Philippe A. Grange

Production of Superoxide Anions by Keratinocytes Initiates P. acnes-Induced Inflammation of the Skin

Acne vulgaris is a chronic inflammatory disorder of the sebaceous follicles. Propionibacterium acnes (P. acnes), a gram-positive anareobic bacterium, plays a critical role in the development of these inflammatory lesions. This study aimed at determining whether reactive oxygen species (ROS) are produced by keratinocytes upon P. acnes infection, dissecting the mechanism of this production, and investigating how this phenomenon integrates in the general inflammatory response induced by P. acnes. In our hands, ROS, and especially superoxide anions (O2 •−), were rapidly produced by keratinocytes upon stimulation by P. acnes surface proteins. In P. acnes-stimulated keratinocytes, O2 •− was produced by NAD(P)H oxidase through activation of the scavenger receptor CD36. O2 •− was dismuted by superoxide dismutase to form hydrogen peroxide which was further detoxified into water by the GSH/GPx system. In addition, P. acnes-induced O2 •− abrogated P. acnes growth and was involved in keratinocyte lysis through the combination of O2 •− with nitric oxide to form peroxynitrites. Finally, retinoic acid derivates, the most efficient anti-acneic drugs, prevent O2 •− production, IL-8 release and keratinocyte apoptosis, suggesting the relevance of this pathway in humans.

Nicotinamide inhibits Propionibacterium acnes-induced IL-8 production in keratinocytes through the NF-κB and MAPK pathways

BACKGROUND Propionibacterium acnes (P. acnes) has been implicated in the inflammatory phase of acne vulgaris. It has been shown to activate interleukin-8 (IL-8) secretion by interacting with Toll-like receptor 2 (TLR-2) on the surface of keratinocytes. Nicotinamide has been shown to be an effective treatment for skin inflammation in various conditions, including acne vulgaris. OBJECTIVE To investigate the molecular mechanisms underlying the anti-inflammatory properties of nicotinamide in keratinocytes stimulated by P. acnes. METHODS HaCaT cells and primary keratinocyte cell lines were stimulated by P. acnes in the presence of nicotinamide. IL-8 production was monitored by ELISA on the cell culture supernatant and by qRT-PCR on total RNA extract. A luciferase reporter system assay was used to assess nicotinamide activity with the IL-8 promoter in transfected keratinocytes. We used western blotting to analyze the effect of nicotinamide on activation of the NF-kappaB and MAPK pathways. RESULTS Nicotinamide significantly decreased IL-8 production in a dose-dependent manner, decreasing both mRNA and protein levels for this chemokine in immortalized HaCaT cells and primary keratinocytes. P. acnes-induced IL-8 promoter activation seemed to be downregulated by nicotinamide, which inhibited IkappaB degradation and the phosphorylation of ERK and JNK MAP kinases. CONCLUSION Our results indicate that nicotinamide inhibits IL-8 production through the NF-kappaB and MAPK pathways in an in vitro keratinocytes/P. acnes model of inflammation. Keratinocytes involved in the innate immune response may be a suitable target for treatment during the early phase of inflammation.

Targeting ADAM-17/notch signaling abrogates the development of systemic sclerosis in a murine model

OBJECTIVE Systemic sclerosis (SSc) is characterized by the fibrosis of various organs, vascular hyperreactivity, and immunologic dysregulation. Since Notch signaling is known to affect fibroblast homeostasis, angiogenesis, and lymphocyte development, we undertook this study to investigate the role of the Notch pathway in human and murine SSc. METHODS SSc was induced in BALB/c mice by subcutaneous injections of HOCl every day for 6 weeks. Notch activation was analyzed in tissues from mice with SSc and from patients with scleroderma. Mice with SSc were either treated or not treated with the γ-secretase inhibitor DAPT, a specific inhibitor of the Notch pathway, and the severity of the disease was evaluated. RESULTS As previously described, mice exposed to HOCl developed a diffuse cutaneous SSc with pulmonary fibrosis and anti-DNA topoisomerase I antibodies. The Notch pathway was hyperactivated in the skin, lung, fibroblasts, and splenocytes of diseased mice and in skin biopsy samples from patients with scleroderma. ADAM-17, a proteinase involved in Notch activation, was overexpressed in the skin of mice and patients in response to the local production of reactive oxygen species. In HOCl-injected mice, DAPT significantly reduced the development of skin and lung fibrosis, decreased skin fibroblast proliferation and ex vivo serum-induced endothelial H(2)O(2) production, and abrogated the production of anti-DNA topoisomerase I antibodies. CONCLUSION Our results show the pivotal role of the ADAM-17/Notch pathway in SSc following activation by reactive oxygen species. The inhibition of this pathway may represent a new treatment of this life-threatening disease.

