In Vivo Expansion of Regulatory T Cells with IL-2/IL-2 Antibody Complex Protects against Transient Ischemic Stroke

Abstract

Regulatory T cells (Tregs) are known to protect against ischemic stroke. However, the low frequency of Tregs restricts their clinical utility. This study investigated whether expanding the number of Tregs in vivo with the IL-2/IL-2 antibody complex (IL-2/IL-2Ab) could improve stroke outcomes and further elaborated the mechanisms of protection in male mice. C57BL/6 mice received IL-2/IL-2Ab or isotype IgG (IsoAb) intraperitoneally for 3 d before (pretreatment) or starting 2 h after (posttreatment) 60 min middle cerebral artery occlusion (MCAO). IL-2/IL-2Ab selectively increased the number of Tregs in the blood, spleen, and lymph nodes. The IL-2/IL-2Ab treatment significantly reduced infarct volume, inhibited neuroinflammation, and improved sensorimotor functions, as manifested by rotarod test and foot fault test, compared with IsoAb-treated stroke mice. Treg depletion was then achieved by diphtheria toxin (DT) injection into transgenic mice expressing the DT receptor under the control of the Foxp3 promoter (DTR mice). The depletion of Tregs completely eliminated IL-2/IL-2Ab-afforded neuroprotection. Interestingly, adoptive transfer of Tregs collected from IL-2/IL-2Ab-treated mice demonstrated more potent neuroprotection than an equal number of Tregs prepared from IsoAb-treated mice, suggesting that IL-2/IL-2Ab not only elevated Treg numbers, but also boosted their functions. Mechanistically, IL-2/IL-2Ab promoted the expression of CD39 and CD73 in expanded Tregs. CD73 deficiency diminished the protective effect of IL-2/IL-2Ab-stimulated Tregs in stroke mice. The results show that IL-2/IL-2Ab expands Tregs in vivo and boosts their immunomodulatory function. The activation of CD39/CD73 signaling in Tregs may participate as a potential mechanism underlying IL-2/IL-2Ab-afforded neuroprotection against ischemic brain injury.SIGNIFICANCE STATEMENT Regulatory T cells (Tregs) are known to protect against ischemic stroke. However, the low frequency of Tregs restricts their clinical utility. This study reported that systemic administration of the IL-2/IL-2 antibody complex (IL-2/IL-2Ab) robustly and selectively expanded the number of Tregs after stroke. IL-2/IL-2Ab pretreatment or posttreatment significantly improved stroke outcomes in a rodent model of ischemic stroke. We further discovered that IL-2/IL-2Ab not only elevated Treg numbers, but also boosted their functions and enhanced the expression of CD39 and CD73. Using CD73-deficient mice, we confirmed the importance of CD73 in the protective effect of IL-2/IL-2Ab-stimulated Tregs in stroke mice. These results shed light on IL-2/IL-2Ab as a clinically feasible immune therapy to boost endogenous Treg responses and ameliorate ischemic brain injury.

Keywords: IL-2/IL-2 antibody complex; cerebral ischemia; neuroprotection; regulatory T cells.