| Objective:To explore the roles and mechanisms of the Hemopexin (Hx)about the brain edema after intracerebral hemorrhage,In this subject theautologous whole blood from the rats will be removed or in addition tothe Hx and injected to the brain of the rats to produce experimentalintracerebral hemorrhage (ICH) model,and then observe the relationsbetween Hx and water content of brain tissue around the hematoma andcontent of hememetabolites in the hematoma,brain tissue inflammationreact and the formation of free radicals around the hematoma. Method：1.To divide SD rats into groups:100SD rats were randomly divided intofour groups(25per group) named Sham operation group,ICH controlgroup,Hx removal group and Hx intervention group. Every group hadfive time points:1d,3d,7d,14d,21d after the surgery.2.Production model:(1) ICH control group: The right caudate nucleus of the rats would beinjected50μl autologous whole blood;(2) the Sham operation group: thesame surgical procedure as ICH control group, but do not injectautologous blood, only inject50μl Saline;(3) Hx removal group: Tailblood1ml,the right caudate nucleus of the rats would be injected theautologous whole blood of removal Hx50μl;(4) Hx intervention group:tail blood100μl, the right caudate nucleus of the rats would be injected autologous whole blood of plus1μl (5μg/μl) Hx50μl.3.It wile evaluatethe model whether successful or not after two hours by Bederson’smethod and the neurological dysfunction scores would be used toestimate by Garcia’s mothod；Water contents of brain tissue around thehematoma would be detected by dry-wet weighmethod;Spectrophotometric would be used to estimate content of theheme degradation products;The SOD activity would be measured by thexanthine oxidase method;the content of the MDA would be measured bythe thiobarbituric acid method;The levels of the inflammatory(IL-1β,IL-6,TNF-α) of the brain tissue would be measured by theRadioimmunoassay;Pathological changes of brain tissue around thehematoma would be detected by HE staining pathology andimmunohistochemical method at each observation time points;The data ofresults were analyzed statistically by the SPSS13.0. Results:1.Theneurological function disability scores at each time points:Theneurological dysfunction scores of the Sham operation group was nosignificant difference(P>0.05) at all time points;There was no significantdifference (P>0.05) in the ICH control group,Hx removal group and Hxintervention group at1d;The Neurobehavioral scores in the ICH controlgroup, Hx removal group and Hx intervention group was significantlylower than that in the Sham operation group (P<0.05)within1d～21d atall time points; The Neurobehavioral scores of the Hx removal group was significantly lower than the ICH group (P<0.05)within3d～21d at eachtime points;The Neurobehavioral scores of the Hx intervention group wassignificantly higher than the ICH group (P<0.05) within3d～21d at eachtime points.2.The water content of the brain tissue around thehematoma:The water content of the brain tissue in the Sham operationgroup was no significant difference (P>0.05) at all time points;The watercontent of the brain tissue in the ICH control group,Hx removal groupand Hx intervention group were significantly higher than the Shamoperation group at corresponding time points(P<0.05)within1d～21d；The water content of the brain tissue in the ICH control group,Hxremoval group and Hx intervention group was no significant difference(P>0.05) at1d; The water content of the brain tissue in the Hx removalgroup were significantly higher than ICH control group (P<0.05)within3d～21d；The water content of the brain tissue in the Hx interventiongroup were significantly lower than ICH control group (P<0.05) within3d～21d;3.The Inflammatory factors (IL-1β,IL-6,TNF-α) changes atvarious stages:There was no significant difference (P>0.05) in the Shamoperation group at each time points about the levels of the IL-1β,IL-6andTNF-α within1d～21d; The levels of the IL-1β,IL-6and TNF-α in ICHcontrol group,Hx removal group and Hx intervention group wassignificantly higher than the Sham operation group (P<0.05) at thecorresponding points within1d～21d; The levels of the IL-1β,IL-6and TNF-α at1d～21d in the Hx removal group was significantly higher thanthe ICH group (P<0.05) at the corresponding points; The levels of theIL-1β,IL-6and TNF-α in the Hx intervention group was significantlylower than the ICH group (P<0.05) within1d～21d;4.The changes ofSOD and MDA:4.1The changes of the SOD activity: There was nosignificant difference (P>0.05) in the Sham operation group at each timepoints; The levels of the SOD activity in the ICH group, Hx removalgroup, and the Hx intervention group was significantly lower than that inthe Sham operation group(P<0.05) within1d～21d;The levels of theSOD activity in the Hx removal group was significantly lower than thecorresponding time points in the ICH control group (P<0.05)at1d～21d;The levels of the SOD activity around the hematoma in