Transcriptomic analysis of the antimicrobial activity of prodigiosin against Cutibacterium acnes.

Prodigiosin, a red pigment produced by Hahella chejuensis, a marine-derived microorganism, has several biological functions, including antimicrobial activity and inflammatory relief. In this study, the antibacterial activity of prodigiosin against skin microorganisms was explored. Paper disc assay on skin bacterial cells revealed that Cutibacterium acnes related to acne vulgaris highly susceptible to prodigiosin. MIC (Minimal Inhibitory Concentration) and MBC (Minimal Bactericidal Concentration) were determined on Cutibacterium species. The RNA-seq analysis of prodigiosin-treated C. acnes cells was performed to understand the antibacterial mechanism of prodigiosin. Among changes in the expression of hundreds of genes, the expression of a stress-responsive sigma factor encoded by sigB increased. Conversely, the gene expression of cell wall biosynthesis and energy metabolism was inhibited by prodigiosin. Specifically, the expression of genes related to the metabolism of porphyrin, a pro-inflammatory metabolite, was significantly reduced. Therefore, prodigiosin could be used to control C. acnes. Our study provided new insights into the antimicrobial mechanism of prodigiosin against C. acnes strains.

Increased 1-Deoxysphingolipids and Skin Barrier Dysfunction in the Skin of X-ray or Ultraviolet B Irradiation and Atopic Dermatitis Lesion Could Be Prevented by Moisturizer with Physiological Lipid Mixture.

BACKGROUND: Skin diseases characterized by epithelial barrier dysfunction show altered sphingolipid metabolism, which results in changes in the stratum corneum intercellular lipid components and structure. Under pathological conditions, 1-deoxysphingolipids form as atypical sphingolipids from de novo sphingolipid biosynthesis. OBJECTIVE: This study investigated the potential role of 1-deoxysphingolipids in skin barrier dysfunction secondary to X-ray and ultraviolet B (UVB) irradiation in vitro and in vivo. It was also evaluated changes in the expression of 1-deoxysphingolipids in lesional human skin of atopic dermatitis. METHODS: In this study, the changes in these 1-deoxysphingolipids levels of skin and serum samples were investigated in skin barrier dysfunction associated with X-ray and UVB irradiation in vitro and in vivo. RESULTS: Increased 1-deoxysphingolipids were observed in cultured normal human epidermal keratinocytes after X-ray irradiation. X-ray or UVB irradiation increased the production of 1-deoxysphingosine in a reconstituted 3-dimensional (3D) skin model. Interestingly, treatment with a physiological lipid mixture (multi-lamellar emulsion contained pseudoceramide), which can strengthen the epidermal permeability barrier function, resulted in decreased 1-deoxysphingosine formation in a reconstituted 3D skin model. Further investigation using a hairless mouse model showed similar preventive effects of physiological lipid mixture against 1-deoxysphingosine formation after X-ray irradiation. An increased level of 1-dexoysphingosine in the stratum corneum was also observed in lesional skin of atopic dermatitis. CONCLUSION: 1-deoxysphingosine might be a novel biomarker of skin barrier dysfunction and a physiological lipid mixture treatment could prevent 1-deoxysphingosine production and consequent skin barrier dysfunction.

Stimulation of Sphingosine Kinase 1 (SPHK1) Is Beneficial in a Huntington's Disease Pre-clinical Model.

Although several agents have been identified to provide therapeutic benefits in Huntington disease (HD), the number of conventionally used treatments remains limited and only symptomatic. Thus, it is plausible that the need to identify new therapeutic targets for the development of alternative and more effective treatments is becoming increasingly urgent. Recently, the sphingosine-1-phosphate (S1P) axis has been reported to be a valid potential novel molecular target for therapy development in HD. Modulation of aberrant metabolism of S1P in HD has been proved to exert neuroprotective action in vitro settings including human HD iPSC-derived neurons. In this study, we investigated whether promoting S1P production by stimulating Sphingosine Kinase 1 (SPHK1) by the selective activator, K6PC-5, may have therapeutic benefit in vivo in R6/2 HD mouse model. Our findings indicate that chronic administration of 0.05 mg/kg K6PC-5 exerted an overall beneficial effect in R6/2 mice. It significantly slowed down the progressive motor deficit associated with disease progression, modulated S1P metabolism, evoked the activation of pro-survival pathways and markedly reduced the toxic mutant huntingtin (mHtt) aggregation. These results suggest that K6PC-5 may represent a future therapeutic option in HD and may potentially counteract the perturbed brain function induced by deregulated S1P pathways.

