Podoplanin-CLEC-2 Axis Participates In Regulating Inflammation After Cerebral Ischemia-Reperfusion In Mice

Objective:Observe the changes of the podoplanin-C lectin-like receptor-2 axis after cerebral ischemia-reperfusion.Observe the protective effect of podoplanin(PDPN)antibody on nerve function after after cerebral ischemia-reperfusion.And explore the main mechanism of the involvement of podoplanin-C lectin-like receptor-2 axis in cerebral ischemia-reperfusion injury.Methods:We introduced the model of middle cerebral artery occlusion in mice and the changes of related proteins after cerebral ischemia-reperfusion in mice were measured by western blot.Immunofluorescence staining was used to detect the cell location of the podoplanin-CLEC-2 axis.TTC staining was used to detect the changes of cerebral infrarct volume in mice.The neurological deficits and recovery after the PDPN blocking antibody administration(clone 8.1.1)were evaluated by various behavioral tests such as cylinder test,foot faults test and grip strength test.Enzyme-linked immunosorbent assay(ELISA)was used to detect the changes of various inflammatory factors.Results:The expression of C-type lectin-like receptor 2(CLEC-2,CLEC-1b)in cerebral ischemia-reperfusion injury increased significantly,peaked at 24 hours,and then decreased gradually.Both of them were distributed in neurons and microglia of cerebral cortex in the form of PDPN-CLEC-2 axis,and participated in cerebral ischemia-reperfusion injury.Admission of PDPN antibody into lateral ventricle could prevent the up-regulation of CLEC-2 after stroke,and neutralization of PDPN has an inhibitory effect on the axis of podoplanin-CLEC-2.In the acute stage after stroke,the inhibition of podoplanin-CLEC-2 axis by administration the PDPN blocking antibody could reduce the infrarct volume,reduce the neurological deficits in the acute stage of reperfusion,and have the protective effect on stroke;in the process of rehabilitation,the asymmetry of the forelimb in the antibody treatment group was reduced,the difference of the times of left and right mistakes was reduced,and the strength is gradually increased,which indicates the sensory function,motor coordination ability and muscle strength were gradually improved,and PDPN blocking antibody promoted the recovery of neural function after stroke.PDPN blocking antibody could selectively reduce the expression of inflammatory factors such as IL-18 and IL-1? in brain injury area of mice,but had no significant effect on IL-6 and IL-10.Conclusions:The podoplanin-CLEC-2 axis was involved in cerebral ischemia-reperfusion injury in mice by neurons and microglia cells.PDPN blocking antibody has neuroprotective effect on ischemia stroke.The podoplanin-CLEC-2 axis was involved in cerebral ischemia-reperfusion injury in mice by regulating the inflammatory response,which provides a new potential target site for clinical treatment.