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The study on the expression of neurolobin in rats during the process of brain edema induced by endotoxin

LIU C;SUN S Q;YANG M;CHEN H (Institute of Neuroscience;Chongqing Medical University;Chongqing 400016;China)

  Endotoxin-induced brain lesion(EIBL) is commonly encountered in clinic.However,the pathological mechanism of EIBL has not yet been fully clarified.Endotoxi n(gram-negative cell wall lipopoly saccharide,LPS) is a major factor resulting in brain lesion.The papers showed that the LPS-induced brain lesion will activate the endogenous protective mechanism in the Patholo gical process of ischemia and hypox ia.Neuroglobin(Ngb),the most important Oxygen-carrying protein in the neurons which is able to remove the free Radical in neurons likely participates in endogenous protective mechanism.Objectives: The aim of this study was to observe the change of Ngb expression,so as to verify the endogenous nuroprotec tive mechanisms.The models of endotoxin shock and LPS brain lesion were established.Changes of Ngb expression in the frontal cortex, hippocampus,cerebrospinal fluid and plasma were examined by ELISA,Western Blot and IHC. EXPERIMENTⅠ:Methods:Seventy adult Sprague-Dawley rats were randomly divided into seven groups (10 rats in each group):the control group(0.5 mL normal saline was injected via vena caudalis of animals) and the experimental groups(0.5 mL Endotoxin was injected via vena caudalis of animals,0.016 g/kg) The specime ns were properly taken at 3 h,6 h, 12 h,24 h,48 h and 72 h after endotoxin injection).The expression of Ngb in each time points was measured by ELISA,WesternBlot and IHC.The brain water content was measured by dry and wet method.The brain water content was measured by dry method.The swelling of the organelles in the hippocampus and cortex of frontal lobe of rats was detected by the transmission Electron Microscope (TEM).Results:①ELISA showed that the Ngb concentrations in the plasma,CSF,hippocampus and cortex of frontal lobe were significan tly higher than those of control group (P<0.01 vs control) after endotoxin injection,and peaked at 48 h.②Western Blot showed that Ngb with relative molecular mass 17 kD was observed in all groups.The Ngb expression in experimental groups was significantly higher than that in the the control group.The expression of Ngb was the highest in 48 h group and 72 h group,and the lowest in 3 h group.③IHC showed that Ngb-po sitive cells were distributed in neurons of pyramidal layer,and Ngb-immunoreactive product was located in the cytoplasm of the pyramidal neurons,which were yellow-brown in color.④After endotoxin injection,the brain water content was significantly higher (P<0.01 vs control),and peaked at 48 h.Conclusions:Our results indicate that there was a positive correlation between the up-regulatio n of Ngb expression and the severity of brain lesion.And the expressions of Ngb were up-regulated with a time-dependent manner.The up-reg ulation of Ngb maybe one of the endogenous nuroprotective mechanis ms under endotoxic shock condition. EXPERIMENTⅡMethods:One hundred adult Sprague-Dawley rats were randomly divided into the control group(n=10,0.1 mL normal saline was injected via ventriculus quartus of animals) and the experimental group(n=90).In the experimental group,ninety adult Sprague-Dawley rats were randomly divided into 9 groups(n=10,in each group;Endotoxin was injected via ventriculus quartus of animals) by factorial design.The dose factor (lower-dose,middle-dose and higher-dose) and the time factor (12h,24h,48h) were analysised in this experiment.The expression of Ngb over different periods was meas ured by ELISA and Western-Blot. Results:①ELISA showed that Ngb expression in the cortex of frontal lobe and hippocampus of rats was increased with the dose and the duration of time after endotoxin injection.②Western Blot showed that Ngb with relative molecular mass 17 kD was observed in all groups.Conclusions:The tendency of Ngb expression change in the cortex of frontal lobe and hippocampus was the same as ELISA.Our results indicate that the changes of Ngb expression were correlated with the endotoxin dose and the duration of time.……