Selective inactivation or reconstitution of adenosine A2A receptors in bone marrow cells reveals their significant contribution to the development of ischemic brain injury

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Abstract

Inactivation of the adenosine A2A receptor (A2AR) consistently protects against ischemic brain injury and other neural insults, but the relative contribution of A2ARs on peripheral inflammatory cells versus A2ARs expressed on neurons and glia is unknown. We created a chimeric mouse model in which A2ARs on bone marrow–derived cells (BMDCs) were selectively inactivated or reconstituted by bone marrow transplantation. Selective reconstitution of A2ARs on BMDCs (A2AR knockout mice transplanted with wild-type bone marrow cells) largely reinstates ischemic brain injury in global A2AR knockout mice. Conversely, selective inactivation of A2ARs on BMDCs (wild-type mice transplanted with A2AR knockout bone marrow cells) attenuates infarct volumes and ischemia-induced expression of several proinflammatory cytokines in the brain, but exacerbates ischemic liver injury. These results indicate that the A2AR-stimulated cascade in BMDCs is an important modulator of ischemic brain injury and that ischemic brain and liver injuries are regulated distinctly by A2ARs on BMDCs.

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Figure 1: Generation of chimeric mice with selective inactivation or reconstitution of the A2AR on BMDCs.
Figure 2: Selective reconstitution of A2ARs on BMDCs largely reinstates MCAO-induced ischemic brain injury, as shown by infarct volumes and infarct areas in brains of three groups of chimeric A2AR knockout mice (WT→WT, black; KO→KO, light gray; and WT→KO, white).
Figure 3: Selective inactivation of A2ARs on BMDCs attenuates MCAO-induced brain injury.
Figure 4: Neutrophil infiltration and microglial activation in ischemic brain were indistinguishable between the WT→WTcr and KO→WTcr mice.
Figure 5: Selective inactivation of A2ARs on BMDCs alters MCAO-induced cytokine mRNA expression in brain.
Figure 6: Selective inactivation of the A2AR on BMDCs exacerbates ischemic liver injury and enhanced proinflammatory cytokine expression, as shown by activity of serum GPT (a) and abundance of mRNA (b) for proinflammatory cytokines.

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Acknowledgements

We thank B.B. Fredholm and M.A. Schwarzschild for discussion and critical reading of the manuscript and D. Xu for assistance with FACS analysis. This work was supported in part by grants NS37403, NS41083 (J-F.C.) and NS-10828 (M.M.) from the US National Institutes of Health and by Bumpus Foundation (J-F.C.).

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Correspondence to Jiang-Fan Chen.

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Supplementary information

Supplementary Table 1

Cerebrovascular and systemic physiology before, during and after MCAO-induced ischemia in WT→WT and KO→WT mice (PDF 19 kb)

Supplementary Table 2

Primers sequences for the genotyping and qPCR analysis (PDF 21 kb)

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