keratinocyte melanocyte co culture

BACKGROUND: Exosomes have been demonstrated to carry proteins, membrane lipids, mRNAs and microRNAs which can be transferred to surrounding cells and regulate the functions of those recipient cells. OBJECTIVES: The objective of the study was to investigate the effects of exosomes released by keratinocytes and fibroblasts on the proliferation, tyrosinase activity and melanogenesis of melanocytes. METHODS: Melanocytes, keratinocytes and fibroblasts obtained from human foreskin were cultured and exosomes secreted by keratinocytes and fibroblasts were harvested from the culture supernatants by ultracentrifugation...

Chronic wounds have become an increasing medical and economic problem of aging societies because they are difficult to manage. Tissue engineering provides new perspectives for the clinically applicable skin substitutes. Epidermal keratinocytes play an important role in wound epithelization and construction of tissue-engineered skin substitutes. How to obtain a large number of autologous epidermal keratinocytes in a short time is the main problem that limits the application of tissue-engineered skin and epidermal cell membranes...

Melanoma is a lethal form of skin cancer triggered by genetic and environmental factors. Excision of early-stage, poorly aggressive melanoma often leads to a successful outcome; however, left undiagnosed these lesions can progress to metastatic disease. This research investigates whether the exposure of poorly aggressive melanoma to certain normal skin cells can explain how non-metastatic melanoma becomes more aggressive while still confined to the skin. To this end, we used a serial co-culture approach to sequentially expose cells from two different, poorly aggressive human melanoma cell lines against normal cells of the skin beginning with normal melanocytes, then epidermal keratinocytes, and finally dermal fibroblasts...

Kynurenine (KYN), the most abundant metabolite of tryptophan, is classically associated with immune tolerance and tumor immune escape. In the last years, KYN is in the spotlight in other biological processes. Here, we showed that KYN inhibited tyrosinase expression and melanin content in primary human melanocyte and keratinocyte co-cultures. Furthermore, KYN decreased melanosome content in a 3D human skin reconstruction model. In these experiments, we used tyrosine + NH4 Cl to induce pigmentation. We compared the inhibitory effect of KYN on melanogenesis with the already known inhibitory effect promoted by IFN-γ...

Familial melanoma accounts for 10% of cases, being CDKN2A the main high-risk gene. However, the mechanisms underlying melanomagenesis in these cases remain poorly understood. Our aim was to analyze the transcriptome of melanocyte-keratinocyte co-cultures derived from healthy skin from familial melanoma patients vs. controls, to unveil pathways involved in melanoma development in at-risk individuals. Accordingly, primary melanocyte-keratinocyte co-cultures were established from the healthy skin biopsies of 16 unrelated familial melanoma patients (8 CDKN2A mutant, 8 CDKN2A wild-type) and 7 healthy controls...

Reconstructed human skin models are a valuable tool for drug discovery, disease modelling and basic research. In the last decades, major progress has been made in this field leading to the development of full thickness skin models (FTSms) better representative of the native human skin by including the cellular crosstalk between the dermal and epidermal layers. However, current available FTSms still present important limitations since they are only suitable for short-term studies, include non-human extracellular matrix components and have a weak skin barrier function compared to in vivo human skin...

Opsin-3 (OPN3) is a potential key regulator of human melanocyte melanogenesis. How OPN3-mediated regulation of melanocyte melanogenesis is triggered is largely unclear. TGFβ can inhibit the growth of human melanocytes and reduce melanin synthesis in melanocytes. However, whether TGFβ2 can modulate pigmentation in normal human primary melanocytes through OPN3 is entirely unknown. In this study, we constructed a coculture model with human epidermal melanocytes and keratinocytes. OPN3, tyrosinase (TYR), tyrosinase-related protein (TRP)-1, and TRP-2 expression and TYR activity were detected to be higher in cocultured cells than in monocultured cells...

Melanin granules cluster within supra-nuclear caps in basal keratinocytes (KCs) of the human epidermis, where they protect KC genomic DNA against ultraviolet radiation (UVR) damage. While much is known about melanogenesis in melanocytes (MCs) and a moderate amount about melanin transfer from MC to KC, we know little about the fate of melanin once inside KCs. We recently reported that melanin fate in progenitor KCs is regulated by rare asymmetric organelle movement during mitosis. Here, we explore the role of actin, microtubules, and centrosome-associated machinery in distributing melanin within KCs...

Introduction: Melanocytes are engaged in synthesis, transport, and release of pigments at the epidermal-melanin units in response to the finely regulated melanogenic pathway. A multifaceted combination of both intrinsic and extrinsic factors - from endocrine and paracrine dynamics to exogenous stimuli such as sunlight and xenobiotics - modulates expression and activity of proteins involved in pigmentation, including the rate-limiting enzyme tyrosinase. As well as playing critical physiological functions comprising skin photoprotection, melanins define hair and skin pigmentation which in turn have impacted considerably to human social communication since time immemorial...

