ALA-PDT promotes IL-1ß secretion from human SZ95 sebocytes via activation of the NLRP3 inflammasome.

BACKGROUND: 5-Aminolevulinic acid photodynamic therapy (ALA-PDT) is an effective treatment for pilosebaceous inflammatory diseases, such as acne vulgaris. In this study, we explored ALA-PDT's mechanisms against acne in vitro. METHODS: We treated human SZ95 sebocytes with ALA (0.2 mM) and subjected them to varied PDT doses (0, 5, 10, 20 J/cm²) over 12 h. We assessed cell viability post-treatment using the Annexin V FITC/PI apoptosis kit. ROS accumulation in the sebocytes was detected with a DCFDA probe. We quantified NLRP3 and caspase-1 mRNA via quantitative PCR and determined IL-1ß release following ALA-PDT by ELISA. Western blotting helped identify the levels of proteins associated with pyroptosis (NLRP3, caspase-1, and IL-1ß). To elucidate the mechanisms, we re-evaluated these parameters after administering various concentrations of NAC antioxidants (0, 0.4, 2, 10 mM) and the caspase inhibitor Z-VAD-FMK (0, 5, 10, 20 µM). RESULTS: Increasing PDT dose inversely affected SZ95 sebocyte survival, with a corresponding rise in ROS and pyroptosis-related proteins (NLRP3, caspase-1, and IL-1ß). Furthermore, NAC and Z-VAD-FMK modulated the expression and secretion of these molecules in a dose-responsive manner. CONCLUSION: Our findings suggest ALA-PDT's potential mechanism of action on sebaceous glands could involve ROS induction, leading to NLRP3 inflammasome assembly, thereby heightening caspase-1 activation and IL-1ß secretion. This cascade may amplify the local inflammatory response to break chronic inflammation in acne vulgaris treatment.

Urolithin A protects human dermal fibroblasts from UVA-induced photoaging through NRF2 activation and mitophagy.

Photoaging, caused by exposure to sunlight and especially UVA, has been identified as one of the culprits for age-related skin deterioration. Here, we initially demonstrated that urolithin A (UroA), a metabolite derived from intestine microflora, possessed sufficient photoprotective capacity and attenuated UVA-induced senescent phenotypes in human fibroblasts, such as growth inhibition, senescence-associated ß-galactosidase activity, breakdown of extracellular matrix, synthesis of senescence-associated secretory phenotypes and cell cycle arrest. Furthermore, UroA lessened the accumulation of intracellular reactive oxygen species, which promoted the phosphorylation and afterwards nuclear translocation of NRF2, subsequently driving the activation of downstream antioxidative enzymes. In parallel, we proved that UroA restored mitochondrial function by induction of mitophagy, which was regulated by the SIRT3-FOXO3-PINK1-PARKIN network. Taken together, our results showed that UroA protected dermal fibroblast from UVA damage through NRF2/ARE activation and mitophagy process, thus supporting UroA as a potential therapeutic agent for photoaging.

Hydrogen Sulfide Promotes Cell Proliferation and Melanin Synthesis in Primary Human Epidermal Melanocytes.

BACKGROUND/AIM: Hydrogen sulfide (H2S) has been found to act as a physiological intercellular messenger to regulate cell survival. In this study, we evaluated whether H2S could promote cell proliferation and melanin synthesis in human epidermal melanocytes (HEMs). METHODS: Primary HEMs were cocultured with sodium hydrosulfide (NaHS, the most widely used H2S donor) or endogenously overexpressed with cystathionine-γ-lyase (CSE) gene, which is the most predominant H2S-producing enzyme. Then, cell viability, intracellular melanin content, tyrosinase (TYR) activity, and expression of microphthalmia-associated transcription factor (MITF), TYR, together with TYR-related protein 1 (TRP-1) in both transcript and protein levels, were detected. RESULTS: We first confirmed that NaHS (10-100 µm) increased cell viability, intracellular melanin content, and TYR activity in a dose-dependent manner. Then, we found that endogenous H2S production also promoted cell proliferation, intracellular melanin content, and TYR activity. In addition, we observed the mRNA and protein expression of MITF, TYR, and TRP-1 was significantly up-regulated after NaHS treatment and CSE gene transfection. CONCLUSIONS: This study demonstrates that H2S promotes cell proliferation and melanin synthesis in HEMs, which indicates pharmacologic regulation of H2S may be potential treatment for skin disorders caused by loss of melanocytes or dysfunction of melanogenesis.

