Tacrolimus reverses pemphigus vulgaris serum-induced depletion of desmoglein in HaCaT cells via inhibition of heat shock protein 27 phosphorylation.

BACKGROUND: Glucocorticoids are the first-line treatment for Pemphigus vulgaris (PV), but its serious side effects can be life-threatening for PV patients. Tacrolimus (FK506) has been reported to have an adjuvant treatment effect against PV. However, the mechanism underlying the inhibitory effect of FK506 on PV-IgG-induced acantholysis is unclear. OBJECTIVE: The objective of this study was to explore the effect of FK506 on desmoglein (Dsg) expression and cell adhesion in an immortalized human keratinocyte cell line (HaCaT cells) stimulated with PV sera. METHODS: A cell culture model of PV was established by stimulating HaCaT cells with 5% PV sera with or without FK506 and clobetasol propionate (CP) treatment. The effects of PV sera on intercellular junctions and protein levels of p38 mitogen-activated protein kinase (p38MAPK), heat shock protein 27 (HSP27), and Dsg were assayed using western blot analysis, immunofluorescence staining, and a keratinocyte dissociation assay. RESULTS: PV sera-induced downregulation of Dsg3 was observed in HaCaT cells and was blocked by FK506 and/or CP. Immunofluorescence staining revealed that linear deposits of Dsg3 on the surface of HaCaT cells in the PV sera group disappeared and were replaced by granular and agglomerated fluorescent particles on the cell surface; however, this effect was reversed by FK506 and/or CP treatment. Furthermore, cell dissociation assays showed that FK506 alone or in combination with CP increased cell adhesion in HaCaT cells and ameliorated loss of cell adhesion induced by PV sera. Additionally, FK506 noticeably decreased the PV serum-induced phosphorylation of HSP 27, but had no effect on p38MAPK phosphorylation. CONCLUSION: FK506 reverses PV-IgG induced-Dsg depletion and desmosomal dissociation in HaCaT cells, and this effect may be obtained by inhibiting HSP27 phosphorylation.

Viaminate Inhibits Propionibacterium Acnes-induced Abnormal Proliferation and Keratinization of HaCat Cells by Regulating the S100A8/S100A9- MAPK Cascade.

BACKGROUND: Viaminate, a vitamin A acid drug developed in China, has been clinically used in acne treatment to regulate epithelial cell differentiation and proliferation, inhibit keratinization, reduce sebum secretion, and control immunological and anti-inflammatory actions; however, the exact method by which it works is unknown. METHODS: In the present study, acne was induced in the ears of rats using Propionibacterium acnes combined with sebum application. RESULTS: After 30 days of treatment with viaminate, the symptoms of epidermal thickening and keratin overproduction in the ears of rats were significantly improved. Transcriptomic analysis of rat skin tissues suggested that viaminate significantly regulated the biological pathways of cellular keratinization. Gene differential analysis revealed that the S100A8 and S100A9 genes were significantly downregulated after viaminate treatment. The results of qPCR and Western blotting confirmed that viaminate inhibited the expression of S100A8 and S100A9 genes and proteins in rat and HaCat cell acne models, while its downstream pathway MAPK (MAPK p38/JNK/ERK1/2) protein expression levels were suppressed. Additional administration of the S100A8 and S100A9 complex protein significantly reversed the inhibitory effect of viaminate on abnormal proliferation and keratinization levels in acne cell models. CONCLUSION: In summary, viaminate can improve acne by modulating S100A8 and S100A9 to inhibit MAPK pathway activation and inhibit keratinocyte proliferation and keratinization levels.

CK2 inhibitor CX4945 inhibits collagen degradation of HaCaT human keratinocyte cells via attenuation of MMP-1 secretion.

