Abstract W P89: Adoptive Regulatory T Cell Therapy Preserves Systemic Immune Homeostasis Following Cerebral Ischemia

Abstract

Background and Purpose: Cerebral ischemia has been shown to result in peripheral inflammatory responses followed by long-lasting immunosuppression. Our recent study demonstrated that intravenous delivery of Tregs markedly protected against transient cerebral ischemia by suppressing neutrophil-derived matrix metallopeptidase-9 production in the periphery. The impact of Tregs on systemic inflammatory responses and immune status, however, has not been fully characterized. Methods: Cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) for 60 minutes in mice or 120 minutes in rats. Tregs were isolated from donor animals by CD4 and CD25 double selection and transferred intravenously into ischemic recipients at 2 hours after MCAO. Animals were sacrificed on different days after reperfusion. The effects of Tregs on systemic inflammation and immune status were evaluated using flow cytometry, ELISAs, and immunohistochemistry. Results: Flow cytometry showed the percentage of CD25 and Foxp3 double positive cells were 2.8±0.2% in the blood, 13.9±1.7% in the lymph node and 11.2±0.5% in the spleen after MCAO; intravenous administration of 2х106 purified Tregs raises functional Tregs up to 4.6±0.6% in the blood, 18.5±1.9% in the lymph node and 15±0.9% in the spleen. The exogenous Tregs as labeled by CD45.1 remain in the blood and peripheral organs for at least 12 days. Functionally, Treg adoptive transfer markedly inhibits MCAO-induced elevation of inflammatory cytokines (IL-6 and TNF-α) in the blood as measured at 1 day, 3 days and 7days after ischemia. Furthermore, Treg treatment corrects long-term lymphopenia and improves cellular immune functions after ischemic brain injury. As a result, Treg-treated animals exhibited decreased bacterial loads in the blood during the recovery from cerebral ischemic attack. Conclusions: Treg treatment did not exacerbate post-stroke immunosuppression. On the contrary, Treg-treated animals displayed improved immune status after focal cerebral ischemia.