榄香烯对人晶状体上皮细胞增生及细胞周期的影响

The effects of elemene on proliferation and cell cycle of human lens epithelial cells

目的探讨中药单体榄香烯(Ele)对人晶状体上皮细胞株(HLE-B3)增生及细胞周期的影响。方法利用重组人碱性成纤维细胞生长因子(rhbFGF)诱导HLE-B3增生,将80mg/L的Ele作用在处于增生状态下的HLE-B324h后,四甲基偶氮唑蓝法(MTT)检测Ele对HLE-B3增生的抑制作用;苏木精-伊红染色观察Ele作用后HLE-B3细胞形态的改变;流式细胞术(FCM)检测Ele对HLE-B3细胞周期的影响。结果MTT检测显示:rhbFGF组HLE-B3吸光度值(0.5990±0.0531)较正常组(0.4091±0.0422)显著升高,Ele组HLE-B3吸光度值(0.4500±0.0614)较rhbFGF组显著降低,抑制率达24.90%(P<0.01)。苏木精-伊红染色后光学显微镜下观察到rhbFGF组较正常组细胞数量增多,细胞形态清晰,胞浆丰富,胞核清晰,交织呈网状结构;Ele组细胞数量明显减少,胞浆减少,轮廓不清,交织的网状结构减少,甚至有的细胞变圆,细胞核凝集,见核固缩现象,胞浆嗜酸性染色。FCM检测细胞周期变化时发现:G1期的细胞在rhbFGF组(42.062%±1.270%)较正常组(46.422%±3.765%)减少(P<0.05),Ele组(60.665%±2.069%)较rhbFGF组明显增加(P<0.01);S期的细胞在rhbFGF组(51.647%±1.123%)较正常组(31.842%±2.798%)明显增加(P<0.01),Ele组(30.222%±3.429%)较rhbFGF组明显减少(P<0.01);G2期的细胞在rhbFGF组(6.288%±0.966%)较正常组(21.735%±3.806%)明显减少(P<0.01),Ele组(9.112%±1.659%)较rhbFGF组明显增加(P<0.01)。结论Ele能通过改变HLE-B3细胞形态及细胞周期的进程而有效抑制rhbFGF诱导的HLE-B3增生,有望成为防治后发性白内障的理想药物。

Objective To suppress the proliferation of lens epithelial cells （LECs） is a primary goal in prevention of after cataract. Recent study demonstrated an effective inhibition of elemene （Ele） on tumor cells. Present study was to investigate the effects of Ele on proliferation and cell cycle of human LECs B3 （ HLE-B3 ）. Methods Recombinant human basic fibroblast growth factor（rhbFGF） was utilized to induce proliferation of HLE-B3. Proliferative HLE-B3 was incubated with 80 mg/L Ele in CO2 incubator for 24 hours. Then the inhibitory effects of Ele on proliferation of HLE-B3 was evaluated by methyl thiazolyl tetrazolium（MTT）. The effect of Ele on HLE-B3 morphology was observed under the optical microscope. The effect of Ele on HLE-B3 cell cycle was analyzed by flow cytometer（FCM）. Results MTT test showed that the optical density（OD） value of rhbFGF group was remarkably higher than that of control group（0. 599 0 ±0. 053 1 versus 0. 409 1 ± 0. 042 2 ） （ P 〈 0.01 ） , and that of Ele group（ 0. 450 0 ±0. 061 4） was obviously declined in comparison to rhbFGF group （P 〈 0. 01 ）. The inhibitory rate of Ele was 24.90 % . In proliferation group, the number of HLE-B3 was increased with the normal cell structure and abundant cytoplasm under the optical microscope. However, in Ele group, the number of HLE-B3 was evidently decreased with less cytoplasm,undistinguished cell structure, condensed and aggregated nucleuses. The result of flow cytometer showed that the percentage of HLE-B3 in G, phase in rhbFGF group was 42. 062% ±1. 270% and that in control group was 46. 422% ±3. 765% with a significant difference（P 〈 0. 05 ）. HLE-B3 in G~ phase in Elc group（60. 665% ± 2. 069% ） was evidently increased in comparison with rhbFGF group（ P 〈 0. 01 ）. HLE-B3 in S phase in rhbFGF group compared with control group was increased （51. 647% ±1.123% versus 31. 842% ±2.798%）（P〈0.01）,but that in S phase in Ele group（30.222% ±3.429%） was lower than rhbFGF group （P 〈 0.01 ）. HLE-B3 in Gz phase in rhbFGF group was decreased in comparison with control group （6.288% ±0.966% versus 21.735% ± 3. 806% ,P 〈 0.01 ）, and that in G2 phase in Ele group （9. 112% ± 1.659% ） compared with proliferation group was increased （ P 〈 0.01 ）. Conclusion Ele could alter the cell cycle of HLE-B3 and effectively inhibit the HLE-B3 proliferation induced by rhbFGF. Ele may be a reliable and effective drug for prevention and treatment of after cataract.