INTRODUCTION: During skin aging, the extracellular matrix (ECM) concomitantly breaks down. Out of the various protein components that comprise ECM, collagen is the most abundant one. Matrix metalloproteinase-1 (MMP-1) is a major collagenase that can degrade collagen. Therefore, the inhibition of MMP-1 may be critical for skin aging prevention. CX4945 is an inhibitor of casein kinase 2 and shows anticancer effects on various types of cancer cells. METHODS AND RESULTS: In this report, we investigated the MMP-1-inhibiting effect of CX4945 in HaCaT human keratinocyte cells. We performed zymography assays, Western blot analysis and immunoprecipitation assay to investigate the anti-MMP-1 effects of CX4945. CX4945 was found to inhibit collagen degradation via attenuation of the MMP-1 secretion out of HaCaT cells. This activity of CX4945 may be mediated by the induction of MMP-1 ubiquitylation via c-Jun N-terminal kinase (JNK) signaling. In wound healing cell migration assay, CX4945 also showed suppressive effect on the migration of HaCaT cells. This finding was closely related to the attenuation of CREB transcription factor via the downregulation of ERK mitogen-activated protein kinase as observed in Western blot analysis. CONCLUSION: Our report suggests that the inhibitory effects of CX4945 on MMP-1 in epidermal cells may offer a basis for further studying its therapeutic potential as an anti-wrinkle agent.

Anti-inflammatory, anti-proliferative and anti-psoriatic potential of apigenin in RAW 264.7 cells, HaCaT cells and psoriasis like dermatitis in BALB/c mice.

AIMS: Psoriasis is an immune-mediated skin disease characterized by keratinocytes hyperproliferation, abnormal differentiation and inflammation. Therefore, this study aimed to investigate in-vitro and in-vivo anti-inflammatory and anti-proliferative activity to evaluate anti-psoriatic potential of apigenin. MAIN METHODS: For in-vivo study, 5 % imiquimod cream was used to induce psoriasis-like skin inflammation in BALB/c mice to mimic human psoriatic conditions. PASI score, CosCam score, histopathology, immunohistochemistry, qRT-PCR, and ELISA were done to evaluate the anti-psoriatic potential of topically applied apigenin. For in-vitro studies, LPS-induced inflammation in RAW 264.7 was done, and qRT-PCR, ELISA, and immunofluorescence were conducted to evaluate the anti-inflammatory activity of apigenin. Migration and cell doubling assay in HaCaT cells were performed to assess the anti-proliferative effect of apigenin. Acute dermal toxicity profile of apigenin has also been done as per OECD guidelines. KEY FINDINGS: Results showed that apigenin significantly reduce the PASI and CosCam scores, ameliorate the deteriorating histopathology, and effectively downregulated the expression of CCR6, IL-17A, and NF-κB. Apigenin effectively downregulated the expression and secretion of pro-inflammatory cytokines through IL-23/IL-17/IL-22 axis. Apigenin suppressed nuclear translocation of NF-κB in LPS-induced RAW 264.7 cells. Cell migration and cell doubling assay in HaCaT cells showing the anti-proliferative potential of apigenin and it was found safe in acute dermal toxicity study. SIGNIFICANCE: Apigenin was found effective against psoriasis in both in-vitro and in-vivo models suggesting apigenin as a potential candidate for the development of anti-psoriatic agent.

Rg3-loaded P407/CS/HA hydrogel inhibits UVB-induced oxidative stress, inflammation and apoptosis in HaCaT cells.

UVB radiation can damage human skin, whereas Ginsenoside Rg3, the active ingredient in red ginseng that is processed from ginseng (Panax ginseng C.A. Meyer), could inhibit UVB induced cell damage through anti-oxidation. Meanwhile, P407/CS/HA hydrogel has significant biomedical applications as carriers of drugs. However, the beneficial effects of Rg3-loaded hydrogel (Rg3-Gel) on human HaCaT keratinocytes induced by UVB have rarely been reported. In our study, Rg3 was loaded into hydrogel and the effect of Rg3-Gel against UVB­induced Hacat cells damages was determined by measuring its ability to alleviate UVB­induced elevation of oxidative stress, pro-inflammatory and apoptotic response. We found that the treatment with Rg3-Gel inhibited the generation of intracellular ROS and MDA and upregulated the expression of antioxidant enzymes SOD and GSH-Px which were inhibited by UVB exposure. Increased levels of pro-inflammatory cytokines TNF­α, COX­2, iNOS and IL­1ß following UVB irradiation were suppressed by the introduction of Rg3-Gel. Additionally, the level of Bcl-2 was decreased and the expression of Bax and Caspase3 were enhanced by Rg3-Gel treatment. In conclusion, Rg3-Gel equipped with the synergistic effect of Rg3 and hydrogel has an effective inhibitory effect on UVB-induced oxidative stress, inflammatory and apoptosis.

