| Objective:In This experiment,A rat model of acute carbon monoxide poisoning was prepared by modified interval intraperitoneal injection CO,and we use the PDTC to intervene before modeling for observing the behavioral and pathological changes of rats at different time points after carbon monoxide poisoning.Determine whether PDTC can improve the neurological function of rats after acute carbon monoxide poisoning and analyze the possible mechanism of NF-KB/iNOS signal pathway in acute carbon monoxide poisoning delayed encephalopathy,and find the possible protective mechanism of PDTC in delayed encephalopathyMethods:1.120 male SD rats with an average weight of 250g were randomly divided into three groups:air control group(AC group n=40),carbon monoxide poisoning group(CO group n=40)and carbon monoxide poisoning+PDTC intervention group(PD group n=40),using water maze test to eliminate mice with cognitive impairment.then each group were randomly divided into 5 subgroups.each group was classified according to 1 day,3 days,7 days,14 days and 21 days after exposure.2.The model of carbon monoxide poisoning was prepared by modified interval intraperitoneal injection in CO group of rats,The same volume of air was injected into the abdomen of AC group of rats.The rats in PD group were given 100 mg/kg of PDTC intraperitoneal injection of body weight before modeling,which lasted once a day until the rats were killed.3.The HbCO concentration of rat tail tip blood was measured after intraperitoneal injection,and the Morris water maze was used to detect the change of memory learning ability in rats.The NF-?B?iNOS expression of hippocampus was determined by immunofluo-rescence and Western Blot method.The changes of cell morphology were observed by HE staining,and the apoptosis of hippocampus was detected by Tunel stainingResults:1.The results of the Morris water maze showed that the average escape latency,the number of times of crossing the platform and the total time of platform movement did not change significantly in each group after exposure within 7days,and there was no statistical difference between the groups(P>0.05),Since the 14th days,the incubation period in the CO group was significantly prolonged compared with AC?PD group,and the AC group also changed significantly compared with the PD group,There were statistically significant differences between the two groups(P<0.05),The results suggest that the rats begin to develop behavioral cognitive changes around 14 days.PD group of rat platform in quadrant exercise time,the number of times through the platform is the same as above,and the statistical differences between groups is significant(P<0.05),With the prolongation of time,the difference became more and more obviously,which indicated that the behavioral cognitive loss associated with delayed encephalopathy gradually increased with the progression of time symptoms.2.The changes of arterial blood HbCO concentration in rats:The HbCO concentration in CO and PD groups were above 50%maintenance time last over 16h,confirmed the successful exposure.3 HE staining results:CO group of rat hippocampal CA1 neurons showed nuclear constriction,enlarged intercellular space,unclear enlarged intercellular space,unclear cytoplasm and nucleus,and disorderly arrangement of cells;The neurons in the CA1 area of hippocampus of AC group rats were arranged neatly and tightly,the nuclear membrane was intact,the nucleolus was clear,and no obvious neuronal cell damage was observed;PD group of rat hippocampal CA1 neuronal cells may be protected by PDTC intervention.4.Immunofluorescence results:Only a small amount expression of NF-?B?iNOS were detected at each time mode in the hippocampal CA1 region of the AC group of rats,and did not show dynamic changes at time points.The difference was not statistically significant(P>0.05);The expression of NF-kB in CO group increased after exposure(1d),increased rapidly at 3d and then slowly increased;the expression of iNOS increased first after exposure,and gradually decreased after reaching peak at 3d.The expression of NF-?B?iNOS was lower in PD group than in CO group,The difference was statistically significant(P<0.05)5.Western Blot test results:the NF-?B in the hippocampal CA1 area of CO group of rats was significantly expressed at 3 d and gradually increased over time;its iNOS expression reached a peak on the 3rd day and then gradually decreased.only a small amount of expression NF-?B?iNOS in the AC group.The expression of NF-?B and iNOS in PD group decreased compared with that in the group,The differences between groups were statistically significant(P<0.05);6.Tunel results:Only a very small number of apoptotic cells were observed at each time point in AC group in CA1 area of rat hippocampus,And the group comparison was not statistically significant(P>0.05).apoptotic cells were observed in the hippocampus from the first day of exposure in the CO group.on the seventh day,the apoptosis index reached a peak,and then decreased slowly with time,and the intra-group comparison was statistically significant(P<0.05).The results of PD group showed the same trend as those of CO group,but the apoptosis index of each time point was lower than that of CO group,The comparison between the two groups was statistically significant(P<0.05)Conclusion:1.multiple intraperitoneal interval injection CO method can successfully establish an acute carbon monoxide delayed encephalopathy model,and has the characteristics of simple operation,low equipment requirements and high success rate.2.The changes of memory behavior in rats were evaluated by Morris water maze experiment after infection.The success of DEACMP model was determined by combining the changes of HbCO concentration in rats and histopathological changes in hippocampus.3.NF-?B/iNOS signaling pathway is involved in the pathogenesis of DEACMP,and by intraperitoneal injection PDTC it can effectively inhibit the expression of NF-?B,reduce the transcriptional expression of downstream iNOS inflammatory factors alleviate the clinical symptoms of DEACMP,reduce the memory behavior damage of rats,and provide some experimental basis and theory for the later clinical DEACMP treatment. |
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