Research On The Effect Of Blood Se On Brain Tissue Damage Of Rats With Severeacute Carbon Monoxide Poisoning

ObjectiveTo analyze the effect of blood Se on brain tissue damage of rats with severe acutecarbon monoxide poisoning, and discuss whether Se is the adjuvant therapy for ratswith severe acute carbon monoxide poisoning.Method1.105of107rats(each weighs450±20g) were selected through Morris watermaze training,10of which were randomly selected into control group (BC group).The remaining95rats were treated with repeated intraperitoneal injections of purecarbon monoxide gas to build the model of acute CO-P; and45rats having survivedthe modeling and resumed normal diet were divided equally into3groups whichincluded low-dose selenium treatment group (LSe group), high-dose seleniumtreatment group (HSe group) and carbon monoxide exposure group (CO-P group).2. Morris water maze test was performed once every three days to evaluateneurobehavioral changes and escape latency was recorded.3. Sodium selenite injection: intramuscular injection of0.1μg/g/d per day foranimals of LSe group, intramuscular injection of0.2μg/g/d per day for animals ofHSe group. Intramuscular injection of corresponding amount of normal saline foranimals of both control group and CO-P group.4. Three blood samples (0.5ml for each) from each group were collected at tailon a daily basis to monitor dynamic changes of blood selenium and glutathione-peroxidase activity.5.21days after carbon monoxide exposure, brain tissues of rats were sampled toobserve brain structure and neurocyte morphology by hematoxylin-eosin (HE) staining method. The structure and cellular morphology of white matter myelin wereobserved by Luxol fast blue (LFB) staining method. Neurocyte apoptosis was stainedby Tunnel terminal transferase method.6. Myelinbasicprotein(MBP) level in hippocampi was observed byimmunohistochemistry(IHC) staining method in which brown was considered aspositive signal. Three fields were selected for each section of gyrus hippocampi witha high power microscope (amplification:10×20). Average intergral optical density(IOD) at positive site was measured with image-pro plus6.0image analysis softwareand statistical analysis was performed based on average data.7. Statistical Method: SPSS13.0software was used for statistical analysis;quantitative data were analyzed with the method ofX S; data between each groupat each time point were compared by repeated-measures ANOVA; pairwisecomparison test was applied in data analysis for4groups, in which LSD test wereapplied for homogeneous variance while Dunnett’s T3and Kruskal-Wallis tests wereused for non-homogeneous variance. Significance level: α <0.05indicates asignificant difference.Result1. Establishment of Rat model of severe acute CO-P: After maintainingCOHB(concentration>65%) for16hours, among the comatose rats which were fedintragastrically for48hours,49.5%survived and49.4%resumed normal diet.2. Neurobehavioral assessment: Morris water maze test and repeated-measuresANOVA were employed. Escape latency of the rats in BC group shortened graduallyas opposed to the gradual prolongation observed in CO-P group, while escape latencyof the rats in both LSe group and HSe group shortened at first but prolonged at last.There was statistically significant difference in escape latency between each group. 3. Dynamic changes of blood Se: The Se level in the four groups were comparedin pairs by statistical methods: LSD was used in data with equal variance whileDunnett’ s T3was used in data with unequal variances. Se level in the two treatmentgroups were higher than in poisoning group. The low-dose treatment group reached atthe same level of the blank group in Day20while the high-dose treatment group usedonly15days to reach at the level, and the differences of daily average Se levelbetween groups were statistically significant (P<0.001).4.Activity changes of GSH-PX: The activity of GSH-PX in the four groups werecompared in pairs by statistical methods: LSD was used in data with equal variancewhile Dunnett’ s T3was used in data with unequal variances. The GSH-PX level inthe blank group was higher than the other three groups. The GSH-PX level in the twotreatment groups reached nearly up to the level in the blank group in Day4-8. Theactivity changes of GSH-PX varied over groups, time point and time, and thedifferences were statistically significant (P<0.001).5. Pearson correlation method was used in analyzing the enzyme activity, resultsshowed that Se level in eight days before Se supplement in poisoning groups hadpositive linear correlation to the low-dose Se treatment group（r=0.755，p<0.05）andthe high-dose Se treatment group (r=0.841，p<0.01).6. HE staining: Severe hydropic degeneration of brain tissue of the rats in CO-Pgroup was observed. Though microscopic changes observed in both two treatmentgroups were consistent with that in CO-P group, the degree of injury was lower inboth treatment groups.7. Demyelination in cerebral white matter was found in the poisoned ratsthrough Luxol Fast Blue(LFB) staining while it was relatively mild in both LSe groupand HSe group.8. Apoptosis in the hippocampi was found in the poisoned rats through Tunnel terminal transferase staining; and it was the same with the cerebellum. Similarpathological changes were observed in both two treatment groups and no significantdifference in degree of changes was found between CO-P group and two treatmentgroups.9.1Under microscope, nerve cell necrosis, and degenerative changes and nervefiber loss at the MBP-positive site of hippocampi of the poisoned rats were observedthrough immunohistochemistry staining. Microscopic pathologic observation showedno significant difference between three groups.9.2IOD values between4groups showed a non-homogeneous variance (F=10.271, P <0.001). IOD values between4groups showed statistical significance (H=21.753, P <0.001) by Kruskal-Wallis rank sum test. In general, IOD values of LSegroup, HSe group and control group were different via pairwise comparison.Difference of IOD values of LSe group, HSe group and control group was found to bestatistically significant via pairwise comparison. However, IOD values between LSegroup and HSe group showed no statistically significant difference.ConclusionBy increasing the Se level in rats, the activity of GSH-PX can be enhanced whilethe brain damage of rats with severe acute carbon monoxide poisoning can bereduced.