Subretinal Pigment Epithelial Deposition of Drusen Components Including Hydroxyapatite in a Primary Cell Culture Model.
Subretinal Pigment Epithelial Deposition of Drusen Components Including Hydroxyapatite in a Primary Cell Culture Model.
复制标题
DOI：
10.1167/iovs.16-21060
复制
发表时间：
2017-02-01
影响因子：
4.4
通讯作者：
Curcio CA
中科院分区：
文献类型：
--
作者：
Pilgrim MG;Lengyel I;Lanzirotti A;Newville M;Fearn S;Emri E;Knowles JC;Messinger JD;Read RW;Guidry C;Curcio CA
在RPE的基底层和布鲁赫膜的内部胶原层之间形成的含有羟基磷灰石、脂质、蛋白质和微量金属的细胞外沉积物是早期AMD的标志。我们研究了培养的RPE细胞是否可以产生含有所有这些分子成分的细胞外沉积物。从新鲜摘除的猪眼分离的视网膜色素上皮细胞在Transwell膜上培养长达6个月。存款的组成和结构进行了表征，使用光，荧光和电子显微镜;同步加速器X射线衍射和X射线荧光;二次离子质谱;和免疫组织化学。当在具有0.4 μ m直径孔的10 μ m厚插入物上培养时，明显具有功能的原代RPE细胞可以在12周内产生含有羟基磷灰石、脂质、蛋白质和微量元素的亚RPE沉积物，而无需外部节补充。这些数据表明，亚RPE存款的形成是由RPE，以及与年龄和早期AMD相关的布鲁赫膜和脉络膜毛细血管复合体的渗透性的丧失引发的，并可能受到其调节。这种早期AMD病变的细胞培养模型提供了用于测试针对RPE下存款形成的新治疗干预的新系统，所述RPE下存款形成是在视力丧失发作之前发生的事件。
Extracellular deposits containing hydroxyapatite, lipids, proteins, and trace metals that form between the basal lamina of the RPE and the inner collagenous layer of Bruch's membrane are hallmarks of early AMD. We examined whether cultured RPE cells could produce extracellular deposits containing all of these molecular components. Retinal pigment epithelium cells isolated from freshly enucleated porcine eyes were cultured on Transwell membranes for up to 6 months. Deposit composition and structure were characterized using light, fluorescence, and electron microscopy; synchrotron x-ray diffraction and x-ray fluorescence; secondary ion mass spectroscopy; and immunohistochemistry. Apparently functional primary RPE cells, when cultured on 10-μm-thick inserts with 0.4-μm-diameter pores, can produce sub-RPE deposits that contain hydroxyapatite, lipids, proteins, and trace elements, without outer segment supplementation, by 12 weeks. The data suggest that sub-RPE deposit formation is initiated, and probably regulated, by the RPE, as well as the loss of permeability of the Bruch's membrane and choriocapillaris complex associated with age and early AMD. This cell culture model of early AMD lesions provides a novel system for testing new therapeutic interventions against sub-RPE deposit formation, an event occurring well in advance of the onset of vision loss.