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目的探讨低分子肝素(low molecular weight heparins,LMWH)对大鼠脑缺血再灌注后NF-κB活化和表达的影响及其可能机制。方法通过线栓法对大鼠建立大脑中动脉栓塞(middle cerebral artery occlusion,MCAO)模型,将总计60只大鼠分为3组:假手术组(Sham,n=20)、模型组(I/R,n=20)、低分子肝素干预组(LMWH,n=20),并用1.5 mg/kg的低分子肝素液进行干预。观察大鼠脑缺血再灌注后神经功能评分、梗死灶体积,并用Western blot、免疫组化、免疫荧光检测各组IL-1β、NF-κB P65及SUMO2/3蛋白表达的变化。结果 (1)神经功能评分结果显示:MCAO后大鼠出现严重的神经功能障碍,经LMWH干预后能明显减轻MCAO导致的神经功能障碍(P<0.01);(2)大鼠在脑缺血再灌注后出现严重的脑梗死(P<0.01),而LMWH能明显减小梗死体积(P<0.01),尤其能够有效改善MCAO模型梗死敏感区纹状体的损伤程度(P<0.01);(3)Western blot检测结果显示:再灌注24 h后I/R组大鼠脑组织中IL-1β、NF-κB P65蛋白表达均高于Sham组(P<0.01),而经LMWH干预后的大鼠IL-1β、NF-κB P65蛋白表达相比I/R组均下降(P<0.01);再灌注3 h后,与Sham组相比,I/R组大鼠脑组织中SUMO2/3蛋白表达增强(P<0.01),而LMWH干预后SUMO2/3蛋白表达进一步增强(P<0.01);(4)免疫组化和荧光检测结果显示:再灌注3 h后,与Sham组相比,I/R组大鼠脑组织中蛋白SUMO2/3水平增加,出现核内移现象(P<0.01),并且主要发生在神经元细胞内,而LMWH干预后发生SUMO2/3核内移现象的神经元细胞进一步增多(P<0.01)。结论 LMWH在大鼠脑缺血再灌注损伤模型中能够促进SUMO2/3表达及核内移,这可能对抑制NF-κB活化减少炎症因子生成产生影响。
Objective To investigate the effects of low molecular weight heparins (LMWH) on the activation and expression of NF-κB after cerebral ischemia-reperfusion in rats and its possible mechanism. Methods The middle cerebral artery occlusion (MCAO) model was established by thread occlusion in rats. A total of 60 rats were divided into 3 groups: sham operation group (n = 20), model group (I / R, n = 20), low molecular weight heparin intervention group (LMWH, n = 20) and intervention with 1.5 mg / kg low molecular weight heparin solution. Neurological scores and infarct volume were observed after cerebral ischemia-reperfusion in rats. The expressions of IL-1β, NF-κB P65 and SUMO2 / 3 were detected by Western blot, immunohistochemistry and immunofluorescence. Results (1) The results of neurological function showed that severe neurological dysfunction was found in rats after MCAO, and the neurological dysfunction caused by MCAO could be obviously alleviated by LMWH intervention (P <0.01). (2) Severe cerebral infarction was found after perfusion (P <0.01), while LMWH could significantly reduce the infarct volume (P <0.01), especially in the striatum of MCAO model (P <0.01) ) Western blot results showed that the expression of IL-1β and NF-κB P65 protein in I / R group was higher than that in Sham group (P <0.01) at 24 h after reperfusion, while the rats in LMWH group Compared with Sham group, the expression of IL-1β, NF-κB P65 protein in I / R group decreased significantly (P <0.01), and the expression of SUMO2 / 3 protein in I / R group (P <0.01). (3) Compared with Sham group, the expression of SUMO2 / 3 protein increased further after LMWH intervention (P <0.01); (4) Immunohistochemistry and fluorescence detection showed that the expression of SUMO2 / The level of protein SUMO2 / 3 in brain tissue of rats in R group increased (P <0.01), and mainly occurred in neurons, while the neurons in SUMO2 / 3 nucleus migrated after LMWH intervention Further increase (P <0.01) . Conclusion LMWH can promote the expression of SUMO2 / 3 and its nuclear translocation in the rat model of cerebral ischemia-reperfusion injury, which may play an important role in inhibiting the activation of NF-κB and reducing the production of inflammatory cytokines.