Author Correction: Defective Sphingosine-1-phosphate metabolism is a druggable target in Huntington's disease.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Aquatide Activation of SIRT1 Reduces Cellular Senescence through a SIRT1-FOXO1-Autophagy Axis.

Ultraviolet (UV) irradiation is a relevant environment factor to induce cellular senescence and photoaging. Both autophagy- and silent information regulator T1 (SIRT1)-dependent pathways are critical cellular processes of not only maintaining normal cellular functions, but also protecting cellular senescence in skin exposed to UV irradiation. In the present studies, we investigated whether modulation of autophagy induction using a novel synthetic SIRT1 activator, heptasodium hexacarboxymethyl dipeptide-12 (named as Aquatide), suppresses the UVB irradiation-induced skin aging. Treatment with Aquatide directly activates SIRT1 and stimulates autophagy induction in cultured human dermal fibroblasts. Next, we found that Aquatide-mediated activation of SIRT1 increases autophagy induction via deacetylation of forkhead box class O (FOXO) 1. Finally, UVB irradiation-induced cellular senescence measured by SA-ß-gal staining was significantly decreased in cells treated with Aquatide in parallel to occurring SIRT1 activation-dependent autophagy. Together, Aquatide modulates autophagy through SIRT1 activation, contributing to suppression of skin aging caused by UV irradiation.

Defective Sphingosine-1-phosphate metabolism is a druggable target in Huntington's disease.

Huntington's disease is characterized by a complex and heterogeneous pathogenic profile. Studies have shown that disturbance in lipid homeostasis may represent a critical determinant in the progression of several neurodegenerative disorders. The recognition of perturbed lipid metabolism is only recently becoming evident in HD. In order to provide more insight into the nature of such a perturbation and into the effect its modulation may have in HD pathology, we investigated the metabolism of Sphingosine-1-phosphate (S1P), one of the most important bioactive lipids, in both animal models and patient samples. Here, we demonstrated that S1P metabolism is significantly disrupted in HD even at early stage of the disease and importantly, we revealed that such a dysfunction represents a common denominator among multiple disease models ranging from cells to humans through mouse models. Interestingly, the in vitro anti-apoptotic and the pro-survival actions seen after modulation of S1P-metabolizing enzymes allows this axis to emerge as a new druggable target and unfolds its promising therapeutic potential for the development of more effective and targeted interventions against this incurable condition.

Antibody to FcεRIα Suppresses Immunoglobulin E Binding to High-Affinity Receptor I in Allergic Inflammation.

PURPOSE: High-affinity receptor I (FcεRI) on mast cells and basophils plays a key role in the immunoglobulin E (IgE)-mediated type I hypersensitivity mediated by allergen cross-linking of the specific IgE-FcεRI complex. Thus, prevention of IgE binding to FcεRI on these cells is an effective therapy for allergic disease. We have developed a strategy to disrupt IgE binding to FcεRI using an antibody targeting FcεRIα. MATERIALS AND METHODS: Fab fragment antibodies, which lack the Fc domain, with high affinity and specificity for FcεRIα and effective inhibitory activity against IgE-FcεRI binding were screened. IgE-induced histamine, ß-hexosaminidase and Ca²âº release in basophils were determined by ELISA. A B6.Cg-Fcer1a(tm1Knt) Tg(FCER1A)1Bhk/J mouse model of passive cutaneous anaphylaxis (PCA) was used to examine the inhibitory effect of NPB311 on allergic skin inflammation. RESULTS: NPB311 exhibited high affinity to human FcεRIα (KD=4 nM) and inhibited histamine, ß-hexosaminidase and Ca²âº release in a concentration-dependent manner in hFcεRI-expressing cells. In hFcεRIα-expressing mice, dye leakage was higher in the PCA group than in controls, but decreased after NPB311 treatment. NPB311 could form a complex with FcεRIα and inhibit the release of inflammation mediators. CONCLUSION: Our approach for producing anti-FcεRIα Fab fragment antibody NPB311 may enable clinical application to a therapeutic pathway in IgE/FcεRI-mediated diseases.