Sialylation levels of anti–proteinase 3 antibodies are associated with the activity of granulomatosis with polyangiitis (Wegener's)

OBJECTIVE To investigate whether the glycosylation and sialylation levels of anti-proteinase 3 (anti-PR3) antibodies could affect their pathogenicity, and whether these levels could be correlated with the activity of granulomatosis with polyangiitis (Wegeners) (GPA). METHODS Forty-two serum samples positive for anti-PR3 antibodies from 42 patients with active or weakly active/inactive GPA were included. Anti-PR3 antibodies were assayed by enzyme-linked immunosorbent assay, and their levels of glycosylation and sialylation were assessed by enzyme-linked lectin assay. The glycosylation and sialylation levels of IgG purified from the serum of healthy donors and patients with active, remitted, or weakly active disease were assessed by permethylation and mass spectrometry analysis of glycans, following neuraminidase digestion. The neutrophil oxidative burst induced by purified IgG was assayed by spectrofluorimetry. RESULTS The mean sialylation ratio of anti-PR3 antibodies was significantly lower in patients with active disease than in patients with weakly active or inactive disease, and this was inversely correlated with the Birmingham Vasculitis Activity Score (BVAS) (P < 0.0001). Similar results were obtained using the BVAS/GPA. The area under the receiver operating characteristic curve for the sialylation ratio of anti-PR3 antibodies, as a test to determine the activity of GPA, was 0.82 (P = 0.0006). The characterization of N-glycans showed a decrease in 2,6-linked sialylated N-glycans and an increase in dHex₁ Hex₃ HexNAc₄ (mass/charge 1,836) agalactosylated structures in purified IgG from patients with active disease compared with controls. The anti-PR3 antibody-induced oxidative burst of neutrophils was inversely correlated with the sialylation levels of anti-PR3 IgG. CONCLUSION The sialylation level of anti-PR3 antibodies contributes to the clinical activity of GPA, by modulating the oxidative burst of neutrophils induced by these autoantibodies.

Evaluation of a PCR Test for Detection of Treponema pallidum in Swabs and Blood

ABSTRACT Syphilis diagnosis is based on clinical observation, serological analysis, and dark-field microscopy (DFM) detection of Treponema pallidum subsp. pallidum, the etiological agent of syphilis, in skin ulcers. We performed a nested PCR (nPCR) assay specifically amplifying the tpp47 gene of T. pallidum from swab and blood specimens. We studied a cohort of 294 patients with suspected syphilis and 35 healthy volunteers. Eighty-seven of the 294 patients had primary syphilis, 103 had secondary syphilis, 40 had latent syphilis, and 64 were found not to have syphilis. The T. pallidum nPCR results for swab specimens were highly concordant with syphilis diagnosis, with a sensitivity of 82% and a specificity of 95%. Reasonable agreement was observed between the results obtained with the nPCR and DFM methods (kappa = 0.53). No agreement was found between the nPCR detection of T. pallidum in blood and the diagnosis of syphilis, with sensitivities of 29, 18, 14.7, and 24% and specificities of 96, 92, 93, and 97% for peripheral blood mononuclear cell (PBMC), plasma, serum, and whole-blood fractions, respectively. HIV status did not affect the frequency of T. pallidum detection in any of the specimens tested. Swab specimens from mucosal or skin lesions seemed to be more useful than blood for the efficient detection of the T. pallidum genome and, thus, for the diagnosis of syphilis.

Autoimmune Bullous Skin Diseases Occurring under Anti-Tumor Necrosis Factor Therapy: Two Case Reports

Background: Anti-tumor necrosis factor (TNF) agents are increasingly being used for a rapidly expanding number of rheumatic and systemic diseases. As a result of this use, and of the longer follow-up periods of treatment, there are a growing number of reports of the development of autoimmune processes related to anti-TNF agents. The use of anti-TNF agents has been associated with more and more cases of autoimmune diseases, principally cutaneous vasculitis, lupus-like syndrome, systemic lupus erythematosus and interstitial lung disease. Observations: We report 2 cases of autoimmune bullous skin disease occurring in patients undergoing TNF-targeted therapy: a bullous pemphigoid and a pemphigus foliaceus. Both patients were treated by anti-TNF agents for rheumatoid arthritis and showed improvement following interruption of that treatment. Here, we discuss the relationship between anti-TNF therapy and the occurrence of autoimmune bullous disease. Conclusion: Anti-TNF agents should be considered as a potential cause of drug-induced autoimmune bullous skin disease.

