Mast cells are widely distributed in the connective tissue of the body, but are particularly prominent in tissues such as skin. An increased number of mast cells can be found in the dermis under inflammatory conditions and ultraviolet (UV) exposed skin. Previous investigations have identified matrix metalloproteinases (MMPs) as key enzymes in the degradation of extra cellular matrix (ECM). This study reports about the potential contribution of human mast cell tryptase as a new triggering enzyme in matrix degradation process. Recent studies suggest that mast cell-derived proteases can activate MMPs. We investigated both the degradation of cellular matrix components and activation of MMPs by human tryptase. Mast cells are increased in photoaged skin and the increase of mast cell tryptase in UV irradiated skin was confirmed. Human mast cell tryptase was purified from human tonsils by a series of standard chromatographic procedures. Degradation of collagen type I was achieved by incubation of human type I collagen with tryptase and the fragments were quantified by SDS-PAGE and staining with Coomassie Brilliant Blue 250-R (CBB). Treatment with tryptase resulted in the activation of proMMP-9 as revealed by gelatinolytic activity in type IV collagen zymography. When tryptase was incubated with human type IV collagen, gradual degradation of intact collagen was detected by Western blotting. Furthermore, type IV collagen degradation was observed in the basement membrane (BM) of a three-dimensional (3D) skin model. Degranulation of mast cells, which release tryptase, can activate MMPs and causes direct damage to ECM proteins. These findings strongly implicate that tryptase either alone or in conjunction with activation of MMPs, can participate in ECM damage and the possible destruction of BM leading to photoaging.

Skin aging is the aging process of skin tissue due to elastin and collagen breakdown. Collagen and elastin are protein of connective tissue in skin dermis that serves to regenerate the skin for firmness and flexibility maintained. Sunlight is composed by three main parts according to their wavelengths, namely UVA, UVB, and UVC where the intensity of UVB sunlight is most active. UVB generates the production superoxide anion (O 2-) which is main free radical in the skin surface. It attacked the cell membrane and subsequently form a new ROS and decrease antioxidants enzymatic excesively. Excessive ROS production resulting overexpression of AP-1 which activated Matrix metalloproteinases lead to collagen breakdown and photoaging. This paper discusses the affinity and interaction of isorhamnetin to CYP1A1 causing inactivation of AP-1 using molecular docking program. CYP1A1 is protein member of cytochrome P450 superfamily of enzymes encoded by the CYP1A1 gene. Cytochrome P450 used as oxidazing catalist in metabolic pathways steroids, fatty acids, xenobiotics, including drugs, toxins and carcinogens. This study consist of two stages: (1) isorhamnetin-CYP1A1 docking and α-naphthoflavone-CYP1A1 redocking (2) ∆G scores, inhibitory constant, and bonding interaction between ligan-reseptor analysis. Gibbs energy and inhibitory (∆G) constant (Ki) showed stability interaction between isorhamnetin and CYP1A1. Based on the ∆G score isorhamnetin has higher potential as CYP1A1 inhibitor than-naphthoflavoneα. Isorhamnetin-CYP1A1 get-10,3 kcal/mol and 11,34 µM and-naphthoflavoneα get-9,1 kcal/mol and 228,4 µM. Isorhamnetin-CYP1A1 binded by 4 π bondingand2hydrogen bonding. The result presented the potential of isorhamnetin to decreased AP-1 expressioan through CYP1A1 inhibition.

GPNMB is involved in multiple cellular functions including cell adhesion, stress protection and stem cell maintenance. In skin, melanocyte-GPNMB is suggested to mediate pigmentation through melanosome formation, but details of keratinocyte-GPNMB have yet to be well understood. We confirmed the expression of GPNMB in normal human epidermal keratinocytes (NHEKs) by reducing the expression using siRNA. A higher calcium concentration of over 1.25 mM decreased the GPNMB expression. Histological staining showed that GPNMB was expressed in the basal layer of normal skins but completely absent in vitiligo skins. the normal expression of GpnMB in nevus depigmentosus skin suggested that lack of GPNMB is characteristic of vitiligo lesional skins. IFN-γ and IL-17A, two cytokines with possible causal roles in vitiligo development, inhibited GPNMB expression in vitro. Approximately 4-8% of the total GPNMB expressed on NHEKs were released possibly by ADAM 10 as a soluble form, but the process of release was not affected by the cytokines. The suppressive effect of IFN-γ on GpnMB was partially via IFN-γ/JAK2/STAT1 signaling axis. Decreased GPNMB expression in keratinocytes may affect melanocyte maintenance or survival against oxidative stress although further studies are needed. These findings indicate a new target for vitiligo treatment, focusing on the novel role of IFN-γ and IL-17 in downregulating keratinocyte-GPNMB. GPNMB (Glycoprotein nonmetastatic melanoma protein B)-also known as osteoactivin, dendritic cell-heparin integrin ligand (DC-HIL), or hematopoietic growth factor inducible neurokinin-1 type-is a type I transmembrane glycoprotein. GPNMB has 2 transcript variants encoding 560 and 572 amino acid isoforms in human and shares a 25% amino acid sequence homology with PMEL-17, a melanocyte-specific melanosomal protein 1. The extracellular part of GPNMB mainly contains an RGD motif that binds to integrin in the process of maintaining cell-cell adhesion, and an Ig-like polycystic kidney disease (PKD) domain involved in protein-protein and protein-carbohydrate interactions. The cytoplasmic tail contains an immunoreceptor tyrosine-based activation motif (ITAM) that takes part in the intracellular signaling via Src and Syk cytoplasmic kinases, and a di-leucine motif required for its endosomal⁄melanosomal sorting signal 2. GPNMB is widely expressed in various tissues such as the skin, brain, breasts, muscle, and bone 3-7. The known functions of GPNMB include cellular adhesion through integrin 1 , regulation of the degeneration/regeneration of the extracellular matrix in skeletal muscles 6 , the mineralization of bone 2 , the differentiation of osteoclasts 8 and osteoblasts 9 , the impairment of T-cell activation 10 , the regulation of inflammatory responses in macrophages 11 , the suppression of motor neuron degeneration in amyotrophic lateral sclerosis 12 , and the invasion and metastasis of several cancers 13-17. GPNMB has also been widely demonstrated to increase the endoplasmic reticulum (ER) stress response by inducing th...

The production of melanin is regulated by α-melanocyte-stimulating hormone (α-MSH), which is produced from proopiomelanocortin (POMC). Keratinocytes release POMC along with lower levels of α-MSH and ACTH. To clarify the mechanism of melanogenesis after ultraviolet (UV)-irradiation, this study focused on the expression of POMC and POMC-derived peptides after UV-irradiation. Western blot analysis and immunoassays indicated that both POMC and α-MSH-like immunoreactivity (α-MSH-LI) increased after UV-irradiation. However, other POMC-derived products were very low. In hypophysectomized mice, α-MSH-LI increased to the same level as in control mice after UV-irradiation. Structural analysis revealed that the major α-MSH-LI product was ACTH(1–8). Furthermore, ACTH(1–8) competed with [125I]-α-MSH for receptor binding and increased melanin production via a melanocortin-1 receptor. These results suggested that melanin was produced through ACTH(1–8) after UV-irradiation. Trypsin-like enzymatic activity, which is responsible for POMC activation, increased after UV-irradiation and was identified as tryptase. In mast cell-deficient mice, which do not produce tryptase, α-MSH-LI levels were unchanged after UV-irradiation. The present study demonstrates the production of ACTH(1–8) from POMC by tryptase, which is a novel peptide-processing mechanism in the extracellular compartment of the skin.