Selective Cannabinoid Receptor-1 Agonists Regulate Mast Cell Activation in an Oxazolone-Induced Atopic Dermatitis Model.

BACKGROUND: Many inflammatory mediators, including various cytokines (e.g. interleukins and tumor necrosis factor [TNF]), inflammatory proteases, and histamine are released following mast cell activation. However, the endogenous modulators for mast cell activation and the underlying mechanism have yet to be elucidated. Endogenous cannabinoids such as palmitoylethanolamide (PEA) and N-arachidonoylethanolamine (anandamide or AEA), were found in peripheral tissues and have been proposed to possess autacoid activity, implying that cannabinoids may downregulate mast cell activation and local inflammation. OBJECTIVE: In order to investigate the effect of cannabinoid receptor-1 (CB1R) agonists on mast cell activation, AEA-derived compounds were newly synthesized and evaluated for their effect on mast cell activation. METHODS: The effects of selected compounds on FcεRI-induced histamine and ß-hexosaminidase release were evaluated in a rat basophilic leukemia cell line (RBL-2H3). To further investigate the inhibitory effects of CB1R agonist in vivo, an oxazolone-induced atopic dermatitis mouse model was exploited. RESULTS: We found that CB1R inhibited the release of inflammatory mediators without causing cytotoxicity in RBL-2H3 cells and that CB1R agonists markedly and dose-dependently suppressed mast cell proliferation indicating that CB1R plays an important role in modulating antigen-dependent immunoglobulin E (IgE)-mediated mast cell activation. We also found that topical application of CB1R agonists suppressed the recruitment of mast cells into the skin and reduced the level of blood histamine. CONCLUSION: Our results indicate that CB1R agonists down-regulate mast cell activation and may be used for relieving inflammatory symptoms mediated by mast cell activation, such as atopic dermatitis, psoriasis, and contact dermatitis.

Pseudoceramide stimulates peroxisome proliferator-activated receptor-α expression in a murine model of atopic dermatitis: molecular basis underlying the anti-inflammatory effect and the preventive effect against steroid-induced barrier impairment.

Topical pseudoceramides are successfully used in skin barrier repair therapy for atopic dermatitis (AD) and demonstrated to reduce the adverse effects of topical glucocorticoids (GC). However, the molecular mechanisms involved are not fully understood. We investigated whether PC-9S (myristoyl/palmitoyloxostearamide/arachamide MEA, Neopharm, Daejeon, Korea), one of the synthetic pseudoceramides, could stimulate peroxisome proliferator-activated receptor (PPAR)α expression in a hapten [oxazolone (oxa)]-induced AD murine model (oxa-AD mice) and subsequently improved permeability barrier, reduced inflammation, and increased antimicrobial peptides (AMPs) expression. Normal hairless mice and oxa-AD mice were topically treated twice daily with either PC-9S-containing physiologic lipid mixture (PLM), vehicle (PLM), or PPARα agonist for 4 days. Topical PC-9S significantly increased PPARα expression in mouse epidermis in vivo and in oxa-AD mice skin comparable with PPARα agonist. Topical PC-9S-containing PLM significantly reduced basal trans-epidermal water loss (TEWL), surface pH, and mast cell infiltrates and prevented the decline of AMPs expression in oxa-AD mice, which were abrogated by PPARα antagonist. Then, oxa-AD mice were treated with super-potent topical GC twice daily for 4 days with or without PC-9S co-applications. Co-treatment with PC-9S-containing PLM suppressed GC-induced increase in basal TEWL, epidermal thinning, reduced loricrin expression, and impaired barrier recovery and these effects were attenuated by PPARα antagonist. Collectively, our findings suggest that pseudoceramide PC-9S-induced stimulation of PPARα expression provides a new mechanism by which pseudoceramides show anti-inflammatory property, improve the permeability and antimicrobial barrier function, and prevent the negative effects of topical GC.

