Mitochondrial ROS-dependent CD4+PD-1+T cells are pathological expansion in patients with primary immune thrombocytopenia

Abstract

Objective: Aberrant-activated T cells, especially CD4⁺T cells, play a crucial part in the pathogenetic progress of immune thrombocytopenia (ITP). PD-1-mediated signals play a negative part in the activation of CD4⁺T cells. However, knowledge is limited on the pathogenic characteristics and function of CD4⁺PD-1⁺T cells in ITP.

Materials and methods: The frequency and phenotype including cell activation, apoptosis, and cytokine production of CD4⁺PD-1⁺T cells were evaluated by flow cytometry. PD-1 Ligation Assay was performed to assess the function of PD-1 pathway in CD4⁺T cells. Mitochondrial reactive oxygen species (mtROS) were detected by MitoSOX Red probe.

Results: Compared with healthy controls (HC), the frequencies of CD4⁺PD-1⁺T cells were significantly increased in ITP patients. However, these cells are not exhausted despite PD-1 expression. Besides retaining cytokine-producing potential, these CD4⁺PD-1⁺T cells also had a possible B-cell helper function including expressing ICOS, CD84, and CD40L. Moreover, the CD4⁺PD-1⁺T cell subset contained higher levels of mitochondrial ROS than CD4⁺PD-1^-T cell subset in patients with ITP. And mtROS inhibition could reduce the secretion of the inflammatory cytokines and regulate the function of CD4⁺PD-1⁺T cells. Upon in-vitro T cell receptor (TCR) stimulation of CD4⁺T cells in the presence of plate-bound PD-L1 fusion protein (PD-L1-Ig), CD4⁺T cells from ITP patients appeared resistant to such PD-1-mediated inhibition of interferon (IFN)-γ secretion.

Conclusions: The CD4⁺PD-1⁺T cells were more abundant in patients with ITP. Additionally, this CD4⁺PD-1⁺T cell subset may be a potential etiology of ITP and a potential immune therapeutic target for ITP patients in the future.

Keywords: CD4(+)T cell; Immune thrombocytopenia (ITP); Mitochondrial reactive oxygen species (mtROS); PD-1.