Reactive oxygen homeostasis – the balance for preventing autoimmunity

Being mainly known for their role in the antimicrobial defense and collateral damage they cause in tissues as agents of oxidative stress, reactive oxygen species were considered “the bad guys” for decades. However, in the last years it was shown that the absence of reactive oxygen species can lead to the development of immune-mediated inflammatory diseases. Animal models of lupus, arthritis and psoriasis revealed reactive oxygen species-deficiency as a potent driver of pathogenesis. On the contrary, in chronic stages oxidative stress can still contribute to progression of inflammation. It seems that a neatly adjusted redox balance is necessary to sustain an immune state that both prevents the development of overt autoimmunity and attenuates chronic stages of disease

No evidence of pathogenic involvement of cathelicidins in patient cohorts and mouse models of lupus and arthritis

Apart from their role in the immune defence against pathogens evidence of a role of antimicrobial peptides (AMPs) in autoimmune diseases has accumulated in the past years. The aim of this project was to examine the functional impact of the human cathelicidin LL-37 and the mouse cathelicidin-related AMP (CRAMP) on the pathogenesis of lupus and arthritis. Serum LL-37 and anti-LL-37 levels were measured by ELISA in healthy donors and patients with Systemic Lupus Erythematosus (SLE) and Rheumatoid arthritis (RA). Pristane-induced lupus was induced in female wild type (WT) and cathelicidin-deficient (CRAMP-/-) mice. Serum levels of anti-Sm/RNP, anti-dsDNA, and anti-histone were determined via ELISA, cytokines in sera and peritoneal lavages were measured via Multiplex. Expression of Interferon I stimulated genes (ISG) was determined by real-time PCR. Collagen-induced arthritis (CIA) was induced in male WT and CRAMP-/- mice and arthritis severity was visually scored and analysed histomorphometrically by OsteoMeasure software. Serum levels of anti-LL-37 were higher in SLE-patients compared to healthy donors or patients with RA. However, no correlation to markers of disease activity or organ involvement was observed. No significant differences of autoantibody or cytokine/chemokine levels, or of expression of ISGs were observed between WT and CRAMP-/- mice after pristane-injection. Furthermore, lung and kidney pathology did not differ in the absence of CRAMP. Incidence and severity of CIA and histological parameters (inflammation, cartilage degradation, and bone erosion) were not different in WT and CRAMP-/- mice. Although cathelicidins are upregulated in mouse models of lupus and arthritis, cathelicidin-deficiency did not persistently affect the diseases. Also in patients with SLE, autoantibodies against cathelicidins did not correlate with disease manifestation. Reactivity against cathelicidins in lupus and arthritis could thus be an epiphenomenon caused by extensive overexpression in blood and affected tissues. In addition, other cationic AMPs could functionally compensate for the deficiency of cathelicidins

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Complexes of cathelicidins and RNA or DNA induce low production of IFNα.

<p>Bars show means and SEM of IFNα concentrations in supernatants from isolated human (A) or mouse (B) pDCs after incubation with free genomic DNA, free self-RNA, or DNA/RNA complexed to LL-37 (A) or (CRAMP). IFNα production from unstimulated pDCs or pDCs incubated with LL-37/CRAMP only was determined as a negative control, CpG2216 and R848 (Resiquimod) were used as a positive control for stimulation of TLR9 and TLR7/8, respectively. *p<0.05, as determined by Student’s t-test. N = 4–7.</p

Autoantibodies against LL-37 in SLE are not linked to disease activity.

<p>(A) In patients with SLE (n = 185) but not with RA (n = 92) or normal healthy donors (NHD, n = 74) serum autoAbs to LL-37 are detected. Shown are optical density (OD) indices, which are the ratios of the OD in the individual donor to the mean OD in NHD sera. One dot represents one patient, horizontal bars show the means. ***P<0.001, as determined by Kruskal-Wallis test with Dunn’s post-hoc test. Dashed line shows cutoff OD (1,93). (B) Sera of SLE patients (n = 21) were preincubated with saturating concentrations of LL-37 peptide before reactivity to plate-bound LL-37 was determined by ELISA. IgG reactivity to LL-37 can be blocked by preincubation with LL-37 peptide. ***P<0.001, as determined by Mann-Whitney U test. (C) Serum levels of LL-37 peptide are raised in a subgroup of SLE and RA patients. One dot represents one donor. Dashed line shows cutoff OD (2,77 ng/ml). (D) No correlation is found between SLE anti-LL-37 autoAbs and disease parameters of SLE. X-Y scatter plots of correlations between OD indices of anti-LL-37 autoAbs in SLE patients and SLEDAI (n = 163), anti-dsDNA autoAbs (n = 73), or serum levels of C-reactive protein (CRP, n = 74) or of IFNα (n = 28), respectively. Spearman coefficients (r) are depicted within the graphs. N.s., not significant. (E) Serum levels of LL-37 peptide are not linked to SLE disease activity. X-Y scatter plot of correlation between serum LL-37 levels and SLEDAI in 110 patients with SLE. Spearman coefficient (r) is depicted within the graph. N.s., not significant.</p

Percentage of CRAMP-expressing cells is higher in pristane-injected animals.

<p>Percentage of CRAMP+ cells in peritoneal lavage and peripheral blood of naïve mice and mice 7 days after pristane-injection. N = 5. *P<0.001, as determined by Student’s t-test.</p