Hovenia dulcis Thunb. Fruit Extract Attenuates Psoriatic Skin Inflammation in Tumor Necrosis Factor-α-Stimulated Human Keratinocyte HaCaT Cells In Vitro.

Hovenia dulcis Thunb. fruit (HDF) is traditionally used for treating liver diseases and alcohol poisoning. The purpose of this study was to explore the effects of HDF on hyperproliferation, levels of inflammatory cytokines, and signaling mechanisms in human psoriatic keratinocyte HaCaT cells. HDF showed a preventive effect on tumor necrosis factor-α (TNF-α)-induced abnormal proliferation of psoriatic keratinocytes. Furthermore, real-time reverse transcription-PCR analysis showed that HDF suppressed the expressions of inflammatory cytokines; interleukin (IL)-1α and IL-1ß and chemokines; CCL-20 and CXCL-8 in TNF-α-induced HaCaT cells. Western blotting revealed that HDF suppressed the levels of phosphorylated IκB and STAT3 together with a decline in the levels of phosphorylated mitogen-activated protein kinases (MAPKs). These outcomes indicate that HDF prevents the abnormal proliferation of keratinocytes and modulates inflammatory responses by suppressing nuclear factor-κB (NF-κB) and STAT3 activation through downregulation of the MAPK signaling pathway in TNF-α-induced psoriatic keratinocytes. Our study demonstrates that HDF is prospective and beneficial for psoriatic skin inflammation.

Qinzhu Liangxue inhibits IL-6-induced hyperproliferation and inflammation in HaCaT cells by regulating METTL14/SOCS3/STAT3 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Psoriasis, an immune-mediated chronic inflammatory skin condition, is treatable with Qinzhu Liangxue (QZLX), a therapeutic medicinal plant formula used in clinical practice. However, further investigation is needed to clarify its molecular mechanisms of action. AIM OF THE STUDY: The potential biological mechanisms of QZLX to alleviate psoriasis involving IL-6-induced hyperproliferation and inflammation by regulating METTL14/SOCS3/STAT3 axis. MATERIALS AND METHODS: HaCaT cell model was induced by IL-6, and dealt with serum containing QZLX. In addition, shRNAs and siRNAs were used for gene silencing, viruses were collected 48 h post-transfection and infected HaCaT cells. Cell viability was detected by CCK-8 assay, cell cycle was determined by flow cytometry. Finally, psoriasis mice model was induced by IMQ cream, then back skin tissue was used for hematoxylin and eosin (H&E). The content of IL-1ß, IL-6, and IL-8 in cell supernatants were analyzed using ELISA kits. Analysis of SOCS3 was used by quantitative RT-PCR, the expression level of SOCS3, METTL3, METTL14, WTAP, SOCS3, YTHDF2, p-STAT3 and STAT3 in HaCaT cells transduced with METTL14 overexpression was detected by Western blot. RESULTS: All results indicated that QZLX could significantly alleviate IL-6-induced HaCaT cell viability, cell cycle progression, and inhibit the level of IL-1ß, IL-6, and IL-8. The m6A levels and level of METTL14 in HaCaT cells treated with IL-6 were enhanced, while it was reversed by QZLX. METTL14 silencing could inhibit IL-6-induced HaCaT cell viability, cell cycle progression and inflammation response, while SOCS3 overexpression also suppressed METTL14-induced HaCaT cell viability, cell cycle progression and inflammation. QZLX could significantly enhance the expression level of SOCS3, while inhibit the level of METTL14, and p-STAT3/STAT3. In addition, QZLX inhibits METTL14-induced HaCaT cell viability, cell cycle progression, and inhibits the level of IL-1ß, IL-6, and IL-8. CONCLUSIONS: Our finding suggested that QZLX ameliorated the inflammation response of psoriasis and performed the potential anti-psoriasis effect by regulating METTL14/SOCS3/STAT3 axis in both mice and HaCaT cells psoriasis model. Therefore, our study demonstrated a significant strategy for inhibiting psoriasis inflammation via targeting METTL14/SOCS3/STAT3 axis.

