| ObjectiveTo explore the neuroprotective mechanism of xenon,we established a model of white matter damage in 3-day-old neonatal rats by injecting intraperitoneally with lipopolysaccharide combined with hypoxia-ischemia and detected the expression of CLIC4 and Bcl-2 in the white matter.MethodsThree-day-old SD rat pups(n=120)were randomly divided into blank control group(n = 24),brain injury group(n = 24)and xenon intervention group(n = 72).The rats in the blank control group were intraperitoneally injected with normal saline without hypoxia-ischemia and xenon intervention.The rats in the brain injury group were intraperitoneally injected with lipopolysaccharide and hypoxia-ischemia without xenon intervention.The rats in the xenon intervention group were intraperitoneally injected with lipopolysaccharide(LPS,0.05 mg / kg),the right common carotid artery was dissected and were treated with hypoxia-ischemia and xenon intervention(the rates were placed in a closed circuit container within 50% xenon mixed with 30% oxygen and 20% nitrogen for3 hours).According to the intervention time of xenon(0-hour,2-hour and 5-hour)after injury,the xenon intervention group was divided into three subgroups(C1?C2 and C3,n=24).Pathological changes in the cerebral tissues of rat pups were detected using hematoxylin and eosin stain.The expressions of CLIC4 and Bcl-2 were detected using RT-PCR and Western blot technique.Results(1)After intraperitoneal injection of lipopolysaccharide and hypoxia-ischemic,rat pups in the brain injury group and the xenon intervention group showed limbs shaking,dysphoria,cyanosis,tachypnea,circling and convulsions.After dealing with xenon,symptoms of rats in Group C1 were disappeared about 2 hours later,while symptoms in Group B,Group C2 and Group C3 had almost disappeared after 5 hours.Rat pups in the blank control group did not show significant abnormal reactions.(2)Our study shown that in the brain injury group the white matter which stained by HE was pale,the structure was loose,the cell swelling,the nuclear membrane was unclear,the nuclear pyknosis which indicate the cell degeneration and necrosis,and the number of glial cells increased;the white matter of the xenon intervention group was stained lightly,the structure was relatively complete and the degenerative and necrotic cells were little;and there was no significant difference between the subgroups.The brain structure of the blank control group was normal,the staining was clear and the cells were neatly arranged.(3)The expression of Bcl-2 was markedly increased in Group C1 and Group C2 when compared with the brain injury group at all of the time points(P < 0.05),but in Group C3 in which xenon-treatment was delayed for 5 hours,the the expression of Bcl-2at 48 h and 72 h when compared with Group B did not reach statistical significance(P >0.05).The expression of Bcl-2 in Group C2 and Group C3 was significantly lower at 48 h and 72 h when compared with Group C1(P < 0.05).(4)The expression of CLIC4 m RNA at different time points in the white matter were increased in the brain injury group when compared with in the blank control group,and the difference was statistically significant(P < 0.05).There was a marked reduction of the expression of CLIC4 m RNA in the subgrops C1?C2?C3 compared with the brain injury group,and the difference was statistically significant(P < 0.05).Conclusion(1)Intraperitoneal injection of LPS combined with hypoxia-ischemia can create white matter damage model in 3-day-old neonatal rats.(2)The inhalation of xenon could reduce the damage of white matter tissue in the brain and reduce cell necrosis.(3)The expression of Bcl-2 protein after white matter injury is increased,and the expression of Bcl-2 protein can be decreased after xenon intervention,showing that xenonmay play a neuroprotective role by inhibiting apoptosis.(4)Xenon intervention can down-regulate the expression of CLIC4 gene,indicating that xenon may play a neuroprotective role by inhibiting the expression of CLIC4 gene in the Bcl-2 protein-related pathway which can inhibit neuronal apoptosis.(5)Xenon application against white matter damage has a therapeutic time window which at least 5 hours,and the sooner of the xenon intervention the better the of the effect. |
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