BMP-2 regulates the expression of myosin Va via smad in melan-a melanocyte.

Myosin Va (Myo Va) is one of three protein complexes involved in melanosome transport. In this study, we identified BMP-2 as an up-regulator of Myo Va expression using 2-methyl-naphtho[1,2,3-de]quinolin-8-one (MNQO). Our results showed that MNQO reduced the mRNA and protein expression of Myo Va and BMP-2 in melanocytes. Knockdown of BMP-2 by siRNA also affected Myo Va mRNA and protein expression, confirming that MNQO regulates Myo Va through BMP-2. Furthermore, phosphorylation of Smad1/5/8 by BMP2 treatment confirmed that the BMP-2/Smad signaling pathway regulates Myo Va expression in Melan-a melanocytes. Smad-binding elements were found in the Myo Va promoter and phosphorylated Smad1/5/8 bind directly to the Myo Va promoter to activate Myo Va transcription and BMP-2 enhances this binding. These findings provide insight into a new role for BMP-2 in Melan-a melanocytes and a mechanism of regulation of Myo Va expression that may be beneficial in the treatment of albinism or hyperpigmentation disorders.

Comprehensive Analysis of Exosomal MicroRNAs Derived from UVB-Irradiated Keratinocytes as Potential Melanogenesis Regulators.

The exosomes derived from keratinocytes can have a substantial impact on melanogenesis by influencing melanocytes. MicroRNAs (miRNAs) encapsulated within exosomes are implicated in the control of melanogenesis, particularly when under the influence of UVB irradiation. This investigation explores UVB-induced exosomal miRNAs from keratinocytes as potential regulators of melanogenesis. UVB-irradiated, keratinocyte-derived exosomes were observed to augment melanogenesis in melanocytes, resulting in an upregulation of MITF, TRP1, TRP2, and TYR expression compared to non-UVB-irradiated exosomes. Additionally, a subset of exosomal miRNAs was differentially selected and confirmed to exert both enhancing and inhibitory effects on melanogenesis through functional assays. Notably, hsa-miR-644a, hsa-miR-365b-5p, and hsa-miR-29c-3p were found to upregulate melanogenesis, while hsa-miR-18a-5p, hsa-miR-197-5p, and hsa-miR-4281 downregulated melanogenesis. These findings suggest the involvement of keratinocyte-derived exosomal miRNAs in melanogenesis regulation within melanocytes. The expression levels of exosomal miRNAs from keratinocytes exhibited a UVB-dependent increase, indicating a potential role for these miRNAs as regulators of melanogenesis in response to UVB irradiation. Furthermore, melanogenesis was found to be dependent on exosomes derived from keratinocytes. This underscores the potential of UVB-induced exosomal miRNAs derived from keratinocytes as regulators of melanogenesis. Moreover, this study unveils a significant role for exosomes in melanocyte pigmentation, presenting a novel pathway in the intricate process of melanogenesis.

Phosphorylation of glucocorticoid receptor induced by 16-kauren-2-beta-18, 19-triol decreases expression of Melanophilin through JNK signalling.

16-kauren-2-beta-18, 19-triol (16-kauren) is a natural diterpenoid substance derived from Asteraceae psiadia punctulata, a small tropical shrub in Africa and Asia, and it can reduce Mlph expression without affecting the expression of Rab27a and MyoVa in melanocytes. Melanophilin (Mlph) is an important linker protein in the melanosome transport process. However, the signal transduction pathway for the regulation of Mlph expression has not been fully established. We examined the mechanism of 16-kauren on Mlph expression. Murine melan-a melanocytes were used for in vitro analysis. Western blot analysis, quantitative real-time polymerase chain reaction, and luciferase assay were performed. The inhibition of Mlph expression by 16-kauren-2ß-18,19-triol (16-kauren) occurs through the JNK signal and is reversed following glucocorticoid receptor (GR) activation by dexamethasone (Dex). Especially, 16-kauren activates JNK and c-jun signalling, part of the MAPK pathway, with subsequent Mlph repression. When the JNK signal is weakened by siRNA, the inhibition of Mlph expression by 16-kauren was not seen. JNK activation by 16-kauren induces GR phosphorylation, which leads to Mlph repression. These results demonstrate that 16-kauren regulates Mlph expression through the phosphorylation of GR via the JNK signalling pathway.

Inhibition of Melanosome Transport by Inducing Exon Skipping in Melanophilin.