The expression of prolyl isomerase Pin1 is expanded in the skin of patients with atopic dermatitis and facilitates IL-33 expression in HaCaT cells.

Atopic dermatitis (AD) is attributable to both a genetic predisposition and environmental factors. Among numerous cytokines involved in the pathogenesis of AD, interleukin-33 (IL-33), reportedly escaping exocytotically in response to a scratch, is abundantly expressed in the skin tissues of patients with AD and is postulated to induce inflammatory and autoimmune responses. In this study, we first demonstrated that peptidylprolyl cis/trans isomerase, NIMA-interacting 1 (Pin1), a unique enzyme which isomerizes the proline residues of target proteins, is abundantly expressed in keratinocytes, and that the areas where it is present in the skin tissues of AD patients became expanded due to hyperkeratosis. Thus, we investigated the effects of Pin1 on the regulation of IL-33 expression using the human keratinocyte cell line HaCaT. Interestingly, silencing of the Pin1 gene or treatment with Pin1 inhibitors dramatically reduced IL-33 expressions in HaCaT cells, although Pin1 overexpression did not elevate it. Subsequently, we showed that Pin1 binds to STAT1 and the nuclear factor-kappaB (NF-κB) subunit p65. Silencing the Pin1 gene with small interfering RNAs significantly reduced the phosphorylation of p65, while no marked effects of Pin1 on the STAT1 pathway were detected. Thus, it is likely that Pin1 contributes to increased expression of IL-33 via the NF-κB subunit p65 in HaCaT cells, at least modestly. Nevertheless, further study is necessary to demonstrate the pathogenic roles of Pin1 and IL-33 in AD development.

Reduction of DNA damage repair efficiency and accumulation of residual damage following chronic UVB-irradiation of HaCaT cells.

Absorption of ultraviolet radiation (UVR) by DNA leads to the predominant formation of cyclobutane pyrimidine dimers (CPD). Since those CPD are responsible for the driver mutations found in skin cancers, their efficient repair is critical. We previously showed that pre-stimulation of fibroblasts with chronic low doses of UVB (CLUV) increases CPD repair efficiency. Since skin cancers are not arising from dermal fibroblasts, this observation is not directly relevant to cutaneous carcinogenesis. We have now exposed HaCaT keratinocytes to a CLUV irradiation protocol to determine whether this pre-stimulation influences CPD removal rate. Similar to fibroblasts, CLUV treatment leads to the accumulation of residual CPD in keratinocytes, which are not repaired but rather tolerated and diluted through DNA replication. In contrast to fibroblasts, in keratinocytes we find that CLUV pre-treatment reduces CPD removal of newly generated damage without inducing a higher sensitivity to UVR-induced cell death. Using our experimental data, we derived a theoretical model to predict CPD induction, dilution and repair that occur in keratinocytes when chronically UVB-irradiated. Altogether, these results suggest that the accumulation of unrepaired CPD and the reduction in repair efficiency caused by chronic UVB exposure might lead to an increase in skin cancer driver mutations.

Photoprotective Effects of Dendrobium nobile Lindl. Polysaccharides against UVB-Induced Oxidative Stress and Apoptosis in HaCaT Cells.

Acute ultraviolet (UV)-B radiation is the major external factor causing photodamage. In this study, we aimed to determine the effects of Dendrobium nobile Lindl. polysaccharides (DNPs) on photodamage in HaCaT keratinocytes after UVB irradiation and the underlying mechanisms. We found that DNPs significantly attenuated the decline in the viability and proliferation of HaCaT cells after UVB irradiation. Moreover, DNPs scavenged reactive oxygen species (ROS), improved the activities of endogenous antioxidant enzymes, including superoxide dismutase, catalase, and glutathione peroxidase, and reduced the levels of malondialdehyde, while partially attenuating cell cycle arrest, suggesting their antioxidant and anti-apoptotic properties. The mitogen-activated protein kinase (MAPK) pathway was found to be important for the attenuation of UVB-induced photodamage in the HaCaT cells. Furthermore, DNPs exerted cytoprotective effects by downregulating UVB-induced ROS-mediated phosphorylation of MAPKs, including p38, c-Jun N-terminal kinase, and extracellular signal-regulated kinase, and by inhibiting p53 expression as well as the apoptotic cascade response. Therefore, DNPs ameliorated UVB-induced oxidative damage and apoptosis in HaCaT cells via the regulation of MAPKs. Our findings thus highlight the Dendrobium nobile Lindl polysaccharides as promising therapeutic candidates for UVB-induced photodamage.

