Changes Of CD8~+T Lymphocyte Subsets In The Peripheral Blood And Injured Spinal Cord Of SD Rats After Spinal Cord Injury

Background：Immune response after spinal cord injury (SCI) is complex, many immune cellsand cytokines are involved in this process.These cells and factors together determine therecovery process after spinal cord injury. So far, the function of CD8+T lymphocytesubsets in SCI is still unclear. Understanding the changes of these cell subsets afterspinal cord injury will be very beneficial to make clear immune mechanism and providescientific reference for the treatment of spinal cord injury.Objective:To investigate the changes of CD8+T lymphocyte subsets in the peripheral bloodand injured spinal cord in SD rats after spinal cord injury.Method:Spinal cord contusion injury of adult female SD rats was prepared using New YorkUniversity (NYU) impactor. The peripheral blood was collected from the tail and thelocalized mononuclear cells were separated by density gradient centrifugation method.The T lymphocyte subsets in the peripheral blood and injured spinal cord were detectedusing flow cytometry.Results:1. Changes of the total T cells in peripheral blood of SD rats before and afterspinal cord injuryThe percentage of T cells in total lymphocytes of peripheral blood in normal SD rats was59.71±3.24. One day after spinal cord injury, the percentage was46.89±4.44,is the lowest value of all experimental observation points. After3days, the percentagereached to53.44±3.97, still at a low level, there was no significant differencecompared to the first day (P>0.05). Five days after SCI, the percentage returned tonormal levels, and maintained to the21days after injury.The percentage was69.30±2.28and70.27±2.64in28d and56d after spinal cordinjury, respectively. There was no significant difference between the two time points (P>0.05). However, compared with other time points, the result of28d was increased, andwas highest in all observation points of its former, the difference was statisticallysignificant (P <0.05). The result of56days was statistically significant differencecompared to normal SD rats,1day,3days,14days and21days after SCI (P <0.05).2. Changes of CD3+CD8+T cells in peripheral blood before and after of spinal cordinjuryBefore SCI, the percentage of CD3+CD8+T cells in CD3+T cells was45.02±3.82.One day after spinal cord injury it decreased to39.86±3.89, the difference wasstatistically significant (P <0.05). However, its value restored to normal levels at the3rd,, and continued up to28days after injury.3.Changes of CD3+CD8+CD28-and CD3+CD8+CD28+subsets in CD3+CD8+T cellsin peripheral blood after spinal cord injuryBefore SCI, the percentage of CD3+CD8+CD28-and CD3+CD8+CD28+in CD3+CD8+T cells in peripheral blood of SD rat was65.43±4.07and34.57±4.07,respectively. The changes of CD3+CD8+CD28-and CD3+CD8+CD28+had opposite trendafter injury. The percentage of CD3+CD8+CD28-cells has no significant difference in1d,3d,7d,14d,28d and56d after SCI compared with before the injury (P>0.05). Fivedays after SCI, the percentage of CD3+CD8+CD28-dropped to49.50±4.82, the lowestvalue of all time points, and CD3+CD8+CD28+rose to50.50±4.82, the highest value of all time points. Compared to the normal,3days,7days,14days,28days and56daysafter SCI, the difference was statistically significant (P <0.05). Compared the results of1day,3days,5days,14days,21days after SCI, the percentage of CD3+CD8+CD28-cellsincreased at28days after injury, the difference was statistically significant (P <0.05).The percentage of CD3+CD8+CD28-cells also increased at56days after injurycompared to pre-injury,1d,3d,5d and21days after SCI, the difference was statisticallysignificant (P <0.05). While the percentage of CD3+CD8+CD28+cells decreased at28days after injury compared with the results of1day,3days,5days,14days and21daysafter SCI, the difference was statistically significant (P <0.05). The percentage ofCD3+CD8+CD28+cells also decreased at56days after injury compared with pre-injury,1d,3d,5d and21days after SCI, the difference was also statistically significant (P <0.05).4. Changes of local CD3+CD8+T cells after spinal cord injuryOne day after spinal cord injury, the percentage of CD3+CD8+T cells in CD3+Tcells in the injured spinal cord was40.55±1.59. The percentages were38.84±4.71,47.57±15.79,50.07±1.97,37.70±8.85,55.14±5.56and44.78±3.26at3d,5d,7d,14d and28d, respectively. Compared to1d after injury, the differences were notstatistically significant (P>0.05). Fifty-six days after spinal cord injury, its valueincreased to70.85±7.16, and compare to1day,3days,5days,14days and28daysafter SCI, the difference was statistically significant (P <0.05).5. Changes of local CD3+CD8+CD28-and CD3+CD8+CD28+subsets in CD3+CD8+Tcells after spinal cord injuryOne day post of injury, the local infiltrated CD3+CD8+CD28-and CD3+CD8+CD28+were69.87±6.00and30.13±6.00in CD3+CD8+T cells, respectively. Pairwisecomparisons between1d,3d,5d after injury, the difference were not statistically significant (P>0.05).Seven days after spinal cord injury, the percent of CD3+CD8+CD28-rise to95.81±0.90, which was the peak of all observation points, while theCD3+CD8+CD28+decline to4.19±0.90, which was the lowest value of all observationpoints. Compared to1d,3d and5d after injury, the difference was statisticallysignificant (P <0.05). Then, the percent of CD3+CD8+CD28-maintained at a high level,the results were92.52±1.00,89.49±3.40,88.44±4.10and95.11±0.40at14d,21d,28d and56d, respectively. The percent of CD3+CD8+CD28+had been maintained in thelower level, the results were7.48±1.00,10.51±3.40,11.56±4.10and4.89±0.40at14d,21d,28d and56d, respectively. This four observation points were the same to7dafter SCI, but were statistically significant1d,3d and5d after injury (P <0.05).Conclusion:1.After spinal cord injury, CD8+T cells in peripheral blood of SD rat increase and caninfiltrate into the injured spinal cord. These suggest that CD8+T cells may be involvedin the pathological process of spinal cord injury.2.After spinal cord injury, the CD8+CD28+T cells in peripheral blood have a transientrise, but this cell subset is not dominant in the injured spinal cord. However, theCD8+CD28-T cell subset is dominant in the injured spinal cord.These results implythat local infiltrated CD8+Tcells may have immunomodulatory effects.