Phenotypic and functional characteristics of CD39highhuman regulatory B cells (Breg)

CD39 and CD73 are key enzymes in the adenosine (ADO) pathway. ADO modulates pathophysiological responses of immune cells, including B cells. It has recently emerged that a subpopulation of ADO-producing CD39⁺CD73⁺ B cells has regulatory properties. Here, we define the CD39^high subset of these cells as the major contributor to the regulatory network operated by human B lymphocytes. Peripheral blood B cells were sorted into CD39^neg, CD39^inter and CD39^high subsets. The phenotype, proliferation and IL-10 secretion by these B cells were studied by flow cytometry. 5'-AMP and ADO levels were measured by mass spectrometry. Agonists or antagonists of A₁R, A_2AR and A₃R were used to study ADO-receptor signaling in B cells. Inhibition of effector T-cell (Teff) activation/proliferation by B cells was assessed in co-cultures. Cytokine production was measured by Luminex. Upon in vitro activation and culture of B cells, the subset of CD39^high B cells increased in frequency (p < 0.001). CD39^high B cells upregulated CD73 expression, proliferated (approximately 40% of CD39^high B cells were Ki-67⁺ and secreted fold-2 higher IL-10 and ADO levels than CD39^neg or CD39^inter B cells. CD39^high B cells co-cultured with autologous Teff suppressed T-cell activation/proliferation and secreted elevated levels of IL-6 and IL-10. The A₁R and A_2AR agonists promoted expansion and functions of CD39^high B cells. CD39 ectonucleotidase is upregulated in a subset of in vitro-activated B cells which utilize ADO and IL-10 to suppress Teff functions. Proliferation and functions of these CD39^high B cells are regulated by A₁R- and A_2AR-mediated autocrine signaling.