Dysfunction of endothelial progenitor cells is associated with the type I IFN pathway in patients with polymyositis and dermatomyositis

Objective. Alterations in phenotype and function of endothelial progenitor cells (EPCs) have been associated with poor vascular outcomes and impaired vascular repair in various conditions. Our hypothesis was that patients with PM and DM have dysregulation of EPCs driven by type I IFN and IL-18 similar to other autoimmune diseases. Methods. Quantification of circulating EPCs was performed by flow cytometry in patients with PM/DM and matched healthy controls. The ability of EPCs to differentiate into mature endothelial cells was investigated by light and fluorescence microscopy quantification in the presence or absence of PM/DM or control serum, neutralizing antibodies to type I IFN receptor or IL-18. Serum type I IFN activity was quantified by induction of type I IFN-inducible genes in HeLa cells. Circulating IL-18 concentrations were assessed by ELISA. Results. Circulating EPCs were significantly lower in PM/DM patients compared with controls. PM/DM EPCs displayed a decreased capacity to differentiate into mature endothelial cells and PM/DM serum significantly inhibited differentiation of control EPCs. This effect was reversed in the majority of samples with neutralizing antibodies to IL-18 or to type I IFN receptor or by a combination of these antibodies. Patients with associated impairments in EPC function had higher type I IFN serum activity. Conclusion. PM/DM is associated with dysregulation of EPC phenotype and function that may be attributed, at least in part, to aberrant IL-18 and type I IFN pathways. The implication of these vasculopathic findings for disease prognosis and complications remains to be determined

The role of autoantibodies in inflammatory myopathies

Polymyositis (PM), dermatomyositis (DM) and inclusion body myositis (IBM), collectively called myositis, are chronic inflammatory myopathies. These disease subsets are heterogeneous but share some features, such as skeletal muscle weakness and inflammatory cell infiltrates in muscle tissue. In many patients involvement of other organs, in particular skin and lung, also occurs. Another characteristic feature is the presence of autoantibodies which are (1) myositis specific, e.g. anti-histidyl-transfer ribonucleic acid (RNA) synthetase (Jo-1) and anti-Mi-2, and (2) myositis associated e.g. anti-Ro52/SSA, anti-Ro60/SSA, anti-PM/Scl and anti-U1sn ribonucleoprotein (RNP) autoantibodies. Presence of anti-Jo-1 autoantibodies is specifically associated with interstitial lung disease (ILD). Despite the clinical associations it is not known whether these autoantibodies have a pathogenic role or are merely an epiphenomenon in the disease mechanism. Aim: The overall goal of this thesis was to evaluate the role of specific and associated autoantibodies in myositis pathogenesis by combining in vivo and in vitro approaches. Results: We demonstrated in vitro that immune complexes containing anti-Jo-1 or anti-Ro52/Ro60 autoantibodies and RNA may act as endogenous type I interferon (IFNalpha/beta) inducers via plasmacytoid dendritic cells (PDCs). Patients also had increased expression of IFNalpha/beta-inducible human myxovirus resistance 1 (MX-1) protein in muscle tissue which correlated with number of blood dendritic cell antigen (BDCA)-2 positive PDCs. DM patients characterized by skin rash also had increased MX-1 expression in muscle tissue but this was located to capillaries, suggesting another induction mechanism and cellular IFNalpha source such as the skin. The same groups of patients displayed elevated levels of B cell activating factor (BAFF), suggesting an induction by a local (muscle and skin) type I IFN system. Sera from PM patients with anti-Jo-1 autoantibodies and ILD exhibited induced intracellular adhesion molecule (ICAM)-1 expression in lung endothelial cells in vitro, which has clinical relevance as ICAM-1 is upregulated in vivo in endothelial cells of capillaries in muscle tissue. The effects of high dose intravenous immunoglobulin (IVIG) treatment on endothelial cell activation and other immunological molecules investigated in muscle tissue varied between patients and were not correlated with improved muscle function. This could be due to the heterogeneity of the disease mechanisms in myositis. The effects of IVIG treatment were thus considered as being limited. Conclusion: There seems to be a role for anti-Jo-1 autoantibodies in the pathogenesis of myositis patients with lung and skin involvement. The findings in this thesis suggest that this role is related to the endogenous production of type I IFN, which perpetuates disease. Sera with anti-Jo-1 autoantibodies are capable of inducing endothelial cell activation, enabling the influx of inflammatory cells into the muscle tissue. An inflammatory condition sustained by inflammatory cells and their mediators may induce changes in muscle fiber metabolism which can lead to muscle damage

