Recombinant growth arrest-specific protein 6 protects the blood–brain barrier by regulating microglia polarization via the GAS6/Axl/SOCS pathway in post-stroke hemorrhagic transformation model

Abstract

Abstract Background Blood–brain barrier (BBB) disruption is the primary cause of hemorrhagic transformation (HT) after ischemic stroke (IS). Axl is well-known as an essential innate immune regulator in macrophages. Our previous study have reported a negative association between serum Axl level and HT risk in patients after tPA thrombolysis, however, the underlying mechanism remains unclear. The present study was designed to investigate whether Axl activation could suppress BBB disruption and reduce HT in post-stroke HT model and the underlying mechanism.Methods and Results In vivo, the post-stroke HT model was established by an injection of 50% glucose and middle cerebral artery occlusion and reperfusion (MCAO/R) surgery 15min later in rats. Recombinant growth arrest-specific protein 6 (rGAS6) and R428 were injected as Axl-specific agonists and antagonists. Neurobehavioral deficits, infarction and hemorrhage volumes, brain edema, and the degree of BBB disruption were assessed. The expressions of GAS6, Axl, and suppressor of cytokine signaling (SOCS) pathway were measured. And the polarization states of microglia and the levels of inflammatory cytokines were analyzed. Our results showed that rGAS6 significantly improved neurological deficits, decreased infarct and hemorrhage volumes, alleviated brain edema and BBB disruption. Additionally, enhanced M2 polarization of microglia and a reduction in the inflammatory response were observed. Mechanism investigations suggested that rGAS6 upregulated Axl phosphorylation and the expressions of SOCS1/3. However, R428 injection abrogated the neuroprotection caused by rGAS6. The in vitro studies further supported the data of in vivo experiments, that rGAS6 treatment enhanced M2 polarization of microglia after oxygen and glucose deprivation/reoxygenation (OGD/R) stimulation via activating GAS6/Axl/SOCS1/3 pathway, which then influenced endothelial cell function.Conclusions Consequently, these data suggested that rGAS6 can protect BBB function and attenuate HT by enhancing microglial M2 polarization through activation of GAS6/Axl/SOCS signaling, and thus support rGAS6 as an effective immune modulator for the clinical prevention and treatment of IS-induced HT.