C-ski基因转染对兔眼小梁切除术后滤过道瘢痕形成的影响

发布时间：2018-03-08 17:12

本文选题：小梁切除　切入点：抗瘢痕　出处：《第三军医大学》2010年硕士论文　论文类型：学位论文

【摘要】： 青光眼滤过术是青光眼的一种常规手术方式,手术的成功率与术区的瘢痕化密切相关。为了提高手术成功率,术中常应用抗代谢药物来预防术后伤口瘢痕化。虽然丝裂霉素-C(MMC)、5-氟尿嘧啶(5-Fu)等抗代谢药物术中应用提高了手术成功率,但术后发生的持续性低眼压、滤过泡渗漏、眼内炎等并发症也随之增多。为了减少术后滤过道瘢痕形成,各种抗瘢痕化学、生物制剂及物理粘附屏障已经运用于动物滤过术模型中,并取得了不错的治疗效果,但是由于各种原因,都还没有应用于临床。 ski基因是一种原癌基因,在体内作为转录的辅活化子或辅阻遏子调节多个重要转录因子的表达,对创口组织再生起着重要的作用。大坪医院野战外科研究所第七研究室前期研究发现,C-ski能促进体外培养的大鼠皮肤成纤维细胞增殖和降低胶原分泌,对兔耳创口也有促进愈合,降低瘢痕的作用,因此推测C-ski基因可能对促进创伤愈合,降低瘢痕形成有重要意义,为此,我们将其用于兔眼小梁切除术后的抗瘢痕研究,观察其疗效,为以后的临床应用奠定基础。本实验主要进行了以下三个方面的研究:1.对C-ski基因重组pcDNA3.0质粒及pcDNA3.0空载质粒进行扩增、提纯并加以鉴定;2.不同剂量的C-ski基因重组pcDNA3.0质粒及PBS对兔眼小梁切除后滤过道瘢痕形成的影响;3. 500ug剂量C-ski基因重组pcDNA3.0质粒及丝裂霉素、PBS、空载质粒对兔眼小梁切除后滤过道瘢痕形成的影响。主要结果和结论如下: 1.通过对C-ski基因重组pcDNA3.0质粒及pcDNA3.0空载质粒进行扩增、纯化和鉴定,结果证实:采用碱裂解法提取的C-ski基因重组pcDNA3.0质粒及pcDNA3.0空载质粒的质和量,能够达到我们后续动物实验的要求。 2.不同剂量的C-ski基因重组pcDNA3.0质粒能降低眼压及延长滤过泡存留时间,并且能抑制滤过道Ⅰ型胶原生成,剂量增加,效果增强,而对滤过道成纤维细胞及肌成纤维细胞生成及转化无明显影响。 3. 500ug剂量的C-ski基因重组质粒对兔眼小梁切除术后进行抗瘢痕治疗时,未引起炎症反应,能明显减少Ⅰ型胶原生成量,抑制滤过道瘢痕的形成,延长滤过术后低眼压的时间,而无丝裂霉素(MMC)副作用。
[Abstract]:Glaucoma filtration is a routine operation for glaucoma. The success rate of operation is closely related to scar change in operation area. Antimetabolic drugs are often used to prevent scar formation after operation. Although the use of anti-metabolic drugs such as mitomycin CMMC and 5-fluorouracil (5-Fu) improves the success rate of the operation, the persistent intraocular pressure and bleb leakage occur after the operation. In order to reduce the formation of post-operative filtration scar, various anti-scar chemical, biological agents and physical adhesion barrier have been used in animal model of filtration surgery, and have achieved good therapeutic effect. However, due to various reasons, has not been applied to clinical. Ski gene is a proto-oncogene. As a coactivator or repressor of transcription in vivo, it regulates the expression of many important transcription factors. It was found that C-ski could promote the proliferation and decrease of collagen secretion of rat skin fibroblasts cultured in vitro, and promote the healing of rabbit ear wounds. Therefore, we speculate that C-ski gene may play an important role in promoting wound healing and reducing scar formation. Therefore, we used it to study the anti-scar effect after trabeculectomy in rabbits. To lay a foundation for clinical application in the future. This experiment mainly carried out the following three aspects of research: 1.Amplification of C-ski gene recombinant pcDNA3.0 plasmid and pcDNA3.0 empty plasmid, Effect of different doses of C-ski Gene Recombinant pcDNA3.0 plasmid and PBS on the formation of Trabeculectomy scar in Rabbit eyes after trabeculectomy; 3500ug / g C-ski Gene Recombinant pcDNA3.0 plasmid and mitomycin PBS; No-loaded plasmid for trabeculectomy in Rabbit eyes. The effect of retrofiltration on scar formation in the passageway. The main findings and conclusions are as follows:. 1. The C-ski gene recombinant pcDNA3.0 plasmid and pcDNA3.0 empty plasmid were amplified, purified and identified. The results showed that the quantity and quality of C-ski gene recombinant pcDNA3.0 plasmid and pcDNA3.0 empty plasmid were obtained by alkaline lysis method. To meet the requirements of our subsequent animal experiments. 2. Different doses of C-ski gene recombinant pcDNA3.0 plasmid can reduce intraocular pressure and prolong the retention time of bleb, and can inhibit the formation of collagen type I in the filtering passage, and the dose is increased and the effect is enhanced. However, there was no significant effect on the formation and transformation of fibroblasts and myofibroblasts. After trabeculectomy, 3500ug dose of C-ski gene recombinant plasmid did not cause inflammatory reaction, significantly reduced collagen I production, inhibited the formation of filtering scar, and prolonged the time of low intraocular pressure after filtering. There were no side effects of MMC.
【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R779.6

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