Increased mitochondrial fission induces NLRP3/cGAS-STING mediated pro-inflammatory pathways and apoptosis in UVB-irradiated immortalized human keratinocyte HaCaT cells.

Ultraviolet B (UVB) irradiation causes skin inflammation and apoptosis. Mitochondria are highly dynamic and undergo constant fusion and fission that are essential for maintaining physiological functions of cells. Although dysfunction of mitochondria has been implicated in skin damages, little is known about the roles of mitochondrial dynamics in these processes. UVB irradiation increases abnormal mitochondrial content but decreases mitochondrial volume in immortalized human keratinocyte HaCaT cells. UVB irradiation resulted in marked upregulation of mitochondrial fission protein dynamin-related protein 1 (DRP1) and downregulation of mitochondrial outer membrane fusion proteins 1 and 2 (MFN1 and MFN2) in HaCaT cells. Mitochondrial dynamics was discovered to be crucial for NLRP3 inflammasome and cGAS-STING pathway activation, as well as the induction of apoptosis. Inhibition of mitochondrial fission by treatments with a DRP1 inhibitor, mdivi-1, or with DRP1-targeted siRNA, efficiently prevented UVB-induced NLRP3/cGAS-STING mediated pro-inflammatory pathways or apoptosis in the HaCaT cells, whereas inhibition of mitochondrial fusion with MFN1and 2 siRNA increased these pro-inflammatory pathways or apoptosis. The enhanced mitochondrial fission and reduced fusion caused the up-regulation of reactive oxygen species (ROS). Application of an antioxidant, N-acetyl-l-cysteine (NAC), which scavenges excessive ROS, attenuated inflammatory responses through suppressing NLRP3 inflammasome and cGAS-STING pathway activation, and rescued cells from apoptosis caused by UVB-irradiation. Together, our findings revealed the regulation of NLRP3/cGAS-STING inflammatory pathways and apoptosis by mitochondrial fission/fusion dynamics in UVB-irradiated HaCaT cells, providing a new strategy for the therapy of UVB skin injury.

ATP supplementation suppresses UVB-induced photoaging in HaCaT cells via upregulation of expression of SIRT3 and SOD2.

BACKGROUND: Skin photoaging is the damage caused by excessive exposure to ultraviolet (UV) irradiation. We investigated the effect of adenosine triphosphate (ATP) supplementation on UVB-induced photoaging in HaCaT cells and its potential molecular mechanism. MATERIALS AND METHODS: The toxicity of ATP on HaCaT cells was examined by the MTT assay. The effects of ATP supplementation on the viability and apoptosis of HaCaT cells were determined by crystal-violet staining and flow cytometry, respectively. Cellular and mitochondrial ROS were stained using fluorescent dyes. Expression of Bax, B-cell lymphoma (Bcl)-2, sirtuin (SIRT)3, and superoxide dismutase (SOD)2 was measured via western blotting. RESULTS: ATP (1, 2 mM) exerted no toxic effect on the normal growth of HaCaT cells. UVB irradiation caused the apoptosis of HaCaT cells, and ATP supplementation inhibited the apoptosis induced by UVB significantly, as verified by expression of Bax and Bcl-2. UVB exposure resulted in accumulation of cellular and mitochondrial reactive oxygen species (ROS), but ATP supplementation suppressed these increases. Expression of SIRT3 and SOD2 was decreased upon exposure to UVB irradiation but, under ATP supplementation, expression of SIRT3 and SOD2 was reversed, which was consistent with the reduction in ROS level observed in ATP-treated HaCaT cells after exposure to UVB irradiation. CONCLUSIONS: ATP supplementation can suppress UVB irradiation-induced photoaging in HaCaT cells via upregulation of expression of SIRT3 and SOD2.

Nanofabrication of cobalt-tellurium using Allium sativum extract and its protective efficacy against H2O2-induced oxidative damage in HaCaT cells.

