An Experimental Study On The Expression And Effect Of Necroptosis After Spinal Cord Injury

Objective. To investigate the role of necroptosis in spinal cord injury (SCI). Meanwhile, we wish to investigate whether necrostatin-1(Nec-1), a selective inhibitor of necroptosis, has a neuroprotective effect on the rat model of traumatic SCI, and to explore the potential mechanisms.Methods:One hundred and forty-four adult female SD rats were divided into four different groups:(1) sham group; (2) SCI group; (3) SCI+Veh group; (4) SCI+ Nec-1(20uL,40mg/mL, intrathecal injection) group. First, the dynamic expression of RIP1 and RIP3 were tested using western-blot. After rats were sacrificed 24 h after injury, propidium iodide (PI) was administered by intravenous injection to label necroptotic cells in the spinal cord. Furthermore, Morphological analyses were performed by hematoxylin-eosin and Nissl staining. The effects of Nec-1 on microcirculation disturbance, histological changes, and improvement in motor function were assessed. Finally, the influences of Nec-1 on the expression of proteins closely related to autophagy and apoptosis, proinflammatory cytokines, activation of inflammatory cell, were also examined to explore the underlying mechanisms of the neuroprotective effects of Nec-1.Results:Part â…  The necroptosis related molecules, RIP1 and RIP3, were upregulated with time and reached the peak at 48h after SCI. In addition, administration of Nec-1 produced an decrease in histological destructions (The Nec-1 treated rats showed more surviving neuron at 1w after SCI, P<0.05) and improved the locomotor function of injured rats,which is indicated by the BBB scores.Part â…¡ Nec-1 could inhibit apoptosis by inhibiting the activation of Caspase-3 while activating Bcl-2. Nec-1 could inhibit autophagy mainly through downregulating the expression of LC3B â…¡ at the injury site (P<0.01). Further study indicated that Nec-1 intervention reduced the release of danger-associated molecular patterns (DAMPs), the production of proinflammatory cytokines, inhibited microglial activation, and neutrophil infiltration (P<0.05), and improved blood supply in the spinal cord trauma area(P<0.01).Conclusion:Necroptosis played an important role after traumatic SCI and inhibition of necroptosis can paly neuroprotective effects on rat model of SCI. These results provide new therapeutic targets for SCI treatment.