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Background and Aims

Distinguishing autoimmune myositis from other muscle disorders with prominent intramuscular inflammation such as muscular dystrophies, is often not easy yet highly relevant for guiding therapeutic decisions and for offering founded prognoses. We investigated the biomarker potential of growth differentiation factor-15 (GDF-15), a cytokine associated with inflammation, to differentiate myositis of autoimmune origin.

Methods

Sera from 35 myositis patients were collected, who had been diagnosed with immune-mediated necrotizing myopathy (n=21), sporadic inclusion body myositis (n=10), polymyositis (n=3), and dermatomyositis (n=1). GDF-15 protein levels were determined using a commercial enzyme-linked immunosorbent assay, and compared with the levels in sera from healthy controls (n=10), and patients with limb girdle muscular dystrophy or mitochondrial myopathies (n=8). Muscle tissue GDF-15 expression patterns were studied using double immunofluorescent staining.

Results

Serum levels of GDF-15 were significantly higher in myositis patients (625±358 pg/ml) than in healthy controls (326±204 pg/ml) (Mann-Whitney U test p=0.01), with no difference between patient subgroups (p=0.1). No correlation was found between GDF-15 levels and blood CK values (p=0.2). Patients with dystrophies displayed substantially lower levels (127±41 pg/ml), while patients with mitochondrial disease displayed the highest mean levels (1312±1393 pg/ml). GDF-15 could be localized to skeletal muscle fibers, with strong sarcoplasmic staining observed in subsets of small CD56 positive regenerating or denervated fibers, and in the inclusions present in sporadic inclusion body myositis fibers.

Conclusions

GDF-15 serum levels could represent a potential biomarker for myositis, with expression by skeletal muscle fibers being a probable source of the cytokine.

Background and Aims

Non-infectious uveitis comprises sight-threatening intraocular inflammatory conditions with unknown etiology. Corticosteroid are the mainstay of therapy. However, tailoring therapies on each individual molecular profile and developing new targeted-therapies are unmet clinical needs. We thus aimed to identify cytokines dysregulated in aqueous humor (AH) from patients with uveitis associated with Behcet’s (BD) and Vogt-Koyanagi-Harada (VKH) disease, two systemic inflammatory diseases.

Methods

Concentrations of 27 cytokines were measured in AH samples from patients with active panuveitis (n=10 BD, n=10 VKH) versus healthy controls (HC, n=10) using a multiplex immunoassay. Leukocytes in AH were counted and characterized by flow cytometry.

Results

IL-6, IP-10, G-CSF, IFNγ showed higher concentrations in AH from both BD and VKH patients while IL-2, IL-4, IL-8, IL-13, TNFα, MIP-1α, eotaxin, IL-1ra showed higher concentrations only in AH from BD patients compared with HC. GM-CSF was the sole cytokine with lower levels in AH from BD patients. No statistically significant differences were found between BD and VKH patients. The concentrations of IL-6, IL-4, IL-8, IP-10, G-CSF, IFNγ, TNFα, MIP-1α, eotaxin, IL-1ra positively correlated with the concentrations of leukocytes in AH, mainly with those of monocytes and granulocytes, suggesting that such cytokines were produced by and/or attracted and/or promoted proliferation and survival of immune cells in these types of uveitis. The AH over plasma cytokine ratios intra-individuals suggested that IL-2, IL-6, IP-10, GM-CSF were produced also intraocularly.

Conclusions

AH sampling followed by cytokine profiling can highlight which cytokines are dysregulated intraocularly in each individual, providing a rationale for tailoring treatments in patients with non-infectious uveitis.

Background and Aims

CCR6 is a G protein-coupled receptor, and ulcerative colitis patients show strong positive with the severity of the disease. How does CCR6 intrinsic signaling in CD4 T cells affect the pathogenicity of Th17 cells during gut inflammation and autoimmunity is not known.

Methods

Immune cells were phenotypically characterized using flow cytometry. FACS purified naïve CD4 T cells were in vitro differentiated into Th17 in the presence or absence of CCL20, and proteomic analysis was performed using mass spectroscopy. Gut inflammation in wild-type C57BL/6 or CCR6^-/- mice was induced by giving dextran sodium sulfate in the drinking water.

