Aldose Reductase Regulates Microglia/Macrophages Polarization Through CREB After Spinal Cord Injury In Mouse

Spinal cord injury (SCI) is a devastating disease of the center nervous system (CNS).It is a severe trauma and can damage motor, sensory, and autonomic function distal to thelevel of injury. SCI produces enormous physical and emotional burdens for bothindividuals and society. Currently, there aren’t any effective therapies available for SCI.There will be multiple larger scars than the primary injury due to the secondary cascadeprocesses. Inflammation contributes greatly to the formation of the glial scar. On theother hand, inflammation is also good for axon regeneration in some degree.Microglia/macrophages are the main inflammatory cells at injury site of the spinal cord.Activated microglia/macrophages have dual nature. They can exacerbate the CNS injury, especially the acute injury stage. They can also promote CNS repair and function recovery.Microglia and macrophages can polarize to two main subsets,“classically activated”(M1)and“alternatively activated”(M2) cells [1,2]. M1cells can express pro-inflammatorycytokines which cause oxidative stress and is neurotoxic. M2cells secreteanti-inflammatory cytokines and can repress inflammation, promote tissue repair andwound healing [3].Aldose reductase (AR, AKR1B1) is one member of the aldo-keto reductase (AKR)superfamily. AR is the first limit enzyme of the polyol pathway and catalyzes the glucoseto sorbitol. Polyol pathway is the alternative way of glucose metabolism underhyperglycemia. Accumulating evidences manifest that AR attributes a significant role inthe regulation of several inflammatory pathologies. AR plays a centrol role in theinflammatory pathogenesis of diabetes, asthma, sepsis, lung diseases and uveitis and canpromote their inflammatory processes. ARIs (Aldose reductase inhibitors) efficientlyalleviate these inflammatory pathologies [5]. We found that expression of AR isupregulated in injured spinal cord after SCI. The upregulation of AR is mainly found inmicroglia/macrophages and neurons but not the astrocytes and oligodendrocytes in theadjacent injury site. AR deficiency mouse has better functional recovery than wide type(WT) mouse after SCI. My research will continue to study the role of AR in SCI. How canAR deficiency influence the functional recovery? And what’s the role of AR in regulationof microglia/macrophages? This will provide some evidences for AR in the regulation ofneuroinflammation.In this study, WT or AR knockout (KO) mice were subjected to SCI by spinal crushinjury model and sacrificed at3d,7d,14d after injury. The injured spinal cord tissues werecut longitudinally at10μm. Sections were stained with Luxol Fast Blue (LFB) andimmunohistochemistry. First, we examine the lesion area and the subsets of microglia/macrophages at the injury site. Then using Western blot detects the oxidative stressmarkers and the expression of symbolic polarization proteins of M1/M2subsets after SCI.In vitro experiments，microglia cell lines （BV2/N9） were treated with different stimulation. Cells were dealt with immunohistochemistry staining and Western blot tofurther explore the mechanism of how AR influences the polarization of microglia andmacrophages.Our study found that LFB staining of the lesion area is smaller in AR ko mice thanWT mice after SCI. In AR ko mice, there was a larger number of M2cells than those inWT, while the number of M1cells in AR ko mice were less compared with WT mice afterSCI. Western blot results show AR deficiency mice expressed more Arginase1andp-CREB than WT mice after SCI. In contrast, WT mice expressed more inducible nitricoxide synthase（iNOS）and NF-κB thanAR ko mice post-injury. The expression of iNOSand NF-κB last longer in WT mice than AR ko mice after SCI. when AR was inhibited byfidarestat in BV2, it caused4-Hydroxynonenal (4-HNE) accumulation. The ARI and4-HNE can induce the phosphorylation of cAMP-responsive binding element (CREB) topromote Arg1expression. Using KG501to inhibite the phosphorylated CREB couldabolish the upregualtion of Arg1by ARI or4-HNE stimulation. Our results suggest thatAR works as a switch which can regulate microglia by polarizing cells to either the M1orthe M2phenotype under M1stimulation based on its states of activity.