Effects of dextran sulfate, 4-t-butylcyclohexanol, pongamia oil and hesperidin methyl chalcone on inflammatory and vascular responses implicated in rosacea.

BACKGROUND: Rosacea is a chronic facial skin disorder characterized by inflammation and vascular abnormalities. The pathophysiology of rosacea involves increased activation of the capsaicin receptor, TRPV1, the vascular endothelial growth factor (VEGF) pathway, and cathelicidin LL-37, MMP-9, and KLKs. We evaluated the activity of four compounds (dextran sulfate, 4-t-butylcyclohexanol [BCH; TRP-regulin®], pongamia oil, and hesperidin methyl chalcone [HMC]) on inflammatory and vascular responses implicated in rosacea. MATERIALS AND METHODS: The anti-inflammatory activity of dextran sulfate was evaluated on PGE2 production after PMA stimulation of NCTC-2544 keratinocytes, and on normal human epidermal keratinocytes (NHEKs) after proinflammatory stimulation to mimic a rosacea environment. The anti-angiogenic activity of dextran sulfate was measured by analyzing pseudotube formation in co-cultured human microvascular endothelial cells/normal human dermal fibroblasts. HMC modulation of vascular responses and IL-8 cytokine production after SP stimulation was evaluated in human skin explants. We also assessed the effect of BCH on TRPV1 activation, and the effect of combined BCH and pongamia oil on the inflammatory response of NHEKs. RESULTS: Dextran sulfate strongly and significantly inhibited PMA-induced PGE2 production, inhibited KLK5 and MMP-9 mRNA expression, and IL-8, IL-1α and VEGF production, and displayed a highly significant inhibitory effect on VEGF-induced pseudotube formation. In SP-stimulated human skin explants, HMC significantly decreased the proportion of dilated vessels, total vessel area, and IL-8 production. BCH significantly and dose-dependently inhibited TRPV1 activation, and BCH and pongamia oil inhibited CXCL1 and CXCL6 mRNA expression and IL-8 production in NHEKs. Combined BCH/pongamia oil inhibited IL-8 production synergistically. CONCLUSION: These in vitro results showed that dextran sulfate, BCH, pongamia oil and HMC, possess complementary soothing and anti-redness properties, supporting their combination in Avène redness-relief cosmetic products for sensitive skin prone to redness, and for topical adjunctive rosacea treatment.

Anti-inflammatory and immunomodulatory effects of Aquaphilus dolomiae extract on in vitro models.

BACKGROUND: Atopic dermatitis (AD) is a common skin disease characterized by recurrent pruritic inflammatory skin lesions resulting from structural and immune defects of the skin barrier. Previous studies have shown the clinical efficacy of Avène thermal spring water in AD, and a new microorganism, Aquaphilus dolomiae was suspected to contribute to these unique properties. The present study evaluated the anti-inflammatory, antipruritic, and immunomodulatory properties of ES0, an original biological extract of A. dolomiae, in immune and inflammatory cell models in order to assess its potential use in the treatment of AD. MATERIALS AND METHODS: An ES0 extract containing periplasmic and membrane proteins, peptides, lipopolysaccharides, and exopolysaccharides was obtained from A. dolomiae. The effects of the extract on pruritus and inflammatory mediators and immune mechanisms were evaluated by using various AD cell models and assays. RESULTS: In a keratinocyte model, ES0 inhibited the expression of the inflammatory mediators, thymic stromal lymphopoietin, interleukin (IL)-18, IL-4R, IL-8, monocyte chemoattractant protein-3, macrophage inflammatory protein-3α, and macrophage-derived chemokine and induced the expression of involucrin, which is involved in skin barrier keratinocyte terminal differentiation. In addition, ES0 inhibited protease-activated receptor-2 activation in HaCaT human keratinocytes stimulated by stratum corneum tryptic enzyme and T helper type (Th) 1, Th2, and Th17 cytokine production in Staphylococcal enterotoxin B-stimulated CD4+ lymphocytes. Lastly, ES0 markedly activated innate immunity through toll-like receptor (TLR) 2, TLR4, and TLR5 activation (in recombinant human embryonic kidney 293 cells) and through antimicrobial peptide induction (psoriasin, human beta-defensin-2, and cathelicidin), mainly through TLR5 activation (in normal human keratinocytes). CONCLUSION: Overall, these in vitro results confirm the marked regulatory activity of this A. dolomiae extract on inflammatory and immune responses, which may be of value by virtue of its potential as an adjunctive treatment of AD inflammatory and pruritic lesions.

