Inhibition of proliferative vitreo retinopathy by controling the related transcription factors.

目的。既往研究表明，视网膜色素上皮细胞的增殖和迁移与增生性玻璃体视网膜病变(PVR)的发病密切相关。最近有报道称，诱骗分子，即双链硫代寡核苷酸，具有与转录因子相同的结合位点序列。本研究检测了E2F诱骗和NFkappaB诱骗对培养的人视网膜色素上皮(RPE)细胞和活体动物模型PVR增殖的影响。用Hangai等人1996年的脂质体中的磷脂酰丝氨酸取代DC胆固醇制备了阳离子脂质体。结果。结果表明，E2F诱骗分子与人成纤维细胞核提取液高度结合，NFkappaB诱骗分子也与IL-1β刺激的人RPE细胞核提取液结合。RT-PCR结果显示，人成纤维细胞中引入E2F诱骗基因后，调控细胞周期的重要因子表达减少，提示NFkappaB诱骗基因可降低IL-1β刺激的RPE细胞中IL-1β的转录水平。BrdU标记指数和DNA合成(H-胸腺嘧啶核苷摄取量)表明，E2F诱骗对细胞增殖有高度抑制作用。在含有NFkappaB诱饵的培养皿中，细胞的迁移明显受到抑制。体内实验：将经HVJ阳离子脂质体导入人成纤维细胞的NFkappaB诱骗基因注入兔PVR模型玻璃体腔内。兔PVR的表现按Blumenkraz等人的分类进行评分。NFkappaB诱饵与加扰诱饵的PVR发生率差异无统计学意义。结论。这些结果提示E2F诱骗和NFkappaB诱骗在体外具有治疗增殖性玻璃体视网膜病变的潜力，体外应用还需要更多的研究。

Purpose. Previous studies suggested that proliferation and migration of retinal pigment epithelial cells are deeply involved in pathogenesis of proliferative vitreo retinopathy (PVR). Recently it is reported that decoys, double-stranded phosphorothioate oligonucleotides, have the same sequence of binding site of transcription factors. In this study we tested the effect of E2F decoys and NFkappaB decoys in the proliferation of cultured human retinal pigment epithelial (RPE) cells and in vivo animal model of PVR.Methods. HVJ cationic liposomes were prepared by replacing phosphatidylserine of HVJ liposomes (Hangai, et al, 1996) to DC cholesterol. Results. It was shown that E2F decoys highly combined to nuclear extracts from human fibroblast cells and NFkappaB decoys also combined to nuclear extracts from human RPE cells stimulated by interleukin 1beta (IL-1beta). RT-PCR indicated that E2F decoys introduced into human fibroblast decreased expression of important factor that is regulating cell cycle, it showed that NFkappaB decoys introduced into human RPE cells reduced transcripts of IL-1beta in IL-1beta stimulated RPE cells. BrdU labeling index and DNA synthesis (H-thymidine uptake) clarified that cell proliferation was highly inhibited with E2F decoys. Cells on culture dishes with NFkappaB decoys were significantly inhibited cell migrations compared with scrambled decoys. In vivo experiment NFkappaB decoys, transferred by HVJ cationic liposomes into human cultured fibroblast, were injected in the vitreous cavity of rabbit PVR model. The appearances of rabbit PVR were scored by a classification of Blumenkraz et al. The incidences of PVR were not significantly different between with NFkappaB decoys and with scrambled decoys. Conclusions. These results suggest that E2F decoys and NFkappaB decoys have the potential to treat proliferative vitreoretinopathy in vitro and it is needed more study to vitro use.