Allium sativum (A. sativum)is well known for its therapeutic and culinary uses. Because of their high medicinal properties, the clove extract was selected to synthesize cobalt-tellurium nanoparticles. The aim of the study was to evaluate the protective activity of the nanofabricated cobalt-tellurium using A. sativum (Co-Tel-As-NPs) against H2O2-induced oxidative damage in HaCaT cells. Synthesized Co-Tel-As-NPs were analyzed using UV-Visible spectroscopy, FT-IR, EDAX, XRD, DLS, and SEM. Various concentrations of Co-Tel-As-NPs were used as a pretreatment on HaCaT cells before H2O2 was added. Then, the cell viability and mitochondrial damage were compared between pretreated and untreated control cells using an array of assays (MTT, LDH, DAPI, MMP, and TEM), and the intracellular ROS, NO, and antioxidant enzyme production were examined. In the present research, Co-Tel-As-NPs at different concentrations (0.5, 1.0, 2.0, and 4.0µg/mL) were tested for toxicity using HaCaT cells. Furthermore, the effect of H2O2 on the viability of HaCaT cells was evaluated using the MTT assay for Co-Tel-As-NPs. Among those, Co-Tel-As-NPs at 4.0 µg/mL showed notable protection; with the same treatment, cell viability was discovered to be 91% and LDH leakage was also significantly decreased. Additionally, the measurement of mitochondrial membrane potential was significantly decreased by Co-Tel-As-NPs pretreatment against H2O2. The recovery of the condensed and fragmented nuclei brought about by the action of Co-Tel-As-NPs was identified using DAPI staining. TEM examination of the HaCaT cells revealed that the Co-Tel-As-NPs had a therapeutic effect against H2O2 keratinocyte damage.

SPOP is essential for DNA replication licensing through maintaining translation of CDT1 and CDC6 in HaCaT cells.

Speckle-type pox virus and zinc finger (POZ) protein (SPOP), a substrate recognition receptor for the cullin-3/RING ubiquitin E3 complex, leads to the ubiquitination of >40 of its target substrates. Since a variety of point mutations in the substrate-binding domain of SPOP have been identified in cancers, including prostate and endometrial cancers, the pathological roles of those cancer-associated SPOP mutants have been extensively elucidated. In this study, we evaluated the cellular functions of wild-type SPOP in non-cancerous human keratinocyte-derived HaCaT cells expressing wild-type SPOP gene. SPOP knockdown using siRNA in HaCaT cells dramatically reduced cell growth and arrested their cell cycles at G1/S phase. The expression of DNA replication licensing factors CDT1 and CDC6 in HaCaT cells drastically decreased on SPOP knockdown as their translation was inhibited. CDT1 and CDC6 downregulation induced p21 expression without p53 activation. Our results suggest that SPOP is essential for DNA replication licensing in non-cancerous keratinocyte HaCaT cells.

Anti-psoriasis activities of hydroxytyrosol on HaCaT cells under psoriatic inflammation in vitro.

BACKGROUND: Psoriasis is a prevalent chronic inflammatory dermatosis, which can significantly impact life quality of patients. The treatment of psoriasis is no cure and novel therapeutic options are urgently needed. Hydroxytyrosol (HT) possesses multiple biological activities, such as anti-inflammatory and anti-proliferation properties, suggesting its potential to counteract hallmarks of psoriasis. However, its role in the regulation of psoriasis remains unknown. OBJECTIVE: In the current study, we explored the anti-proliferative activity and anti-inflammatory responses of HT in psoriatic keratinocytes in vitro. METHODS: We used M5 cytokines cocktail, which includes tumor necrosis factor (TNF)-α, oncostatin-M, interleukin (IL)-17A, IL-1α, and IL-22, to simulate HaCaT cells to establish the cell model of psoriasis and explore the effects of HT on psoriasis in vitro. RESULTS: This study showed that HT exerted potent anti-inflammatory effect via influencing the expression of IL-6, IL-8, and TNF-α in M5-induced cell model of psoriasis. Moreover, it suppressed the expression of antimicrobial proteins in psoriatic keratinocytes. Additionally, it inhibited cell proliferation in psoriasis cell model. CONCLUSIONS: Altogether, our results suggested that HT has anti-psoriasis effects in vitro and HT may be a promising therapeutic agent in psoriasis treatment.