Pharmacological properties of Myrtacine® and its potential value in acne treatment.

This study aimed at evaluating the antiproliferative, antibacterial, and anti-inflammatory properties of an ethanolic myrtle extract (Myrtacine®) in vitro, characterising its potential active compounds (myrtucommulones A and B') by structural analysis, and evaluating their biological activity. Antiproliferative activity was assessed by the BrdU incorporation assay in HaCat keratinocytes and inhibitory and bactericidal activities against P. ACNES strains by measuring the minimal inhibitory concentration (MIC) and D value. Anti-inflammatory effect was evaluated by measuring 6-keto-prostaglandin F1 α and [³H]-arachidonic acid metabolite production in keratinocytes stimulated for inflammation. Myrtacine® inhibited keratinocyte proliferation by 27 % and 76 % at 1 and 3 µg/mL, respectively (p < 0.001). A comparable effect, though less marked, was observed with 5 µg/mL myrtucommulones A and B' (-36 % and -28 %, respectively). Myrtacine® inhibited erythromycin-sensible and -resistant P. ACNES strains growth with MICs of 4.9 µg/mL and 2.4 µg/mL, respectively. Myrtucommulone B' and myrtucommulone A displayed a similar inhibitory activity against both strains (for both strains, MIC = 1.2 µg/mL and about 0.5 µg/mL, respectively). At 3 and 10 µg/mL, Myrtacine® significantly decreased all metabolite production from cyclooxygenase (81 % and 107 %, p < 0.0001) and lipoxygenase (52 % and 95 %, p < 0.001) pathways. Finally, Myrtacine® exhibited a concentration-dependent anti-lipase activity at 100 µg/mL and 1 mg/mL, as it decreased lipase activity by respectively 53 % and 100 % (p < 0.01 for both). In conclusion, in vitro, Myrtacine® demonstrated antiproliferative, antibacterial, and anti-inflammatory properties that may be of value to exert a global action in the treatment of acne lesions.

Rhealba® oat plantlet extract: evidence of protein-free content and assessment of regulatory activity on immune inflammatory mediators.

Owing to their high content of flavonoids and saponins, plantlets of Avena sativa L. (Poaceae) are likely to possess anti-inflammatory and immunoregulatory properties of value in the treatment of atopic dermatitis (AD). With a view to its potential use in atopic subjects at risk of developing sensitisation to dietary proteins, we prepared a plantlet extract without proteins and isolated 2 flavonoids, isoorientin-2''- O-arabinoside (1) and isovitexin-2''- O-arabinoside (2), and two saponins, avenacosides A (3) and B (4). The absence of protein in this extract was evidenced by electrophoresis and Western immunoblotting. Furthermore, Western immunoblotting demonstrated the absence of cross-reaction between grain and plantlet proteins. We evaluated the anti-inflammatory activity of the plantlet extract and its compounds IN VITRO in a model of keratinocyte inflammation: 6-keto prostaglandin F1 α production was inhibited by the plantlet extract (- 35 % and - 57 % at 10 and 30 µg/mL, respectively; p < 0.001) and isoorientin-2''- O-arabinoside (- 31 %, - 51 %, and - 56 % at 3, 10, and 30 µg/mL, respectively; p < 0.001). Intracellular interleukin-2 production in activated T lymphocytes was also inhibited by 16 %, 27 %, and 31 % with 3, 10, and 30 µg/mL plantlet extract, respectively, and by 23 % and 32 % with 3 and 10 µg/mL avenacoside A, respectively, (p < 0.001), demonstrating their immunoregulatory activity IN VITRO. The plantlet extract was also effective on the phenotype and function of dendritic cells (DC) differentiated from monocytes. It decreased the expression of major histocompatibility complex class II molecules on DC and significantly impaired their stimulatory activity on autologous T-cell proliferation (-25 %, p < 0.05). In conclusion, this protein-free oat plantlet extract exhibits anti-inflammatory and immunoregulatory activities in vitro.