Bench-to-bedside axis of therapeutic product development is currently being oriented towards minimum invasiveness on both ends-not only clinical application but harvesting of the starting biological material as well. This is particularly relevant for Advanced Therapy Medicinal Products and their specific legislative requirements, even more so in skin regeneration. It is precisely the skin equivalents and grafts that benefit from the minimum-to-noninvasive approach to a noteworthy extent, taking in account the sensitive nature of both skin harvesting and grafting...

The coat color of mammals is primarily determined by the type, quantity, and distribution of melanin in the skin and hair. As an endogenous gas molecule, nitric oxide (NO) regulates tyrosinase production by modulating the cGMP-dependent protein kinase (PKG) pathway, which enhances melanin synthesis. However, some interrelationships have not been fully elucidated. In the present study, mouse melanocytes co-cultured with mouse keratinocytes in vitro, or as monocultures, were used as research models. The results indicated that ultraviolet B irradiation increased nitric oxide synthase (NOS) activity and NO production, and increased PKG, p21-activated kinase 4 (PAK4), and microphthalmia-associated transcription factor (MITF) levels, as well as tyrosinase (TYR), tyrosinase-related protein 1 and 2 expression, and melanin synthesis...

As the outermost barrier of the body, skin is a major target of oxidative stress. In the brain, estrogen has been reported synthesized locally and protects neurons from oxidative stress. Here, we explored whether estrogen is also locally synthesized in the skin to protect from oxidative stress and whether aberrant local estrogen synthesis is involved in skin disorders. Enzymes and estrogen receptor expression in skin cells were examined first by quantitative real-time PCR and Western blot analyses. Interestingly, the estrogen synthesis enzyme was mainly localized in epidermal keratinocytes and estrogen receptors were mainly expressed in melanocytes among 13 kinds of cultured human skin cells...

With the rapid growth of wireless communication devices, the influences of electromagnetic fields (EMF) on human health are gathering increasing attention. Since the skin is the largest organ of the body and is located at the outermost layer, it is considered a major target for the health effects of EMF. Skin pigmentation represents one of the most frequent symptoms caused by various non-ionizing radiations, including ultraviolet radiation, blue light, infrared, and extremely low frequency (ELF). Here, we investigated the effects of EMFs with long-term evolution (LTE, 1...

Access to complex in vitro models that recapitulate the unique markers and cell-cell interactions of the hair follicle is rather limited. Creation of scalable, affordable, and relevant in vitro systems which can provide predictive screens of cosmetic ingredients and therapeutic actives for hair health would be highly valued. In this study, we explore the features of the microfollicle, a human hair follicle organoid model based on the spatio-temporally defined co-culture of primary cells. The microfollicle provides a 3D differentiation platform for outer root sheath keratinocytes, dermal papilla fibroblasts, and melanocytes, via epidermal-mesenchymal-neuroectodermal cross-talk...

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Research on melanogenesis, its regulation in health and disease, and the discovery of new molecules with pigmenting and depigmenting activities use different models. Here we standardize a protocol based on previous ones using primary human melanocytes and keratinocytes in co-cultures, in which melanogenesis was induced under mild conditions by the addition of tyrosine plus ammonium chloride (NH4 Cl). The expression of MITF, TYR, TYRP1, and Melan-A as well as melanin content were measured. Furthermore, we extended this study to a reconstructed 3D model...

BACKGROUND: 585 nm light-emitting diodes have been proven to suppress melanogenesis in melanocytes. However, whether LEDs will influence normal human epidermal keratinocytes (NHEKs) and paracrine effect of LEDs-irradiated NHEKs in melanogenesis remains unknown. OBJECTIVE: To elucidate the possible mechanisms in vitro of anti-melanogenic activity of 585 nm LEDs on paracrine effect of NHEKs and its exosomes. METHODS: NHEKs irradiated with different fluences of 585 nm LEDs were evaluated the cell viability by CCK8 assay...

Clinical studies have proven that ultraviolet B (UVB) based phototherapy can induce perifollicular and marginal repigmentation patterns in the skin of vitiligo patients. It is, however, difficult to conceive how melanocytes can easily exit from their tightly interconnected epidermal microenvironment to re‑enter a different location in the skin to establish a new network with neighboring keratinocytes. While it is known that matrix metalloprotease 9 (MMP9) is involved in the degradation of the extracellular matrix in physiological or pathological processes, little is known about whether MMP9 affects melanocyte migration in vitiligo repigmentation...

The content and distribution of melanin in the epidermis determines the wide variety of skin colors associated with ethnic/racial diversity. Although it was previously reported that qualitative changes in keratinocyte-derived exosomes regulate melanocyte pigmentation in vitro, their practical involvement, especially in skin color development in vivo, has remained unclear. To address this unexplained scientific concern, the correlation of epidermal exosomes isolated from human skin tissues with melanosomal protein expression levels was demonstrated in this study for the first time...

BACKGROUND: Rab27a, Mlph, and MyoVa form a tripartite complex and relate to melanosome distribution. Melanophilin (Mlph) acts as a linker protein between Rab27a and MyoVa. The biological activity and function of 16-kauren on the expression of Mlph has not yet been studied. OBJECTIVE: We examined the effect of 16-kauren on melanosome transport and skin pigmentation. METHODS: Murine Melan-a melanocytes and SP-1 keratinocytes were used for in vitro analysis...