Dysfunction of endothelial progenitor cells is associated with the type I IFN pathway in patients with polymyositis and dermatomyositis

Objective. Alterations in phenotype and function of endothelial progenitor cells (EPCs) have been associated with poor vascular outcomes and impaired vascular repair in various conditions. Our hypothesis was that patients with PM and DM have dysregulation of EPCs driven by type I IFN and IL-18 similar to other autoimmune diseases. Methods. Quantification of circulating EPCs was performed by flow cytometry in patients with PM/DM and matched healthy controls. The ability of EPCs to differentiate into mature endothelial cells was investigated by light and fluorescence microscopy quantification in the presence or absence of PM/DM or control serum, neutralizing antibodies to type I IFN receptor or IL-18. Serum type I IFN activity was quantified by induction of type I IFN-inducible genes in HeLa cells. Circulating IL-18 concentrations were assessed by ELISA. Results. Circulating EPCs were significantly lower in PM/DM patients compared with controls. PM/DM EPCs displayed a decreased capacity to differentiate into mature endothelial cells and PM/DM serum significantly inhibited differentiation of control EPCs. This effect was reversed in the majority of samples with neutralizing antibodies to IL-18 or to type I IFN receptor or by a combination of these antibodies. Patients with associated impairments in EPC function had higher type I IFN serum activity. Conclusion. PM/DM is associated with dysregulation of EPC phenotype and function that may be attributed, at least in part, to aberrant IL-18 and type I IFN pathways. The implication of these vasculopathic findings for disease prognosis and complications remains to be determined. <br/

Expression of interleukin-18 in muscle tissue of patients with polymyositis or dermatomyositis and effects of conventional immunosuppressive treatment

Objectives: To investigate the expression of IL-18 in symptomatic and asymptomatic muscle tissues of patients with PM and DM and the effects of conventional immunosuppressive treatment on such expression. Methods: Two cohorts of patients were included in this study. The first cohort consisted of 10 new-onset myositis patients. IL-18 expression was compared between symptomatic and asymptomatic muscle biopsies that were taken prior to treatment. The second cohort consisted of another 10 patients with repeated muscle biopsies before and after 8 months with conventional immunosuppressive treatment. Using immunohistochemistry, IL-18 expression in muscle tissues was compared before and after treatment. Biopsies from seven healthy individuals were included as controls. Results: IL-18 expression was predominantly localized to inflammatory cells and capillaries in patients and mostly to capillaries in healthy controls. Total IL-18 expression in muscle tissues from the new-onset patients, at both symptomatic and asymptomatic sites, was significantly higher compared with healthy controls (P = 0.007 and P = 0.002) with no statistical difference in appearances between symptomatic and asymptomatic sites. The number of IL-18 positive capillaries was not different among symptomatic, asymptomatic and healthy muscles. Total IL-18 expression appeared lower in biopsies from patients receiving and improving with immunosuppressive treatment, particularly the number of IL-18 positive inflammatory cells but not the number of IL-18 positive capillaries, which was consistent with significantly decreased expression of CD68+ macrophages (P = 0.04). Conclusion: IL-18 is highly expressed in muscle tissue in the context of inflammatory myopathies and based on its plausible effector functions could provide a novel therapeutic target in future

Sera From Anti-Jo-1-Positive Patients With Polymyositis and Interstitial Lung Disease Induce Expression of Intercellular Adhesion Molecule 1 in Human Lung Endothelial Cells

