豚鼠早期实验性近视眼视网膜色素上皮细胞前部及后极部TGF—β2表达变化的研究

Altered TGF-β2 expression in retinal pigment epithelial（RPE）cells from an experimentally-induced myopia guinea pig model

目的研究豚鼠早期实验性近视眼视网膜色素上皮（RPE）细胞前部及后极部转化生长因子β2（TGF-β2）的改变，探索近视的发病机制。方法2周龄豚鼠30只随机分为A、B、C组，每组10只，再随机选取5只（10眼）正常2周龄豚鼠不作任何干预，作为正常对照眼。选取任意眼戴一10DS凹透镜，分别饲养6天、15天、30天后除去镜片，验光及测眼轴长度确定近视形成后，采用细胞酶消化法培养豚鼠的前部及后极部RPE细胞，取3～6代RPE细胞进行免疫细胞化学、实时荧光定量PCR、western—blot蛋白印迹法检查前部及后极部RPE细胞TGF-β2及其mRNA、蛋白表达变化。结果TGF-β2的表达定位于细胞浆和细胞核。实验组中前部RPE细胞于15天阳性表达率较高，30天阳性表达率最高；后极部RPE细胞于6天表达率开始较高，至15天阳性表达率最高，以后保持较高的阳性表达率，差异有显著性（P〈0．05）；SC组各组自身前部及后极部比较，差异无显著意义（P〉0．05）。结论体外培养的豚鼠RPE细胞稳定表达TGF-β2、TGF—β2 mRNA及其蛋白。且前部RPE细胞于诱导15天开始阳性表达率较高；后极部RPE细胞于诱导6天开始阳性表达率较高。后极部RPE细胞较前部RPE细胞表达差异有显著性。

Objective To observe the expression of transforming growth factor-β2 （TGF-β2）in cultured guinea pig anterior and posterior retinal pigment epithelial （RPE）cells in lens-induced myopia （LIM）. Methods Three groups （n = 10）of two-week-old guinea pigs were used to develop concave lensinduced myopia（LIM）in one eye via the out-of-focus method for 6,15 or 30 d, respectively, while the other eye in each guinea pig served as the self-control（SC）. After myopia induction,lenses were removed,and RPE cells were cultured and passaged twice. TGF-β2 expression levels of retinal pigment epithelial（RPE）cells in LIM and SC groups were compared by immunocytochemistry, quantitative real-time PCR（qRT-PCR）and Western blot analyses. All datas were statistically analyzed by SPSS 13. 0 system. Results The TGF-132 expression of the anterior portion of the RPE cells in the LIM group was significantly higher at 15 d and the highest at 30 d after myopia induction compared with the SC group（P〈0.05）. The TGF-β2 staining of the posterior RPE cells in the LIM group began to rise significantly at 6 d, peaked at 15 d and remained significantly higher than that of the anterior part, as well as the SC group, even at 30 d after myopia induction （P 〈 0.05）. Conclusion During myopia development in lens-induced guinea pigs,the increase in TGF-β2 activity of RPE eells initiated at the posterior pole. Along with the induction time,the TGF-β2 activity in all RPE cells became elevated.