/bcl-2比值升高,这可能是调衡方抗肿瘤诱导Lewis肺癌细胞凋亡的机制。

Objective To investigate the effect of Tiaoheng prescription medicated serum on apoptosis of Lewis lung cancer cell line and explore the possible molecular mechanism. Methods 32 experimental animals were randomly divided into four groups. High - , middle - , low - dose Tiaoheng prescription and normal saline were a- dopted for preparation of different medicated serum. Ceils were divided into five groups, High - , middle - , low - dose Tiaoheng prescription groups, AG490 treatment group and JAK ＋ STAT signal path specified inhibitor group. Lewis lung cancer ceils were cultured and treated with 101~1 serum of the Tiaoheng Prescription and normal saline for 24, 48 and 72 h. Changes of cell proliferation were analyzed by MTr assay and alteration of cell cycles were deter- mined by flow cytometry after propidium iodide staining. Expression levels of Bax mRNA and Bcl - 2 mRNA were ex- amined by real time PCR method. Results The rate of cell proliferation was significantly slowed down by Tiaoheng Prescription in a time and dose dependent manners. Flow cytometry showed that Lewis lung cancer cells were blocked at S stage after drug treatment （ P 〈 O. 05, P 〈 0.01 ）. Tiaoheng Prescription serum induced apoptosis of Lewis lung cancer cells in a dose dependent manner, Middle dose group, high dose group and middle dose ＋ AG490 group ob- viously（ P 〈 0.01 ）. Besides, the expression level of bax mRNA was significantly increased by Tiaoheng Prescription serum（ P 〈0.05, P 〈0. O1 ）, while the expression level of bc1-2 mRNA was reduced in a dose dependent manner.Conclusion Tiaoheng prescription medicated serum can decrease Bcl -2 content of Lewis lung cancer cell and enhance the expression of Bax, increase Bax / Bcl -2ratio, which may be mechanism for inducing apoptosis of Lewis lung carcinoma and anti -tumor.