Age-related differences in sensitivity of peripheral blood monocytes to lipopolysaccharide and Staphylococcus aureus toxin B in atopic dermatitis.

As shown by atopy patch tests, atopic dermatitis (AD) is dominated in its acute phase by the development of a specific T(H)2 response after exposure of the skin to common environmental antigens. Relying on our previous data showing that Staphylococcus aureus enterotoxin B (SEB) induced the activation of monocyte-derived dendritic cells (DCs) through Toll-like receptor (TLR)2 and that SEB-pulsed DCs commit allogenic naive T cells into T(H)2, we assessed monocytes sensitivity to SEB and lipopolysaccharide (LPS) in a group of children and adult patients with AD. Monocytes from AD patients (15 adults with mostly severe disease and 15 children with mild to moderate disease) exhibited an activated and tolerant state as supported by (i) secretion of large amounts of IL-6, IL-10, and tumor necrosis factor-alpha even in the absence of stimulation; (ii) their inability to modulate neither HLA-DR and CD54 nor TLR2 and TLR4 expression after in vitro challenge with SEB; (iii) inhibition of IL-12p70 secretion in response to LPS. Interestingly, monocytes from some of the children studied responded to in vitro challenge with LPS, suggesting new hypotheses to explain disease regression. Our data support the notion that monitoring sensitivity of monocytes to bacterial toxins could prove useful to assess disease progression and prognosis in AD.

Skin-infiltrating CD8+ T cells initiate atopic dermatitis lesions.

Skin lesions in the allergic form of atopic dermatitis (AD) are induced by allergen-specific T cells that infiltrate the skin at the site of allergen exposure. Although Th2-type CD4+ T cells appear to be crucial in AD pathophysiology, little is known about the contribution of CD8+ T cells in the development of the allergic skin inflammation. In the present study, we have analyzed the respective role of CD8+ and CD4+ T cells in the development of AD skin lesions in a mouse model of allergen-induced AD. In sensitized mice, CD8+ T cells are rapidly and transiently recruited to the allergen-exposed site and initiate the inflammatory process leading to skin infiltration with eosinophils and Th1/Th2-producing cells. CD8+ T cell-depleted mice show no inflammation, demonstrating that these cells are mandatory for the development of AD. In contrast, CD4+ T cell-depleted mice develop a severe form of eczema. Furthermore, adoptive transfer of CD8+ T cells from sensitized mice into naive recipient mice leads to skin inflammation soon after allergen exposure. These data indicate that allergen-primed CD8+ T cells are required for the development of AD-like lesions in mice.

n-3 and n-6 polyunsaturated fatty acids induce the expression of COX-2 via PPARgamma activation in human keratinocyte HaCaT cells.

Polyunsaturated fatty acids (PUFA) n-3 inhibit inflammation, in vivo and in vitro in keratinocytes. We examined in HaCaT keratinocyte cell line whether eicosapentaenoic acid (EPA) a n-3 PUFA, gamma-linoleic acid (GLA) a n-6 PUFA, and arachidic acid a saturated fatty acid, modulate expression of cyclooxygenase-2 (COX-2), an enzyme pivotal to skin inflammation and reparation. We demonstrate that only treatment of HaCaT with GLA and EPA or a PPARgamma ligand (roziglitazone), induced COX-2 expression (protein and mRNA). Moreover stimulation of COX-2 promoter activity was increased by those PUFAs or rosiglitazone. The inhibitory effects of GW9662 and T0070907 (PPARgamma antagonists), on COX-2 expression and on stimulation of COX-2 promoter activity by EPA and GLA suggest that PPARgamma is implicated in COX-2 induction. Finally, PLA2 inhibitor methyl arachidonyl fluorophosphonate blocked the PUFA effects on COX-2 induction, promoter activity and arachidonic acid mobilization suggesting involvement of AA metabolites in PPAR activation. These findings demonstrate that n-3 and n-6 PUFA increased PPARgamma activity is necessary for the COX-2 induction in HaCaT human keratinocyte cells. Given the anti-inflammatory properties of EPA, we suggest that induction of COX-2 in keratinocytes may be important in the anti-inflammatory and protective mechanism of action of PUFAs n-3 or n-6.

