Effects Of Wnt5b On Regeneration And Functional Recovery After Spinal Cord Injury In Mice

Objective:Repair and functional recovery after spinal cord injury（SCI）are a worldwide medical problem.It is of great scientific and social significance to explore methods to promote nerve regeneration and functional recovery.This topic mainly discusses the expression changes of non-classical Wnt signaling pathway related genes after spinal cord injury,and the influence of Wnt5b knockout on regeneration and repair after spinal cord injury.Methods:（1）Establishment of SCI model and selection of appropriate injury degree:SCI model was established by hitting T10 spinal cord with Louisville Injury System Apparatus（LISA）.The adult mice were randomly divided into three groups:sham group,SCI groups with spinal cord displacement of 0.45mm（SCI 0.45 group）and 0.65mm（SCI 0.65 group）.After injury,Basso mouse scale（BMS）score,Catwalk test,Von Frey test and Cold Hot plate test were conducted on mice in each group at different time points to test the functional recovery of the mice,and the appropriate depth of injury was selected for comparative analysis.The expression of non-classical Wnt signaling pathway and planar cell polarity（PCP）related genes at different time points after SCI at the injury site and lumbar4-5（L4-5）was examined by Real-time PCR after behavior tests.（2）Research on the role of Wnt5b in spinal cord injury in mice:The wild type and Wnt5b knock out mice were divided into Wnt5b^+/+group and Wnt5b^-/-group with spinal cord displacement of 0.45mm.Locomotor function was assessed based on the Basso mouse scale and Catwalk 9.0 apparatus.Neuropathic pain after SCI were measured by Von Frey test and Cold Hot plate test.Immunohistochemical staining was used to analyze the morphology of spinal cord tissue and the proliferation of astrocytes after injury in mice.Results:（1）The hindlimb BMS score of SCI 0.65 group was lower in SCI 0.65 group compared with SCI 0.45 group.Limb coordination rate in both injured groups was worse than sham group.The function of the SCI 0.65 group was difficult to recover due to the deeper degree of injury,so 0.45mm was selected as the depth of SCI in mice.（2）On the 28th day,the mice in the injury group showed mechanical hyperalgesia and lasted for 42 days,and the more severe the injury,the lower the mechanical pain threshold.The mice suffered from abnormal heat and cold pain after SCI but not related to the degree of injury.（3）Changes in Wnt/PCP signaling pathway related genes expression were different at different time points after spinal cord injury,in which Wnt5b down-regulated slightly on day 1 and was up-regulated from day 14 to day 42.（4）The hindlimb BMS score and limb coordination rate of Wnt5b^-/-group were higher than Wnt5b^+/+group after injury.（5）The Wnt5b^-/-group had significantly lower sensitivity to mechanical pain and cold/heat pain than the Wnt5b^+/+group.（6）Tissue damage and astrocyte proliferation in Wnt5b^-/-group were less than that in Wnt5b^+/+group.Conclusion:（1）Neuropathic pain was induced after SCI,Mechanical allodynia but not thermal hyperalgesia showed different sensitivity to different degrees of SCI.（2）After injury,Wnt5b expression in SCI segment of mice decreased first and then was continuously up-regulated.（3）Wnt5b knockout not only improves hindlimb motor dysfunction and sensory disturbance caused by SCI but also promotes spinal cord tissue recovery and attenuates astrocyte proliferation.