| Objective To elucidate the protective effect of limb ischemiapostconditioning on rat focal cerebral ischemia-reperfusion injury and study theinfluence of limb ischemia postconditioning on endogenous janus kinase2/signal transducer and activators of transcription3signal transduction pathway,explore its brain protective mechanism.Methods Using the modified suturemethod to make focal brain artery ischemiaand reperfusion rats model.(Middlecerebral artery occlusion and reperfusion, MCAO/R). sixty healthy maleSpragueDawley rats（240-260g）were randomly divided into four group：sham;ischemia-reperfusion (I/R),ischemia2hours and reperfusion24hours withoutadditional intervention; limb ischemia postconditioning(Lpost), after2hoursischemia, the rats were comprised3cycles of15seconds reperfusion followedby15seconds ischemia, and then the rats were reperfused24hours; andLpost+AG490group(JAK2/STAT3pathway inhibitor)group, rats were treatedwith intraperitoneal injection of AG490lmg/kg5minutes before reperfusion,followed by limb ischemia postconditioning。Neurological function scores wereused to assess the behavior,HE staining was used to examine brain tissueinjury,Cerebral infarction volumes were quantified by TTC staining，Neural cellapoptosis was detected by TUNEL, westernblot assay ischemic penumbraP-STAT3expression,The expressions of Caspase-3were detectedbyimmunohistochemistry.Analysis the relationship between limb ischemia postconditioning and P-STAT3,Caspase-3expression amount in cerebralischemic and reperfusion district. Results1.The effect of limb ischemiapostconditioning on the changement of neuroethology after ischemia/reperfusion:sham-operated group had no significant neurological deficits, cerebralischemia/reperfusion model group and limb ischemia treatment+AG490grouprespectively showed more severe neuronal deficits than Limb ischemiatreatment group(P<0.01), indicating limb ischemia postconditioning mayimprove the symptoms of neurological deficits.2.The effect of limb ischemiapostconditioning on the cerebral parietal cortex pathological changes: Braintissue structures were normal in sham group, endothelial cells and nerve cellsintact structure, clear cytoplasm and nucleus. Ischemia-reperfusion groupshowed cortical cell shrinkage, cytoplasm loose, cytoplasmic concentration,nuclear condensation, chromatin condensation near the nuclear membrane,formation of apoptotic bodies. neurons degeneration and necrosis Lighted inLpost+AG490group than in cerebral I/R group, and necrosis was lighter inLpost group than the previous two, the difference was significant(P<0.05).limbischemia postconditioning can reduced brain pathological tissue damage.3.Theeffect of limb ischemia postconditioning on the cerebral ischemic lesion volumeafter ischemia/reperfusion: Ischemic mainly involving the cortex and basalganglia，TTC staining is shown that the red part are ischemia, ischemic part arethe pale white. Sham group rat brain tissue without infarct. Cerebralischemia-reperfusion group the largest，Lpost+AG490group reduce infarct sizethan the former，The difference was statistically significant compared them(P<0.05). Lpost group reduced infarct size most, they was statistically significantcompared (P <0.05).limb ischemia postconditioning can reduce the infarct lesionvolume.4.The effect of limb ischemia postconditioning on the expression of Caspase-3immunoreactive cells after ischemia/reperfusion: Caspase-3is mainlyexpressed in neurons， immunoreactive cells was brown, microscopecytoplasm, nucleus or particle calm brown colored positive cell, No brownstaining was negative.The Caspase-3positive cells were occasionally observedin sham group， The expression of Caspase-3immunoreactive cells wasincreased in ischemia penumbra zone after focal cerebral ischemia/reperfusion,Contrast with the sham group, the expression of Caspase-3immunoreactive cellswas increased obviously in cerebral I/R group(P<0.05). Compared with I/Rgroup, Lpost group and Lpost+AG490group Caspase-3immunoreactive cellswere significantly reduced, the difference was statistically significant (P<0.05).Lpost group Caspase-3immunoreactive cells decreased significantly thanLpost+AG490group (P<0.05), limb ischemia postconditioning can reduce thenumber of Caspase-3immunoreactive cells.5.The effect of limb ischemiapostconditioning on the expression of P-STAT3expression in Parietal brainischemia after ischemia/reperfusion: Sham group showed no expression ofP-STAT3protein，P-STAT3protein was significantly increased in cerebral I/Rgroup, Lpost group reducing expression of P-STAT3protein and in Lpost+AG490group,P-STAT3protein expression was significantly reduced。Thedifference was statistically significant compared them(P <0.05),limb ischemiapostconditioning can reduce the expression of P-STAT3protein.Conclusions1.Neurologic deficits, ischemic pathological changes of brain tissue, increasesof infarct volume and brain cell apoptosis are typically shown afterischemia/reperfusion.2.cerebral ischemia-reperfusion induce P-STAT3expression, increase expression of Caspase-3positive cells. Caspase-3participate in the formation of apoptosis after ischemia, and STAT3activationand overexpression may regulate the expression of Caspase-33. limb ischemia postconditioning can improve ischemic reperfusion neurological deficits,reduce the extent of necrosis of brain cells, reduce infarct volume and brain cellapoptosis. The protective mechanism may be associated with inhibitionJAK/STAT pathway decreasing P-STAT3expression in brain tissue, reducingCaspase-3expression. |
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