胚胎发育中神经视网膜对血视网膜外屏障结构形成的作用

Effects of neural retina on development of the structure of outer blood-retinal barrier in embryogenesis

目的 研究胚胎发育中神经视网膜对血视网膜外屏障结构形成的作用。 方法 将孵化 7、10、14 d的鸡胚眼各 15 0、12 0和 90只 ,分批分别剥离视网膜神经上皮层 (RNE)和色素上皮层 (RPE)。 RNE用来制备不同条件的培养液 (7drc SF3、 10 drc SF3和 14 drc SF3) ,将 7d和 14 d的 RPE细胞分别培养到12 mm Transwell滤膜上 ,其顶房分别用 SF3或上述条件培养液 ,底房均用 SF2培养液。形成单层 RPE细胞后 ,测定此上皮层的电阻 (TER)。最后固定细胞 ,用细胞免疫组织化学方法和透射电子显微镜检测 RPE细胞间紧密连接形成的情况。 结果 不同培养液 (SF3/ SF2 ,7drc SF3/ SF2 ,10 drc SF3/ SF2 ,14 drc SF3/ SF2 )培养 7d和 14 d鸡胚眼的 RPE细胞 TER之间的差异有非常显著性的意义 (P<0 .0 1)。电子显微镜结果显示神经视网膜条件培养液可以使 RPE细胞产生细胞极性 ,有效地促使紧密连接条带形成连续的网状结构。 结论 神经视网膜对血视网膜外屏障结构的形成起积极的促进作用。

Objective To investigate effects of neural retina on development of the structure of outer blood retinal barrier in embryogenesis. Methods The retinal neural epithelium (RNE) and pigment epithelium (RPE) layers of 150, 120 and 90 embryonic chicken eyes incubated for 7, 10, and 14 days were peeled off. RNE was used to prepare the culture medium with different conditions (7drcSF3, 10drcSF3, 14drcSF3). RPE cells of 7- and 14-incubated chicken embryos were cultured on laminin-coated transwell filter. The SF3, 7drcSF3, 10drcSF3, 14drcSF3 medium were used respectively in the apical chamber and SF2 was used in basolateral chamber. After the formation of monolayer, the transepithelial electrical resistance of the RPE was detected. After the fixation of RPE cells, the condition of the tight junction among the cells was observed by immunohistochemistry and transmission electron microscopy. Results For the RPE cells of 7- and 14-day incubated embryonic eyes, the difference of TER in various medium of SF3/SF2, 7drcSF3/SF2, 10drcSF3/SF2, 14drcSF3/SF2 was statistically significant (P<0.01). The polarity of RPE cells was induced and the netlike tight junctional strands was urged in the retina-conditioned medium. Conclusion The neural retina may actively promote the formation of the structure of outer blood retinal barrier.