壳聚糖复合膜上及移植部位黑素细胞的实验研究

发布时间：2018-07-29 10:54

【摘要】：白癜风是一种常见的色素病，世界人口的发病率为0.5%-4%[1]，以局限性或泛发性皮肤色素脱失为特征。是一种毁容性的疾病，对病人造成很大的精神和心理压力。发病机制复杂，治疗困难。尽管白癜风的治疗手段多种多样，但是均不能达到治愈的目的。目前白癜风的治疗技术主要分为药物治疗、光疗和外科治疗等，对于稳定期的白癜风，，移植治疗是一种快速有效的方法。但是难治易动部位的白癜风如关节、口周等活动部位黑素细胞附着有一定的困难，因此利用组织工程技术，组织工程膜作为载体，应用于白癜风的治疗领域。目的：探讨利用壳聚糖／明胶复合交联膜（ＣＣＧＭ）作为载体进行黑素细胞移植，确定最佳移植时机，明确移植后黑素细胞在动物创伤部位定植的动态情况。方法：将黑色素细胞接种于CCGM上，分别于4h、1d、2d、3d、4d、5d时通过MTT法测定黑素细胞的生长情况、NaOH裂解法测定黑素含量，评估CCGM上黑素细胞的活性，确定载有黑素细胞组织工程膜的最佳移植时机。应用皮肤共聚焦激光扫描显微镜结合免疫组化技术检测CCGM上搭载的黑素细胞向动物皮肤创面转移情况。结果： 4h、1d、2d、3d、4d、5d时黑素细胞在CCGM和普通培养皿上的增殖速度基本一致，黑素制造量均呈现逐渐增多的状态,以黑素细胞接种到CCGM48h黑素细增殖速度最快，黑素制造量最多，48h后黑素细胞的增殖及黑素制造量进入平台期，可知黑素细胞接种CCGM48h黑素细胞活性较好。黑素细胞接种到CCGM和普通培养皿上48h再移植到裸鼠的移植部位，移植后7d、14d、21d、28d皮肤共聚焦激光扫描显微镜和免疫组化检测移植部位均有黑素细胞定植，以移植后28d最为明显。结论：最佳移植时间是黑素细胞接种到CCGM上48h，移植后7d黑素细胞会种植在动物创面基底层部位，后逐步增多。
[Abstract]:Vitiligo is a common pigment disease, the incidence of the world population is 0.5-4%, characterized by localized or generalized skin pigment loss. Is a disfigurement disease, causing great mental and psychological pressure to patients. The pathogenesis is complicated and the treatment is difficult. Although the treatment of vitiligo is varied, none of them can achieve the goal of cure. At present, the treatment of vitiligo is mainly divided into drug therapy, phototherapy and surgical treatment. Transplantation is a rapid and effective method for stable vitiligo. However, it is difficult to attach melanocytes in vitiligo such as joints and perioral areas. Therefore, tissue engineering technology and tissue engineering membrane are used as carriers for the treatment of vitiligo. Aim: to investigate the optimal time for melanocyte transplantation by using chitosan / gelatin composite cross-linked membrane (CCGM) as the carrier to determine the dynamic situation of melanocyte colonization in animal trauma site after transplantation. Methods: melanocytes were inoculated on CCGM. The growth of melanocytes was measured by MTT method and the content of melanin was determined by NaOH lysis method. The activity of melanocytes on CCGM was evaluated. To determine the optimal time for transplantation of tissue engineering membrane containing melanocytes. Skin confocal laser scanning microscopy and immunohistochemical technique were used to detect the metastasis of melanocytes on CCGM to animal skin wounds. Results: the proliferation rate of melanocytes on CCGM was basically the same as that on common culture dishes at 4 h, 1 d, 2 d, 3 d and 4 d, and the production of melanin increased gradually, and melanocytes inoculated into CCGM48h had the fastest fine proliferation rate. The proliferation of melanocytes and the amount of melanin production entered the plateau stage after 48 hours of maximum melanin production. The results showed that melanocytes inoculated with CCGM48h melanocytes had better activity. Melanocytes were inoculated into CCGM and common culture dish for 48 h and then transplanted to nude mice. After 7 days, 14 days, 21 days and 28 days after transplantation, melanocyte colonization was detected by confocal laser scanning microscope and immunohistochemistry, especially 28 days after transplantation. Conclusion: the best time of transplantation is that melanocytes are inoculated into CCGM for 48 hours. The melanocytes will be implanted in the basal layer of animal wound 7 days after transplantation and then increase gradually.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R758.41

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