毛囊混合细胞诱导毛囊重建的探索研究

发布时间：2018-03-14 21:06

本文选题：外根鞘细胞　切入点：毛乳头细胞　出处：《昆明医学院》2010年硕士论文　论文类型：学位论文

【摘要】： 目的:自行分离培养获得和购买ScienCell公司的人毛囊外根鞘细胞株(HHFORSC)及人毛囊毛乳头细胞株(HHDPC),以得到充足的种子细胞。HHDPC、HHFORSC与海藻酸钠3D支架在体外构建三维模型,培养后植入SD大鼠皮下以诱导其形成毛囊,并探讨毛囊的生物学特征和这两种细胞在毛囊形成中的作用。方法:购买ScienCell公司的人毛囊外根鞘细胞株(HHFORSC)和人毛囊毛乳头细胞株(HHDPC),复苏培养;同时利用酶消化法和组织块法获得HHDPC和HHFORSC,培养后比较两种不同途径得到的细胞生长特征,以获得充足的HHFORSC和HHDPC。将上述两种细胞混合后种植于海藻酸钠3D细胞支架,构建毛囊的体外三维模型,培养7周,光镜观察毛囊形成情况。同时将该三维模型移植入SD大鼠皮下,饲养SD大鼠8周,移植部位取材后,行HE染色、免疫组化和电镜观察毛囊形成情况。结果:由ScienCell公司购买的细胞株和我们自行分离培养获得的HHDPC和HHFORSC生长特征没有明显差异。体外毛囊三维模型培养7周后,光镜下可见支架中有均匀散在分布的毛囊混合细胞,未见到角化物质及毛囊结构。SD大鼠饲养8周移植部位取材,HE染色可见细胞聚集成团状的毛囊样结构,CK14,CK15和波形蛋白染色阳性,电镜下可见支架中有散在细胞分布。结论:我们通过酶消化法和组织块法分离获得的HHDPC和HHFORSC保持了良好的生长特性。体外毛囊三维模型培养未能形成毛囊样结构。SD大鼠体内三维模型培养,HE染色可见细胞聚集成毛囊样结构,CK14,CK15染色阳性,波形蛋白染色阳性,证明在该模型中诱导出了具有向毛囊分化倾向的毛囊样结构,并且HHDPC保持了诱导毛囊形成的能力,HHFORSC保持了毛囊干细胞的作用,为以后成功重建毛囊指明了方向。
[Abstract]:Objective: to isolate and culture the human hair follicle outer root sheath cell line HHFORSCand the human hair follicle hair papilla cell line HHDPCC, so as to obtain sufficient seed cells, HHDPC-HHFORSC and sodium alginate 3D scaffolds to construct a three-dimensional model in vitro. After culture, SD rats were implanted subcutaneously to induce the formation of hair follicles. The biological characteristics of hair follicles and the role of these two kinds of cells in hair follicle formation were discussed. Methods: human hair follicle outer root sheath cell line HHFORSC) and human hair follicle dermal papilla cell line HHDPCC were purchased for resuscitation culture, and HHDPC and HHFORSCwere obtained by enzyme digestion and tissue block method. In order to obtain sufficient HHFORSC and HHDPC. the above two kinds of cells were mixed and implanted in sodium alginate 3D cell scaffold. The three-dimensional model of hair follicle was established in vitro and cultured for 7 weeks. The formation of hair follicle was observed by light microscope. At the same time, the model was transplanted subcutaneously into SD rats. SD rats were fed for 8 weeks. After transplantation, he staining, immunohistochemistry and electron microscope were used to observe the hair follicle formation. Results: there was no significant difference between the growth characteristics of HHDPC and HHFORSC obtained by ScienCell and the growth characteristics of HHDPC and HHFORSC obtained by our own isolation and culture. After 7 weeks of culture in vitro hair follicle, there were hair follicle mixed cells scattered in the scaffold under light microscope. No keratocystis and hair follicles were observed in SD rats after 8 weeks of feeding. He staining showed that the cells were clustered into clusters of hair follicle structure, CK14, CK15 and vimentin were positive, and the cells were scattered in the scaffold under electron microscope. Conclusion: the HHDPC and HHFORSC isolated by enzyme digestion and tissue mass method have good growth characteristics. In vitro hair follicle three-dimensional model culture can not form hair follicle-like structure. SD rat three-dimensional model culture can be stained with HE. It was found that the cells gathered to form a follicle-like structure, and the cells were positive for CK14 / CK-15 staining. The positive staining of vimentin showed that hair follicle-like structure was induced in the model, and HHDPC maintained the ability to induce hair follicle formation. HHFORSC maintained the role of hair follicle stem cells. It points out the direction for the successful reconstruction of hair follicles in the future.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R751

【参考文献】