摘要
目的通过建立实验性大鼠蛛网膜下腔出血(subarachnoidhemorrhage,SAH)模型,观察SAH后柔脑膜纤维化情况;并检测柔脑膜中结缔组织生长因子(connective tissue growth factor,CTGF)的表达,探讨CTGF在SAn后柔脑膜纤维化过程中的作用。
方法使用立体定位仪在显微操作下行枕大池两次注射自体血建立大鼠SAH模型。两次注血后第21天,4%多聚甲醛灌注后取脑,异戊烷—液氮快速冰冻脑组织并—80℃保存;冰冻切片后行Masson染色及免疫组织化学染色分别检测柔脑膜纤维及CTGF的表达。应用多媒体彩色图文分析系统对柔脑膜的胶原纤维进行定量分析。
结果1、大鼠SAH动物模型实验成功率为78.95%,且脑膜结构保存良好。2、MASSON染色显示:SAH组柔脑膜厚度及染色深度明显高于生理盐水组(p＜0.01);生理盐水组与空白组无显著差异(p＞0.01)。3、CTGF免疫组织化学染色显示:在柔脑膜的蛛网膜细胞胞浆及硬脑膜的纤维细胞胞浆中,SAH组的染色深度较生理盐水组明显增强(P＜0.01);生理盐水组与空白组比较无显著差异(P＞0.01)。
结论1.枕大池两次注射自体血,冰冻切片后行Masson染色是研究大鼠SAH后柔脑膜纤维化较好的可行的实验方法;2.大鼠SAH后柔脑膜纤维化明显;3.CTGF在大鼠SAH模型中表达增加,提示CTGF在柔脑膜纤维化过程中可能起重要作用。
Objective: Observe leptomeningeal fibrosis and the expression of connective tissue growth factor(CTGF) after subarachnoid hemorrhage(SAH) via animal models of SAH in rats.
Methods: Rats were used to construct animal models of SAH by twice injecting autoblood solution into the cisterna magna with the aid of a surgical microscope on the stereotaxic frame. 21 days later, the animals were anesthetized and poured with 4% Paraform. Thereafter, their brains were removed and frozen quickly by isopentane- liquid nitrogen, and then preserved in 80℃below zero. Using multimedia chromatic colour analytical system, we did quantitative analysis towards the collagen fiber of leptomeningeal, following the frozen section, Masson and immunohistochemistry staining. The Leptomeninges fibrosis and the expression of CTGF on arachnoid cells were tested by immunohistochemistry.
Results: 1. The achievement ratio of the animal models was 78.95%; moreover, leptomeninges structure was preserved well. 2. Displayed by the Masson staining, the film thickness and the staining depth of leptomeninges in experimental group increased predominantly, compared with normal sodium group(p<0.01) . There was no significant difference between normal sodium group and the blank group (p>0.01).3. The result of the test immunohistochemistry of CTGF showed that the staining depth of leptomeningeal arachnoid endochylema and duramatral fibrocyte endochylema in experimental group was increased significantly , compared with normal sodium group(p<0.01) , while no significant difference between normal sodium group and blank group was observed (p>0.01) .
Conclusion: 1. It was a feasible method for studying leptomeninges fibrosis after SAH to inject twice autoblood into the cisterna magna of rats, followed by the frozen section and Masson staining. 2. Leptomeninges fibrosis was significant after SAH. 3. The expression of CTGF on arachnoid cells after SAH was increased significantly , which means CTGF may play an important role in the process of leptomeningeal fibrosis after SAH in rats.
引文
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