Background Systemic sclerosis (SSc) is an autoimmune disease characterized by vascular injury, autoimmune phenomena, inflammation, and fibrosis of the skin and various internal organs. Several lines of evidence indicate that abnormal B-cell function plays a key role in the development of SSc. The anti-CD20 monoclonal antibody therapy seems to show some clinical efficacy in SSc further emphasizing the importance of B cells in the pathomechanism of the disease. The B-cell compartment in peripheral blood of SSc patients contains an elevated number of naive and a decreased number of memory B cells.
Objectives The aim of the present research was to set up an algorithm for the extended analysis of these B-cell subsets and to evaluate the clinical significance of the defined subpopulations.
Methods Peripheral blood samples were obtained from SSc patients and healthy controls, PBMCs were isolated using ficoll gradient centrifugation, followed by magnetic bead separation of CD19+ B cells. Multiparametric flow cytometry was performed with antibodies specific for CD27, IgD, CD80, CD95 molecules. Detection of CD27 and IgD was applied to distinguish between naive (CD27-IgD+) and memory (CD27+) B cells. IgD posivity was also used to separate non-switched (CD27+IgD+) and switched (CD27+IgD-) memory subsets. In addition to expression of CD80, which provides a co-stimulatory signal necessary for T cell activation and survival, expression of CD95 – FAS receptor was also examined to investigate the activation state of the previously identified B cell subpopulations.
Results The ratio of naive B cells was higher, the proportion of memory B cells, including both subsets, was decreased in SSc patients compared to healthy controls. Among SSc patients the ratio of switched memory and CD95+ memory B cells was higher in diffuse cutaneous SSc and in patients with pulmonary fibrosis. The proportion of switched memory B cells was also elevated in the anti-Scl-70 antibody positive group compared to ACA positive patients. In dcSSc patients the ratio of CD95+ memory B cells was also higher.
Conclusions According to our results detailed flow cytometric analysis of naive and memory B-cell subsets could contribute to better distinction between the two SSc subtypes and to the evaluation of disease severity, consequently may be a useful new tool in routine immunological diagnostics.
Acknowledgements This work was supported by Hungarian Scientific Research Fund - OTKA 75912 and 112939.
Disclosure of Interest None declared