| Ischemia stroke is a prevailing nervous system disease, which disturbspeople's life due to high morbidity and mortality as well as high risk to bedisability Salvation therapy in the post-ischemia period seem to be limited torescue the damage from ischemia and reperfusion, although there are lots ofprogress in the study on the theoretic mechanisms. So, how to prevent the injuriesafter stroke becomes a hot topic in neuroscience field. Cerebral ischemiatolerance induced by preconditioning brings us a new hope to protect and cureIschemia brain injuries.A serial of experiments showed that oxygen breathing preconditioning is aneasily handled and clinically practical pretreatment method, which hints a goodclinical practice due to its induction of cerebral ischemia tolerance. Theexperiment had proved that pure oxygen inhalation could activate neurons andglia in the brain, but induction of long term of ischemia tolerance under oxygeninhalational pretreatment still remains elusive. The research on that issue willexplore the mechanisms of brain protection. So we designed the following experiments: observe that effect of different time course oxygen inhalationalpretreatment on cerebral ischemia–reperfusion injury in rat and the effects oflong-term oxygen inhalational pretreatment induced cerebral ischemia toleranceon blood–brain barrier(BBB), then further study the glia and MMP9 whichmainly compose of BBB with HIC. All the works explore the mechanisms ofbrain protection and provide plot of the strategies for the treatments andprotections against brain ischemia injury, which has both theoretical and clinicalsignificance.Part 1 The neuroprotective effects of different time course oxygen-breathingpreconditioning against cerebral ischemia-reperfusion injury in ratsExperiment 1: Effects of different time course oxygen-breathingpreconditioning on the protection against cerebral ischemia-reperfusion injury inratsAim: to identify the optimal strategy of oxygen-breathing preconditioning forneuroprotective effects against cerebral ischemia-reperfusion injury in ratsMethods: Fifty-six male Spraque-Dawley(SD) rats , weighing 280～320 g,were randomly divided into 7 groups (n = 8 in each group), control group withair-breathing; other 6 groups are respectively group A~F pretreated with differentstrategies of oxygen-breathing。The rat middle cerebral artery occlusion (MCAO)models were made and occlusion sustained for 120min with nylon monofilament.After reperfusion for 24h, neurological deficit scores were evaluated with theGarcia and then the animals killed to measure infarct volumes.Results: The neurological scores were higher in Group C(inhaling 100%oxygen for once a day, 8 h/day, three days)and Group D(inhaling 100% oxygenfor 8 h/ day,1 week)than that in other groups; Infarct volume percentage in GroupC and Group D were significantly smaller than that in other groups. Experiment 2 The effect of different time course oxygen-breathingpreconditioning on SOD and MDA in MCAO ratsAim: to evaluate the effect of different time course oxygen-breathingpreconditioning on SOD and MDA in MCAO ratsMethods: 24 male SD rats, were randomly divided into four groups (n=6 ineach) and breathed pure oxygen: group 1 (breath O2 1h, one time/day,1week);group 2 (breath O2 8h, one time/day,1week ); group 3 (breath O2 24h, three times,24h interval ); group 4(breath O2 8h, one time/day, 3days ). The rat middlecerebral artery occlusion (MCAO) models were made and occlusion sustained for120min with nylon monofilament. After reperfusion for 24h, neurological deficitscores were evaluated with the Garcia and then the animals were killed tomeasured SOD and MDA from both sides.Results: in the ischemic ipsilateral hemisphere, SOD of group with breathingO2 8h, one time/day,1week(134.5±26.2) was higher than other groups andsignificantly higher than group with breathing O2 1h, onetime/day,1week(69.5±13.0). While MDA of group with breathing O2 8h, onetime/day,1week(4.3±0.58) was lower than other groups and significantly lowerthan group with breathing O2 1h, one time/day,1week(9.1±1.4) (p<0.01). Thereare no statistical differences on SOD and MDA between ischemia and nonischemiasides of brain in each group.Experiment 3 Duration of cerebral ischemia tolerance induced by oxygenbreathing preconditioningAim: to observe the time-lasting of cerebral ischemia tolerance induced byoxygen breathing preconditioningMethods: from the experiment 1, choosing the best mode of oxygenbreathingprecondition, then according to the different time before MCAO, 40 male SD rats were randomly divided to 5 groups(n=8), Group I: 24h post oxygenbreathingprecondition; Group II: 48h post precondition; Group III: 1w postprecondition; Group IV: 1w post precondition, intensifying with 8h-oxygenbreathing24h before MCAO; Group control: air breathing 1w post precondition.Then, the rat middle cerebral artery occlusion (MCAO) models were made andocclusion sustained for 120min with nylon monofilament. After reperfusion for24h, neurological deficit scores were evaluated with the Garcia and then theanimals killed to measure infarct volumes.Results: Compared with control group, Group I, Group II, Group IV havesignificantly higher Carcia scores and lower cerebral infract volume ratio thancontrol group.Part 2 The effect of long-term oxygen breathing preconditioning on Blood-Brain Barrier permeability in MCAO ratsExperiment 1 The effect of long-term oxygen breathing preconditioning onBlood-Brain Barrier integrity in MCAO ratsAim: To evaluate the effect of long-term oxygen breathing preconditioningon Blood-Brain Barrier integrity in MCAO ratsMethods: 10 male SD rats were randomly divided into two groups as follows(n=5 in each): control group breathing air and experiment group breathing O2, 8h,one time/day, I week. Then, the rat middle cerebral artery occlusion (MCAO)models were made and occlusion sustained for 120min with nylon monofilament.Just at the beginning of reperfusion, 2% Evans blue was injected through tail vein.BBB permeability was measured at 24 h after reperfusion.Results: EB content of the ischemic ipsilateral hemisphere in experimentgroup was significantly lower than that in control group (p<0.05). EB content ofthe ischemic contralateral hemisphere in experiment group was significantly higher than control group (p<0.05).Experiment 2 The effect of long-term oxygen breathing preconditioning onGFAP, MMP9 and cytokine IL-1βin MCAO ratsAim: To evaluate the effect of long-term oxygen breathing preconditioning onGFAP, MMP9 and cytokine IL-1βin MCAO rats.Methods 10 male SD rats were randomly divided into two groups as follows(n=5 in each): control group breathing air and experiment group breathing O2, 8h,one time/day, I week. Then, the rat middle cerebral artery occlusion (MCAO)models were made and occlusion sustained for 120min with nylon monofilament.After 24 h reperfusion, the brain tissues were fixed with PF and usingfluorescence-IHC method to detect GFAP, MMP9 and cytokine IL-1β.Results: Compared with control group, the number of GFAP positive neuronswas significantly greater in experiment group (p<0.05), while the contents ofMMP and IL-1βin oxygen breathing group was less than control group (p<0.05).ConclusionConclusions1. Breathing 100% oxygen for 8 h per day, 1 week, has the best effective neuroprotectionagainst focal cerebral ischemia-reperfusion injury in rats rather thanother time course oxygen breathing preconditioning.2. The tolerance of cerebral ischemia induced by oxygen-breathing preconditioncan last for several days, but this long term protective effect decayed with thetime going, which need"awaking"with short-time oxygen-breathing.3. Long-term oxygen breathing preconditioning reduces Blood-Brain Barrierpermeability in ipsilateral hemisphere in MCAO rats , but in ischemiccontralateral hemisphere Blood-Brain Barrier permeability increases versely.4. Long-term oxygen breathing preconditioning decreases MMP expression and reduces the levels of IL-1βreleasing, increases GFAP positive neurons inMACO rat, which at least partially explain the mechanism for permeability ofblood-brain barrier decreasing. |
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