the Hxintervention group was significantly higher than the corresponding timepoints in the ICH control group (P<0.05) at1d～21d;4.2The levels ofthe MDA at various stages: There was no significant difference (P>0.05)in the Sham operation group at each time points;The levels of the MDAin the ICH control group,Hx removal group and Hx intervention groupwas significantly higher than the corresponding time points in the Shamoperation group(P<0.05) within1d～21d; The levels of the MDA in theHx removal group was significantly higher than the ICH controlgroup(P<0.05) at corresponding time points within1d～21d; The levelsof MDA in the Hx intervention group was significantly lower than the ICH control group(P<0.05) at corresponding time points within1d～21d;5.The changes of the He and its degradation degradation products(CO,Fe,UCB): There was no significant difference (P>0.05)in the Shamoperation group at each time points;The levels of the He and itsdegradation products in the ICH control group, Hx removal group and theHx intervention group was no significant difference (P>0.05) at1d;Thelevels of He and its degradation products in the ICH control group, Hxremoval group and the Hx intervention group was significantly higherthan the Sham operation group(P<0.05) at corresponding time pointswithin3d～21d;The levels of the He and its degradation products in theHx removal group was significantly higher than the ICH controlgroup(P<0.05) within3d～21d;The levels of the He and its degradationproducts in the Hx intervention group was significantly lower than thethe ICH control group(P<0.05) within3d～21d;6.The observation of theHE dyeing and immunohistochemical by Microscopy:There was noobviously pathological changes in the Sham operation group;It wasshowed obviously classic pathological changes in the ICH control group,Hx removal group and Hx intervention group that theneurons,glial,endothlial cells of blood vessel was swelling and the braintissue is loosen,the gap was widen around the neurons and glia,inaddition,it was showed lymphocytic Infiltration and gliosis and smallblood vessel to dilate;The pathological changes in the Hx removal group was significantly more serious than the ICH control group;Thepathological changes in intervention group was significantly less seriousthan the ICH control group;7.Immunohistochemica scoresresults:There was no significant difference （P>0.05) in the Shamoperation group at each time points about within1d～21d;The scores inthe Sham operation group,ICH control group,Hx removal group and Hxintervention group was no significantly difference at1d;The scores in theICH control group,Hx removal group and Hx intervention group washigher than the Sham operation group (P<0.05) at the correspondingpoints within3d～21d; The scores at3d～21d in the Hx removal groupwas significantly higher than the ICH group (P<0.05) at thecorresponding points; The levels of the scores in the Hx interventiongroup was significantly lower than the ICH group (P<0.05) at thecorresponding points;The Immunohistochemistry scores in the ICHcontrol group,Hx removal group and Hx intervention group was highestin3th day.8.The correlation analysis:(1) The He and its degradationproducts，content of IL-1β，IL-6,TNF-α and content of MDA in the ICHcontrol group, Hx removal group and Hx intervention group wasnegatively correlated with the immunohistochemistry scores; SODactivity was positively correlated with the immunohistochemistryscores.(2)The neurological function disability scores in the ICH controlgroup, Hx removal group and Hx intervention group was significantly negatively correlated with water content of the brain tissue，the levels ofthe IL-1β,IL-6,TNF-α and the levels of the He and its degradationproducts and MDA content,But positively correlated with SOD activityand immunohistochemistry scores;(3)The water content of the braintissue in ICH control group,Hx removal group and Hx intervention groupwas positively correlated with the levels of the IL-1β,IL-6,TNF-α and thelevels of the He and its degradation products and MDA content,negatively correlated with SOD activity and immunohistochemistryscores. Conclusion:1.The models of rats intracerebral hemorrhage madeby autologous whole blood is simple, repeatable and easy to operate, thechanges of pathophysiology of rat models were almost the same as thepatients with ICH,so it can be widely used for cerebral hemorrhage study.2.The brain tissue showed obviously edema after ICH which related tothe risen of the He and its degradation products and inflammation factorsand free radical release.3.Increased expression of Hx after ICH and it caninhibit the delayed brain edema formation.4.Hx has anti-oxidativedamage,inhibit inflammatory reaction and free radical generation toprotect brain.4. The combination of Hx and He to form Hx-Henon-covalent complex, it can reduce the accumulation in the neural tissuewithin the He and its metabolism and its neurotoxicity. |
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