LTD4及MK571对人结肠癌细胞SW480、Caco-2凋亡和增殖作用的体外研究

Growth inhibitory effects and apoptosis of LTD4 and MK571 on human colon cancer cell lines SW480 and Caco-2 in vitro

目的 观察半胱氨酰白三烯D4（LTD4）及半胱氨酰白三烯受体1拮抗剂MK571对人结肠癌细胞株SW480、Caco-2增殖和凋亡的影响.方法 MTT法检测细胞生长活性;FITC Annexin V/碘化丙啶（PI）双染流式细胞术检测细胞的凋亡率;PI染色细胞,用流式细胞仪检测细胞周期.结果 12.5 ～ 400 μg/L的LTD4对结肠癌SW480细胞有促增殖作用,且存在时效和量效依赖性;而1.6～12.5μg/L的LTD4处理24、48 h,对SW480细胞的增殖抑制作用无明显影响,作用时间延长至72 h,可促进SW480细胞的增殖.50～ 400 μg/L的LTD4对结肠癌Caco-2细胞有促增殖作用,且存在时效和量效依赖性;而1.6-50 μg/L的LTD4对结肠癌Caco-2细胞的影响与对照组相比差异无统计学意义.6.25～200 μmol/L的MK571对结肠癌SW480细胞有抑制作用,且存在时效和量效依赖性;而0.8 ～ 6.25 μmol/L的MK571对SW480细胞的影响与对照组相比无明显差异.25～ 200 μmol/L的MK571对结肠癌Caco-2细胞有抑制作用,且存在时效和量效依赖性;而0.8～ 25 μmol/L的MK571对结肠癌Caco-2细胞的影响与对照组相比差异无统计学意义.与阴性对照组相比,25 ～ 100 μg/L的LTD4对2种结肠癌SW480细胞和Caco-2细胞有促凋亡作用,且存在时效和量效依赖性;而100、200 μmol/L的MK571对2种结肠癌的凋亡无明显作用.与阴性对照组相比,25 ～ 100 μg/L的LTD4可以降低2种结肠癌细胞株G0/G1期比例,增加G2/M及S期比例;而100、200 μmol/L的MK571增加2种结肠癌细胞G0/G1期的比例,降低G2/M及S期比例.结论 LTD4具有促进人结肠癌SW480、Caco-2细胞株增殖的作用,该作用可能的作用机制是抑制凋亡、增加G2/M及S期细胞比例.MK571可抑制2种结肠癌细胞株的增殖,但对凋亡无明显影响,其抑制增殖作用的可能机制是阻滞细胞于G0/G1期.

Objective To observe the growth inhibitory effects and apoptosis of cysteinyl leukotriene D4 （CysLTD4） and cysteinyl leukotriene receptor 1 （CysLT1 R） antagonists MK571 on human colon cancer SW480 and Ca- co -2 cell lines. Methods The MTF colorimetry was used to detect the cell growth inhibitory rates of LTD4 and MK571 on human colon cancer SW480 and Caco -2 cells. The apoptotic cells were assessed by flow cytometry with FITC Annex- in V/propidium iodide （PI） label. The cell cycle was examined with PI staining by flow cytometry. Results In the range of concentration 12.5 -400 μg/L LTD4 could induce proliferative response on the colon cancer SW480 cells in a dose - and time - dependent manner. After exposed to 1.6 - 12.5 μg/L LTD4 for 24 or 48 h, the proliferation index had no significant difference on SW480 ceils, while prolonged treatment time to 72 h, it could promote the proliferation of SW480 cells. In the range of concentration 50 - 400 μg/L LTD4 could promote proliferative response on Caco - 2 cells ina dose - and time - dependent manner. While compared with control group, 1.6 - 50 txg/L LTIM had no significant difference on the proliferation of Caco -2 cells. In the range of concentration 6.25 -200 Ixmol/L MK571 could inhibit proliferation response on SW480 cells in a dose - and time - dependent manner. While compared with control group, 0.8 - 6.25 txmol/L MK571 had no significant difference on the proliferation of SW480 cells. In the range of concentration 25 -200 txmol/L MK571 could inhibit proliferation response on Caco- 2 cells in a dose- and time- dependent manner. While compared with control group, 0.8 -25 p^mol/L MK571 had no significant difference on the proliferation of Caco - 2 cells. Compared with control group, 25 -100 p^g/L LTD4 could reduce the apoptosis rate of the two colon cancer cell lines in a dose - dependent manner. However, 100 i^mol/L or 200 txmol/L MK571 had no obvious effect on apoptotic rate in both colon cancer cell lines. Treatment of cells with LTD4 in the concentration of 25 -100 Ixg/L, the percentage in G0/G1 - phase was reduced and G2/M S - phase was increased in both colon cancer cell lines. At the concentration of 100 p^mol/L or 200 Ixmol/L MK571 could improve G0/G~ phase and reduce GJM and S phase. Conclusion LTD4 promotes the proliferation of SW480 and Caco -2 cells by activating its receptor. It can be a role by improving cells in G2/M phase, and inhibiting tumor cell apoptosis. MK571 inhibits the growth of SW480 and Caco -2 cells, but does not affect on apoptosis. The inhibition of proliferation by MK571 may achieve by blocking cells in C0/G1 phase.