Sphingosine kinase 1 activation enhances epidermal innate immunity through sphingosine-1-phosphate stimulation of cathelicidin production.

BACKGROUND: The ceramide metabolite, sphingosine-1-phosphate (S1P), regulates multiple cellular functions in keratinocytes (KC). We recently discovered that production of a key innate immune element, cathelicidin antimicrobial peptide (CAMP), is stimulated via a NF-κB-dependent mechanism that is activated by S1P when S1P is generated by sphingosine kinase (SPHK) 1. OBJECTIVE: We investigated whether pharmacological modulation of SPHK1 activity, using a novel synthetic SPHK1 activator, (S)-methyl 2-(hexanamide)-3-(4-hydroxyphenyl) propanoate (MHP), stimulates CAMP expression. METHODS: MHP-mediated changes in both S1P and CAMP downstream mediators were analyzed in normal cultured human KC by qRT-PCR, Western immunoblot, ELISA, confocal microscopy for immunohistochemistry, HPLC and ESI-LC/MS/MS, and microbial pathogen invasion/colonization in a human epidermal organotypic model. RESULTS: Treatment with MHP directly activated SPHK1 and increased cellular S1P content in normal cultured human KC. Because MHP did not inhibit S1P lyase activity, which hydrolyses S1P, augumented S1P levels could be attributed to increased synthesis rather than blockade of S1P degradation. Next, we found that exogenous MHP significantly stimulated CAMP mRNA and protein production in KC, increases that were significantly suppressed by siRNA directed against SPHK1, but not by a scrambled control siRNA. NF-κB activation, assessed by nuclear translocation of NF-κB, occurred in cells following incubation with MHP. Conversely, pretreatment with a specific inhibitor of SPHK1 decreased MHP-induced nuclear translocation of NF-κB, and significantly attenuated the MHP-mediated increase in CAMP production. Finally, topical MHP significantly suppressed invasion of the virulent Staphylococcus aureus into murine skin explants. CONCLUSION: MHP activation of SPHK1, a target enzyme of CAMP production, can stimulate innate immunity.

Topical cannabinoid receptor 1 agonist attenuates the cutaneous inflammatory responses in oxazolone-induced atopic dermatitis model.

BACKGROUND: Even with the widespread clinical use of cannabinoid receptor (CBR) stimulating compounds, such as palmitoylethanolamine, the role of CBR agonists on inflammatory skin diseases is not yet fully understood. This study was performed to investigate the effects of CBR agonists on skin inflammation, using acute and chronic inflammation animal models. METHODS: The effectiveness of the newly synthesized cannabinoid receptor 1 (CB1R) agonists was determined using in vitro assays. Markers for epidermal permeability barrier function and skin inflammation were measured, and histological assessments were performed for evaluation. RESULTS: Topical application of CB1R-specific agonist significantly accelerated the recovery of epidermal permeability barrier function and showed anti-inflammatory activity in both acute and chronic inflammation models. Histological assessments also confirmed the anti-inflammatory effects, which is consistent with previous reports. CONCLUSIONS: All of the results suggest that topical application of CB1R-specific agonist can be beneficial for alleviating the inflammatory symptoms in chronic skin diseases, including atopic dermatitis.

Growth hormone-releasing peptide-biotin conjugate stimulates myocytes differentiation through insulin-like growth factor-1 and collagen type I.

Based on the potential beneficial effects of growth hormone releasing peptide (GHRP)-6 on muscle functions, a newly synthesized GHRP-6-biotin conjugate was tested on cultured myoblast cells. Increased expression of myogenic marker proteins was observed in GHRP-6-biotin conjugate-treated cells. Additionally, increased expression levels of insulin-like growth factor-1 and collagen type I were observed. Furthermore, GHRP-6-biotin conjugate-treated cells showed increased metabolic activity, as indicated by increased concentrations of energy metabolites, such as ATP and lactate, and increased enzymatic activity of lactate dehydrogenase and creatine kinase. Finally, binding protein analysis suggested few candidate proteins, including desmin, actin, and zinc finger protein 691 as potential targets for GHRP6-biotin conjugate action. These results suggest that the newly synthesized GHRP-6-biotin conjugate has myogenic stimulating activity through, at least in part, by stimulating collagen type I synthesis and several key proteins. Practical applications of the GHRP-6-biotin conjugate could include improving muscle condition.

