In Vitro Study on Lethal Effect of Human Choroidal Melanoma OCM-1 Cell Line by Repeating -70℃ Freeze Thawing

Objective: To investigate the effects of repeating-70℃freeze thawing on human choroidal melanoma cell line OCM-1. Methods: OCM-1 cells were frozen by repeating -70℃ freeze thawing with various durations and frequencies. Then the inhibit rate of cells was examined by MTT essay. The cell viability was measured by monoclonal formation assay. We also used the HE staining, immunohistochemistry staining and the laser-scanning confocal microscopy (LSCM) to investigate the morphological changes of the cells. Results: The growth of OCM-1 cells was inhibited by repeating -70℃ freeze thawing in time-dependent and frequency-dependent manners (P < 0.01). Different morphous including necrosis and apoptosis of the cells could be observed after -70℃ freeze thawing by the LSCM. Conclusion: Repeating -70℃ freeze thawing can not only kill cells directly and induce considerable cells to apoptosis, but also inhibit the growth of the survivals. The kill and wound ratio of the cells disposal with different times and frequencies present variance. And the distinction when treated with different frequencies during the same time is much more significant than different times with the same frequency, which guide clinical workers to choose repeating cryotherapy with short term method instead of single cryotherapy with long term in choroidal melanoma treatment.

脉络膜黑色素瘤中整合素α_vβ_3表达的研究

目的:整合素α_vβ_3为整合素家族成员之一,是一类跨膜粘附分子,其在多种肿瘤细胞及新生内皮血管细胞中高表达而在成熟血管内皮、上皮细胞及正常细胞中表达较低或无表达。基于这些特征,整合素α_vβ_3近年来成为分子影像研究的热点之一。鉴于整合素α_vβ_3在人眼脉络膜黑色素瘤中是否存在表达尚不可知,本课题拟研究整合素α_vβ_3在人脉络膜黑色素瘤中表达情况,为以整合素α_vβ_3为基础的分子显像提供理论基础。方法:培养人源脉络膜黑色素瘤细胞株OCM-1,使用蛋白免疫印迹方法检测整合素α_vβ_3在OCM-1中表达水平,并使用免疫组化方法检测人脉络膜黑色素瘤病理切片中整合素α_vβ_3表达分布。结果:蛋白免疫印迹实验表明在OCM-1细胞株中存在整合素α_vβ_3表达,其表达水平介于已知高表达整合素α_vβ_3的头颈鳞癌细胞株HEP-2和较低表达整合素α_vβ_3的头颈鳞癌细胞株CNE-1之间,免疫组化方法检测人脉络膜黑色素瘤病理切片中也存在整合素α_vβ_3表达。结论:人脉络膜黑色素瘤中具有整合素α_vβ_3表达,可以为后期研究基于整合素α_vβ_3的分子显像技术提供依据。

角膜内皮细胞与脉络膜色素瘤细胞培养后细胞生物行为研究

目的观察角膜内皮细胞（CECs）与脉络膜黑色素瘤细胞（OCM-1）共培养后，细胞形态、中心体表达及细胞骨架形态、结构情况。方法对CECs与OCM-1Transwell体系共培养后，通过倒置相差显微镜、HE染色观察CECs生长及形态变化；通过抗中心体γ-tubulin蛋白抗体和抗α-tubulin蛋白抗体免疫荧光染色，观察内皮细胞中心体形态和数量，细胞骨架形态和结构。结果实验组（CECs与OCM-1共培养）CECs形态基本规则，较空白对照组（CECs单独培养）和阳性对照组（CECs与基质细胞共培养）细胞密度高；免疫荧光染色显示，贴近细胞核边缘有清晰的一到两个中心体，细胞显示结构形态规则的细胞骨架。结论CECs与OCM-1共培养后细胞形态、中心体、细胞骨架形态和结构未见明显异常。