Elevated SIRT3 Parkin-dependently activates cell mitophagy to ameliorate TNF-α-induced psoriasis-related phenotypes in HaCaT cells through deacetylating FOXO3a for its activation.

Sirtuin 3 (SIRT3) is reported to be closely relevant to the pathogenesis of psoriasis, but its detailed functions and molecular mechanisms have not been fully studied. Thus, this study aimed to investigate the effects and underlying mechanisms by which SIRT3 regulated the development of psoriasis. Specifically, we verified that SIRT3 was aberrantly downregulated in psoriasis-like skin tissues in mice models in vivo and TNF-α-stimulated HaCaT cells in vitro, compared to their corresponding normal counterparts. Functional experiments confirmed that upregulation of SIRT3 triggered cell mitophagy, restrained oxidative stress and inflammation, and inhibited excessive cell proliferation in the TNF-α-stimulated HaCaT cells in vitro, which were all ablated by co-treating cells with the mitophagy inhibitor 3-MA. Subsequently, the mechanism experiments uncovered that elevated SIRT3 deacetylated forkhead box class o 3A (FOXO3a) for its activation, which further activated the Parkin-dependent cell mitophagy in the HaCaT cells. Next, through performing the rescuing experiments, we validated that SIRT3 ameliorated TNF-α-induced psoriasis-associated phenotypes in the HaCaT cells via modulating the FOXO3a/Parkin signal pathway. Collectively, we concluded that SIRT3 triggered cell mitophagy through activating the FOXO3a/Parkin pathway to ameliorate TNF-α-induced psoriasis in the HaCaT cells, and this study provided evidences to support that SIRT3 could be used as important therapeutic target for the treatment of psoriasis.

Anti-inflammatory effects of amarogentin on 2,4-dinitrochlorobenzene-induced atopic dermatitis-like mice and in HaCat cells.

BACKGROUND: Amarogentin (AMA) is a secoiridoid glycoside extracted from Swertia and Gentiana roots and exhibits many biological effects such as antioxidative, anti-inflammatory, and antitumor activities. Atopic dermatitis (AD) is a chronic inflammatory skin disease caused by disorders in the regulation of multiple inflammatory cytokines. No effective cure has been found for AD now. METHODS: We constructed the HaCat and splenocyte model and tested the inhibitory effect of AMA on IL-4, IL-6, and IL-13 secretions using enzyme-linked immunosorbent assay (ELISA). The AD mouse model was constructed and treated with AMA, the severity of skin lesions was observed, epidermal tissue was collected, and epidermal thickness and mast cell infiltration were observed using hematoxylin and eosin and toluidine blue staining, respectively. The expression of kallikrein-related peptidase 7 (KLK7) and filaggrin (FLG) was detected using immunostaining and Western blot analysis. The mRNA expression of KLK7 and FLG was detected using quantitative polymerase chain reaction (qPCR). Blood immunoglobulin E (IgE) secretion was detected. RESULTS: AMA inhibited IL-6 secreted by tumor necrosis factor (TNF)-α-induced HaCaT cells and reduced IL-4 and IL-13 secreted by phytohemagglutinin (PHA)-induced primary cells in the mice spleen. It was found that the treatment of AMA with 2,4-dinitrochlorobenzene-induced AD-like mice could promote the recovery of dermatitis, reduce the score of dermatitis severity and the scratching frequency, treat the skin lesions, reduce the epidermal thickness, decrease the infiltration of mast cells, reduce the IgE level in serum, decrease the expression levels of AD-related cytokines, increase protein and mRNA expression of FLG, and reduce the protein and mRNA expression of KLK7 in the skin tissues of AD-like mice. CONCLUSION: In conclusion, AMA inhibits inflammatory response at the cellular level, and AMA reduces the validation response of specific dermatitis mice, relieves pruritus, and repairs the damaged skin barrier.

Five Rutaceae family ethanol extracts alleviate H2O2 and LPS-induced inflammation via NF-κB and JAK-STAT3 pathway in HaCaT cells.

