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目的:研究2个不同年龄组供体来源的人视网膜微血管内皮细胞的特性。方法:分离培养2个不同年龄组的人视网膜微血管内皮细胞,分别为出生后30 d内和40-65岁供体组。视网膜微血管内皮细胞具有特异性表达,通过免疫荧光染色CD31、vWF和乙酰化低密度脂蛋白(Dil-Ac-LDL)的摄取来定位内皮细胞的标记。通过使用matrigel基质胶培养细胞,激光共聚焦显微镜和Western blotting分析vWF在细胞内的分布,评估2组细胞管状结构形成能力以及vWF在细胞内的不同分布。结果:从2个不同年龄组中成功地获得高纯度视网膜微血管内皮细胞。2组在种到基质胶后6 h均能形成典型的二维管腔样结构,但婴幼儿组评分比成人组高(27.2%±2.2%),且二维管腔样结构在30 h仍可较好地维持,而成人组已看到部分自发降解。另外可见到vWF在婴幼儿组主要表达在细胞核内,而成人组主要表达在胞浆。结论:不同年龄来源的人视网膜血管内皮细胞具有某些自身特定的性状特征,在与年龄相关的疾病的体外研究中,供体年龄因素可能造成的影响应该得到一定的重视。
Objective: To study the characteristics of human retinal microvascular endothelial cells from two different age groups. Methods: Human retinal microvascular endothelial cells were isolated and cultured in two different age groups, respectively, within 30 days after birth and 40-65 years old donor group. Retinal microvascular endothelial cells were specifically expressed and labeled by immunofluorescent staining of CD31, vWF and uptake of acetylated low-density lipoprotein (Dil-Ac-LDL). The distribution of vWF in the cells was analyzed by confocal microscopy and Western blotting using matrigel matrices, and the ability of tubular cell formation and the distribution of vWF in the cells were evaluated. RESULTS: High purity retinal microvascular endothelial cells were successfully obtained from 2 different age groups. The typical two-dimensional lumen-like structures were formed in both groups 6 h after matrigel injection, but the score of infants and young children was 27.2% ± 2.2% higher than that of adults and the two-dimensional lumen-like structures were still at 30 h Can be better maintained while the adult group has seen partial spontaneous degradation. In addition, it can be seen that vWF is mainly expressed in the nucleus of the infants and young children, while the adult group is mainly expressed in the cytoplasm. CONCLUSION: Human retinal vascular endothelial cells of different ages have certain traits of their own traits. In the in vitro studies of age-related diseases, the possible influence of donor age should be given some attention.