Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, California, USA.
J Virol. 2014 Apr;88(8):4021-39. doi: 10.1128/JVI.03492-13. Epub 2014 Jan 22.
Congenital human cytomegalovirus (HCMV) infection is a major cause of central nervous system structural anomalies and sensory impairments. It is likely that the stage of fetal development, as well as the state of differentiation of susceptible cells at the time of infection, affects the severity of the disease. We used human embryonic stem (ES) cell-derived primitive prerosette neural stem cells (pNSCs) and neural progenitor cells (NPCs) maintained in chemically defined conditions to study HCMV replication in cells at the early stages of neural development. In contrast to what was observed previously using fetus-derived NPCs, infection of ES cell-derived pNSCs with HCMV was nonprogressive. At a low multiplicity of infection, we observed only a small percentage of cells expressing immediate-early genes (IE) and early genes. IE expression was found to be restricted to cells negative for the anterior marker FORSE-1, and treatment of pNSCs with retinoic acid restored IE expression. Differentiation of pNSCs into NPCs restored IE expression but not the transactivation of early genes. Virions produced in NPCs and pNSCs were exclusively cell associated and were mostly non-neural tropic. Finally, we found that viral genomes could persist in pNSC cultures for up to a month after infection despite the absence of detectable IE expression by immunofluorescence, and infectious virus could be produced upon differentiation of pNSCs to neurons. In conclusion, our results highlight the complex array of hurdles that HCMV must overcome in order to infect primitive neural stem cells and suggest that these cells might act as a reservoir for the virus.
Human cytomegalovirus (HCMV) is a betaherpesvirus that is highly prevalent in the population. HCMV infection is usually asymptomatic but can lead to severe consequences in immunosuppressed individuals. HCMV is also the most important infectious cause of congenital developmental birth defects. Manifestations of fetal HCMV disease range from deafness and learning disabilities to more severe symptoms such as microcephaly. In this study, we have used embryonic stem cells to generate primitive neural stem cells and have used these to model HCMV infection of the fetal central nervous system (CNS) in vitro. Our results reveal that these cells, which are similar to those present in the developing neural tube, do not support viral replication but instead likely constitute a viral reservoir. Future work will define the effect of viral persistence on cellular functions as well as the exogenous signals leading to the reactivation of viral replication in the CNS.
先天性人类巨细胞病毒（HCMV）感染是中枢神经系统结构异常和感觉障碍的主要原因。很可能是胎儿发育阶段以及感染时易感细胞的分化状态影响了疾病的严重程度。我们使用人胚胎干细胞（ES）细胞衍生的原始前体神经干细胞（pNSC）和神经祖细胞（NPC）在化学定义的条件下培养，以研究细胞在神经发育早期阶段的 HCMV 复制。与之前使用胎儿衍生 NPC 观察到的情况相反，HCMV 感染 ES 细胞衍生的 pNSC 是非进行性的。在低感染复数时，我们仅观察到一小部分细胞表达即刻早期基因（IE）和早期基因。发现 IE 表达仅限于 FORSE-1 前体标记阴性的细胞，并且用视黄酸处理 pNSC 可以恢复 IE 表达。pNSC 分化为 NPC 可恢复 IE 表达，但不能激活早期基因。在 NPC 和 pNSC 中产生的病毒粒子仅与细胞相关，并且主要是非神经趋向性的。最后，我们发现尽管免疫荧光检测不到可检测的 IE 表达，但病毒基因组仍可在感染后 pNSC 培养物中持续存在长达一个月，并且当 pNSC 分化为神经元时可产生感染性病毒。总之，我们的结果突出了 HCMV 必须克服的一系列复杂障碍，以便感染原始神经干细胞，并表明这些细胞可能成为病毒的储库。
人巨细胞病毒（HCMV）是一种β疱疹病毒，在人群中高度流行。HCMV 感染通常无症状，但可导致免疫抑制个体的严重后果。HCMV 也是先天性发育性出生缺陷的最重要传染性原因。胎儿 HCMV 疾病的表现范围从耳聋和学习障碍到更严重的症状，如小头畸形。在这项研究中，我们使用胚胎干细胞生成原始神经干细胞，并使用这些细胞在体外模拟胎儿中枢神经系统（CNS）的 HCMV 感染。我们的结果表明，这些细胞类似于发育中的神经管中的细胞，不支持病毒复制，但可能构成病毒库。未来的工作将确定病毒持续存在对细胞功能的影响，以及导致 CNS 中病毒复制重新激活的外源性信号。
