The Role Of Immunosuppressive Status In The Occurrence Of Perioperative Neurocognitive Disorders After Cardiopulmonary Bypass In Rats

Objective By intervening Treg cells to regulate the state of immunosuppression after cardiopulmonary bypass,the role of immunosuppressive state in the occurrence of perioperative neurocognitive disorders after cardiopulmonary bypass was explored.Methods 32 clean SD rats(provided by the Animal Experiment Department of the General Hospital of Northern Theater Command)were randomly divided into 4 groups with 8 rats in each group:group S(sham operation group),group C(CPB group),group P(CPB+P300i)and group D(CPB+DMSO).Morris water maze training was performed 4 times a day for 5 consecutive days before operation.Each group of rats was subjected to anesthesia,mechanical ventilation and other operations.The rats in group S were operated only by arterial vein without establishing CPB model,while the other three groups were established with CPB model.The rats in group S and C were intraperitoneally injected with 2.5mg/kg saline every day one week before operation,the rats in group P were intraperitoneally injected with 2.5mg/kg P300i every day one week before operation and the rats in group D were intraperitoneally injected with the same dose of DMSO every day one week before operation.The Morris water maze experiment was performed 3 days after the operation,after collecting data about the average latency of the rat and the number of crossing platforms,the peripheral blood of the rats was collected,the rats were killed,and brain tissue was extracted by cold perfusion.Flow cytometry was used to detect the number of CD4~+T lymphocytes in peripheral blood and the proportion of Treg cells in CD4~+T lymphocytes;the concentrations of inflammatory factors IL-4 and IFN-?in peripheral blood of rats and the levels of brain injury related proteins S100?and NSE in serum were detected by ELISA;the apoptosis of hippocampal neurons was detected by TUNEL staining;the gene expression levels of different inflammatory factors in hippocampal tissue were detected by real-time quantitative PCR.SPSS 25.0 and Graphpad Prism 8.0 software were used for statistical analysis,P<0.05 was considered statistically significant.Results 1.The results of flow cytometry showed that compared with the sham operation group,the number of CD4~+T lymphocytes was significantly reduced at 1 and3 days after CPB(P<0.05),while the proportion of CD4~+CD25~+Foxp3~+Treg cells was significantly increased(P<0.05).The results of ELISA showed that the levels of IL-4and IFN-?in peripheral blood increased significantly 1 day after CPB,but the level of IFN-?returned to the preoperative level 3 days after CPB,while IL-4 remained at a relatively high level within 7 days after CPB.2.The results of flow cytometry showed that p300i can significantly reduce the proportion of Treg cells in peripheral blood after CPB in rats.The results of the water maze experiment showed that compared with group S,the average incubation period of group C,group P and group D was significantly prolonged,and the number of times of crossing the platform was significantly reduced(P<0.05);compared with group D,the average incubation period of group P was significantly shortened and the times of crossing platforms was significantly increased(P<0.05).3.The changes of peripheral adaptive immunosuppressive status reduce inflammatory injury in hippocampus after CPB in rats.The results of ELISA showed that compared with the S group,the S100?levels in the C and D groups were significantly increased(P<0.05),but there was no significant difference between the C and D groups(P>0.05);compared with the group D,the S100?levels in the group P were significantly reduced(P<0.05);there was no significant difference between group P and S(P>0.05).Plasma NSE levels were increased in each group of transfused rats.Compared with group D,the plasma NSE levels in group P decreased slightly,but the results were not statistically significant(P>0.05).TUNEL staining results showed that compared with the group S,the apoptosis rate of hippocampal neurons in the C,P,and D groups was significantly increased(P<0.05);compared with group D,the apoptosis rate of hippocampal neurons in group P was relatively significantly reduced(P<0.05).The results of real-time quantitative PCR showed that compared with the group S,the expression of IL-1?and TNF-?genes in the hippocampus of the C,D,and P group were up-regulated(P<0.05);the TNF-?gene expression of the group P was significantly lower than that in group D(P<0.05).Conclusion CPB can lead to the increase of regulatory T cells in rats and induce the state of immunosuppression,which is not conducive to the maintenance of postoperative cognitive function.Reducing Treg cells and alleviating peripheral immunosuppression can help improve cognitive function after CPB in rats.