| Objective: In this study,a model of perioperative neurocognitive disorders(PND)was established to detect related indicators of pyroptosis and explore the relationship between NLRP3/ Caspase-1-mediated pyroptosis and the pathogenesis of PND.To elucidate the role of neuronal pyroptosis in cognitive dysfunction in aging rats.Methods: PND model was established to detect the expression of pyroptosis related factors,and the pyroptosis level was observed by immunofluorescence staining.Forty-five 18-month-old SD rats were randomly divided into 3 groups: model group(group M),control group(group C)and inhibitor group(group I),with 15 rats in each group.Group I was intraperitoneally injected with NLRP3 inhibitor MCC950 30 minutes before anesthesia.Group M and group I were treated with internal fixation of tibial fracture by inhalation of 3% sevoflurane and 50% air oxygen mixture for 2hours,followed by intraperitoneal injection of buprenorphine for analgesia.Group C only inhaled 50% air oxygen mixture for 2 hours,then intraperitoneal injection of buprenorphine analgesia treatment.One day,three days and seven days after modeling,animal behavior tests were conducted,namely water maze experiment and open field experiment.After behavioral detection,the hippocampus tissues were collected,and the NLRP3,caspase-1,IL-18,IL-1β gene and protein expression were detected by PCR and Western Blot.The pyroptosis level of cells and the production of mitochondrial reactive oxygen species were observed by immunofluorescence staining.Results:1.Animal behavior test results:(1)Water maze experiment: 1 and 3 days after modeling,the escape incubation period and swimming distance in group M were significantly increased compared with group C;In the third quadrant,the activity time,distance,and frequency of crossing the platform were significantly reduced compared with group C.The values of group I are between group C and group M.After 7 days of modeling,the difference of indexes in each group decreased and tended to the level of group C.(2)Open field experiment: 1d,3d and 7d after modeling,the total distance and average speed of group M were significantly lower than those of group C and group I;3d after modeling,the total distance and speed of group I were lower than group C.2.RCR: Compared with group C and group I,m RNA expressions of NLRP3,caspase-1,IL-18 and IL-1β in group M were significantly up-regulated 1 and 3 days after modeling;The m RNA expression of each indicator in group I was between group M and group C,and was significantly up-regulated compared with group C.On7 days after modeling,m RNA expressions of NLRP3,caspase-1,IL-18 and IL-1βwere not significantly different among all groups.3.Western Blot: Compared with groups C and I,the expressions of NLRP3,caspase-1,IL-18 and IL-1β in group M were significantly up-regulated 1 and 3 days after modeling.The expression of each indicator protein in group I was between group M and group C,and was significantly up-regulated compared with group C.On day 7after modeling,there were no significant differences in the expression of NLRP3,caspase-1,IL-18 and IL-1β among all groups.4.Immunofluorescence staining: C ompared with group C and group I,the pyroptosis rate and mitochondrial reactive oxygen species production in group M were significantly increased at 1d,3d and 7d after modeling.The production of mitochondrial reactive oxygen species in group I was higher than that in group C at 1d,3d and 7d after modeling.Conclusion: NLRP3/ caspase-1-mediated pyroptosis may be one of the mechanisms leading to PND.NLRP3 inhibitor MCC950 can block the pyroptosis pathway and reduce the degree of PND damage. |
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