Cell type-specific effects of the integrated stress response pathway after traumatic spinal cord injury - After contusive spinal cord injury (SCI), primary loss of neural tissue is followed by secondary damage that exacerbates functional deficits and can be targeted for neuroprotection. Functionally important components of the secondary injury include destruction of axons, death of oligodendrocytes (OLs) and demyelination that result in white matter damage (WMD). WMD is further amplified by neurotoxic inflammation that is mediated by the microglia (MG) and monocyte-derived macrophages (MDMs). After SCI, MG/MDMs are activated by pattern recognition molecules (PRM) and lipid overload. The integrated stress response (ISR) is triggered by various stimuli including ER stress, mitochondrial damage or PRM-associated cytosolic aggregation of signaling adapter proteins such as ASC. Those stimuli activate ISR kinases such as PERK and HRI that phosphorylate the translation factor eIF2α at the Ser-51 residue (peIF2α) leading to inhibition of general protein synthesis and upregulation of the transcription factors ATF4 and CHOP. Initially, ISR activation attempts to restore cellular homeostasis. If that fails, prolonged ISR follows and cell death may occur. In addition, HRI- mediated ISR may facilitate pro-inflammatory signaling by PRMs. We and others have demonstrated that after SCI, the ISR is activated and global deletion of Chop protects OLs, improves functional recovery and reduces neuroinflammation. Preliminary data show that global deficiency of HRI/EIF2AK1 reduces ISR activation, promotes OL survival, suppresses neuroinflammation and improves functional recovery after SCI. Moreover, the deleterious lipid overload phenotype is reduced in Hri-/- mice in vivo and in bone marrow derived macrophages (BMDM) in culture. Additional cell culture studies reveal partial protection of Hri-/- OLs and a reduced inflammatory potential of Hri-/- MGs after PRM stimulation. These findings support the novel hypothesis that after contusive SCI, the ISR kinase HRI is a major contributor to the secondary injury including death of OLs and cytotoxic neuroinflammation. In OLs, the HRI-mediated ISR promotes cell death downstream of HRI-activating stimuli (mitochondrial/oxidative stress). In MG/MDMs, HRI facilitates cytotoxic neuroinflammation by enhancing PRM signaling and/or foamy macrophage formation. Therefore, aim 1 will test the cell autonomous contributions of HRI to OL death after SCI. Aim 2 will define the role of the HRI as a regulator of the SCI-associated neuroinflammation. Aim 3 will examine whether pharmacological inhibition of HRI improves SCI outcome. Mouse lines with OL- or MG/MDM-specific deletions of Hri or a specific, CNS permeable HRI inhibitor will be used with a moderate contusive T9 SCI in mice. Gene expression changes will be analyzed using the Ribotag system. This proposal will evaluate HRI as a targetable, pleiotropic mediator of the secondary injury after SCI. It will also probe regulation of SCI-associated neuroinflammation by HRI. Importantly, HRI inhibition may become a new therapeutic target for small molecule drugs to promote neuroprotection and/or reduce cytotoxic neuroinflammation following traumatic CNS injury.
