There is increasing evidence that senescent cells are a driving force behind many age-related pathologies and that their selective elimination increases the life- and healthspan of mice. Senescent cells negatively affect their surrounding tissue by losing their cell specific functionality and by secreting a pro-tumorigenic and pro-inflammatory mixture of growth hormones, chemokines, cytokines and proteases, termed the senescence-associated secretory phenotype (SASP). Here we identified an extract from the plant Solidago virgaurea subsp. alpestris, which exhibited weak senolytic activity, delayed the acquisition of a senescent phenotype and induced a papillary phenotype with improved functionality in human dermal fibroblasts. When administered to stress-induced premature senescent fibroblasts, this extract changed their global mRNA expression profile and particularly reduced the expression of various SASP components, thereby ameliorating the negative influence on nearby cells. Thus, the investigated plant extract represents a promising possibility to block age-related loss of tissue functionality.

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f MITF-M and PAX3 are proteins central to the establishment and transformation of the melanocyte lineage. They control various cellular mechanisms, including migration and proliferation. BRN2 is a POU domain transcription factor expressed in melanoma cell lines and is involved in proliferation and invasion, at least in part by regulating the expression of MITF-M and PAX3. The T361 and S362 residues of BRN2, both in the POU domain, are conserved throughout the POU protein family and are targets for phosphorylation, but their roles in vivo remain unknown. To examine the role of this phosphorylation, we generated mutant BRN2 in which these two residues were replaced with alanines (BRN2TS¡BRN2AA). When expressed in melanocytes in vitro or in the melanocyte lineage in transgenic mice, BRN2TS induced proliferation and repressed migration, whereas BRN2AA repressed both proliferation and migration. BRN2TS and BRN2AA bound and repressed the MITF-M promoter, whereas PAX3 transcription was induced by BRN2TS but repressed by BRN2AA. Expression of the BRN2AA transgene in a Mitf heterozygous background and in a Pax3 mutant background enhanced the coat color phenotype. Our findings show that melanocyte migration and proliferation are controlled both through the regulation of PAX3 by nonphosphorylated BRN2 and through the regulation of MITF-M by the overall BRN2 level. POU family transcription factors are expressed in a wide variety of cell types. They are involved in diverse functions, such as cell type determination, proliferation, renewal, invasion, and migration. The members of the POU domain family of transcription factors share the POU DNA-binding domain (DBD) called the POU domain. The POU domain consists of two DNA-binding units (POUs for POU specific and POUh for POU homeodomain) connected by a flexible linker (3). This molecular structure allows POU proteins to recognize a large set of DNA targets and also to bind different transactivator proteins, depending on the spacing and the positioning adopted by the two subdomains of the POU DBD (50).POU transcription factor function can be modulated by posttranslational modifications, including sumoylation, oxidation, ubiquitinylation, glycosylation, and particularly phosphorylation (5,20,32). Two residues in the DBD domain, a threonine and a serine, are conserved in all mammalian POU domains (see Table S1 at http://umr3347.curie.fr/fr/quipes-de -recherche/d-veloppement-normal-et-pathologie-des-m-lanocytes /differential-function-non-pho). These serine and/or threonine residues in Oct-1, Pit-1, and BRN2 are phosphorylated by protein kinase A (PKA) (31,46,55).Several lines of evidence suggest that PKA is involved in melanocyte lineage proliferation. Forskolin stimulates the proliferation of human melanocytes (54). Proliferation of human melanocytes is induced in a dose-dependent manner by alphamelanocyte-stimulating hormone (1,57,58). Dibutyryl adenosine cyclic AMP induces the proliferation of epidermal melanocytes in culture (29). PKA phosphorylates claudin-1 and allows its tr...

Sestrin 2 (SESN2) is an evolutionarily conserved regulator of mechanistic target of rapamycin complex 1 (mTORC1) which controls central cellular processes such as protein translation and autophagy. Previous studies have suggested that SESN2 itself is subjected to regulation at multiple levels. Here, we investigated the expression of SESN2 in the skin and in isolated skin cells. SESN2 was detected by immunofluorescence analysis in fibroblasts and keratinocytes of human skin. Differentiation of epidermal keratinocytes was not associated with altered SESN2 expression and siRNA-mediated knockdown of SESN2 did not impair stratum corneum formation in vitro. However, SESN2 was increased in both cell types when the expression of its paralog SESN1 was blocked by siRNA-mediated knock down, indicating a compensatory mechanism for the control of expression. Irradiation with UVB but not with UVA significantly increased SESN2 expression in both keratinocytes and fibroblasts. Upregulation of SESN2 expression could be completely blocked by suppression of p53. These results suggest that SESN2 is dispensable for normal epidermal keratinization but involved in the UVB stress response of skin cells.

