Neuron-derived CCL17 Exerts Neuroprotection Through Activation of CCR4/mTORC2 Axis in Microglia After Subarachnoid Hemorrhage in Rats

Abstract Background and purposeCCL17 has been proven to play a critical role in immune regulation. However, there is a lack of evidence to address the effect of CCL17 after subarachnoid hemorrhage。 Therefore, the aim of this study was to investigate the therapeutic effects of CCL17 and its underlying mechanism after SAH in rats.MethodsThe levels of secreted CCL17 from primary neurons after stimulation of OxyHb were examined in vitro. Rats were subjected to the endovascular perforation model of SAH and were randomly assigned to receive either recombinant CCL17 or vehicle, which were administered intranasally at 1 h after SAH. AZD2098 and JR-AB2-011 were applied to investigate the CCR4/mTORC2 axis in CCL17-mediated neuroprotection. To elucidate the underlying mechanism, the in vitro kinase assay was performed in primary microglia. Adeno-associated virus-CD68 knockout for mTORC2 in brain microglia was administered via intracerebroventricular injection 21 days before SAH. Brain water content, short-term neurobehavior evaluation, western blot, quantity RT-PCR, and immunofluorescence staining were performed.ResultsEndogenous CCL17 was increased and secreted from neurons, with CCR4 primarily expressed on microglia in both in vivo and in vitro models of SAH. Exogenous rCCL17 significantly alleviated neuronal apoptosis and brain edema, and improved short-term neurofunction in a dose-dependent manner at 24 h and 72 h after SAH in rats. The rCCL17 also increased M2-like polarization of microglia at 72 h after SAH in rats and in primary microglia culture. In addition, the rCCL17 treatment further activated mTORC2 signaling after SAH in rats and in primary microglia cultures. The neuroprotection of rCCL17 was abolished by inhibition of CCR4 or mTORC2.ConclusionNeuron-derived CCL17 could alleviate SAH induced-neurological deficits by promoting M2-like polarization of microglia after SAH. The microglialCCR4/mTORC2 axis-mediated CCL17 involved neuroprotection.