启膈散及其拆方对食管癌细胞Eca109 PLC-γ1介导的细胞信号转导的影响

Effects of qigesan and its constituents on phospholipase C-γ1-mediated signaling in esophageal carcinoma cell line Eca109

目的:观察启膈散及其活血和化痰两个拆方对人食管癌细胞株Eca109细胞磷脂酶C-γ1(PLC-γ1)介导的细胞信号转导的影响,以研究启膈散治疗食管癌的作用机制.方法:用启膈散(W)及其活血(P)和化痰(R)两个拆方的水提物处理体外培养的人食管癌细胞株Eca109细胞,测定细胞PLC-γ1、表皮生长因子受体(EGFR)和PKCα蛋白表达水平、蛋白酪氨酸磷酸化(PY99)水平和豆蔻酰化富丙氨酸C激酶底物(MARCKS)(Ser152/156)磷酸化水平、细胞内游离钙离子[Ca^(2+)]i浓度及蛋白激酶C(PKC)活性.结果:1mg/L的启膈散及其拆方明显抑制Eca109细胞PLCγ1蛋白表达(0.31%,0.42%,0.20%),随着药物浓度增加其抑制作用分别增强,启膈散及其拆方对体外培养的食管癌细胞PLC-γ1、EGFR和PKCα表达、PLC-γ1和EGFR酪氨酸磷酸化、MARCKS磷酸化、[Ca^(2+)]i浓度(169.65±30.54 nmol/L,145.84±24.12 nmol/L.214.94±37.14 nmol/L vs 231.86±51.10 nmol/L,P>0.05,P<0.05,P>0.05)及PKC活性均有不同程度的抑制作用,以P组作用最强.结论:启膈散及其拆方通过不同程度的抑制食管癌细胞PLC-γ1介导的信号转导而抑制肿瘤细胞的生长.

AIM： To study the effects and mechanism of action of qigesan and its constituents on the human esophageal carcinoma cell line Eca 109 in vitro. METHODS： The human esophageal carcinoma cell line Eca 109 was cultured and treated with different concentrations of qigesan as a whole （W group）, water extracts of the constituents of qigesan that promote blood circulation （P group）, and other water extracts of the constituents of qigesan that help prevent stasis （R group）. Protein expression of phospholipase C （PLC）-γ1, epidermal growth factor receptor （EGFR） and protein kinase C （PKC）α, protein tyrosine phosphorylation level （PY99）, intracellular calcium concentration （[Ca^2＋]i）, and PKC activity weremeasured RESULTS： The protein expression levels of PLC-γ1, EGFR and PKCα, EGFR and PLC-γ1 tyrosine phosphorylation level, [Ca^2＋]i （169.65 ± 30.54 nmol/L, 145.84 ± 24.12 nmol/L, 214.94 ± 37.14 nmol/L vs 231.86 ± 51.10 nmol/L; P 〉 0.05, P 〈 0.05, P 〉 0.05） and PKC activity in the cultured esophageal cancer cells were inhibited by qigesan and its individual constituents at different concentrations. The greatest effects were seen in group P. CONCLUSION： Qigesan and its individual constituents inhibit esophageal carcinoma cell growth by inhibiting PLC-γ1 mediated signal transduction.