Objectives To investigate and compare the growth of human fetal retinal pigment epithelial (RPE) cells seeded onto electrospun polyamide nanofibers (EPN) or etched pore polyester (EPP), and, further, to explore their possible use as prosthetic Bruch's membrane. Methods: Human fetal RPE cells were planted onto the EPN, EPP and plastic (control) substrates in Transwells. The cultures were assessed with respect to cell attachment at 2, 4, 8 hours and proliferation at 1, 4, 8 days after seeding. Growth and morphology of the cells were monitored under the phase contrast microscope, and the phenotype was identified by immunofluorescence staining with antibodies against tight junction protein ZO-1. Strips of single EPP coated with nothing or EPP coated with EPN was differently implanted into the subretinal space of two P21 RCS rats for two weeks and the histologic slides of the retina were assessed. Results: Cultured human fetal RPE cells were attached to either EPN or EPP substrates (with seeding on plastic substrate as control). After 8 h, the numbers of adherent cells in the EPN, EPP and control groups were 1.23v105/cm2, 1.70×105/cm2, and 1.64×105/cm2, respectively. The number of RPE cells attached to EPN was obviously less than that to both EPP and control (P<0.05). On the first day, the proliferation of cells on EPN was less than that of EPP and control (P<0.05); but by the 8th day in culture, the proliferation of cells on EPN had increased and was higher than proliferation on both EPP and control (P<0.05). All of the RPE cells cultured on EPN and EPP substrates were in monolayer, and the EPN-attached cells resembled the inner collagenous layer of Bruch's membrane. Immunofluorescence staining showed that the RPE cells cultured on EPN and EPP substrates adopted a higher expression of ZO-1 than that on the plastic control substrate. Subretinal implantation of either EPP alone or EPP as a carrier for free EPN for 2 weeks in P21RCS rats resulted in an expected encapsulation and loss of photoreceptor layer. No toxicity or other adverse reaction was observed in the vicinity of the transplant.rnConclusion: EPN and EPP could maintain human fetal RPE cell attachment and proliferation. Both EPN and EPP appeared to be grossly tolerance and biocompatible with subretinal implantation. EPN represents an intriguing prospect for prosthetic Bruch's membrane replacement because of its similarity in structure to native Bruch's membrane.%目的:比较观察人胚胎视网膜色素上皮(RPE)细胞在微孔聚酰胺纳米纤维膜(EPN)和蚀刻多孔聚酯膜(EPP)上的生长情况,为理想Bruch's膜替代物的研究奠定实验基础.方法:人胚胎RPE细胞培养在底分别为EPN(EPN组)、EPP(EPP组)的Transwell小室以及常规Transwell小室(对照组).在第2.4,8h和第1,4,8天分别评估3组细胞的贴壁和增殖情况.对比观察细胞生长形态及两种替代物的表面结构；免疫荧光染色观察RPE细胞ZO-1的表达.通过组织病理切片观察EPN和EPP植入21天龄(P21) RCS大鼠视网膜下间隙2周后的生物学反应.结果:接种8h(密度为4×105/cm2)后,EPN组,EPP组和对照组RPE细胞贴壁生长数量分别为1.23×105/cm2,1.70×105/cm2和1.64x105/cm2,其中EPN组细胞数量明显少于EPP组及对照组(P＜0.05)；以2.5×105/cm2的密度接种后第1天,EPN组细胞密度明显低于EPP组和对照组组(P＜0.05),但在接种后第8天,EPN组细胞数量显著多于EPP组和对照组(P＜0.05).EPN和EPP均支持RPE单细胞层形成,前者纤维排列类似Bruch's膜内胶原层；与对照组比,EPN和EPP组RPE细胞均呈现较强的ZO-1表达,较好地保持了原代RPE细胞表型；EPP和EPN植入视网膜下间隙后出现纤维包裹和光感受器层破坏、丢失,但在移植物附近未见明显毒性或排斥等其他异常反应.结论:EPN和EPP均支持人胚胎RPE细胞生长,二者植入P21 RCS大鼠视网膜下间隙后存在大体生物相容性.EPN的纤维排列与Bruch's膜内胶原层相近,可能是一种具有广阔发展前途的Bruch's膜替代物.
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