In Vitro Culturing and Harvesting of Human Plucked Hair Follicles

Abstract

Human hair follicles are miniature hair growing organs. A common human hair disorder is androgenetic alopecia (AGA), which becomes a medical problem only when the hair loss is subjectively seen as excessive, premature, and distressing on the scalp. The objectives of the present study were to culture the hair stem cells in vitro and to study the morphology of the cultured cells for the treatment of AGA. The present study proposes that plucked human hairs are a cheap source to treat male baldness and in vitro culturing of hair follicle cells is the potential method to apply the cultured cells back into the balding scalp. It may be possible to create thousands of hair follicles from that original follicle. In this study human hair follicle cells of normal and AGA male groups were taken by plucking the hair follicle cells. The hair follicles cells of normal and AGA were cultured in vitro without a feeder layer, as the feeder layer has many drawbacks. The plucked hair follicles which were in the anagen stage were selected from both groups. These hair follicles were digested by enzymatic disaggregation using trypsin/EDTA. Then these cells were cultured in a FAD medium ( Dulbecco’s Modified Eagle Medium and Ham’s F 12 medium, 3:1) plus a 17% serum and incubated in a CO2 Incubator at 37 °C in 5% CO2 without a feeder layer. The whole procedure was performed under sterile conditions. The morphology of cultured and subcultured cells was observed daily under a phase contrast microscope for 14 days. The viable cultured cells of both groups refracted light, while dead cells appeared black. Keratinocytes appeared after 24 hours, Melanocytes appeared at 48 hours, and stem cells appeared in 7 to 10 days. Shelf life of cultured and subcultured cells of normal and AGA group was 12 and 7 days, respectively. Live cell counting was done by using an improved Neubauer chamber. DNA extraction and optical density (OD) assay of cultured and subcultured cells was undertaken using the plucked hair follicles of normal and AGA human male subject. The plucked human hair follicles cells were harvested and cultured successfully without a feeder layer. Their genomic DNA was extracted successfully. This hair cloning technique is an alternative to the usual method of hair transplantation. The most positive aspect of the new technique compared to hair transplantation is the preservation of the 'donor hair area'. This technique will be cheaper and more 'patient friendly'.