葡萄糖饥饿对胃癌细胞MMP-2、MMP-9表达的影响及意义

Effect and significance of glucose starvation on expressions of MMP-2 and MMP-9 in gastric cancer cells

目的探讨葡萄糖饥饿对胃癌AGS和MKN28细胞MMP-2、MMP-9表达的影响及意义。方法胃癌细胞AGS和MKN28在含不同葡萄糖浓度(5mmol/L、1mmol/L、0.5mmol/L、0.05mmol/L、0mmol/L)培养液中培养24h后,Westernblotting检测细胞内MMP-2、MMP-9蛋白表达变化,荧光定量PCR检测MMP-2、MMP-9mRNA表达变化,明胶酶谱实验检测细胞培养上清中MMP-2、MMP-9活性。CCK-8检测在葡萄糖饥饿环境下外源性脯氨酸(5mmol/L)对胃癌细胞活力的影响。结果随着培养液中葡萄糖浓度降低,胃癌细胞内MMP-2和MMP-9在蛋白水平和转录水平呈浓度梯度依赖上调,且在葡萄糖完全缺乏条件下表达上调达到最大。对培养液上清中酶活性检测呈相同趋势。与未添加脯氨酸组相比,外源性添加脯氨酸(5mmol/L)可改善葡萄糖饥饿环境下胃癌细胞活力,这一效应在12h达到最大。结论葡萄糖饥饿可促使胃癌细胞内MMP-2和MMP-9表达及酶解活性增强,加剧细胞外基质中胶原分解代谢,而细胞外游离脯氨酸可支持乏营养肿瘤微环境下胃癌细胞的存活。

Objective To investigate the effect of glucose starvation on the expressions of MMP-2 and MMP-9 in gastric cancer cell lines AGS and MKN28, and further elucidate its significance. Methods Gastric cancer cells were cultured in RPMI-1640 containing different concentrations of glucose (5 mmol/L, 1 mmol/L, 0.5 mmol/L, 0.05 mmol/L, 0 mmol/L), expressions of MMP-2 and MMP-9 were detected by Western blotting and Real-time PCR, activation of MMP-2 and MMP-9 in FBS-free supernants were determined by gelatin zymography. Effect of exogenous proline (5 mmol/L) on the viability of gastric cancer cells under glucose starvation was detected by CCK-8. Results The expressions of MMP-2 and MMP-9 protein or mRNA were increased with the concentrations of glucose lowered in the medium. A concentration-dependent increase in both enzymes activities was also observed. Maximum increase in expression or activity was detected in the complete absence of glucose in the medium. The addition of exogenous proline (5 mmol/L) significantly improved gastric cancer cells viability under glucose starvation at the time point 12 hours. Conclusion The glucose starvation induced extracellular matrix degradation by upregulating MMP-2 and MMP-9, result in making proline available to support gastric cancer cell growth under nutrient stress in tumor microenvironment.