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The exposure of skin to ultraviolet (UV) radiation can have both beneficial and deleterious effects: it can lead, for instance, to increased pigmentation and vitamin D synthesis but also to inflammation and skin cancer. UVB may induce genetic and epigenetic alterations and have reversible effects associated with post-translational and gene regulation modifications. β-catenin is a main driver in melanocyte development; although infrequently mutated in melanoma, its cellular localization and activity are frequently altered. Here, we evaluate the consequence of UVB on β-catenin in the melanocyte lineage. We report that in vivo, UVB induces cytoplasmic/nuclear relocalization of β-catenin in melanocytes of newborn mice and adult human skin. In mouse melanocyte and human melanoma cell lines in vitro, UVB increases β-catenin stability, accumulation in the nucleus and cotranscriptional activity, leading to the repression of cell motility and velocity. The activation of the β-catenin signalling pathway and its effect on migration by UVB are increased by an inhibitor of GSK3β, and decreased by an inhibitor of β-catenin. In conclusion, UVB represses melanocyte migration and does so by acting through the GSK3-β-catenin axis.

the Republic of) Human skins protect their physiological and structural integrities from various insults such as UVB-irradiation (UV-IR). UV-IR induces multiple downstream signaling pathways that regulate expression of genes. Although human stem cell conditioned mediums show the regenerative effects on photo-aged skin, the molecular mechanism of epithelial precursor cellconditioned media (EPCs-CM) action on photo-aged skin is not elucidated yet. We examined whether EPCs-CM supplementation attenuates UVB-mediated skin photo-aging and regulate UVB-induced ERK signaling on dermal fibroblasts and 3-dimentional human skin equivalents (3D HSE). 3D HSE has been used with the aim of analyzing regenerative photo-protection after UV exposure. EPCs-CM treatment is carried out with various concentrations on dermal fibroblasts (DFs). Protein expressions of MMP1 and procollagen-I in skin aging are evaluated by Western blot, and Immunohistochemistry (IHC). Modulation of ERK signaling by EPCs-CM treatment are examined in DFs and 3D HSE exposed to UVB-IR Our current studies show that UVB-IR up-regulates the expression of MMP-1 and decreases the level of procollagen-1 in primary human dermal fibroblasts respectably. However, EPCs-CM treatment recovers the expression of MMP-1 and procollagen-1 regulated by UVB-IR. Furthermore, EPCs-CM delays UVB-mediated senescence status and inhibits UVB-induced ERK phosphorylation in 3D HSE. Therefore, our studies demonstrate that EPCs-CM may protect skin damages via inhibiting UVB-mediated ERK activation, suggesting that EPCs-CM can be useful material for repair and regeneration of photo-aged skin. 689Silver released from wound dressings damage skin fibroblasts and keratinocytes and ex vivo skin and 2 Cell Physiology Group, Contipro a.s., Dolni Dobrouc, Czech Republic Silver is a favourite antiseptic used in wound healing products, yet it exhibits oxidative damage to eukaryotic cells. We evaluated toxicity of silver dressings to skin cells. Three silverimpregnated bandages were investigated e silver with carboxymethyl cellulose (Ag-CMC), silver with carboxymethyl cellulose and alginate (Ag-CMC-ALG) and nanocrystalline silver on polyester (Ag 0 -PE). Silver was eluted from the bandages and the extracts were assessed in MTT viability test on primary normal human dermal fibroblasts (NHDF) and normal human epidermal keratinocytes (NHEK). Viability of NHEK was reduced by more than 30% and by more than 70% in NHDF after 24 h. After incubation with the bandage extracts, NHDF and NHEK were losing their morphology and were necrotic. Silver dressings induced TNF-a and IL-6 release after 24 h incubation with blood monocytes. Contact toxicity of the bandages on monolayer of mouse 3T3 fibroblasts showed clear halo zones indicating dead cells around the bandages. We investigated, if the silver toxic effects are relevant to intact skin of porcine auricles. The ex vivo skin was incubated in direct contact with the silver bandages for 27 and 51 h. Silver could penetrate the skin as visualised by silver-staining ...