去整合素对人眼晶状体上皮细胞的粘附移行作用

Effects of disintegrin on adhesion and migration of human lens epithelial cells

目的研究去整合素对体外培养的人眼晶状体上皮细胞生长的抑制作用。方法对先天性白内障患者术中环行撕前囊膜,进行原代培养并传代,取第2代细胞用于实验。抗粘附实验:将悬浮的细胞与不同浓度的去整合素(5μg·L-1、10μg·L-1、20μg·L-1、40μg·L-1、80μg·L-1)在37℃、50mL·L-1CO2条件下一起孵育30min。然后接种于涂有鼠尾胶原的96孔培养板。8h后,用HBSS轻洗各孔2次,MTT法测定粘附细胞的吸光度。抗移行实验:对融合的细胞进行划线,80μg·L-1的去整合素进行干预,24h、48h、72h后,细胞计数法观察裸露区内细胞的移行数量。结果不同浓度的去整合素(20μg·L-1、40μg·L-1、80μg·L-1)对晶状体上皮细胞的粘附均具有抑制作用且呈剂量依赖性。与对照组相比,24h、48h、72h内80μg·L-1的去整合素可显著抑制晶状体上皮细胞的移行。结论去整合素抑制体外培养的人眼晶状体上皮细胞的粘附及移行。

Objective To study the inhibitory effect of disintegrin on adhesion and migration in cultured human lens epithelial cells （LECs） in vitro. Methods In the congenital cataract surgery, the anterior capsule membrane sampled by CCC technique was cultured and subcultured and the second generations of LECs were used in this experiment. Anti-adhesion experiment： The LECs suspended in 1640 medium were pre-incubated for 30 minutes in the presence of various concentrations of disintegin（5μg·L^-1, 10μg·L^-1, 20 μg·L^-1, 40μg·L^-1 and 80μg·L^-1 ） at 37℃, 50 mL·L^-1 CO2, then inoculated in 96-well plate coated with rat-tail collagen. After incubation for 8 hours,adherent cells were washed twice with HBSS, then the absorbance was measured at 570 nm to detect the numbers of cells adhering using MTT.Anti-migration experiment：A denuded area was made when the second passage of LECs was confluent into a monolayer. Cells were then exposed to media with（without） 80μg·L^-1 disintegrin at 37℃, 50 mL·L^-1 CO2. After 24 hours, 48 hours and 72 hours, the cells having migrated into the denuded area were enumerated under light microscope. Results Disintegrin （20μg·L^-1, 40μg·L^-1and 80μg·L^-1） inhibited LECs from attaching to collagen in a dose-dependent manner. The migration of LECs was blocked efficiently with 80μg·L^-1 of disintegrin in the 24th hour, 48th hour and 72nd hour. Conclusion Disintegin can inhibit the adhesion and migration of LECs in vitro.