Dysfunction of ATG7-dependent autophagy dysregulates the antioxidant response and contributes to oxidative stress-induced biological impairments in human epidermal melanocytes.

Autophagy is a process involving the self-digestion of components that participates in anti-oxidative stress responses and protects cells against oxidative damage. However, the role of autophagy in the anti-oxidative stress responses of melanocytes remains unclear. To investigate the role of autophagy in human epidermal melanocytes, we knocked down and overexpressed ATG7, the critical gene of autophagy, in normal human epidermal melanocytes. We demonstrated that ATG7-dependent autophagy could affect melanin content of melanocytes by regulating melanogenesis. Moreover, suppression of ATG7-dependent autophagy inhibits proliferation and promotes oxidative stress-induced apoptosis of melanocytes, whereas enhancement of ATG7-dependent autophagy protects melanocytes from oxidative stress-induced apoptosis. Meanwhile, deficiency of ATG7-dependent autophagy results in premature senescence of melanocytes under oxidative stress. Notably, we verified that ATG7-dependent autophagy could alter oxidative stress homeostasis by regulating reactive oxygen species (ROS) production, nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway, and the activity of several antioxidant enzymes in melanocytes. In conclusion, our study suggested that ATG7-dependent autophagy is indispensable for redox homeostasis and the biological functions of melanocytes, such as melanogenesis, proliferation, apoptosis, and senescence, especially under oxidative stress.

Exogenous hydrogen sulfide inhibits human melanoma cell development via suppression of the PI3K/AKT/ mTOR pathway.

BACKGROUND: Melanoma is one of the most aggressive, therapy-resistant skin cancers in the world. Hydrogen sulfide (H2S), a newly discovered gasotransmitter, plays a crucial role in the progression and development of many types of cancers. However, the effect of H2S on human skin melanoma remains to be elucidated. OBJECTIVE: We aimed to explore the effect of exogenous H2S on melanoma cells and its underlying mechanisms. METHODS: In this study, human skin melanoma cell lines, including A375 and SK-MEL-28, were treated with a donor of H2S (NaHS). CCK-8, scratch assay, flow cytometric analysis, western blotting and transmission electron microscopy (TEM) were performed to explore the effects of H2S on cell behaviors. RESULTS: Treatment with NaHS inhibited cell proliferation, migration and division, while it could induce cell apoptosis and autophagy in melanoma cell lines. Moreover, NaHS significantly decreased the expression of p-PI3K, p-Akt and mTOR proteins. Furthermore, insulin-like growth factor-1 (IGF-1), the activator of PI3K/AKT/mTOR pathway, could reverse the cell behaviors caused by NaHS. CONCLUSION: Our results demonstrated that exogenous hydrogen sulfide could inhibit human melanoma cell development via suppression of the PI3K/AKT/mTOR pathway. Hydrogen sulfide might serve as a potential therapeutic option for melanoma.

The biologic effect of hydrogen sulfide and its function in various diseases.

INTRODUCTION: Hydrogen sulfide (H2S), a colorless, water soluble, flammable gas with a characteristic smell of rotten eggs, has been known as a highly toxic gas for several years. However, much like carbon monoxide (CO) and nitric oxide (NO), the initial negative perception of H2S has developed with the discovery that H2S is generated enzymatically in animals under normal conditions. With the result of this discovery, much more work is needed to elucidate the biologic effects of H2S. In recent years, its cytoprotective properties have been recognized in multiple organs and tissues. In particular, H2S plays important roles in combating oxidative species such as reactive oxygen species (ROS) and reactive nitrogen species (RNS) and protect the body from oxidative stress. Therefore, this review discusses the biologic effect of H2S and how it protects cells in various diseases by acting as an antioxidant that reduces excessive amounts of ROS and RNS. ETHICS AND DISSEMINATION: Ethical approval and informed consent are not required, as the study will be a literature review and will not involve direct contact with patients or alterations to patient care. CONCLUSION: H2S has been found to be cytoprotective in oxidative stress in a wide range of physiologic and pathologic conditions, an increasing number of therapeutic potentials of H2S also have been revealed. However, there is still much debate on the clear mechanism of action of H2S, so that the mechanisms of cell signaling that promote cellular survival and organ protection need to be further investigated to provide better H2S-based therapeutics.

