THE ISOLATION AND MAINTENANCE OF THE HUMAN PILOSEBACEOUS UNIT

We have previously developed methods for the isolation and maintenance of human sebaceous glands and hair follicles. However, in long-term c ulture the maintenance of both is suboptimal. This may be due to a lac k of stem cells, which are thought to be located in the bulge area of the hair follicle, and this region is not present in either model. Iso lation of the entire pilosebaceous unit would retain this region, and may lead to improved maintenance of both structures. We describe a met hod for the isolation of viable, individual, pilosebaceous units by mi crodissection from human scalp face-lift skin. The viability of isolat ed pilosebaceous units has been determined by light microscopy, patter ns of DNA synthesis by [methyl-H-3] thymidine autoradiography, and lip ogenesis by [U-C-14] acetate uptake into lipids. When maintained for 7 days in supplemented Williams E medium, isolated pilosebaceous units showed a significant increase in length. This was due to the productio n of a keratinized hair fibre which grew at the in vivo rate of 0.3 mm /day. Light microscopy and [methyl-H-3] thymidine autoradiography conf irmed that after 7 days maintenance the hair follicle retained apparen tly normal morphology and patterns of DNA synthesis. However, the morp hology of the sebaceous gland on maintenance was more variable, genera lly showing luminal keratinization. Moreover [methyl-H-3] thymidine au toradiography of sebaceous glands showed a marked reduction on mainten ance. The rates and patterns of lipogenesis by the whole pilosebaceous unit Were, respectively, lower and different from those seen with iso lated human sebaceous glands; this indicates that the bulk of piloseba ceous lipogenesis is derived from the hair follicle. Rates of recovery of [C-14] from 2 mM-[U-C-14] sodium acetate into thin-layer chromatog raphy plates after 7 days maintenance decreased, although this was not statistically significant, indicating that rates of lipogenesis may f all on maintenance. Pilosebaceous units were maintained for 7 days on Gelfoam (an absorbable gelatin sponge) at the media-air interface. Ini tial results show a marked improvement in sebaceous gland morphology. It is possible, therefore, to obtain viable human pilosebaceous units by microdissection, and to maintain them in vitro for up to 7 days, wi th apparently full retention of hair follicle function, but only parti al retention of sebaceous gland function.