Effects of Lactobacillus rhamnosus on allergic march model by suppressing Th2, Th17, and TSLP responses via CD4(+)CD25(+)Foxp3(+) Tregs.

Allergic march (AM) is characterized by the progression of clinical signs of atopic dermatitis (AD) to allergic asthma or rhinitis, but its pathogenesis is not completely understood. We developed mouse model of AM with three 1-week exposures (separated by 2-week interval) to an OVA or saline (control) followed by OVA challenge. The development of AM was confirmed by phenotypes of AD and allergic asthma. Increases in IL-4, IL-17, and thymic stromal lymphopoietin (TSLP) responses were associated with the progression of AM, and these responses were suppressed by treatment with Lcr35. Moreover, Lcr35 treatment led to an increase in the number of CD4(+)CD25(+) Foxp3(+) regulatory T (Treg) cells in the mesenteric lymph nodes (MLNs) of AM mice. In conclusion, the oral application of Lcr35 prevented the development of AM in this model by suppressing Th2, Th17, and TSLP responses via a mechanism that may involve CD4(+)CD25(+)Foxp3(+) Tregs in MLNs.

A novel synthetic mycolic Acid inhibits bronchial hyperresponsiveness and allergic inflammation in a mouse model of asthma.

PURPOSE: Recognition of microbes is important to trigger the innate immune system. Mycolic acid (MA) is a component of the cell walls of mycobacteria such as Mycobacterium bovis Bacillus Calmette-Guerin. MA has immunogenic properties, which may modulate the innate and adaptive immune response. This study aimed to investigate whether a novel synthetic MA (sMA) inhibits allergic inflammatory responses in a mouse model of asthma. METHODS: BALB/c mice were injected intraperitoneally with sMA followed by sensitization and challenge with ovalbumin (OVA). Mice were examined for bronchial hyperresponsiveness (BHR), the influx of inflammatory cells into the lung tissues, histopathological changes in the lungs and CD4(+)CD25(+)Foxp3(+) T cells in the spleen, and examined the response after the depleting regulatory T cells (Tregs) with an anti-CD25mAb. RESULTS: Treatment of mice with sMA suppressed the asthmatic response, including BHR, bronchoalveolar inflammation, and pulmonary eosinophilic inflammation. Anti-CD25mAb treatment abrogated the suppressive effects of sMA in this mouse model of asthma and totally depleted CD4(+)CD25(+)Foxp3(+) T cells in the spleen. CONCLUSIONS: sMA attenuated allergic inflammation in a mouse model of asthma, which might be related with CD4(+)CD25(+)Foxp3(+) T cell.

Preventive effects of multi-lamellar emulsion on low potency topical steroid induced local adverse effect.

BACKGROUND: Topical steroid treatment induces diverse local Wand systemic adverse effects. Several approaches have been tried to reduce the steroid-induced adverse effects. Simultaneous application of physiological lipid mixture is also suggested. OBJECTIVE: Novel vehicles for topical glucocorticoids formulation were evaluated for the efficacy of reducing side-effects and the drug delivery properties of desonide, a low potency topical steroid. METHODS: Transcutaneous permeation and skin residual amount of desonide were measured using Franz diffusion cells. The in vivo anti-inflammatory activity was evaluated using murine model. RESULTS: Topical steroids formulation containing desonide, in either cream or lotion form, were prepared using multi-lamellar emulsion (MLE), and conventional desonide formulations were employed for comparison. MLE formulations did not affect the anti-inflammatory activity of the desonide in phobol ester-induced skin inflammation model, compared with conventional formulations. While the penetrated amounts of desonide were similar for all the tested formulations at 24 hours after application, the increased lag time was observed for the MLE formulations. Interestingly, residual amount of desonide in epidermis was significantly higher in lotion type MLE formulation. Steroid-induced adverse effects, including permeability barrier function impairment, were partially prevented by MLE formulation. CONCLUSION: Topical desonide formulation using MLE as a vehicle showed a better drug delivery with increased epidermal retention. MLE also partially prevented the steroid-induced side effects, such as skin barrier impairment.