Moreover, the levels of phosphatidylethanolamine-conjugated form of microtubule-associated protein light chain 3 (LC3)-II (LC3-II) are widely used to monitor autophagic activity in cells. ![]() The products of autophagy-related gene (Atg) genes orchestrate every autophagy step ensuring the correct advancing of the process. During autophagy, portions of the cytoplasm are sequestered within characteristic double-membrane vesicles known as autophagosomes and are finally delivered to lysosomes for bulk degradation. Despite the relatively high efficacy of TNF antagonists, approximately one third of patients are intolerant or non-responsive to such drugs, thus the discovery of new therapeutic targets is desirable in RA management.Īutophagy (or macroautophagy) is an evolutionarily conserved lysosomal degradation pathway in eukaryotic organisms. Different immune cell types including fibroblast-like synoviocytes (FLS) and lymphocytes take part in both joints-restricted and extra-articular manifestations of the disease, and tumor necrosis factor alpha (TNFα) is clearly the most important cytokine in RA. Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by autoantibodies production as a result of breakage of immunological tolerance mechanisms. Clinical, demographic and serological characteristics of patients with RA enrolled for sorting experiments ( n = 8). AV on x axis), chosen as representative of five experiments, are also shown. Representative dot plots (PI on y axis vs. Apoptosis is expressed as percentage of AV-positive cells. Flow cytometry analysis of apoptosis in PBMCs after treatment with TNFα. Allophycocyanin = APC Peridinin chlorophyll protein = PerCP Phycoerythrin = PE Fluorescein isothiocyanate = FITC. Flow cytometry gate strategy for purification of CD4+, CD8+ T lymphocytes and B lymphocytes from patients with RA. Changes in autophagy and apoptosis before and after treatment with anti-TNF drugs in relation to anti-CCP in RA responding patients. Correlation between anti-CCP antibodies levels (U/ml) and spontaneous autophagy (expressed as LC3-II) and apoptosis (expressed as percentage of Annexin V-positive cells). Levels of anti-cyclic citrullinated peptide antibodies (anti-CCP Abs) in patients with RA before and after treatment with anti-TNF drugs. Relation between autophagy and citrullination in RA patients. Autophagy and apoptosis levels in PBMCs isolated from responding (A) and non-responding (B) patients to anti-TNF drugs alone or anti-TNF drugs plus methotrexate.
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