This study was designed to investigate the effects of five Rutaceae family ethanol extracts (FRFEE): Citrus medica Linn (CML), Citrus aurantium L. Cv. Daidai (CAD), Citrus medica Linn. var. sarcodactylis (Noot.) Swingle (CMS),Citrus sinensis L. Osbeck (CSO) and Zanthoxylum bungeanum Maxim (ZBM) on retarding the progression of H2O2 and LPS-induced HaCaT cells. Cell inflammatory injury model was established by H2O2 and LPS. The alleviative effects of FRFEE were evaluated by detecting the activity of superoxide dismutase (SOD), glutathione (GSH) and the generation of reactive oxygen species (ROS). The inflammatory signaling pathways of NF-κB and JAK-STAT3 were detected by Western blotting, the mRNA expression levels of inflammatory factors and skin barrier factors were detected by RT-PCR. 50% ethanol extracts of five medicinal and food homologous herbs of Rutaceae family showed different levels of anti-oxidant and anti-inflammatory activities. The FRFEE effectively improved SOD and GSH content and decreased ROS levels. Meanwhile, FRFEE strongly suppressed two inflammatory signaling pathways NF-κB and JAK-STAT3. The RT-PCR examination of inflammatory factors and skin barrier factor revealed significant anti-inflammatory effects of FRFEE. It was worth noting that among the five extracts, Zanthoxylum bungeanum Maxim extract had the best anti-inflammatory and anti-oxidation effects. In addition, it could strongly inhibit the expression of psoriasis factor CCL20. In summary, these results suggested that Zanthoxylum bungeanum Maxim extract could be used as an anti-psoriatic agent in the treatment of psoriasis among FRFEE.

Fermented Taiwanofungus camphoratus Extract Ameliorates Psoriasis-Associated Response in HaCaT Cells via Modulating NF-𝜠B and mTOR Pathways.

Psoriasis is a chronic autoimmune disease, and until now, it remains an incurable disease. Therefore, the development of new drugs or agents that ameliorate the disease will have marketing potential. Taiwanofungus camphoratus (TC) is a specific fungus in Taiwan. It is demonstrated to have anticancer, anti-inflammation, and hepatoprotective effects. However, the effects of TC fermented extract on psoriasis are under investigation. In this research, we studied the ability of TC on antioxidative activity and the efficacy of TC on interleukin-17 (IL-17A)-induced intracellular oxidative stress, inflammation-relative, and proliferation-relative protein expression in human keratinocytes. The results of a DPPH radical scavenging assay, reducing power assay, and hydroxyl peroxide inhibition assay indicated that TC has a potent antioxidant ability. Furthermore, TC could reduce IL-17A-induced intracellular ROS generation and restore the NADPH level. In the investigation of pathogenesis, we discovered TC could regulate inflammatory and cell proliferation pathways via p-IKKα/p-p65 and p-mTOR/p-p70S6k signaling pathways in human keratinocytes. In conclusion, TC showed characteristics such as antioxidant, anti-inflammatory, and anti-psoriatic-associated responses. It is expected to be developed as a candidate for oxidative-stress-induced skin disorders or psoriasis treatment.

Umbilicaria esculenta Extract Exhibits Antiwrinkle Activity by Suppressing ErbB2 Phosphorylation.

Umbilicaria esculenta (UE), an edible lichen, is widespread in northeast Asian countries, including China, Japan, and Korea. In the present study, we examined the antiwrinkle activity of UE. We observed that the UE extract (UEE) suppressed ultraviolet (UV)-induced matrix metalloprotein-1 (MMP-1) expression and reactive oxygen species (ROS) generation in a human keratinocyte cell line (HaCaT) and human skin tissue. In addition, UEE reversed the UV-induced decrease in collagen in the human skin tissue. Excessive and chronic UV exposure is a key factor underlying skin wrinkle formation via MMP-1 expression. As treatment with UEE disrupted the UV-activated mitogen-activated protein kinase (MAPK) signaling pathway, we applied an antibody array to unveil the underlying mechanism of UEE. Interestingly, UEE treatment inhibited ErbB2 phosphorylation, but not epidermal growth factor receptor phosphorylation, a heterodimerization partner with ErbB2. Furthermore, UEE treatment enhanced UV-suppressed phosphatase activity via ROS suppression. Collectively, our findings indicate that UEE enhances ErbB2 dephosphorylation to suppress UV-induced MMP-1 expression.