Molecular and cellular basis of Exogen and Anagen induction

Abstract

The hair follicle has the unique capacity to pass through periods of growth, regression and rest before regenerating itself to restart the cycle. This dynamic cycling capacity enables animals to change their coats, and for hair length to be controlled at different body sites. While progress has been made on some of the outstanding questions in hair follicle biology, many of the cellular and molecular mechanisms involved in driving hair follicle cycling remain undiscovered. The first section of work in this thesis reports on the dermal papilla, an essential component of the hair follicle with a key role in follicle regeneration. Using hanging drop cultures spheres of human dermal papilla cells were created, and the expression of cytoskeletal and extracellular matrix components characteristic of intact papillae were analysed using immunohistochemistry and real time PCR. I found evidence that over time the gene and protein expression profiles of the papilla cell spheres became more representative of intact dermal papillae, although differences were seen between papilla cells strains derived from different individuals. I then demonstrated the inductive capability of human dermal papilla cells using this spherical model in an amputated follicle assay. This data provides some of the first evidence that cultured human dermal papilla cells can retain inductive capacity without having to be combined with other cell types. This induction phenomenon may also have relevance to anagen initiation and cycling. The second section of this thesis was concentrated on the process of club fibre shedding, now widely recognised as a phase of the follicle cycle known as exogen. The vibrissa follicle, with its predictable timing of club fibre loss, was first developed as a new model for exogen, and the structural and adhesive changes associated with the progression towards club fibre release were analysed using SEM and ТЕМ. Combined with plucking experiments these showed club loss to be gradual process. The expression of proteins involved in cell adhesion, differentiation, communication and digestion, all potential mediators of fibre loss were examined around "young “and "old" club fibres using immunohistochemistry. To further assess the mechanism of exogen, and to search for signalling molecules microarray technology was utilised, enabling the identification of 75 genes that were associated with the process. Group analysis highlighted a particular role for proteases and their inhibitors in the retention and final release of the club fibre. Moreover, immunofluorescent analysis of the results from the microarray identified a process of differentiation, specific to the cell layer surrounding the club fibre prior to club fibre release. This data provides evidence that the process of exogen is progressive, associated with maturation of the cells surrounding the club fibre, and terminating with the release of the club fibre. The processes involved in club fibre release are becoming more widely recognised as an important aspect of the hair cycle and this thesis is one of the first comprehensive pieces of work that analyses club retention and release as an active phase.