前列腺毛细血管内皮细胞屏障功能研究  

Barrier function of rat prostate capillary endothelial cells in vitro

作　　者：崔栋[1] 尚永刚[1] 韩广玮[2] 刘城城 李龙坤[1] 易善红[1] 

机构地区：[1]第三军医大学新桥医院泌尿外科,重庆400037 [2]解放军第169医院泌尿外科,湖南衡阳421002 [3]重庆市第四人民医院泌尿外科,重庆400014

出　　处：《第三军医大学学报》2016年第4期385-389,共5页Journal of Third Military Medical University

基　　金：第三军医大学临床科研基金(2014D325)~~

摘　　要：目的 分离培养大鼠前列腺毛细血管内皮细胞,体外研究大鼠前列腺毛细血管内皮细胞的屏障功能。方法 采用细胞生长特性选择法分离培养大鼠前列腺毛细血管内皮细胞,用第Ⅷ因子相关抗原抗体对分离培养的前列腺毛细血管内皮细胞进行免疫学鉴定。免疫组化观察毛细血管内皮细胞之间紧密连接蛋白claudin-1的表达,采用跨膜电阻测量仪检测毛细血管内皮细胞的跨膜电阻,用酚红渗漏实验检测其通透性,以大鼠肺毛细血管内皮细胞、脑毛细血管内皮细胞作为阴性与阳性对照。结果 成功分离大鼠前列腺毛细血管内皮细胞,体外培养可以形成紧密的单层细胞结构,呈铺路石样生长。第Ⅷ因子相关抗原抗体的免疫荧光结果显示：分离培养的大鼠前列腺毛细血管内皮细胞胞浆表达绿色荧光,提示分离的细胞为毛细血管内皮细胞;免疫组化结果显示紧密连接蛋白claudin-1在相邻的毛细血管内皮细胞之间表达;体外培养的大鼠前列腺毛细血管内皮细胞,阴性对照组肺毛细血管内皮细胞,阳性对照组脑毛细血管内皮细胞的跨膜电阻峰值分别可以达到（142.2±3.1）、（43.3±3.5）、（248.2±6.2）Ω/cm2,并稳定保持2-3 d,三者比较差异有统计学意义（P〈0.05）。酚红渗漏实验结果显示前列腺毛细血管内皮细胞、阳性对照组脑毛细血管内皮细胞随着细胞单层跨膜电阻的增加,基底侧的酚红浓度减低,阴性对照组肺毛细血管内皮细胞酚红渗透量随跨膜电阻的改变变化趋势不明显。结论 大鼠前列腺毛细血管内皮细胞与脑毛细血管内皮细胞等屏障内皮相比拥有相似的结构功能,体外培养的大鼠前列腺毛细血管内皮细胞具有屏障特性,可以作为血前列腺屏障研究的体外模型。Objective To culture rat prostate capillary endothelial cells in vitro, and study their barrier function. Methods The rat prostate capillary endothelial cells were isolated and cultured with cell growth feature selection method. Antibody against Ⅷ factor related antigen was used for immunological identification of prostate capillary endothelial cells. Immunohistochemistry was adopted to observe the expression of tight junction protein claudin-1. Millicell epithelial resistance measuring instrument was employed to measure the transepithelial electrical resistance of the capillary endothelial cells, and phenol red leakage test was used to detect the permeability of prostate capillary endothelial cells. The rat pulmonary capillary endothelial cells and brain capillary endothelial cells were taken as negative control and positive control respectively. Results The rat prostate capillary endothelial cells were successfully isolated and cultured to form compact monolayer cell structure in vitro. The immunofluorescence detection of factor Ⅷ showed that the cultured cells were factor Ⅷ-positive and green fluorescence was expressed. Immunohistochemical results showed that the tight junction protein claudin-1 was expressed between adjacent endothelial cells. The transepithelial electrical resistance peaks of in vitro cultured rat prostate capillary endothelial cells, negative control pulmonary capillary endothelial cells and positive control brain capillary endothelial cells reached 142. 20 ± 3.10, 43.3 ± 3.5 and 248.20± 6.0 Ω/cm：, respectively, and stably kept for 2 - 3 d, and there were statistically significant differences between the 3 groups of cells （ P 〈 0.05）. Phenol red leakage test showed that with the increase of the transepithelial electrical resistance, the concentration of phenol red in basal lateral region of prostate capillary endothelial cells and positive control brain capillary endothelial cells was reduced. However, in negative control pulmonary capillary endothelial cells, the change t

关 键 词：前列腺 毛细血管内皮细胞 血前列腺屏障 跨膜电阻 

分 类 号：R322.13[医药卫生—人体解剖和组织胚胎学] R322.64[医药卫生—基础医学]