Clinical and Serologic Baseline and Follow-Up Features of Syphilis According to HIV Status in the Post-HAART Era

There is a lack of large studies appraising the effect of the human immunodeficiency virus (HIV) on the course of syphilis since the advent of highly active antiretroviral therapy (HAART). We aimed to appraise the effect of HIV on clinical and serologic features of syphilis at baseline and during follow-up in the post-HAART era. We designed a retrospective cohort study of consecutive syphilis cases, diagnosed between 2000 and 2007, in an academic venereal disease center. Data were collected using standardized medical forms. Patients were treated according to the European guidelines. Serologic failure was defined as either a 4-fold rise in Venereal Disease Research Laboratory (VDRL) titers 30-400 days posttreatment or a lack of 4-fold drop in VDRL titers at 270-400 days posttreatment. Among 279 syphilis cases with informative baseline clinical and serologic data, HIV infection was significantly associated with men having sex with men, French origin, multiple partners, lesser usage of condom, history of sexually transmitted disease, early syphilis, anal primary chancre, and cutaneous eruption. Median baseline titer from the Treponema pallidum hemagglutination assay (TPHA) was higher in HIV-infected patients (p = 0.02). Among 144 informative syphilis cases, there was a nonsignificant trend for a lower rate of serologic response among HIV-positive patients (91.8% vs. 98.3%, p = 0.14). Serologic failure was significantly associated with a history of previous syphilis (p < 0.05). The median delay to serologic response was similar in HIV-positive (117 d) and in HIV-negative (123 d) patients (p = 0.44). We conclude that for patients under HAART treatment, the effect of HIV on serologic response to syphilis treatment is likely minimal or absent. Abbreviations: CSF = cerebrospinal fluid, FTA-abs = fluorescent treponemal antibody absorption test, HAART = highly active antiretroviral therapy, HIV = human immunodeficiency virus, HR = hazard ratio, MSM = men having sex with men, RPR = rapid plasma reagin, STD = sexually transmitted disease, TPHA = Treponema pallidum hemagglutination assay, VDRL = Venereal Disease Research Laboratory.

Molecular subtyping of Treponema pallidum in Paris, France.

Two major Treponema pallidum subtypes, 14 d/g and 14 d/f, were identified in a population of 119 patients with syphilis in Paris, France, characterized by a high proportion of men who have sex with men. A new subtype named 11 q/j was characterized, and a reinfection case was determined in 1 patient having consecuitve syphilis infection at 19-month interval.

Remodeling of VE-cadherin junctions by the human herpes virus 8 G-protein coupled receptor

Kaposi Sarcoma (KS) are opportunistic tumors, associated with human herpes virus 8 (HHV8) infection. KS development is highly favored by immune-depression and remains the second most frequent tumor in acquired immune deficiency syndrome patients. Although it has been shown that experimental expression of the HHV8 G-protein-coupled receptor (vGPCR) in the endothelial compartment is alone sufficient to recapitulate the formation and progression of KS-like lesions, its functional effects on endothelial homeostasis are not fully understood. Here we show that vGPCR expression in endothelial cells induces an increase in paracellular permeability both in vivo and in vitro. By using pharmacological inhibitors and small interference RNA-based knockdown, we demonstrate an essential role for the PI(3)Kinase-γ/Rac nexus in vGPCR-mediated permeability. This was further accompanied by dramatic remodeling of VE-cadherin-dependent cell–cell junctions. Importantly, this in vitro vGPCR-initiated signaling signature was observed in a large panel of human KS. Altogether, our results support the hypothesis that endothelial vGPCR signaling is co-opted in KS, and unveil new key cellular targets for therapeutic intervention.

Does inflammatory acne result from imbalance in the keratinocyte innate immune response

Acne is a multifactorial chronic disease affecting around 80% of teenage population. The pathogenesis of acne involves inflammatory reactions and colonization by the Propionibacterium acnes (P. acnes) strain. P. acnes stimulates the keratinocytes involved in the innate immune response, the intensity of which could be influenced either by bacterial intrinsic factors or by endogenous factors of the host.