Gressens P, Langston C, Martin J R
Laboratory of Experimental Neuropathology, National Institute of Neurological Diseases and Stroke, NIH, Bethesda, Maryland 20892.
J Neuropathol Exp Neurol. 1994 Sep;53(5):469-82. doi: 10.1097/00005072-199409000-00006.
To more precisely define the role of herpes simplex virus (HSV) in development of nervous system disease in neonates with disseminated infection, an in situ polymerase chain reaction (ISPCR) method was used to detect and localize HSV DNA sequences in paraffin sections of neural and non-neural autopsy tissues. In subregions of adjacent sections corresponding to ISPCR-labeled and unlabeled areas, HSV specificity was verified using solution PCR and Southern blots. In serial sections, ISPCR results were compared to lesions, HSV antigen and, in selected samples, to viral sequence detection by in situ hybridization. By ISPCR, HSV-specific labeling was limited to HSV-infected neonates and experimentally infected mouse controls. ISPCR-labeled cells corresponded to regions that were histologically abnormal and contained HSV antigen or in situ hybridization signal in some foci; in others, labeled cells were in areas with no evident lesions or antigen. Results suggest two routes of HSV spread to the CNS: (i) blood-borne infection, with HSV DNA in splenic lymphocytes, circulating cells, meningeal vessel walls and cells in intraventricular hemorrhage, and (ii) neural spread, with HSV detected in brain stem sensory neurons. In the brain of one neonate surviving acute infection, detection of HSV nucleic acid sequences suggests a latent or persistent viral genome. With other methods, this highly sensitive ISPCR technique permits a more complete definition of HSV infection in these infants and provides new insights into disease mechanisms. Fuller understanding of HSV persistence and recurrent neurological disease in survivors will require further studies using these and other techniques in human tissues and in animal models.
为了更精确地确定单纯疱疹病毒（HSV）在患有播散性感染的新生儿神经系统疾病发展中的作用，采用原位聚合酶链反应（ISPCR）方法检测并定位神经和非神经尸检组织石蜡切片中的HSV DNA序列。在与ISPCR标记和未标记区域相对应的相邻切片的亚区域中，使用溶液PCR和Southern印迹法验证HSV特异性。在连续切片中，将ISPCR结果与病变、HSV抗原进行比较，并在选定样本中与原位杂交检测的病毒序列进行比较。通过ISPCR，HSV特异性标记仅限于HSV感染的新生儿和实验感染的小鼠对照。ISPCR标记的细胞对应于组织学上异常且在某些病灶中含有HSV抗原或原位杂交信号的区域；在其他区域，标记的细胞位于无明显病变或抗原的区域。结果提示HSV传播至中枢神经系统有两条途径：（i）血行感染，脾脏淋巴细胞、循环细胞、脑膜血管壁和脑室内出血中的细胞中有HSV DNA；（ii）神经传播，在脑干感觉神经元中检测到HSV。在一名急性感染存活的新生儿大脑中，HSV核酸序列的检测提示存在潜伏或持续的病毒基因组。使用其他方法，这种高度敏感的ISPCR技术能够更完整地定义这些婴儿中的HSV感染，并为疾病机制提供新的见解。要更全面地了解幸存者中HSV的持续存在和复发性神经系统疾病，需要在人体组织和动物模型中使用这些及其他技术进行进一步研究。
