Abstract
BackgroundSpinal cord injury (SCI) favors a persistent pro-inflammatory macrophages/microglia-mediated response with only a transient appearance of anti-inflammatory phenotype of immune cells. However, the mechanisms controlling this special sterile inflammation after SCI are still not fully elucidated. It is known that damage-associated molecular patterns (DAMPs) released from necrotic cells after injury can trigger severe inflammation. High mobility group box 1(HMGB1), a ubiquitously expressed DNA binding protein, is an identified DAMP, and our previous study demonstrated that reactive astrocytes could undergo necroptosis and release HMGB1 after SCI in mice. The present study aimed to explore the effects and the possible mechanism of HMGB1on macrophages/microglia polarization, as well as the neuroprotective effects by HMGB1 inhibition after SCI.MethodsIn this study, the expression and the concentration of HMGB1 was determined by qRT-PCR, ELISA, and immunohistochemistry. Glycyrrhizin was applied to inhibit HMGB1, while FPS-ZM1 to suppress receptor for advanced glycation end products (RAGE). The polarization of macrophages/microglia in vitro and in vivo was detected by qRT-PCR, immunostaining, and western blot. The lesion area was detected by GFAP staining, while neuronal survival was examined by Nissl staining. Luxol fast blue (LFB) staining, DAB staining, and western blot were adopted to evaluate the myelin loss. Basso-Beattie-Bresnahan (BBB) scoring and rump-height Index (RHI) assay was applied to evaluate locomotor functional recovery.ResultsOur data showed that HMGB1 can be elevated and released from necroptotic astrocytes and HMGB1 could induce pro-inflammatory microglia through the RAGE-nuclear factor-kappa B (NF-κB) pathway. We further demonstrated that inhibiting HMGB1 or RAGE effectively decreased the numbers of detrimental pro-inflammatory macrophages/microglia while increased anti-inflammatory cells after SCI. Furthermore, our data showed that inhibiting HMGB1 or RAGE significantly decreased neuronal loss and demyelination, and improved functional recovery after SCI.ConclusionsThe data implicated that HMGB1-RAGE axis contributed to the dominant pro-inflammatory macrophages/microglia-mediated pro-inflammatory response, and inhibiting this pathway afforded neuroprotection for SCI. Thus, therapies designed to modulate immune microenvironment based on this cascade might be a prospective treatment for SCI.
Highlights
Spinal cord injury (SCI) initiates primary injury and a cascade of secondary injury, leading to transient or permanent functional disorder even paralysis [1]
We further demonstrated that inhibiting High mobility group box 1 (HMGB1) or receptor for advanced glycation end products (RAGE) effectively decreased the numbers of detrimental pro-inflammatory macrophages/ microglia while increased anti-inflammatory cells after SCI
Immunohistochemistry detected cytoplasmic, nuclear, and both cytoplasmic and nuclear HMGB1-immunoreactivity around lesion center (Fig. 1b), and quantification showed that approximately 60% of the HMGB1-positive cells were GFAP-positive (Fig. 1c), which was consistent with our previous result in mice of SCI [19]
Summary
Spinal cord injury (SCI) initiates primary injury and a cascade of secondary injury, leading to transient or permanent functional disorder even paralysis [1]. That the relationship between neuronal or myelin loss and pro-inflammatory response after SCI has been proven by other research and our previous study [4, 5]. The pathological mechanism of the pro-inflammatory macrophages/microglia-mediated detrimental microenvironment after SCI has been not fully clarified. Spinal cord injury (SCI) favors a persistent pro-inflammatory macrophages/microglia-mediated response with only a transient appearance of anti-inflammatory phenotype of immune cells. High mobility group box 1(HMGB1), a ubiquitously expressed DNA binding protein, is an identified DAMP, and our previous study demonstrated that reactive astrocytes could undergo necroptosis and release HMGB1 after SCI in mice. The present study aimed to explore the effects and the possible mechanism of HMGB1on macrophages/microglia polarization, as well as the neuroprotective effects by HMGB1 inhibition after SCI
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