Background and Objectives Recent studies in our laboratory have revealed that a significant portion of SS patients manifests significantly impaired phagocytosis of apoptotic cells (ApoCell-phagocytosis), in a manner similar to SLE, a fact which probably leads to the inflammatory and autoimmune responses that characterise these two disorders. Furthermore, our data indicate that approximately 80% of sera from patients with SS and SLE, but not those with RA and healthy individuals, inhibit the clearance of early apoptotic cells from healthy monocytes. In the present study, we sought to investigate the role of IgG immunoglobulins from patients with SLE and SS in the phagocytosis of apoptotic cells.
Materials and Methods Total IgG immunoglobulin was isolated from the serum of patients with SS (n = 24), SLE (n = 12) and RA (n = 8) and healthy donors (n = 11) using Melon Gel Resin columns. Apoptotic Jurkat cells (induction of apoptosis by UV-radiation) were incubated with purified IgG (50 µg/ml) in PBS/BSA 1%. The binding of IgG on the surface of apoptotic cells was assessed by flow cytometry and analysed with the binding index (% positive cells × mean fluorescence intensity). ApoCell-phagocytosis was assessed by flow cytometry using monocyte-derived macrophages (MDM) from healthy individuals (n = 3) that were incubated (90 min) with CFSE-labelled early apoptotic Jurkat cells which had been opsonised with IgG from patients or healthy individuals or with PBS (control).
Results IgG from patients with SS and SLE exhibited high binding levels in apoptotic cells compared to healthy donors and patients with RA (all for p < 0.0001). Pre-incubation of apoptotic cells with IgG from patients with both SS and SLE had a significant inhibitory effect on the phagocytosis process (comparative ApoCell-phagocytosis: after pre-incubation with PBS [x = 100%], with IgG from healthy individuals [n = 11, x = 100%], with IgG from RA patients [n = 4, x = 100%], with IgG from SS patients [n = 17, x = 75%, p < 0.0001], with IgG from SLE patients [n = 10, x = 71%, p = 0.0002]). Finally, negative correlation was found between the apoptotic cell binding levels of IgG from patients and healthy individuals and the results of ApoCell-phagocytosis assays using apoptotic cells pre-incubated with the corresponding IgG (n = 42, p < 0.0001).
Conclusions Our results indicate that the impaired phagocytosis of apoptotic cells observed in patients with SS and SLE is primarily due to the presence of antibodies reactive with the surface of apoptotic cells. Further studies need to focus on the mechanism by which immunoglobulins exert such blocking effect, as well as its immunological consequences.
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