Tsutomu Soma

Control of murine hair follicle regression (catagen) by TGF-β1 in vivo

The regression phase of the hair cycle (catagen) is an apoptosis‐driven process accompanied by terminal differentiation, proteolysis, and matrix remodeling. As an inhibitor of keratinocyte proliferation and inductor of keratinocyte apoptosis, transforming growth factor β1 (TGF‐β1) has been proposed to play an important role in catagen regulation. This is suggested, for example, by maximal expression of TGF‐β1 and its receptors during late anagen and the onset of catagen of the hair cycle. We examined the potential involvement of TGF‐β1 in catagen control. We compared the first spontaneous entry of hair follicles into catagen between TGF‐β1 null mice and age‐matched wild‐type littermates, and assessed the effects of TGF‐β1 injection on murine anagen hair follicles in vivo. At day 18 p.p., hair follicles in TGF‐β1 —/— mice were still in early catagen, whereas hair follicles of +/+ littermates had already entered the subsequent resting phase (telogen). TGF‐β1 — /— mice displayed more Ki‐67‐positive cells and fewer apoptotic cells than comparable catagen follicles from +/+ mice. In contrast, injection of TGF‐β1 into the back skin of mice induced premature catagen development. In addition, the number of proliferating follicle keratino‐cytes was reduced and the number of TUNEL + cells was increased in the TGF‐β1‐treated mice compared to controls. Double visualization of TGF‐β type II receptor (TGFRII) and TUNEL reactivity revealed colocalization of apoptotic nuclei and TGFRII in catagen follicles. These data strongly support that TGF‐β1 ranks among the elusive endogenous regulators of catagen induction in vivo, possibly via the inhibition of keratinocyte proliferation and induction of apoptosis. Thus, TGF‐βRII agonists and antagonists may provide useful therapeutic tools for human hair growth disorders based on premature or retarded catagen development (effluvium, alopecia, hirsutism).—Foitzik, K., Lindner, G., Mueller‐Roever, S., Maurer, M., Botchkareva, N., Botchkarev, V., Handjiski, B., Metz, M., Hibino, T., Soma, T., Dotto, G. P., Paus, R. Control of murine hair follicle regression (catagen) by TGF‐β1 in vivo. FASEB J. 14, 752–760 (2000)

Hair‐inducing ability of human dermal papilla cells cultured under Wnt/β‐catenin signalling activation

Abstract:  It is well known that dermal papilla cells (DPCs) play crucial roles in hair follicle induction. In this study, we examined whether Wnt/β‐catenin activation results in maintenance of the hair‐inducing ability of human DPCs. Expression of DPC marker genes was maintained under Wnt/β‐catenin signalling stimulation by GSK‐3β inhibition. Furthermore, human DPCs showed constant hair induction when transplanted with murine epidermal cell fraction. Alu‐positive human DPCs were essentially detected adjacent to the reconstructing epidermal structure positive for P‐cadherin immunoreactivity. The transplanted human DPCs were abundant in the surrounding dermal sheath portion of the fully regenerated hair follicles. These results support the importance of Wnt/β‐catenin signalling in hair follicle induction. This study may provide valuable information to establish a culture method of human DPCs for cell‐based therapy.