Results

Our results showed that ulcerative colitis patients have a significantly high frequency of RORgt⁺T-bet⁺CD4⁺ T cells in PBMCs as compared to healthy individuals. In the presence of CCL20, in vitro differentiation of purified naïve human CD4⁺ T cells into Th17 cells significantly increased the frequency of pathogenic IL-23R⁺Th17 cells (IL-17⁺IFNγ⁺GMCSF⁺ Th17 cells) in the AKT/mTOR/STAT3 signaling dependent manner. Furthermore, CCR6^-/- mice showed reduced inflammatory colitis and lower frequency of pathogenic Th17 in the gut-associated lymphoid tissues as compared to wild-type mice. Deficiency of CCR6^-/- abrogated the CCL20 driven differentiation of pathogenic Th17 cells. Proteomics analysis of Th17 cells differentiated in the presence of CCL20 showed significant alteration in several proteins belonging to various cellular metabolic pathways.

Conclusions

Signaling through the CCL20-CCR6 axis drive the differentiation of pathogenic Th17 cells by altering the cellular metabolism, and a strategy to block the non-chemotactic function of CCR6 may help in controlling the gut inflammation and autoimmunity.

Background and Aims

Rheumatoid Arthritis (RA) is a chronic immune -mediated disease characterized by pro-inflammatory cytokine production, synovial hypertrophy, cartilage and bone destruction resulting in severe disability. Cytokine-mediated immunity plays a major role in the pathogenesis of RA. Interleukin-33 (IL-33) is a pro-inflammatory cytokine from the IL-1 family that is expressed by tissue damage. IL-33 has been detected in high levels and has profound effects in experimental inflammatory arthritis.

The aim of the present study was to investigate the effect of IL-33 in the serum of RA patients and to evaluate the correlation of serum IL-33 with disease activity, laboratory parameters and clinical manifestations.

Methods

Sixty (44 females, 16 males) RA patients, mean age 43.53(10.88) years diagnosed according to the ACR/EULAR criteria for RA and 40 age/sex-matched healthy controls were enrolled in this study. Information on disease duration, disease severity and activity, comorbidities and medications (conventional disease modifying anti-rheumatic drugs and biological therapies) was collected. Serum levels of Il-33 were measured using an enzyme-linked immunoabsorbent assay (ELISA). Disease activity was assessed by the Disease Activity Score 28 (DAS-28).

Results

Serum Il-33 levels were significantly higher in RA patients compared to controls and correlated with disease severity, DAS28, erythrocyte sedimentation rate, C-reactive protein, rheumatoid factor (RF)-IgM, RF-IgG, anticitrullinated peptide antibodies (ACPA), TNFα and IL-6. Serum IL-33 levels decreased significantly in RA patients that had received anti-TNF and IL-6 inhibitors.

Conclusions

IL-33 appears to play a pivotal role in the pathogenesis of RA and could be a new therapeutic target. IL-33 is a reliable biomarker of disease activity and severity..

Background and Aims

Systemic lupus erythematosus is a severe autoimmune disease caused by a combination of genetic, stochastic as well as environmental factors. A number of studies have demonstrated the influence of epigenetic mechanisms in the development of autoimmune diseases. Bregs are of particular importance for the control of autoimmune diseases – deficiency in Bregs can lead to pathological autoimmunity. As changes in the population of regulatory B-cell have been identified in patients with systemic lupus their number is a determining factor for the proper functioning of the immune system.

The aim of the study is to investigate the influence of overmethylation on the function and on the number of human regulatory B cells (Breg).

Methods

PBMCs from lupus patients were isolated and cultured in the presence of different concentrations of folic acid. The level of intracellular IL10 and the percentage of Breg were determined by flow cytometry.

Results

10 lupus patients and 10 healthy donors participated in the study. Two of the patients showed an increase in the IL10 producing Bregs after incubation with folic acid. No differences were observed in the samples of healthy volunteers.

Conclusions

A number of scientific studies confirm the involvement of epigenetic changes in the etiology of systemic lupus. Numerous scientific developments point to the role folic acid as the main modulator of gene expression. Modulation of development of systemic lupus by epigenetic pathways would be a novel scientific approach for modulation of genetic defects at molecular level.