ALA-PDT suppressed the cell growth by Akt-/Erk-mTOR-p70 s6k pathway in human SZ95 sebocytes in vitro.

BACKGROUND: Topical 5-aminolevulinic acid mediated photodynamic therapy (PDT) is known to be an effective method in treating acne vulgaris and other sebaceous gland-related diseases. The therapeutic mechanisms of ALA-PDT still remain undetermined. Our team has reported that ALA-PDT suppressed the cell growth in SZ95 sebocytes by mTOR-p70 S6K signaling. In this study, we aimed to investigate upstream of the mammalian target of rapamycin (mTOR) signaling cascade after ALA-PDT on cell growth of human SZ95 sebocytes. MATERIAL AND METHODS: Human SZ95 sebocytes were treated with different concentration of 5-ALA PDT. Western blotting was used to detect and analyze the protein expression level of P-Akt (T308)/Akt, P-Akt (S473)/Akt, P-Erk/Erk, P-AMPKα (T172)/AMPK, P-AMPKα1 (S485)/AMPKα2 (S491)/AMPK, P-PRAS40/PRAS40, RagC. Meanwhile, mTOR pathway activator IGF-1 and mTORC1 inhibitor rapamycin were added to observe the interferences of P-p70 S6K/p70 S6K after ALA-PDT. RESULTS: mTOR pathway inhibitor rapamycin decreased the level of P-p70 S6K reduced by ALA-PDT. Conversely, mTOR pathway activator IGF-1. ALA-PDT reduced the level of P-Akt (T308), P-Erk, P-AMPKα (T172), P-AMPKα1 (S485)/AMPKα2 (S491) and P-PRAS40, and no change was observed in the level of Rag C. CONCLUSION: ALA-PDT suppresses the cell growth in SZ95 cells through Akt-/Erk- mTOR -p70 s6k pathway rather than PRAS40-/RagC- mTOR pathway.

ALA-PDT suppressing the cell growth and reducing the lipogenesis in human SZ95 sebocytes by mTOR signaling pathway in vitro.

BACKGROUND: 5-Aminolevulinic acid mediated -photodynamic therapy (ALA-PDT) is known to be effective in treating acne vulgaris and other sebaceous gland-related diseases. However, the therapeutic mechanisms of ALA-PDT still remain undetermined. In this study, we aimed to investigate the effects and mechanisms of ALA-PDT on the cell growth and lipogenesis of human SZ95 sebocytes. MATERIAL AND METHODS: Human SZ95 sebocytes were treated with different concentration of ALA-PDT.CCK-8 assay was used to detect cell proliferation activity. Fluorescence microscope and flow cytometry were used to observe the secretion of lipids in SZ95 cells after Nile red staining. Western blotting was used to detect and analyze the protein expression level of P-p70 S6K/p70 S6K, P-4E-BP1/4E-BP1, SREBP-1, PPARγ, P-mTOR/mTOR, and P-Raptor/Raptor. Mean while, mTOR pathway activator IGF-1 and mTORC1 inhibitor rapamycin were added to observe the interferences on the ALA-PDT treatment of SZ95 cells. RESULTS: ALA-PDT suppressed the cell growth and reduced the secretion of lipids in a dose-dependent manner in SZ95 cells. ALA-PDT reduced the protein levels of P-p70 S6K (T389), SREBP-1, PPARγ, P-mTOR and P-Raptor. IGF-1 had counter effects on ALA-PDT, and rapamycin enhanced the effects of ALA-PDT in SZ95 cells in suppressing the cell growth and reducing the secretion of lipids. CONCLUSION: ALA-PDT suppressed the cell growth in SZ95 cells by mTOR-p70 S6K(T389) signaling and reduced the lipogenesis in SZ95 cells by mTOR-SREBP-1/PPARγ signaling. Sebaceous glands atrophy and reduction of sebum secretion after ALA-PDT may be caused by the suppression of lipogenesis and cell growth in sebocytes.