脊髓损伤模型大鼠软脊膜下注射2型腺相关病毒免疫荧光染色分析.

BACKGROUND: The method of injecting adeno-associated virus subpial can limit the virus to a small area centered on the injection point,with less systemic toxicity and side effects, and save the amount of virus. However, there are no reports on the application of this method in spinal cord injury rats, and the transfection of astrocytes and neurons at the injured site is still unclear.OBJECTIVE: To investigate the transfection effect of subpial injection of adeno-associated virus 2 on astrocytes and neurons in the spinal cord injury site and the technical points of the surgical procedure.METHODS: The study was approved by the Experimental Animal Ethics Committee of Jilin University. Twenty-four Wistar rats were randomly divided into two groups: adeno-associated virus group and model group. Rats in both groups were used to establish the spinal cord injury model using the clamp method. The adeno-associated virus group received 60 μL of virus solution by subpial injection, and the model group received an equal amount of 5% dextran solution. At postoperative 7, 14, and 21 days, spinal cord specimens were taken for immunofluorescence staining to observe the distribution of astrocytes and the transfection of adeno-associated virus 2.RESULTS AND CONCLUSION:(1) After subpial injection of the virus, the virus solution evenly distributed in the range of about 1 cm above and below the injured site.(2) At 7 and 14 days after surgery, only a small amount of green fluorescent protein was expressed in the spinal cord injury area. Until day 21, a large number of reactive astrocytes were observed around the spinal cord injury cavity. The strongest expression was in the periphery of the spinal cord cavity, with obvious glial boundary membrane. It was also found that adeno-associated virus 2 could stably transfect astrocytes and neurons, in which astrocytes were concentrated around the periphery of the spinal cord cavity,while neurons were scattered within the scope of virus transfection. Only trace adeno-associated virus 2 transfection was found in normal tissues outside the scope of injection.(3) The method has the advantages of confining the virus to the injured site, targeting astrocyte and nerve cell transfection, low viral dosage, low systemic toxicity and side effects, and low cost. [ABSTRACT FROM AUTHOR]