补肾疏肝方对人肺癌A549细胞及其相关干性因子的影响

目的:探讨补肾疏肝方对人肺癌A549细胞及其相关干性因子的影响。方法:SD雄性大鼠随机分为6组,每组10只,分别为补肾疏肝方低、高剂量组(15,30 g·kg-1),顺铂组(8 g·kg-1),低剂量联合顺铂组(联合低剂量组),高剂量联合顺铂组(联合高剂量组),正常组,顺铂组ip给予相应药物,补肾疏肝方ig给予相应药物,正常组给予等体积的生理盐水,制备大鼠含药血清;分别配制10%含补肾疏肝方低、高剂量、顺铂、低剂量联合顺铂、高剂量联合顺铂,分别与肺癌A549细胞共同培养,CCK8法检测补肾疏肝方含药血清对A549增殖的影响;流式细胞术检测含药血清对A549细胞周期的影响;Annexin V/PI双染流式细胞术观察含药血清对A549细胞凋亡的影响;荧光定量PCR检测含药血清对相关m RNA表达的影响;流式细胞术检测含药血清对CD133蛋白表达的影响。结果:补肾疏肝方以时间、剂量依赖方式抑制A549细胞增殖,与顺铂联合有协同增效作用,24,48,72 h高剂量联合顺铂组的抑制率分别为45.39%,54.76%及59.94%;与正常组比较,补肾疏肝方、联合、顺铂含药血清可促进A549细胞凋亡,随着浓度增加,G2/M期细胞的百分比明显增加,明显下调CD133,SOX-2,OCT-4 m RNA的表达,明显降低CD133蛋白表达量,均具有明显统计学差异(P0.05,P0.01)。结论:补肾疏肝方可抑制A549细胞增殖,促进细胞凋亡,与顺铂联合可下调CD133,SOX-2,OCT-4等干性因子的表达。

Effects of Bushen Shugan Formula on Human Lung Cancer A549 Cells and Related Stem Genes

Objective: To study the effects of Bushen Shugan formula (BSSGF) on human lung cancer A549 cells and related stem genes. Method: SD rats were randomly divided into 6 groups:the normal group (normal saline), the cisplatin group (8 g · kg^-1), the low-and high-dose BSSGF groups (15, 30 g · kg^-1), the low-dose BSSGF plus cisplatin group and the high-dose BSSGF plus cisplatin group of 10 rats each. The rats received the corresponding medicines and the serum containing drugs were prepared. A549 cells were cultured in the drug serum with different concentrations. The cell proliferation was detected by CCK8 assay.The cell cycle was tested by the flow cytometry. The cell apoptosis was detected by using Annexin V/PI staining with flow cytometry. The related gene expression was detected by using quantitative PCR. The protein expression of CD133 was tested by using flow cytometry. Result: BSSGF could inhibit the proliferation of A549 in a time-and dose-dependent manner and had synergies when combined with cisplatin. The inhibition rate was 45.39%, 54.76% and 59.94%, respectively, after 24, 48 and 72 h incubation BSSGF could induce the cell apoptosis, the cell percentage in G₂/M increased with the increasing of concentration. BSSGF combined with cisplatin could reduce the gene expressions of CD133, SOX-2, OCT-4, and lowered the protein expression of CD133. Conclusion: BSSGF could inhibit the cell proliferation and promote the cell apoptosis of A549 cells. Combined with cisplatin, BSSGF could reduce the expression of stem factors, such as CD133, SOX-2 and OCT-4.