Exon skipping is an efficient technique to inhibit specific gene expression induced by a short-sequence peptide nucleic acid (PNA). To date, there has been no study on the effects of PNA on skin pigmentation. In melanocytes, the tripartite complex is responsible for the transport of mature melanosomes from the nucleus to the dendrites. The tripartite complex is composed of Rab27a, Mlph (Melanophilin), and Myosin Va. Defects in the protein Mlph, a melanosome transport-related protein, are known to cause hypopigmentation. Our study shows that Olipass peptide nucleic acid (OPNA), a cell membrane-permeable PNA, targets exon skipping in the Mlph SHD domain, which is involved in Rab27a binding. Our findings demonstrate that OPNA induced exon skipping in melan-a cells, resulting in shortened Mlph mRNA, reduced Mlph protein levels, and melanosome aggregation, as observed by microscopy. Therefore, OPNA inhibits the expression of Mlph by inducing exon skipping within the gene. These results suggest that OPNA, which targets Mlph, may be a potential new whitening agent to inhibit melanosome movement.

The Effect of Lactobacillus plantarum Extracellular Vesicles from Korean Women in Their 20s on Skin Aging.

Extracellular vesicles, which are highly conserved in most cells, contain biologically active substances. The vesicles and substances interact with cells and impact physiological mechanisms. The skin is the most external organ and is in direct contact with the external environment. Photoaging and skin damage are caused by extrinsic factors. The formation of wrinkles is a major indicator of skin aging and is caused by a decrease in collagen and hyaluronic acid. MMP-1 expression is also increased. Due to accruing damage, skin aging reduces the ability of the skin barrier, thereby lowering the skin's ability to contain water and increasing the amount of water loss. L. plantarum suppresses various harmful bacteria by secreting an antimicrobial substance. L. plantarum is also found in the skin, and research on the interactions between the bacteria and the skin is in progress. Although several studies have investigated L. plantarum, there are only a limited number of studies on extracellular vesicles (EV) derived from L. plantarum, especially in relation to skin aging. Herein, we isolated EVs that were secreted from L. plantarum of women in their 20s (LpEVs). We then investigated the effect of LpEVs on skin aging in CCD986sk. We showed that LpEVs modulated the mRNA expression of ECM related genes in vitro. Furthermore, LpEVs suppressed wrinkle formation and pigmentation in clinical trials. These results demonstrated that LpEVs have a great effect on skin aging by regulating ECM related genes. In addition, our study offers important evidence on the depigmentation effect of LpEVs.

Regulation of Melanophilin (Mlph) gene expression by the glucocorticoid receptor (GR).

Mlph plays a crucial role in regulating skin pigmentation through the melanosome transport process. Although Mlph is a major component involved in melanosome transport, the mechanism that regulates the expression of the Mlph gene has not been identified. In this study, we demonstrate that Mlph expression is regulated by the glucocorticoid receptor (GR). Alteration of GR activity using a specific GR agonist or antagonist only regulated the expression of Mlph among the 3 key melanosome transport proteins. Translocation of GR from the cytosol into the nucleus following Dex treatment was confirmed by separating the cytosol and nuclear fractions and by immunofluorescence staining. In ChIP assays, Dex induced GR binding to the Mlph promoter and we determined that Dex induced the GR binding motif on the Mlph promoter. Our findings contribute to understanding the regulation of Mlph expression and to the novel role of GR in Mlph gene expression.

Antiaging Potential of Fucoxanthin Concentrate Derived from Phaeodactylum tricornutum.

The aim of the study was to investigate the potential of a fucoxanthin concentrate prepared from Phaeodactylum tricornutum as a wrinkle care cosmetic agent. The concentrate (up to 25 µg/ml) did not affect the proliferation of human fibroblasts. In addition, the concentrate significantly increased procollagen synthesis in the fibroblasts at 12.5 and 25 µg/ml; however, it significantly decreased the expression of matrix metalloproteinase (MMP)-1, MMP-2, and MMP-9 at 25 µg/ml. In a follow-up study, a wrinkle care cream containing 0.03% of fucoxanthin concentrate was prepared and tested in women (aged 35-50 years, n = 21) for 8 weeks. The cream was applied twice daily. Safety assessment of the cream was carried out visually. In addition, interviews were conducted to investigate if adverse events such as erythema, edema, scaling, itching, stinging, burning, tightness, or prickling had occurred. No symptoms that threaten skin safety were reported. Evaluation of wrinkles around the eyes using the replica method showed a statistically significant decrease in wrinkles at week 8. Moreover, skin moisture and elasticity increased significantly from week 4. These results suggest that the fucoxanthin concentrate has no adverse effects on the skin and can be used as an active ingredient in wrinkle care cosmetics.

