Myeloid Pannexin-1 mediates acute leukocyte infiltration and leads to worse outcomes after brain trauma.

BACKGROUND:Neuroinflammation is a major component of secondary damage after traumatic brain injury (TBI). We recently reported that pharmacological inhibition of Pannexin-1 (Panx1) channels markedly reduced the inflammatory response after TBI. Panx1 channels have been shown to be important conduits for adenosine 5'-triphosphate (ATP) release and are associated with leukocyte infiltration and pyroptosis. Because Panx1 blockers significantly decrease ATP release and migration of activated microglia and other myeloid cells (such as monocyte-derived macrophages and dendritic cells) in vitro, we hypothesized that myeloid Panx1 channels play a specific role in immune cell infiltration promoting tissue damage following TBI. METHODS:The murine-controlled cortical impact (CCI) model was used on myeloid-specific Panx1 conditional knockout (Cx3cr1-Cre::Panx1^fl/fl) mice to determine whether myeloid Panx1 mediates neuroinflammation and brain damage. Immune cell infiltration was measured using flow cytometry. Locomotor and memory functions were measured using the rotarod and Barnes maze test, respectively. The levels of biomarkers for tissue damage and blood-brain barrier leakage were measured using western blot and magnetic resonance imaging. Panx1 channel activity was measured with ex vivo dye uptake assays, using flow cytometry and confocal microscopy. RESULTS:CCI-injured Cx3cr1-Cre::Panx1^fl/fl mice showed markedly reduced immune cell infiltration to the brain parenchyma compared with Panx1^fl/fl mice. As expected, Panx1 dependent activity, assessed by dye uptake, was markedly reduced only in myeloid cells from Cx3cr1-Cre::Panx1^fl/fl mice. The expression of biomarkers of tissue damage was significantly reduced in the CCI-injured Cx3cr1-Cre::Panx1^fl/fl mice compared with Panx1^fl/fl mice. In line with this, magnetic resonance imaging showed reduced blood-brain barrier leakage in CCI-injured Cx3cr1-Cre::Panx1^fl/fl mice. There was also a significant improvement in motor and memory function in Cx3cr1-Cre::Panx1^fl/fl mice when compared with Panx1^fl/fl mice within a week post-CCI injury. CONCLUSION:Our data demonstrate that CCI-related outcomes correlate with Panx1 channel function in myeloid cells, indicating that activation of Panx1 channels in myeloid cells is a major contributor to acute brain inflammation following TBI. Importantly, our data indicate myeloid Panx1 channels could serve as an effective therapeutic target to improve outcome after TBI.

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