New Methods of Gene Transfer to the Retina

基因转移到视网膜的新方法

• 批准号: 12557145
• 负责人: NAKAZAWA Mitsuru
• 金额: $ 8万
• 依托单位: Hirosaki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12557145/
• 关键词: retinitis pigmentosa; gene transfer; gene therapy; electroporation; conjunctiva; glaucoma; retina

Retinitis pigmentosa is a complex of hereditary progressive retinal degenerations that is nominated as the third commonest cause of legal blindness in adult population in Japan with an incidence of 1 out of 3,500 people, and therefore is an important disease in terms of the policy against blindness. Gene therapy, retinal transplantation and visual prosthesis have been currently studied by many researchers in the world, and expected to be developed as effective treatment. In this study, we investigated the technical possibility and effect of in vivo gene transfer to the retina by electroporation as a method of gene therapy for retinal degeneration. To test the technical possibility, we employed pars plana vitrectomy on the rabbit eyes to approach the retina, injected DNA solution in the subretinal space, and applied electroporation with a needle electrodes. The result indicated that this method could transfer DNA only in a small area of the retina, which might be insufficient to cause t … More herapeutic effects. It also indicated that we needed to develop another type of electrodes such as cup-shaped one. As the second part of the study, we studied the expression and effect of electroporatic gene transfer to the ocular tissue. First of all, we employed conjunctival tissue as a target of gene transfer prier to the retina, because it seemed easier to transfer DNA to the conjunctiva than to the retina. The result showed that the green fluorescent protein transferred to the conjunctiva by electroporation had been apparently expressed in the target tissue 30 days after transfer. Then we tried to transfer metalloproteinase 3 cDNA to the conjunctival flap during trabeculectomy on the rabbit eyes to examine the effect of the gene on intraocular pressure in the postoperative period. The result indicated that postoperative intraocular pressure of the eye treated with gene transfer showed as low as the eye treated by trabeculectomy with MMC, and that there was no pathological reaction in the area where DNA had been injected. All these results have suggested that we successfully transferred DNA fragments to the conjunctiva prier to the retinal, that it is possible for us to perform gene transfer to the retina using electroporation with some modification, and that gene therapy can be applied to glaucoma filtration surgery. Less

色素性视网膜炎是一种遗传性进行性视网膜变性的复杂疾病，是日本成年人法定失明的第三大常见原因，发病率为1 / 3500，因此是防治失明政策的重要疾病。基因治疗、视网膜移植和视觉假体是目前世界上许多研究者研究的，并有望发展成为有效的治疗方法。在本研究中，我们探讨了通过电穿孔将体内基因转移到视网膜作为一种基因治疗视网膜变性的方法的技术可能性和效果。为了检验技术上的可能性，我们对兔眼采用平面部玻璃体切除术接近视网膜，在视网膜下间隙注射DNA溶液，并用针电极进行电穿孔。结果表明，这种方法只能在视网膜的一小块区域内转移DNA，可能不足以产生更大的治疗效果。这也表明我们需要开发另一种类型的电极，如杯形电极。作为研究的第二部分，我们研究了电穿孔基因在眼组织中的表达和作用。首先，我们将结膜组织作为基因转移到视网膜之前的目标，因为将DNA转移到结膜似乎比转移到视网膜更容易。结果表明，通过电穿孔转移到结膜的绿色荧光蛋白在转移30天后已在靶组织中明显表达。然后我们尝试在兔眼小梁切除术期间将金属蛋白酶3cdna转移到结膜瓣，观察该基因对术后眼压的影响。结果显示，基因移植术后眼内眼压与MMC小梁切除术后眼内眼压一样低，且注射DNA部位无病理反应。这些结果表明，我们成功地将DNA片段先于视网膜转移到结膜上，我们可以通过电穿孔进行一些修饰的视网膜基因转移，并且基因治疗可以应用于青光眼滤过手术。少

项目成果

Matsumoto, M, Matsuhashi, H., Nakazawa, M: "Normal tension glaucoma and primary open angle glaucoma associated with increased platelet aggregation"Tohoku J. Exp. Med.. 193(4). 293-299 (2001)

Matsumoto, M、Matsuhashi, H.、Nakazawa, M：“与血小板聚集增加相关的正常眼压性青光眼和原发性开角型青光眼”Tohoku J. Exp。

Tanimoto N, et al.: "Electroretinographic findings in three family members with X-linked juvenile retinoschisis associated with a novel Pro192Thr mutation of the XLRS1 gene"Japanese Journal of Ophthalmology. 46 (5). 566-576 (2002)

Tanimoto N 等人：“三个患有 X 连锁青少年视网膜劈裂症的家庭成员的视网膜电图检查结果与 XLRS1 基因的新型 Pro192Thr 突变相关”，《日本眼科杂志》。

中沢 満: "眼科診療Q&A31"加齢黄斑変性の原因遺伝子とその臨床応用の可能性(分担). 1000 (2002)

Mitsuru Nakazawa：“眼科问答31”负责年龄相关性黄斑变性的基因及其临床应用的可能性（共享）1000（2002）。

Ohguro H, Ogawa K, Maeda T, Maruyama I, Maeda A, Takano Y, Nakazawa M: "Retinal dysfunction in cancer-associated retinopathy is improved by Ca2+ anatagonist administration and dark adaptation"Investigative Ophthalmology & Visual Science. 41(10). 2589-2595

Ohguro H、Okawa K、Maeda T、Maruyama I、Maeda A、Takano Y、Nakazawa M：“通过 Ca2 拮抗剂给药和暗适应可改善癌症相关视网膜病变中的视网膜功能障碍”调查眼科

Yamazaki H, et al.: "Nilvadipine, a Ca2+ antagonist preserves retinal morphology and functions in RCS rat"In vest Ophthalmol Vis Sci. 43(4). 919-926 (2002)

Yamazaki H 等人：“尼伐地平 (Nilvadipine)，一种 Ca2+ 拮抗剂，可保留 RCS 大鼠的视网膜形态和功能”，投资于眼科 Vis Sci。