脑出血免疫抑制大鼠模型的构建及评估

Immunosuppressed rat model of cerebral hemorrhage: construction and assessment

背景:研究表明,大鼠脑出血后经治疗可以有效抑制其免疫功能。已有文献研究对缺血性脑卒中后免疫抑制现象进行详细研究,但脑出血后机体免疫功能研究鲜见报道。目的:构建脑出血后免疫抑制大鼠模型并进行稳定性评估。方法:将60只Wistar大鼠随机分为3组,即对照组、假手术组、脑出血组,每组20只。采用自身动脉血50μL注入大鼠基底节方法建造急性脑出血模型;假手术模型注入50μL生理盐水,手术过程同脑出血模型;对照组不做任何处理。分别于造模后24,48,72,96 h用血液分析仪检测白细胞、淋巴细胞、淋巴细胞百分数;采用ELISA测定大鼠血清中促炎细胞因子白细胞介素6和抗炎细胞因子转化生长因子β的水平;剖取大鼠脾组织进行组织学及病理学检查;通过RT-PCR及Western blotting检测转化生长因子β、白细胞介素6基因和蛋白在脾组织中的表达变化。结果与结论:(1)与假手术组及对照组相比,脑出血组大鼠于造模后24,48,72,96 h白细胞数明显升高,淋巴细胞百分数逐渐降低,差异有显著性意义(P<0.05);(2)与假手术组及对照组相比,脑出血组大鼠血液、脾的白细胞介素6在24 h时高表达,72 h达高峰,96 h时表达降低,差异有显著性意义(P<0.05);(3)与假手术组及对照组相比,脑出血组大鼠血液、脾的转化生长因子β于24 h时低表达,72 h逐渐升高,96 h时高表达,差异有显著性意义(P<0.05);(4)结果提示,大鼠在脑出血后先呈现免疫功能亢奋,随即96 h后出现免疫功能抑制,说明脑出血后免疫抑制大鼠模型造模成功,并具有良好的稳定性。

BACKGROUND： Treatment after intracerebral hemorrhage can effectively suppress immune function. The immune suppression after ischemic stroke has been studied in detail. OBJECTIVE： To construct an immunosuppressed rat model after cerebral hemorrhage, and assess its stability. METHODS： Sixty Wistar rats were randomly divided into control group, sham group, and cerebral hemorrhage group, with 20 rats in each group. Rat models of acute cerebral hemorrhage were established by 50 μL arterial blood injection in the rat basal ganglia. Rats in the sham group were injected with 50 μL of saline, and the operation was identical to cerebral hemorrhage model. Rats in the control group received no treatment. At 24, 48, 72 and 96 hours after model establishment, leukocytes, lymphocytes, and lymphocyte percentage were analyzed by blood analyzer. Enzyme linked immunosorbent assay was used to determine the levels of serum proinflammatory cytokine interleukin-6 and anti-inflammatory cytokine transforming growth factor β in rats. Dissected rat spleen tissue was subjected to histological and histopathological detection. RT-PCR and western blotting were utilized to measure changes in transforming growth factor β, interleukin-6 gene and protein expression in the spleen. RESULTS AND CONCLUSION：（1） Compared with the sham group and control group, leukocyte number was significantly higher, but lymphocyte percentage gradually reduced in the cerebral hemorrhage group at 24, 48, 72 and 96 hours（P〈0.05）.（2） Compared with the sham group and control group, interleukin-6 levels in the blood and spleen were higher at 24 hours, peaked at 72 hours, and decreased at 96 hours in the cerebral hemorrhage group（P〈0.05）.（3） Compared with the sham group and control group, transforming growth factor β expression was lower at 24 hours, gradually increased at 72 hours, and higher at 96 hours in the rat blood and spleen of the cerebral hemorrhage group（P〈0.05）.（4） These findings indicate that immune function excitement first appeared after cerebral hemorrhage, and immune suppression appeared at 96 hours, indicating successful model establishment and good stability.