Human dendritic cells conditioned with Staphylococcus aureus enterotoxin B promote TH2 cell polarization.

BACKGROUND: Immune surveillance against microbes at sites of interface with environment involves immediate recognition of pathogen-associated molecular patterns by dendritic cells (DCs). According to their first-line position, DCs are key parameters for the establishment of an appropriate innate and adaptive response against pathogens to avoid disease development. Even though their role in pathogenesis is well known, bacterial toxins have been less examined for their ability to drive DC activation and T-cell polarization. OBJECTIVE: We made the assumption that early conditioning of DCs with Staphylococcus aureus enterotoxins could take part in T-cell polarization. METHODS: Human monocyte-derived DCs were stimulated with S aureus enterotoxin B (SEB) and characterized with respect to secretion of inflammatory cytokines and their ability to drive polarization of naive allogenic T cells. RESULTS: We demonstrated that SEB induced maturation of DCs and that SEB-activated DCs secreted high levels of IL-2 but no IL-12p70, contrary to LPS-activated ones. Accordingly, we further showed that SEB-activated DCs were able to drive polarization of naive T cells into the T(H)2 subset. By using highly purified SEB and Toll-like receptor (TLR)-2 stably transfected Human Embryonic Kidney (HEK) 293 cells, we demonstrated for the first time the ability of SEB to induce TLR2 signaling. Furthermore, the involvement of SEB-TLR2 interaction in activation of dendritic cells was supported by neutralizing activity of anti-TLR2 antibodies. CONCLUSION: Altogether, our findings reinforce the notion that bacterial toxins may appear as new pathogen-associated molecular patterns, which could play a major role in inflammation and bacterial pathologies.

Isolation and absolute configuration of new bioactive marine steroids from Euryspongia n. sp.

The apolar fraction of the crude alcoholic extract of the sponge Euryspongia n. sp. was shown to display anti-inflammatory activity. Bioassay guided chromatographic purification led to the isolation of a known compound petrosterol (1) of 3beta-hydroxy-24-norchol-5-en-23-oic acid (2), which has never yet been found as a natural substance, and of a new steroid, 3beta-hydroxy-26-norcampest-5-en-25-oic acid (3). The absolute configurations of 2 and 3 were deduced from comparative 1H NMR data of the (S)- and (R)-phenylglycine methyl ester derivatives. These compounds were evaluated for their anti-inflammatory activity against 6-keto-prostaglandinF1alpha release in a human keratinocyte cell line HaCaT.

Avena Rhealba inhibits A23187-stimulated arachidonic acid mobilization, eicosanoid release, and cPLA2 expression in human keratinocytes: potential in cutaneous inflammatory disorders.

The aim of the present study was to examine the effects of Avena Rhealba (AR) oatmeal extract on the metabolism of arachidonic acid (AA) and eicosanoids as well as on the expression of cytosolic phospholipase A(2 )(cPLA(2)) in the human keratinocyte cell line HaCaT. For this purpose, we examined the effects of AR on basal and A23187-triggered release of [(3)H]-AA from phospholipids and on the production of [(3)H]-labeled metabolites of the cyclooxygenase (CO) and 5-lipoxygenase (LO) pathways. AR was found to inhibit A23187-triggered [(3)H]-AA mobilization from phospholipids (p<0.05) and production of [(3)H]-labeled metabolites of CO (p<0.05) and LO (p<0.05) pathways. These results suggest AR decreases PLA(2)-dependent mobilization of AA from phospholipids. A closer examination of the effects of AR on prostaglandin 6KF1alpha (6KPGF1alpha), the stable metabolite of prostacyclin, revealed dose-dependent inhibition of this AA metabolite. AR also decreased A23187- and tumor necrosis factor alpha-induced cPLA(2) overexpression, as shown by cPLA(2) immunodetection and mRNA expression. These results demonstrate the high potential of AR in the treatment of inflammatory diseases of the skin.