Influential Factors Of Skin Wound Transplantation With Autogenous Hair Follicle In Rats

ObjectiveBy establishing a rat model of autologous hair follicle transplantation to skin wound,to observe the effects of different treated hair follicle autologous grafts on wound healing,so as to prove that there may be physical or biological barriers in hair follicles that affect wound healing.Through the space-time expression of stem cell marker CK15,CK17 and stem cell activation-related signaling pathways Wnt/Lef1,to observe and explore the regulation patterns of hair follicle stem cells migration and differentiation of and their possible mechanisms,in order to confirm the feasibility of using hair follicle stem cells for skin wounds repair and to lay experimental and theoretical basis for its clinical application.Besides,to propose an economical and efficient treatment for skin defects by comparing the repairing efficiency of hair follicle grafts and autologous skin grafts.Methods1.A wedge of skin was cut from one of the whisker pads of rats,and the whisker follicles were separated from the skin.The intact whisker follicles and whisker follicles respectively with the papilla removed,the hair shaft removed,the hair shaft and the papilla removed were prepared.2.A spindle skin strip（about 1.0cm×0.3-0.4cm）was cut from one side foot pads of rats,and 9small strips（about 0.2cm×0.2cm）were cut from it with micro instruments.3.A full-thickness skin wound with a diameter of 3cm was made on the back of rats.4.Grouping and wound treatment:Rats were randomly divided into 6 groups and the following interventions were performed:（1）Physical-chemical-barrier removal group:9 whisker hair follicles with hair shaft removed and papilla removed were attached to autologous dorsal skin wound（2）Physical-barrier removal group:9 whisker hair follicles with removed hair shaft were attached to the skin wound of autologous back（3）Chemical-barrier removal group:9 whisker hair follicles with dermal papilla removed were attached to autologous dorsal skin wound（4）Intact hair follicle group:9 untreated rat whisker hair follicles were attached to autologous dorsal skin wounds（5）Positive control group:one side foot pad was trimmed into about 9 0.2cm×0.2cm skin strips,which were pasted on the autologous back skin wound（6）The blank group will not undergo any migration processing.5.Binding and fixing the wounds;the rats were kept separately in cages to prevent them from biting each other’s wounds.6.On the 3rd,7th and 14th day after surgery,the wound repair was observed,samples were collected,paraffin sections were prepared,morphological changes were observed by HE staining,and the expression of stem cell markers CK15 and CK17 and downstream transcription factor Lef1 of Wnt signaling pathway was observed by fluorescence immunohistochemistry.Results1.Compared with the blank control group,the transplantation of different treated rat whisker hair follicles into the autologous back skin wound could promote the healing of the back wound in rats.2.New epidermal cell layer formed around the upper end of the transplanted papilla free hair follicles on 14^thday,and CK17 and Lef1 were positively expressed,while CK15 was positively expressed on the migration track of hair follicle stem cells.3.With the removal of dermal papilla,the hair follicle stem cells of the transplanted hair follicles were more likely to migrate to the direction of the epidermis during wound repair.On the opposite,with the dermal papilla reserved,the hair follicle stem cells were more likely to differentiate to form new hair follicles near the wound during wound repair.4.Efficiency of wound healing:skin film≈group with physical-chemical barrier removed/group with chemical barrier removed>group with physical barrier removed/group with complete hair follicle>blank group.Conclusions1.With the dermal papilla removed,the whisker hair follicles of the rats transplanted into the autologous back wound could lead to new epidermis formation,and the new epidermis was formed by the migration and differentiation of hair follicle stem cells.2.Compared with the hair follicles with dermal papilla,the stem cells of dermal papilla free hair follicles promote wound repair more efficiently,which indicated that the repair efficiency may be associated with hair follicle structure itself and gave indirect proof to a"chemical"barrier affects migration and differentiation of hair follicle bulge stem cells in wound repair.3.The healing efficiency of hair follicle transplantation removing the dermal papilla was similar to that of autologous skin grafts.In the case of insufficient autologous skin grafts,hair follicle transplantation could also provide an economical and efficient method to repair skin defects in clinic,so as to improve the clinical feasibility of using hair follicle stem cells to repair skin wounds.