Regeneration of pigment cells and basic and clinical study for treatment of age-related macular degeneration

老年性黄斑变性是老年人最严重的疾病之一。即使可以手术切除黄斑下和脉络膜新生血管膜，手术区域视网膜色素上皮的严重损伤或脱落是不可避免的。色素营养不良也是一种具有严重视力障碍的破坏性遗传性眼病。到目前为止，我们对这两种疾病都没有有效的治疗方法。我们进行了视网膜色素上皮（RPE）培养的基础实验，并将其移植到动物视网膜下间隙，特别是皇家外科学院（RCS）大鼠作为遗传性视网膜变性模型，观察其对光感受器细胞死亡的拯救作用。1)用视网膜营养不良的RCS大鼠，分别移植Long Evans大鼠、人和牛的RPE，观察其光感受器细胞死亡的延缓。2)从眼库眼或胎眼中获得RPE细胞并进行培养。它们作为新鲜的移栽，或用原代或多代培养。我们还尝试将这些细胞冷冻保存3个月。至少在几个基因表达的观点上，它们的特征被很好地保留了下来。在未来，如果我们能够将这些RPE细胞保存在深度冷冻的状态下，我们可以在适当的时间和数量为患者提供临床使用，就像“RPE银行”一样。3)在实验动物中，它们没有表现出任何免疫反应。而将人RPE细胞与胶原片移植到兔前房后，不仅在手术眼，而且在对侧未手术眼均表现出视网膜电图抑制和视网膜下间隙巨噬细胞浸润的明显反应。提示异体RPE移植的临床应用应谨慎。有趣的是，在RCS大鼠的光感受器细胞退化过程中，MHC II类细胞的表达被观察到，但如果通过移植人培养的RPE来挽救这些动物，则MHC II类细胞的表达被抑制。4)如果我们考虑到该技术的临床应用，自然自体移植比异种或同种异体移植要好得多。虹膜色素上皮（IPE）是一种神经源性的色素细胞，且虹膜周围切除术容易获得大量的虹膜色素上皮，因此我们尝试使用IPE进行移植。培养大鼠视网膜内皮细胞瘤（IPE），用大鼠bFGF cDNA转染载体（pCNX2），移植到RCS大鼠视网膜下间隙。这些转染后的细胞表达了较强的bFGF mRNA。光感受器保存完好，无免疫反应。如果我们可以使用IPE，无论是野生的还是经过分子技术修饰的，它们都适用于患者，更安全，更容易。少

Age-related macular degeneration is one of the most serious diseases in the elderly people. Even if submacular and choroidal neovascular membrane could be surgically excised, severe damage or evacuation of retinal pigment epithelium are innevitable in the operated area. Pigmentary dystrophy is also a devastating hereditary eye disease with severe visual disturbance. Up to now, we have no effective treatments for both of them. We have conducted basic experiments of retinal pigment epithelium (RPE) culture, their transplantation to subretinal space of animals, especially, the royal college of surgeon's (RCS) rat, a model of hereditary retinal degeneration and observed their rescue effects for photoreceptor cell death.1) With the retinal dystrophic RCS rat, we transplanted RPE from Long Evans rat, human, and bovine and could observe the retardation of the photoreceptor cell death.2) RPE cells from eye bank eye or fetus eye were easily obtained and cultured. They were transplanted as fresh … More or cultured with primary or multiple passage. We also tried cryopreservation of these cells up to 3 months. Their characteristics were well preserved at least in the viewpoints of several gene expression. In the future, if we could keep these RPE cells in deep-freezed condition, we could use them clinically in appropriate time and number of cells for patients just as "RPE bank".3) In experimental animals, they did not show any immunological reaction. But transplantation of human RPE cells with collagen sheet into the anterior chamber in rabbit showed difinite reaction like suppression of electroretinogram and macrophage infiltration in the subretinal space, not only in the operated but contralateral non-operated eye. These results suggest we must be cautious in clinical application of heterogenous RPE transplantation. It was an interesting observation that the expression of MHC class II cells were observed in the course of photoreceptor cell degeneration in the RCS rat but supressed if they were rescued by the transplantation of human cultured RPE in these animals.4) If we cpnsider clinical application of this technique, it is natural the autograft is much better than the xeno-or allograft. We tried to use the iris pigment epithelium (IPE) for transplantation because they were neural origin, pigmented cells and easily and enough amount of them were obtained by peripheral iridectomy. IPE of rat were cultured the vector (pCNX2) with cDNA of rat bFGF were transfected and transplanted into the subretinal space of RCS rat. These transfected cells expressed strong mRNA of bFGF.The photoreceptors were well preserved and no immunological reaction. If we could use IPE,either wild or modified with molecular technique, they were applicable for patients, much safer and easier. Less