Objective. To investigate whether sera or purified IgG from patients with polymyositis (PM) and patients with dermatomyositis (DM), with or without interstitial lung disease (ILD), can activate endothelial cells (ECs). Methods. Patients' sera were selected based on the presence or absence of anti-Jo-1, anti-SSA, or anti-U1 small nuclear RNP autoantibodies. The presence of autoantibodies was determined by line blot assays. Cultured human microvascular ECs derived from lung tissue (HMVEC-L) were incubated with sera or purified IgG from 22 patients with PM, 7 patients with DM, and 1.0 healthy individuals as controls. Assessment of intercellular adhesion molecule 1 (ICAM-1) expression was conducted by immunofluorescence (n = 22) and by cell-based enzyme-linked immunosorbent assay (ELISA) (n = 20). Serum levels of soluble ICAM-1 (sICAM-1) were determined by ELISA. Results. Sera from PM patients with ILD who were positive for anti-Jo-1 autoantibodies had a significantly stronger effect on the expression of ICAM-1 by HMVEC-L in comparison with sera from healthy controls and patients with other autoantibodies. Purified IgG did not induce ICAM-1 expression. Higher serum levels of sICAM-1 were found in patients with myositis compared with healthy controls. Conclusion. EC activation with ICAM-1 expression could contribute to the multiorgan involvement, including the development of myositis and ILD, in patients carrying anti-Jo-1 autoantibodies. The EC-activating factors are not the autoantibodies themselves, but might be systemic factors associated with these autoantibodies

Intravenous immune globulin suppresses angiogenesis in mice and humans

Human intravenous immune globulin (IVIg), a purified IgG fraction composed of ~ 60% IgG1 and obtained from the pooled plasma of thousands of donors, is clinically used for a wide range of diseases. The biological actions of IVIg are incompletely understood and have been attributed both to the polyclonal antibodies therein and also to their IgG (IgG) Fc regions. Recently, we demonstrated that multiple therapeutic human IgG1 antibodies suppress angiogenesis in a target-independent manner via FcγRI, a high-affinity receptor for IgG1. Here we show that IVIg possesses similar anti-angiogenic activity and inhibited blood vessel growth in five different mouse models of prevalent human diseases, namely, neovascular age-related macular degeneration, corneal neovascularization, colorectal cancer, fibrosarcoma and peripheral arterial ischemic disease. Angioinhibition was mediated by the Fc region of IVIg, required FcγRI and had similar potency in transgenic mice expressing human FcγRs. Finally, IVIg therapy administered to humans for the treatment of inflammatory or autoimmune diseases reduced kidney and muscle blood vessel densities. These data place IVIg, an agent approved by the US Food and Drug Administration, as a novel angioinhibitory drug in doses that are currently administered in the clinical setting. In addition, they raise the possibility of an unintended effect of IVIg on blood vessels

Intravenous immune globulin suppresses angiogenesis in mice and humans

Human intravenous immune globulin (IVIg), a purified IgG fraction composed of ~ 60% IgG1 and obtained from the pooled plasma of thousands of donors, is clinically used for a wide range of diseases. The biological actions of IVIg are incompletely understood and have been attributed both to the polyclonal antibodies therein and also to their IgG (IgG) Fc regions. Recently, we demonstrated that multiple therapeutic human IgG1 antibodies suppress angiogenesis in a target-independent manner via FcγRI, a high-affinity receptor for IgG1. Here we show that IVIg possesses similar anti-angiogenic activity and inhibited blood vessel growth in five different mouse models of prevalent human diseases, namely, neovascular age-related macular degeneration, corneal neovascularization, colorectal cancer, fibrosarcoma and peripheral arterial ischemic disease. Angioinhibition was mediated by the Fc region of IVIg, required FcγRI and had similar potency in transgenic mice expressing human FcγRs. Finally, IVIg therapy administered to humans for the treatment of inflammatory or autoimmune diseases reduced kidney and muscle blood vessel densities. These data place IVIg, an agent approved by the US Food and Drug Administration, as a novel angioinhibitory drug in doses that are currently administered in the clinical setting. In addition, they raise the possibility of an unintended effect of IVIg on blood vessels