脑动静脉畸形Ⅳ型胶原酶信使核糖核酸的原位杂交研究

In situ hybridization of mRNA for type Ⅳ collagenase(MMP-2,MMP-9) and their tissue inhibitors in cerebral arteriovenous malformations

目的 应用原位杂交技术检测脑动静脉畸形 (AVM)及正常对照组大脑皮质动脉标本中相对分子质量为 92 0 0 0的IV型胶原酶 (MMP 9)、72 0 0 0的IV型胶原酶 (MMP 2 )及其它们的组织抑制因子 1和 2 (TIMP 1和TIMP 2 )的信使核糖核酸 (mRNA) ,并定位产生它们的细胞 ;用透射电镜观察AVM的超微结构 ,以探讨大脑AVM的发病机制。方法 用地高辛标记的MMP 2、MMP 9、TIMP 1和TIMP 2反义RNA(antisenseRNA)探针与AVM和对照组 (各 5例 )皮质动脉组织进行原位杂交 ,定位产生它们的细胞 ,并用正义RNA (senseRNA)探针作阴性对照 ;对杂交组织的相邻切片 ,进行HE染色和Weigert弹力和胶原纤维染色 ,透射电镜观察。结果 5例大脑AVM标本中均见MMP 9mRNA阳性杂交信号。杂交阳性信号位于血管内膜、中膜和外膜 ,且以外膜最为密集。外膜部分断裂、厚薄不一 ,甚至分层。可见平滑肌细胞退变 ,甚至分解成细胞碎片。阳性杂交信号定位于单核细胞、成纤维细胞和平滑肌细胞。对照组动脉标本中未发现MMP 9mRNA阳性杂交信号。在AVM和供血动脉组织中未发现MMP 2、TIMP 1和TIMP 2mRNA的阳性杂交信号。结论 血管内膜的破坏与大脑AVM形成有关 ;外膜也参与了脑AVM的形成过程 ;外膜胶原纤维的破坏与外膜中MMP 9mRNA高含量呈正相关。

Objective To identify the cells responsible for mRNA production within the cerebral AVMs and their feeding arteries by in situ hybridization, and to study the cerebral AVMs by transmission electron microscopy(TEM). Methods In situ hybridization was carried out with digoxigenin RNA probes to localize the cells responsible for MMP 2, MMP 9, TIMP 1and TIMP 2 mRNA production in the histologic sections of the cerebral AVMs ( n =4) and their feeding arteries ( n =7). Results The levels of MMP 9 mRNA were found rather high in all the lamellae of the cerebral AVMs, especially in the adventitia. But none of the MMP 9 mRNA was found in normal cerebral arteries. None of MMP 2, TIMP 1, TIMP 2 mRNAs was found in either the cerebral AVMs or the normal cerebral arteries. In situ hybridization studies colocalized MMP 9 expression to monocyte, fibroblast, smooth muscle cell in the AVM lesions. Weigert studies showed the partial lesions of internal elastic lamina. TEM disclosed missing of almost all the endothelial cells. Conclusions The missing of the endothelial cells was likely to be the foundation of cerebral AVM formation. The adventitia might also play an important role in the formation of cerebral AVM formation. The destruction of collagen in the adventitia might be due to the high level of MMP 9 mRNA.