Several observations highlight the relevance of regulatory B cells (Breg) to clinical transplantation. We have identified a new Breg population expressing the CD9, able to block T cell proliferation through IL-10, that is missing in a mouse model of chronic lung inflammation. The aim of our study was to determine the molecular mechanism of regulation of these CD9+ Bregs on effectors T cells in Balb-c mouse.

CD19+CD9+ and CD19+CD9- B cells and CD3+CD4+CD25- effector T cell from the spleen were sorted using a FACSAria III and activated for 48h (a-CD40+LPS for B cells and IL-2 for T cells). T cells were then co-culture with CD9+ or CD9- for 48h. T cells alone were used as control. Cell death, cell cycle arrest, apoptosis, and mitochondrial depolarization were determined by Yellow Dye, propidium iodide, Annexin V and JC-1 staining respectively. Caspases cleavage and protein expression of the Bcl-2 family members were estimated by western blotting.

We confirmed that CD9+, but not CD9- cells, induce effector T cells death, and inhibit proliferation with a cycle arrest in G0/G1. CD9+ Bregs activate T cell apoptosis through both intrinsic and extrinsic apoptosis pathways as illustrated by the mitochondrial depolarization and the cleavage of caspase 8-9 and PARP. We also showed that CD9+ Bregs induce expression of pro-apoptotic members of the Bcl-2 family such as Bax and downregulated expression of anti-apoptotics such as Bcl-2, Bcl-XL and Mcl-1. Finally, we confirmed the presence of CD9+ Bregs in human.

These data clearly show that CD9+ Bregs are able to exert a powerful anti-inflammatory function by inducing effector T cell apoptosis. Thus, an uncontrolled inflammation such as chronic lung allograft dysfunction, could results from a deficiency in CD9+ Bregs.

CITATION INFORMATION: Brosseau C, Durand M, Braza F, Durand E, Loy J, Lacoste P, Royer P, Cheminant M.-A, Magnan A, Brouard S. CD9+ Regulatory B Lymphocytes Control Chronic Lung Inflammation by Inducing Effector T Cell Apoptosis. Am J Transplant. 2016;16 (suppl 3).

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