Data Availability StatementThe datasets used and/or analyzed through the current study are available from the corresponding author on reasonable request. was evaluated, and the results revealed that SU6656 and PP2 inhibited the phosphorylation of Src in G361 cells. Src inhibition by these chemical inhibitors induced melanogenesis in G361 cells and upregulated the mRNA expression levels of melanogenesis-associated genes encoding microphthalmia-associated transcription factor, tyrosinase-related protein 1 (TRP1), TRP2, and tyrosinase. In addition, Src inhibition by small interfering RNA induced melanogenesis and upregulated the mRNA expression levels of melanogenesis-associated genes. As the p38 mitogen-activated protein kinase (MAPK) and cyclic adenosine monophosphate response element binding (CREB) pathways serve key functions in melanogenesis, the present study further examined whether Src mediates melanogenesis via these pathways. As expected, Src inhibition via SU6656 or PP2 administration induced the phosphorylation of p38 or CREB, as determined by western blotting analysis, and increased the levels of phosphorylated p38 or CREB, as determined by immunofluorescence staining. In addition, the induced pigmentation and melanin content of G361 cells by Src inhibitors was significantly inhibited by p38 or CREB inhibitors. Taken together, these data indicate that Src is usually associated with melanogenesis, and COL24A1 Src inhibition induces melanogenesis via the MAPK and CREB pathways in G361 cells. Keywords: Src inhibition, melanin, G361 cell, p38, cyclic adenosine monophosphate response element binding Introduction Melanin is an important factor in determining the color of the human skin, hair and eyes (1,2). It is produced in the melanosome through a complex process known as melanogenesis (3C5). In addition, melanin serves an important role in photoprotection from ultraviolet (UV) radiation and external stress (1,3). Growth factors, cytokines, hormones and other receptor ligands exert their function by interacting with their receptors around the cell surface, generating a signaling cascade and leading to unique patterns of protein phosphorylation. Melanocytes express several unique receptor tyrosine kinases (RTKs) that bind bone morphogenic protein (BMP), hepatocyte growth factor (HGF) and c-Kit ligand. For example, BMP-2 stimulates tyrosinase gene expression and melanogenesis in differentiated melanocytes, and BMP signaling controls hair pigmentation via cross-talk with the melanocortin receptor-1 pathway (6). The activation of Met in response to HGF acts as a mitogen for melanocytes and synergistically contributes to malignant progression with the aberrant expression of basic fibroblast growth factor in malignant melanocytes (7). Normal human melanocytes and melanoma cells express the c-Kit gene and stem cell factor (SCF), a ligand of the c-Kit receptor that upregulates the expression of melanogenic proteins (8). In addition, SCF/c-Kit signaling is required for cyclic regeneration of the locks pigmentation device (9). Phosphorylation of the RTKs eventually activates some kinases referred to as mitogen-activated proteins kinases (MAPKs) or various other intracellular signaling substances such as for example cyclic adenosine monophosphate (10). After that, following phosphorylation of protein such as for example microphthalmia-associated transcription aspect (MITF), the transcription of genes that take part in melanocyte proliferation and Azacitidine price melanogenesis is certainly turned on (11). The Src kinase family members (SKF) is certainly a family group of non-receptor tyrosine kinases that’s made up of nine associates including Src, And Fyn Yes. SKF interacts numerous cellular cytosolic, membrane and nuclear proteins, changing these proteins by phosphorylating tyrosine residues and adding to the development of cellular change and oncogenic activity (12). Of the, C-terminal Src Azacitidine price kinase (c-Src) is certainly Azacitidine price encoded with the SRC gene in human beings; it phosphorylates particular tyrosine residues in various other proteins. c-Src could be turned on by many transmembrane protein including RTKs, such as for example platelet-derived growth aspect receptor, epidermal development aspect receptor, and c-Kit. As a result, c-Src is certainly closely from the RTK pathways (13). As RTKs serve a crucial function in the advancement and development of several types of cancers (12), an increased activity degree of c-Src tyrosine kinase is certainly from the development of various kinds of cancers, such as for example pancreatic and breasts cancers (14). Therefore, diverse Src inhibitors have been developed to prevent cancer progression, and drugs against RTKs are widely used in malignancy therapy. However, there is little to no evidence on the effects of SKF or its inhibitors on melanocytes. Therefore, the present study investigated the effect of a c-Src inhibitor on melanocytes and its associated signaling pathways. Materials and methods Cell cultures and chemical treatment Human G361 melanoma cells (cat. no. ATCC? CRL-1424?; American Type Culture Collection, Manassas, VA, USA) were cultured in low glucose Dulbecco’s altered Eagle’s medium with 10% fetal bovine serum (FBS; Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA) and 1% penicillin/streptomycin (Gibco; Thermo Fisher Scientific, Inc.). G361 cells were managed at 37C in a humidified 5% CO2 incubator. -Melanocyte-stimulating hormone (-MSH; Sigma-Aldrich; Merck KGaA, Darmstadt, Germany), SU6656 (cat. no. S7774; Selleck Chemicals, Houston, TX, USA), PP2 (cat. no. S7008; Selleck Chemicals), H-89 [protein kinase A (PKA) inhibitor; cat. no. B1427; Sigma-Aldrich; Merck KGaA], SB203580 (p38 inhibitor; cat. no. S8307; Sigma-Aldrich; Merck KGaA) dasatinib (cat. no. CDS023389; Sigma-Aldrich; Merck KGaA) and nilotinib (cat. no. CDS023093; Sigma-Aldrich; Merck.