Identification of novel hair-growth inducers by means of connectivity mapping

The aim of this study was to identify novel inducers of hair growth using gene expression profiling at various stages of hair‐growth induction. First, we analyzed gene expression at the onset of hair growth in mice induced by cyclosporin A (CsA), a well‐known hair‐growth inducer, using DNA microarray analysis. The results unveiled genes involved in the step‐by‐step progression of hair growth, including increases in melanin biosynthesis and decreases in immune response at d 2 and the subsequent stimulation of cell proliferation at d 4, followed by the up‐regulation of hair specific keratins at d 7 after CsA treatment. With the use of the connectivity map (Cmap), agents that had a similar “gene signature” to that of the profiles of CsA‐treated mice were identified. Several agents, including CsA, were identified by the Cmap and were evaluated for hair induction activity in vivo. One of the proposed agents, fluphenazine (from the d 2 signature) actually induced hair growth in vivo (ED50: 2 mM for single application), and the subsequent application of 5 mM iloprost (from the d 4 signature) significantly enhanced the hair‐growth effect of fluphenazine. From these results, Cmap analysis was proven to be a useful method that connects gene expression profiles of complicated biological processes, such as hair‐growth induction, to effective agents.—Ishimatsu‐Tsuji, Y., Soma, T., Kishimoto, J. Identification of novel hair‐growth inducers by means of connectivity mapping. FASEB J. 24, 1489–1496 (2010). www.fasebj.org

Perivascular localization of dermal stem cells in human scalp.

Abstract:  In mammalian skin, the existence of stem cells in the dermis is still poorly understood. Previous studies have indicated that mesenchymal stem cells (MSCs) are situated as pericytes in various mammalian tissues. We speculated that the human adult dermis also contains MSC‐like cells positive for CD34 at perivascular sites similar to adipose tissue. At first, stromal cells from adult scalp skin tissues showed colony‐forming ability and differentiated into mesenchymal lineages (osteogenic, chondrogenic and adipogenic). Three‐dimensional analysis of scalp skin with a confocal microscope clearly demonstrated that perivascular cells were positive for not only NG2, but also CD34, immunoreactivity. Perivascular CD34‐positive cells were abundant around follicular portions. Furthermore, CD34‐positive cell fractions collected with magnetic cell sorting were capable of differentiating into mesenchymal lineages. This study suggests that dermal perivascular sites act as a niche of MSCs in human scalp skin, which are easily accessible and useful in regenerative medicine.

Cultured human dermal papilla cells secrete a chemotactic factor for melanocytes

Large numbers of pigmented melanocytes are located in human hair follicles, predominantly around the dermal papillae, and the number of melanocytes and the melanogenic activity of the hair follicles are closely related to the hair cycle. We found that cultured human dermal papilla cells secreted a melanocyte chemoattractant into the medium. Skin fibroblasts also showed weak chemoattraction of melanocytes, while skin keratinocytes and melanocytes did not. Since this chemotactic activity was heat-and protease-sensitive and was present in the relatively high molecular weight fraction (130-200 kDa), it may be due to extracellular matrix (ECM) that proteins secreted from the cultured dermal papilla cells. This chemotactic signal between dermal papillae and melanocytes may control the localization and migration of hair melanocytes in vivo.

Isolation of mesenchymal stem cells from human dermis.

Recent studies revealed that mammalian dermis contains multipotent stem cells such as skin-derived precursors (SKPs). SKPs grow in suspension as spheres. In contrast, mesenchymal stem cells (MSCs) are adherent fibroblastic cells. Here, we describe the procedure to isolate MSCs under low-serum culture conditions. In addition, we explain the method to collect MSCs using magnetic cell sorting.

Marked Changes in Lamellar Granule and Trans-Golgi Network Structure Occur During Epidermal Keratinocyte Differentiation

Epidermal lamellar granules transport various lipids, proteins, and protein inhibitors from the trans-Golgi network to the extracellular space, and play an important role in skin barrier formation. We elucidated the 3-dimensional structure of lamellar granules and the trans-Golgi network in normal human skin by focused ion beam scanning electron microscopy. Reconstructed focused ion beam scanning electron microscopy 3-dimensional images revealed that the overall lamellar granule structure changed from vesicular to reticular within the second layer of the stratum granulosum. Furthermore, the trans-Golgi network was well developed within this layer and spread through the cytoplasm with branched, tubular structures that connected to lamellar granules. Our study reveals the unique overall 3-dimensional structure of lamellar granules and the trans-Golgi network within the cells of the epidermis, and provides the basis for an understanding of the skin barrier formation.