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Background Keratinocytes can function as innate immune cells under oxidative stress and aggravate the cutaneous T-cell response that undermines melanocytes in the setting of vitiligo. The NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is a regulator of innate immunity that exists in keratinocytes. However, the role of the NLRP3 inflammasome in the pathogenesis of vitiligo has not been investigated. Objective We sought to explicate the contribution of the activated NLRP3 inflammasome in keratinocytes to the autoimmune response in patients with vitiligo. Methods Perilesional and serum samples from patients with vitiligo were collected to examine the status of the NLRP3 inflammasome in the setting of vitiligo. Cultured keratinocytes were treated with H2O2 to investigate the mechanism for NLRP3 inflammasome activation under oxidative stress. Peripheral blood T cells were extracted from patients with vitiligo to explore the influence of the NLRP3 inflammasome on the T-cell response in patients with vitiligo. Results Expressions of NLRP3 and downstream cytokine IL-1β were consistently increased in perilesional keratinocytes of patients with vitiligo. Notably, serum IL-1β levels were increased in patients with vitiligo, correlated with disease activity and severity, and decreased after effective therapy. Furthermore, oxidative stress promoted NLRP3 inflammasome activation in keratinocytes through transient receptor potential cation channel subfamily M member 2 (TRPM2), a redox-sensitive cation channel, which was dependent on TRPM2-mediated calcium influx. More importantly, blocking TRPM2-induced NLRP3 inflammasome activation in keratinocytes impaired chemotaxis for CD8+ T cells and inhibited the production of cytokines in T cells in patients with vitiligo. Conclusion Oxidative stress–induced NLRP3 inflammasome activation in keratinocytes promotes the cutaneous T-cell response, which could be targeted for the treatment of vitiligo.

Background Oxidative stress plays important roles in the pathogenesis of vitiligo. The removal of hydrogen peroxided (H2O2) has been established to be beneficial to vitiligo patients. Berberine (BBR), a natural isoquinoline alkaloid, has antioxidant activity, however, whether BBR can defend human melanocytes against oxidative injury remains to be elucidated. Objective In the present study, we investigated the potential protective effect of BBR against oxidative stress on an immortalized normal human melanocyte cell line PIG1. Methods Generally, PIG1 cells were pretreated with various concentrations of BBR for 1 h followed by exposure to 1.0 mM H2O2 for 24 h. Cell apoptosis, intracellular reactive oxygen species (ROS) levels were assessed through flow cytometry. Cell apoptosis, melanogenesis and the activation of Nrf2-ARE and Mitf signaling pathway were assayed. Results Our results showed that cell viability rose and intracellular ROS generation, cell apoptosis of melanocytes decreased significantly in response to H2O2 through pretreatment with BBR. Furthermore, We found that BBR can dramatically induce Nrf2 nuclear translocation, increase total Nrf2 levels and enhance ARE activity. Besides, Nrf2-siRNA transfection can abrogate the protection of BBR in melanocytes against oxidative injury. At last, we verified that BBR could facilitate melanogenesis function via modulation of Mitf and its target proteins. Conclusion The results above suggest that BBR can protect melanocytes against oxidative stress via its anti-oxidative activity. Also, we found H2O2-induced activation of NFκB was inhibited by BBR. Therefore, it is worthy of investigation BBR as a potential drug for treatment of vitiligo.

Vitiligo is a complex disorder characterized by patchy loss of skin pigmentation due to abnormal melanocyte function. Overwhelming evidences have suggested that oxidative stress plays a major role in the loss of melanocytes thereby mediating the onset and progression of vitiligo. The nuclear factor erythroid 2-like factor 2 (Nrf2) is a master regulator of cellular redox homeostasis and the activation of Nrf2 signaling pathway is impaired in the vitiligo melanocytes. Baicalein, as flavonoid extracted from the Scutellaria baicalensis, has been proved to possess the ability to activate Nrf2 signaling pathway in other cell types and mouse model. Our previous data found that baicalein exerts a cytoprotective role in H2O2-induced apoptosis in human melanocytes cell line (PIG1). Based on these founding, we hypothesized that baicalein activates Nrf2 signaling pathway, alleviates H2O2-induced mitochondrial dysfunction and cellular damage, thereby protecting human vitiligo melanocytes from oxidative stress. In the present study, we found that baicalein effectively inhibited H2O2-induced cytotoxicity and apoptosis in human vitiligo melanocytes (PIG3V). Further results demonstrated that baicalein promoted Nrf2 nucleus translocation as well as up-regulated the expression of Nrf2 and its target gene, heme oxygenase-1 (HO-1). Moreover, the protective effects of baicalein against H2O2-induced cellular damage and apoptosis as well as mitochondrial dysfunction were abolished by Nrf2 knockdown. Additionally, we observed that Nrf2 knockdown suppressed proliferation and increased the sensitivity of PIG3V cells to H2O2 treatment. Finally, we explored the mechanism of baicalein associated with Nrf2 activation and found that the phosphorylation of Nrf2 as well as ERK1/2and PI3K/AKT signaling were not involved in the baicalein-induced activation of Nrf2. Taken together, these data clearly suggest that baicalein enhances cellular antioxidant defense capacity of human vitiligo melanocytes through the activation of the Nrf2 signaling pathway, providing beneficial evidence for the application of baicalein in the vitiligo treatment.

