Single-cell analysis of autoreactive B cells in patients with systemic sclerosis revealed that responses to therapy are determined by effector and regulatory B cell balance

Abstract

Abstract Objective Systemic sclerosis (SSc) is a connective tissue disease with poor prognosis. Although the standard treatment for skin and lung fibrosis is cyclophosphamide (CYC), there are not a few non-responders to the treatment. Recent studies showed that autoreactive B cells play a critical role in SSc. We hypothesized that these B cells determine the effectiveness of CYC treatment. We focused on cytokine profiles of single autoreactive B cells in patients with SSc. Methods We simulated the interaction of B cells and endothelial cells in microfluidic cell culture systems. We cultured endothelial cells in a microchannel. Then, we loaded B cells of patients with SSc. We collected single B cells which adhere to endothelial cells by trypsin treatment. We stimulated these B cells, with PMA and ionomycin, and measured the levels of produced interleukin (IL)-10 and IL-6 to detect the percentages of regulatory and effector B cells, by our original micro fluidic-ELISA system. Results In SSc patients, the number of B cells adhering to endothelial cells significantly increased compared with healthy controls. Most of these B cells produced anti-endothelial cells antibodies. The frequencies of IL-10-producing regulatory B cells after the CYC treatment increased in responders and decresed in non-responders. Those of IL-6-producing effector B cells after the treatment decreased in responders and increased in non-responders. Conclusions The effectiveness of CYC treatment to SSc patients is associated with cytokine profiles of B cells, which interact with endothelial cells. Checking the profiles of these B cells, after the first few months on from initiation of CYC therapy, may help to decide whether or not we should continue the treatment.