The Effects Of Monocyte Derived Macrophage On Postoperative Cognitive Dysfunction In Rats Undergoing Cardiopulmonary Bypass

Objective Through observing the effects of monocyte derived macrophage(Mo-Mφ)on postoperative cognitive dysfunction in rats undergoing cardiopulmonary bypass,this research aims to explore the mechanism of action of this cell and how to exert its brain protective effect were explored.Methods Clean and healthy adult SD rats,40 males,weighing 350-400 g,were provided by the Animal Experiment Department of the Northern Theater Gereral Hospital.Randomly divided into 4 groups of 10: sham operation+saline group(S group),CPB+saline group(C group),CPB+PBS liposome group(P group),CPB+clodronate liposome group(L group).Rats undergoing water maze were trained 4 times a day for 5 consecutive days before surgery.Rats were fasted for 6 hours before surgery and not fasted in drinking water,Intraperitoneal anesthesia with 2% sodium pentobarbital(50mg/kg)and tracheal intubation using light transmission method for mechanical ventilation were conducted,group S only performed venipuncture and catheterization without extracorporeal circulation;group C received arteriovenous puncture and extracorporeal circulation for 60min;group P was subjected to extracorporeal circulation for60 min and PBS liposome were injected 4ul/g via tail vein 48 h and 24 h before CPB;group L was injected with 4ul/g clodronate liposome via tail vein at the same time in group P;Group S and group C were injected with saline before tail vein.The Morris water maze was used to collect experimental data on the 7th day after CPB.The behavioral changes of each group of rats were analyzed,and then the rats were killed by bleeding from the abdominal main vein after anesthesia.Serum was collected,cardiac perfusion was performed with 4℃ice-cold saline,and hippocampus tissue and brain were collected on an ice.Use ELISA to detect changes in blood inflammatory factors such as IL-6 、IL-1β 、TNF-α and the expression level of brain injury marker proteins such as S100βand NSE;Flow cytometry to detect Mo-Mφ in peripheral blood before and after administration Clearance rate;use TUNEL to observe hippocampal neuron apoptosis;real-time quantitative PCR was used to detect the expression level of IL-1β、IL-10、TNF-α、TGF-β and Arg1、i NOS in hippocampus;Western blot was used to detect the expression level of i NOS in hippocampus.SPSS 22.0software was used for statistical analysis of the experimental data.The normal distribution measurement data were expressed as mean ± standard deviation(`x ± s).Comparison between groups was performed by one-way analysis of variance.P<0.05 indicated that the difference was statistically significant.Results1.Morris water maze results of rats in each group Compared with the S group,the average latency of the rats in the C,P,and L groups was prolonged,and the number of times to cross the platform and Target quadrant dwell time was reduced.The difference was statistically significant(P<0.05).Compared with the C group,the average latency of the L group was longer.The number of crossing platforms and dwell time decreased,and the difference was statistically significant(P<0.05).There was no significant difference between group P and group C.2.ELISA to detect changes in serum IL-6,IL-1β,TNF-αinflammatory factors of rats in each group at 7 day after operation Compared with the S group,the serum levels of IL-6,IL-1β,and TNF-α in the serum of the C,P,and L groups were all increased,and the differences were statistically significant(P<0.05);the C,P,and L group There were no significant differences in IL-6,IL-1β,TNF-α in serum of rats.3.ELISA to detect the expression levels of brain injury markers S100β and NSE in plasma of rats in each group at 7 day after operation Compared with group S,the levels of S100β in the plasma of rats in group C,P and L were significantly increased,and the difference was statistically significant(P<0.05).Compared with group C,the levels of S100β in rats in group L were significantly increased.The difference was statistically significant(P<0.05).Compared with group C,the difference between group P and group C was not statistically significant(P>0.05).The level of NSE in plasma was slightly higher in the rats in each group than in the other groups.The S group was diverted,but the results were not statistically significant(P>0.05).4.Changes in the number of Mo-Mφ in the brain detected in flow cytometry Compared with the C group,the number of Mo-Mφ in the L group was reduced,and the difference was statistically significant(P<0.05).There was no statistically significant difference between the P group and the C group.5.Changes in hippocampal neuron apoptosis rate through TUNEL detection Compared with group S,the apoptosis of hippocampal neurons in group C,P and L was significantly increased,and the difference was statistically significant(P<0.05).Compared with group C,the number of hippocampal neurons in group L significantly increased,the difference was statistically significant(P<0.05),and the difference between group P and group C was not statistically significant.6.IL-1β、IL-10、TNF-α、TGF-β inflammatory factor and Arg1、i NOS gene expression in the hippocampus through rt-PCR detection Compared with the S group,the expressions of inflammatory factor and Arg1、i NOS in the hippocampal of the rats in the C,P,and L groups increased,and the difference was statistically significant(P<0.05);Compared with group C,the hippocampal levels of IL-1β 、 TNF-α and i NOs genes in rats in group L increased,and the difference was statistically significant(P<0.05),and IL-10、Arg1 gene expression decreased.There was no significant difference in group P compared with group C.7.The expression of i NOS protein in the hippocampus through Western blot detection Compared with group S,the expressions of i NOS protein in hippocampus of rats in group C,P and L were increased,and the differences were statistically significant(P<0.05).Compared with group C,The expressions of i NOS protein in group L increased,and the difference was statistically significant(P<0.05).There was no significant difference between group P and group C.Conclusions1.Clearing peripheral monocyte derived macrophage can aggravate CPB-induced POCD.2.Elimination of peripheral monocyte derived macrophage may affect local polarization of microglia in the brain,thereby aggravating local neuroinflammatory damage and thus POCD.