The prevention of hydrogen peroxide (H2O2)-induced oxidative stress has proved to be beneficial to vitiligo patients. Simvastatin possesses antioxidative capacity and has shown protective effect in various oxidative stress-related diseases. However, whether simvastatin can protect human melanocytes against oxidative stress has not been investigated. In this study, we initially found that pretreatment with 0.1 μmol/L to 1.0 μmol/L simvastatin led to increased cell viability and decreased cell apoptosis of melanocytes in response to H2O2. In addition, simvastatin was able to potentiate the activity of antioxidant enzymes and lessen intracellular reactive oxygen species accumulation. Furthermore, we found that simvastatin promoted the activation of nuclear erythroid 2-related factor (Nrf2) and that knockdown of Nrf2 abolished the protective effect of simvastatin against H2O2-induced oxidative damage. More importantly, the mutual enhancement between mitogen-activated protein kinase pathways and p62 contributed to simvastatin-induced Nrf2 activation in melanocytes. Finally, simvastatin showed more antioxidative capacity and better protective effect than aspirin in H2O2-treated melanocytes. Taken together, our results show that simvastatin protects human melanocytes from H2O2-induced oxidative stress by activating Nrf2, thus supporting simvastatin as a potential therapeutic agent for vitiligo.

Oxidative stress plays important roles in the pathogenesis of vitiligo. The removal of hydrogen peroxided (HIn the present study, we investigated the potential protective effect of BBR against oxidative stress on an immortalized normal human melanocyte cell line PIG1.Generally, PIG1 cells were pretreated with various concentrations of BBR for 1 h followed by exposure to 1.0 mM HOur results showed that cell viability rose and intracellular ROS generation, cell apoptosis of melanocytes decreased significantly in response to HThe results above suggest that BBR can protect melanocytes against oxidative stress via its anti-oxidative activity. Also, we found H

In vitiligo, melanocytes are particularly vulnerable to oxidative stress owing to the pro-oxidant state generated during melanin synthesis and to the genetic antioxidant defects. Autophagy is a controlled self-digestion process which can protect cells against oxidative damage. However, the exact role of autophagy in vitiligo melanocytes in response to oxidative stress and the mechanism involved are still not clear. To determine the implications of autophagy for melanocyte survival in response to oxidative stress, we first detected the autophagic flux in normal melanocytes exposure to H2O2, and found that autophagy was significantly enhanced in normal melanocytes, for protecting cells against H2O2-induced oxidative damage. Nevertheless, vitiligo melanocytes exhibited dysregulated autophagy and hypersensitivity to H2O2-induced oxidative injury. In addition, we confirmed that the impairment of Nrf2-p62 pathway is responsible for the defects of autophagy in vitiligo melanocytes. Noteworthily, upregulation of the Nrf2-p62 pathway or p62 reduced H2O2-induced oxidative damage of vitiligo melanocytes. Therefore, our data demonstrated that dysregulated autophagy owing to the impairment of Nrf2-p62 pathway increase the sensitivity of vitiligo melanocytes to oxidative stress, thus promote the development of vitiligo. Upregulation of p62-dependent autophagy may be applied to vitiligo treatment in the future.

The prevention of hydrogen peroxide (H

In vitiligo, melanocytes are particularly vulnerable to oxidative stress owing to the pro-oxidant state generated during melanin synthesis and to the genetic antioxidant defects. Autophagy is a controlled self-digestion process which can protect cells against oxidative damage. However, the exact role of autophagy in vitiligo melanocytes in response to oxidative stress and the mechanism involved are still not clear. To determine the implications of autophagy for melanocyte survival in response to oxidative stress, we first detected the autophagic flux in normal melanocytes exposure to H2O2, and found that autophagy was significantly enhanced in normal melanocytes, for protecting cells against H2O2-induced oxidative damage. Nevertheless, vitiligo melanocytes exhibited dysregulated autophagy and hypersensitivity to H2O2-induced oxidative injury. In addition, we confirmed that the impairment of Nrf2-p62 pathway is responsible for the defects of autophagy in vitiligo melanocytes. Noteworthily, upregulation of the Nrf2-p62 pathway or p62 reduced H2O2-induced oxidative damage of vitiligo melanocytes. Therefore, our data demonstrated that dysregulated autophagy owing to the impairment of Nrf2-p62 pathway increase the sensitivity of vitiligo melanocytes to oxidative stress, thus promote the development of vitiligo. Upregulation of p62-dependent autophagy may be applied to vitiligo treatment in the future.