A novel function of Prohibitin on melanosome transport in melanocytes.

Prohibitin (PHB, also known as PHB1 or BAP32), is a highly conserved 31kDa protein that expressed in many cellular compartments, such as mitochondria, nucleus, cytosol, and plasma membrane, and plays roles in regulating the transcription of genes, apoptosis, and mitochondrial biogenesis. There is a report that Prohibitin expression is required for the stimulation of pigmentation by melanogenin. However, no studies have been published on the function of PHB in melanocytes, especially in melanosome transport. Methods: Immunofluorescence was performed to confirm the localization of PHB. siRNA transfections, Co-immunoprecipitation, western blotting and proximity ligation assay were performed to find binding state between proteins and demonstrate functions of PHB on melanosome transport. Results: PHB is located in the melanosome and perinuclear aggregation of melanosome is induced when expression of PHB is reduced with no influence on melanin contents. PHB binds directly to Rab27a and Mlph but not Myosin-Va. Rab27a and Mlph bind to specific domains of PHB. Reduced expression of PHB led to the impaired binding affinity between Rab27a and Mlph. Conclusion: PHB regulates melanosome transport by linking to Rab27a and Mlph in melanocytes. Targeting and regulating PHB not only manages pigmentation in melanocytes, but also controls hyperpigmentation in melanoma.

Particulate Matter-Induced Aryl Hydrocarbon Receptor Regulates Autophagy in Keratinocytes.

Particulate matter (PM), which refers to the mixture of particles present in the air, can have harmful effects. Damage to cells by PM, including disruption of organelles and proteins, can trigger autophagy, and the relationship between autophagy and PM has been well studied. However, the cellular regulators of PM-induced autophagy have not been well characterized, especially in keratinocytes. The Aryl Hydrocarbon Receptor (AhR) is expressed in the epidermis and is activated by PM. In this study, we investigated the role of the AhR in PM-induced autophagy in HaCaT cells. Our results showed that PM led to AhR activation in keratinocytes. Activation of the AhR-target gene CYP1A1 by PM was reduced by co-treatment with α-naphthoflavone (α-NF), an AhR inhibitor. We also evaluated activation of the autophagy pathway in PM-treated keratinocytes. In HaCaT cells, treatment with PM treatment led to the induction of microtubules-associated proteins light chain 3 (LC3) and p62/SQSTM1, which are essential components of the autophagy pathway. To study the role of the AhR in mediating PM-induced autophagy, we treated cells with α-NF or used an siRNA against AhR. Expression of LC3-ІІ induced by PM was decreased in a dose dependent manner by α-NF. Furthermore, knockdown of AhR with siAhR diminished PM-induced expression of LC3-ІІ and p62. Together, these results suggest that inhibition of the AhR decreases PM-induced autophagy. We confirmed these results using the autophagy-inhibitors BAF and 3-MA. Taken together, our results indicate that exposure to PM induces autophagy via the AhR in HaCaT keratinocytes.

The absence of Rab27a accelerates the degradation of Melanophilin.

Melanophilin (Mlph) forms an interaction with Rab27a and the actin-based motor protein MyosinVa (MyoVa) on mature melanosome membranes and the tripartite complex regulates melanosome transport in melanocytes. In this study, we found that Rab27a siRNA decreased Mlph and Rab27a protein levels, but Mlph mRNA levels were not changed. Other Rab27a siRNA sequences also showed the same results. When Rab27a siRNA was treated with melan-a melanocytes, Rab27a protein was decreased within 6 hours and Mlph protein was decreased within 24 hours. To determine whether the absence of Rab27a promotes Mlph degradation, we inhibited protein degradation by treatment with proteasome (MG132) and lysosomal enzyme (E64D and Pepstatin A) inhibitors in melan-a melanocytes. MG132 inhibited the degradation of Mlph, but E64D and Pepstatin A had no effect on Mlph. The absence of Rab27a enhanced ubiquitination of Mlph and induced proteasomal degradation. From these results, we concluded that Mlph interaction with Rab27a is important for Mlph stability and melanosome transport.