| Acute spinal cord injury is a common and devastating disease in spinal surgeryfield. The treatment of spinal cord injury become serious medical care problem witha gradually increased incidence. The great harm on the physical and psychologicalaspects of victims, high costs to healthcare bring to the family and social heavyburdens. The complex pathological mechanism of acute spinal cord injury makes itdifficult for finding an effective therapy. Primary insult to the spinal cord willdirectly destroy tissue and cause the unrecoverable injury, followed by a series ofself-destructive factors release into tissue. The self-destructive process promotespinal cord tissue ischemia hypoxia, edema, degeneration and necrosis. Trauma tospinal cord cause secondary injury, edema is one of the important pathophysiologicalprocess of the damage，and has great effect on prognoses. In addition to theapplication of drug therapy, however, the mechanism underlying the spinal cordedema are not fully understood. This study was designed to evaluate the changes ofcritical factors involved in the process of spinal cord edema and to explore thepossible mechanism of spinal cord edema at cellular and molecular levels. Lookingforward to find the corresponding targets for drug treatment and provide theoreticalbasis of spinal cord injury therapy.Objective: To observe the pathological changes and spinal cord water content atdifferent time points in acute spinal cord injury model, we made a model of acutespinal cord injury. In order to explore the mechanism of spinal cord edema andco-expression of AQP4and Kir4.1, we investigated the variation of inwardlyrectifying potassium channel4.1(Kir4.1) and aquaporin-4(AQP4) during spinal cordedema. The interaction between AQP4and Kir4, the role of each factor with edemawere estimated by observing the variation of AQP4, Kir4.1, P38, p-P38, IκB andNF-κB p65during spinal cord edema. It will provide the theoretical basis and new drug treatment targets to repair spinal cord injury.Methods: Acute spinal cord injury was induced by clip compression. A total of66adult SD rats (weighing,250-300g) were divided into control, sham-operatedgroup (surgery without spinal cord injury), MP intervention group (ASCI,1h,injection with10ml MP) and ASCI group (acute spinal cord injury). Sham-operatedgroup and ASCI group were both divided into four subgroups at8h,24h,3d,7dwith different time points. MP intervention group was divided into two subgroups at3d,7d. Rats were kept in separated cages and assisted with urination and defecationafter the surgery.The neurologic symptoms of the rats were scored by Tarlov after the surgery.Dry-wet weight method was used to measure water content in spinal cord. RoutineHE staining showed pathologic changes of tissues. Immunofluorescence doublelabeling was conducted to detect changes of AQP4and Kir4.1. Real-time PCR wasperformed to detect mRNA variation of AQP4and Kir4.1. Western blot analyseswere performed to determine expression levels of AQP4, Kir4.1, p-P38, P38, IκB,NF-κB p65in the spinal cord of each group. Data were analyzed using the SPSSsoftware ver.18.0. Measured data were compared with analysis of variance and t-test.The p-values<0.05were considered to indicate statistical significance.Results: The motor neuron injury and motor dysfunction tended to grow worseover time after spinal cord injury in rat. Methylprednisolone could relieve motordysfunction after spinal cord injury.The water content of spinal cord was increased over time after spinal cordinjury in rat. The methylprednisolone could reduce spinal cord edema after spinalcord injury.Routine HE staining showed that the central canal and gray matter bleeded at8h after spinal cord injury in rat. Tissue had mild edema around the blood vessels. Thebleeding was severe in the gray matter at24h. Cyst formation, necrosis and neuronsswelling were observed. All pathological changes became worse at3d. Tissue had severe edema around the blood vessels. Intercellular spaces were obvious. Neuronsswelling and nucleus pycnosis tended to worse. Most of the spinal cord becamenecrosis and the gray matter degenerated at7d. The cystic cavity and cavity wereformed. The bound between the gray matter and white matter was clear at3,7d aftertreated with methylprednisolone. The bleeding and nucleus pycnosis were decreased.The edema around the blood vessels was relieved and the volume of edema wasdecreased.The expression level of AQP4and Kir4.1were consistent. Their protein andmRNA expression level were increased over time after spinal cord injury. Thevariation of AQP4and Kir4.1correlated with spinal cord edema.The co-expression of AQP4and Kir4.1was observed on the cell membrane ofspinal pia mater, central canal and astrocytes in the spinal cord. The expression ofthem were increased over time and significantly reduced after treated withmethylprednisolone.The expression of AQP and Kir4.1were upregulated after spinal cord injury inrat. The expression level of AQP4and Kir4.1were decreased after treated withmethylprednisolone. The increased expression of p-P38was observed in the spinalcord edema after spinal cord injury. The expression of p-P38was decreased aftertreated with methylprednisolone. The expression of IκB was decreased and NF-κBwas increased in the spinal cord edema after spinal cord injury. The expression ofIκB was upregulated and NF-κB was decreased after treated withmethylprednisolone.Conclusions: spinal cord injury damaged the motor neuron and leaded to motordysfunction. Methylprednisolone could relieve motor dysfunction irreversibly.A serious of pathologic changes, such as spinal cord edema, were observed afterspinal cord injury in rat. The pathologic changes relieved and recovered slowly afterinterference with methylprednisolone.The correlation of edema with AQP4and Kir4.1suggests the variation of AQP4 and Kir4.1directly affected the swelling after spinal cord injury in rat.The co-expression of AQP4and Kir4.1was observed in the grey matter ofspinal cord, and less existed in the white matter. The expression of them wereproportional to spinal cord edema.AQP4and Kir4.1participated in the process of spinal cord edema, after spinalcord injury. Suggesting that the synergy between AQP4and Kir4.1promoted theformation of spinal cord edema. Both P38MAPK and NF-κB signal pathwayparticipated the formation of spinal cord edema by regulating AQP4and Kir4.1. |
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