γδT细胞受体在小鼠心肺脑复苏后全脑缺血再灌注损伤中的作用机制

Mechanism ofγδT cell receptor in global cerebral ischemia-reperfusion injury after cardiopulmonary-cerebral resuscitation in mice

为探究γδT细胞受体在小鼠心肺脑复苏(cardiopulmonary-cerebral resuscitation,CPCR)后全脑缺血再灌注损伤中的作用机制,将成年雄性C57BL/6小鼠随机分为3组:空白组、CPCR组和CPCR+γδT受体拮抗组。采用窒息法建立CPCR后全脑缺血再灌注模型,制作脑组织石蜡切片,进行H-E和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记(terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling,TUNEL)染色,分别观察各组小鼠脑组织受损和神经元凋亡程度。用Western blotting和免疫组化法检测各组小鼠脑组织内IL-17A、IL-23p19蛋白的表达,用RT-PCR检测各组小鼠脑组织内IL-17A、IL-23p19 mRNA表达。结果显示,与空白组比较,CPCR组小鼠脑缺血灶中心区出现大量死细胞或细胞呈不规则状态肿大,核膜破裂,细胞结构消失,且神经元细胞凋亡程度增加,而CPCR+γδT受体拮抗组小鼠较CPCR组上述现象变轻;CPCR组小鼠较空白组脑组织损伤评分、细胞凋亡率、IL-17A和IL-23p19表达水平均显著升高(P<0.01),而CPCR+γδT受体拮抗组小鼠较CPCR组脑组织损伤评分、细胞凋亡率、IL-17A和IL-23p19表达水平均显著降低(P<0.05)。该研究提示,γδT细胞受体可能通过影响γδT细胞IL-17A及IL-23p19的水平在小鼠CPCR后全脑缺血再灌注损伤的致病机制中发挥促炎作用。

To explore the mechanism ofγδT cell receptor in global cerebral ischemia-reperfusion injury after cardiopulmonary-cerebral resuscitation(CPCR)in mice,adult male C57BL/6 mice were randomly divided into three groups:blank group,CPCR group,and CPCR+γδT receptor antagonist group.The model of global cerebral ischemia-reperfusion injury in mice after CPCR was established by asphyxia and then paraffin sections of mouse brain tissue were made.H-E staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling(TUNEL)were used to observe the injury and apoptosis of cerebral tissue.Western blotting and immunohistochemistry were used to evaluate the protein expression levels of IL-17A and IL-23p19.The mRNA expression levels of IL-17A and IL-23p19 were evaluated by RT-PCR.The results showed that compared to those of the blank group,large numbers of cells in the central area of cerebral ischemia foci in the CPCR group were dead or showed irregular state of enlargement,rupture of nuclear membrane,and disappearance of cell structure.And the apoptosis in the CPCR group was more severe than that in blank group.In contrast,all the above manifestations were less severe in CPCR+γδT receptor antagonist group.Compared to those in the blank group,the brain injury score,apoptosis rate,IL-17A and IL-23p19 expressions were significantly increased in the CPCR group(P<0.01).Compared to the CPCR group,the CPCR+γδT receptor antagonist group had significantly lower brain injury score,apoptosis rate,IL-17A and IL-23p19 expressions(P<0.05).These results suggest that theγδT cell receptor plays a proinflammatory role in the pathogenesis of global cerebral ischemia-reperfusion injury after CPCR in mice by regulating the levels of IL-17A and IL-23p19 inγδT cells.