Essential role of program death 1-ligand 1 in regulatory T-cell-afforded protection against blood-brain barrier damage after stroke.
OBJECTIVE
Our recent research revealed that adoptively transferred regulatory T cells (Tregs) reduced acute ischemic brain injury by inhibiting neutrophil-derived matrix metalloproteinase-9 (MMP-9) and protecting against blood-brain barrier damage. The mechanisms underlying Treg interactions with neutrophils remain elusive. This study evaluates the contribution of program death 1-ligand 1 (PD-L1) to Treg-mediated neutrophil inhibition and neuroprotection after cerebral ischemia.


METHODS
In vitro experiments were performed using a transwell system or a coculture system allowing cell-to-cell contact. Focal cerebral ischemia was induced in mice for 60 minutes. Tregs (2×10(6)) isolated from donor animals (wild-type or PD-L1-/-) were intravenously injected into ischemic recipients 2 hours after middle cerebral artery occlusion (MCAO). MMP-9 production, blood-brain barrier permeability, and brain infarct were assessed at 1 or 3 days after MCAO.


RESULTS
In vitro experiments reveal that Treg-mediated inhibition of neutrophil MMP-9 required direct cell-to-cell contact. The suppression of MMP-9 was abolished when Tregs were pretreated with PD-L1 neutralizing antibodies or when neutrophils were pretreated with PD-1 antibodies. In vivo studies confirmed that intravenous administration of Tregs pretreated with PD-L1 antibodies or Tregs isolated from PD-L1-deficient mice failed to inhibit MMP-9 production by blood neutrophils 1 day after 60 minutes MCAO. Furthermore, the blood-brain barrier damage after MCAO was greatly ameliorated in PD-L1-competent Treg-treated mice but not in PD-L1-compromised Treg-treated mice. Consequently, PD-L1 dysfunction abolished Treg-mediated brain protection and neurological improvements 3 days after MCAO.


CONCLUSIONS
PD-L1 plays an essential role in the neuroprotection afforded by Tregs against cerebral ischemia by mediating the suppressive effect of Tregs on neutrophil-derived MMP-9.
From the State Key Laboratory of Medical Neurobiology and Institute of Brain Sciences, Department of Anesthesiology, Huashan Hospital, Fudan University, Shanghai, China (P.L., L.M., X.L., Y. Gao, J.C., X.H.); Center of Cerebrovascular Disease Research (P.L., L.M., Y. Gan, J.Z., J.C., X.H.) and Departments of Surgery and Immunology, Starzl Transplantation Institute (A.W.T.), University of Pittsburgh School of Medicine, PA; Geriatric Research, Educational and Clinical Center, Veterans Affairs Pittsburgh Health Care System, PA (P.L., X.H.); and Cerebrovascular Diseases Research Institute, Xuanwu Hospital of Capital Medical University, Beijing, People's Republic of China (X.L.).
Peiying
P
Li
Leilei
L
Mao
Xiangrong
X
Liu
Yu
Y
Gan
Jing
J
Zheng
Angus W
AW
Thomson
Yanqin
Y
Gao
Jun
J
Chen
Xiaoming
X
Hu
D000818:Animals; D060890:B7-H1 Antigen / Q000502:physiology*; D001812:Blood-Brain Barrier / Q000473:pathology*; D001921:Brain / Q000473:pathology; D002545:Brain Ischemia / Q000473:pathology; D002452:Cell Count; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D007150:Immunohistochemistry; D020244:Infarction, Middle Cerebral Artery / Q000473:pathology; D002546:Ischemic Attack, Transient / Q000473:pathology; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009422:Nervous System Diseases / Q000503:physiopathology; D020556:Neutrophil Infiltration / Q000502:physiology; D009504:Neutrophils / Q000502:physiology; D020521:Stroke / Q000473:pathology*; D050378:T-Lymphocytes, Regulatory / Q000502:physiology*
D060890:B7-H1 Antigen; D016207:Cytokines; D020780:Matrix Metalloproteinase 9
matrix metalloproteinase 9; neutrophils; programmed cell death 1 ligand 1; regulatory T-cells; stroke