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The purpose of this study was to evaluate human hair follicle melanogenic activity using the [C-14]-2-thiouracil, which was known to incorporate into nascent melanins. Results obtained on pigmented, grey and non-pigmented hair follicles demonstrated that [C-14]-2-TU incorporation was restricted to the melanogenic compartment with a strong accumulation located around dermal papilla and within the fibre of pigmented hair follicles. Quantitative analysis of [C-14]-2-TU incorporation showed a significant increase in pigmented hair follicles upon stimulation with 1 mu m forskolin concomitant to an increase in tyrosinase levels. A strong significant decrease in [C-14]-2-TU incorporation was noted, when hair follicles were incubated with the tyrosinase competitive inhibitor kojic acid (200 mu m). Incubation with the MC1-R agonist alpha-MSH (0.2 mu m) did not induce a significant stimulation of hair melanogenesis. The present model could thus represent a useful new tool to identify modulators of human hair pigmentation.

NAD(+) dependent 15-hydroxyprostaglandin dehydrogenate (15-PGDH) catalyses oxidation of 15(S)-hydroxyl group of prostaglandins and as a result inactivates their physiological potential. Positive effects of prostaglandins or prostaglandin analogues were reported on terminal hair, vellus hair or eyelash growth and a complex prostaglandin network was recently described in human hair follicle. In the present study, we showed that 15-PGDH was expressed in human hair follicle mainly in melanocytes and keratinocytes, which brought us to consider this enzyme as a possible target to sustain local prostaglandin production. Using a recombinant enzymatic strategy, specific 15-PGDH inhibitors were screened. We identified a thiazolidine dione derivative exhibiting efficacy on follicular outer root sheath keratinocytes, since it concomitantly decreased the production of deactivated 13,14 dihydro 15-ketoprostaglandin F(2alpha) and sustained prostaglandin F(2alpha)in vitro production. In the context of recent interest in prostaglandins and prostaglandin analogues as hair regrowth agents, we postulated that the use of selected 15-PGDH inhibitors could reinforce or prolong the effect of these physiological mediators on hair and skin.

The hair follicle dermal papilla is composed primarily of extracellular matrix (ECM) proteins secreted by resident fibroblasts. Dermal papilla is endowed with hair morphogenic properties, yet its composition is poorly characterized. In an attempt to understand its specificity better, we compared the protein composition of ECM secreted by cultured dermal papilla fibroblasts with that of dermal fibroblasts. ECM proteins are generally large, difficult to solubilize, and abundantly post-translationally modified. We thus implemented an original protocol for analyzing them: ECM samples were enzymatically digested directly in the culture flasks and analyzed by LC-MS/MS. Sequencing of proteolytic peptides by MS/MS yielded protein identification. The relative abundance of a given protein in dermal fibroblast versus dermal papilla samples was estimated by comparing proteolytic peptide intensities detected by MS. Using this approach, several matrix proteins were found to be present at markedly different levels in each ECM type; in particular, thrombospondin 1 and fibronectin appeared to be overrepresented in the dermal papilla fibroblast ECM. MS results were supported by Western blot and immunostaining experiments. In addition, peptide intensities were processed in two ways, which proved to favor either the quantification accuracy or the information precision at the sequence level.

Une nouvelle strategie utilisant le couplage entre la chromatographie en phase liquide et la spectrometrie de masse en tandem a ete developpee et appliquee a l'analyse de melanges complexes de proteines digerees. Cette strategie a ete mise en oeuvre pour l'analyse comparative des proteines de matrice extracellulaire synthetisees par des fibroblastes de derme et des fibroblastes de papille dermique en culture. Ces analyses ont permis de mettre en evidence des proteines plus abondamment et specifiquement exprimees dans chaque type d'echantillon. Cette etude, realisee sur des echantillons obtenus in vitro, a revele des marqueurs proteiques probables de la papille dermique, compartiment situe a la base du follicule pileux, et qui est specifiquement capable d'induire la pousse des cheveux.