The Primary Study Of Tissue Engineered Hair Follicle

BackgroundAt present, autograft of hair follicles is the most important treatment for hair loss,especially for male pattern baldness. But there are some problems for this treatment,such as formation of scar in donor site, scantiness of donor site, low density oftransplantations, low survival rate of transplantations, so the tissue engineered hairfollicles provide a promising method for repairing hair loss.A hair follicle is composed of epithelial cells including matrix, out root sheathinner root sheath and dermal cells including dermal papilla and deral sheath.Dermal papilla cells are a kind of fibroblast cells, that can aggregate during growingin mediums containing FCS. According to the different phages of hair cycle, theshape of the dermal papilla change periodically. It plays an important role in theformation of hair follicle and hair cycle, so establish the culture model of dermalpapilla cells in vitro is very important to study the characteristics of dermal papillacells and induction of hair follicles. The epithelial cells is the base for producinghair fibers, so it is absolutely necessary for reconstructing hair follicles. But themethod of isolating these cells is just fit to be used in a lab, it can not meet the needsrequired by tissue engineering.Isolation and cultivation of vibrissa hair follicles of rat is very important for the complex experiments, such as gene labeling and gene expressing, but themicrodissecting method is hard to manipulate, cells are hard to adhere and growslowly, it is also easy to be contaminated, this is a laboring task.Objective1. To establish the method for isolating dermal papilla cells and epithelial cells ofhuamn scalp hair follicles in large scale rapidly and efficiently.2. To improve the method for isolating dermal papilla cells and epithelial cells ofrat vibrissa hair follicles.3. To establish the method of reconstructing hair follicles.Methods1. Dermal papilla cells of human scalp hair follicles were isolated bydigesting-filtrating method, and cultured in DMED medium containing 15%FCS,the bioligical behavior in vitro was observed, the adhering rate was measured andcompared with the cells isolated by microdisseting method. The immunohisto-chemical stain ofα-SMA was performed on these cells.2. Epithelial cells of human scalp hair follicles are isolated by step-by-stepdigesting method, and cultured in K-SFM medium, the bioligical behavior in vitrowas observed. The immunohistochemical stain of K-19 was performed on these cells.3. Dermal papilla cells of rate vibrissa hair follicles were isolated bywhole-follicle-digesting method, and cultured in DMED medium containing 15%FCS,the bioligical behavior in vitro was observed.4. Epithelial cells of rate vibrissa hair follicles were isolated by two-stepwhole-follicle-digesting method,and cultured in K-SFM medium, the bioligicalbehavior in vitro was observed.5. Follicular cells were implanted onto the back skin of nude mouse byhypodermal injection, the histologic examination of the skin was performed after 14days.Results1. Digesting-filtrating method succeeded in isolating dermal papilla cells ofhuman scalp hair follicless. Cells proliferated rapidly in vitro, the adhering rate washigh and can reached 96%, they grew in a aggregating way and were positive forimmunohistochemical stain ofα-SMA.2. Step-by-step digesting method succeeded in isolating epithelial cells of humanscalp hair follicless. Cells proliferated rapidly in vitro in the medium of K-SFM, likepaving stones,and were positive for immunohistochemical stain of K-19.3. Whole-follicle-digesting method succeeded in isolating dermal papilla cells ofrat vibrissa hair follicless. Cells proliferated rapidly in vitro, the adhering rate washigh and can reached 95%, they grew in a aggregating way.4. Two-step whole-follicle-digesting method succeeded in isolating epithelialcells of rat vibrissa hair follicless. Cells proliferated rapidly in vitro in the medium ofK-SFM, like paving stones.5. New formed hair follicles can be seen under microscopy.Conclusion1. Digesting-filtrating method and Step-by-step digesting method can isolatedermal papilla cells and epithelial cells of human scalp hair follicless on a large scalerapidly and efficiently. Adhering rate of dermal papilla cells isolated bydigesting-filtrating method is higher than that isolated by microdisseting method.2. Whole-follicle-digesting method is an efficient method to isolate dermalpapilla cells and epithelial cells of rat vibrissa hair follicles, it solve the problemscaused by microdissecting, such as hard adhering, low migration of cells and lowgrowth.3. A mixture composed of single follicular cells can form new hair follicles under particular conditions.