Mouse Embryonic Stem Cells Inhibit Murine Cytomegalovirus Infection through a Multi-Step Process

Kawasaki, Hideya; Kosugi, Isao; Arai, Yoshifumi; Iwashita, Toshihide; Tsutsui, Yoshihiro

doi:10.1371/journal.pone.0017492

Cited by 20 publications

(37 citation statements)

References 78 publications

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“…It has been reported that the susceptibility of mouse stem cells to CMV may be determined by the concentration of heparan sulfate, presence of β1-integrin and vimentin, and nuclear pore density (Kawasaki et al 2011). It is possible that one of these factors may be related to the acquisition of susceptibility to HCMV in human cells, including hNS-1 cells, which will be the object of further investigation.…”

Section: Discussionmentioning

confidence: 98%

“…The increased susceptibility to infection after differentiation induction may be a consequence of changes in one or more steps of the infection process, such as attachment, entry, trafficking, nuclear entry, and promoter activity (Kawasaki et al 2011). It has been reported that the susceptibility of mouse stem cells to CMV may be determined by the concentration of heparan sulfate, presence of β1-integrin and vimentin, and nuclear pore density (Kawasaki et al 2011).…”

Section: Discussionmentioning

confidence: 99%

“…It was shown that the concentration of heparan sulfate in the cell surface, the presence of β1-integrin and vimentin, and the nuclear pore density is important for the attachment, penetration, and internalization to the nucleus (Kawasaki et al 2011). In mouse models, it has been observed that embryonic stem cells cannot be infected with CMV and they acquire susceptibility through differentiation (Matsukage et al 2006;Kawasaki et al 2011). However, differentiated neurons are also refractory to the infection and this may suggest that only during a precise period during differentiation, the cells can be infected (Cheeran et al 2005).…”

Section: Introductionmentioning

confidence: 99%

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Effects of cytomegalovirus infection in human neural precursor cells depend on their differentiation state

et al. 2015

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Cytomegalovirus (CMV) is the most common cause of congenital infection in developed countries and a major cause of neurological disability in children. Although CMV can affect multiple organs, the most important sequelae of intrauterine infection are related to lesions of the central nervous system. However, little is known about the pathogenesis and the cellular events responsible for neuronal damage in infants with congenital infection. Some studies have demonstrated that neural precursor cells (NPCs) show the greatest susceptibility to CMV infection in the developing brain. We sought to establish an in vitro model of CMV infection of the developing brain in order to analyze the cellular events associated with invasion by this virus. To this end, we employed two cell lines as a permanent source of NPC, avoiding the continuous use of human fetal tissue, the human SK-N-MC neuroblastoma cell line, and an immortalized cell line of human fetal neural origin, hNS-1. We also investigated the effect of the differentiation stage in relation to the susceptibility of these cell lines by comparing the neuroblastoma cell line with the multipotent cell line hNS-1. We found that the effects of the virus were more severe in the neuroblastoma cell line. Additionally, we induced hNS-1 to differentiate and evaluated the effect of CMV in these differentiated cells. Like SK-N-MC cells, hNS-1-differentiated cells were also susceptible to infection. Viability of differentiated hNS-1 cells decreased after CMV infection in contrast to undifferentiated cells. In addition, differentiated hNS-1 cells showed an extensive cytopathic effect whereas the effect was scarce in undifferentiated cells. We describe some of the effects of CMV in neural stem cells, and our observations suggest that the degree of differentiation is important in the acquisition of susceptibility.

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Section: Discussionmentioning

confidence: 98%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Effects of cytomegalovirus infection in human neural precursor cells depend on their differentiation state

et al. 2015

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“…The fetal brain and auditory system are the main sites of the clinical manifestations of congenital HCMV (cCMV) infection (1)(2)(3)(4), and sensorineural hearing loss is the most common long-term sequela in congenitally infected infants (4)(5)(6). In the fetal brain, the bilateral subventricular zone (SVZ), where neural progenitor/stem cells (NPCs) are a predominant cell type, is a site of virus-induced damage that has been well described in infants with severe congenital HCMV infection (7)(8)(9)(10)(11). NPCs are fully permissive for HCMV infection (12)(13)(14)(15)(16)(17), and the infection has been shown to perturb NPC proliferation and differentiation (18)(19)(20)(21).…”

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confidence: 99%

Human Cytomegalovirus Infection Dysregulates the Localization and Stability of NICD1 and Jag1 in Neural Progenitor Cells

Liu

Yang

et al. 2015

J Virol

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Human cytomegalovirus (HCMV) infection of the developing fetus frequently results in major neural developmental damage. In previous studies, HCMV was shown to downregulate neural progenitor/stem cell (NPC) markers and induce abnormal differentiation. As Notch signaling plays a vital role in the maintenance of stem cell status and is a switch that governs NPC differentiation, the effect of HCMV infection on the Notch signaling pathway in NPCs was investigated. HCMV downregulated mRNA levels of Notch1 and its ligand, Jag1, and reduced protein levels and altered the intracellular localization of Jag1 and the intracellular effector form of Notch1, NICD1. These effects required HCMV gene expression and appeared to be mediated through enhanced proteasomal degradation. Transient expression of the viral tegument proteins of pp71 and UL26 reduced NICD1 and Jag1 protein levels endogenously and exogenously. Given the critical role of Notch signaling in NPC growth and differentiation, these findings reveal important mechanisms by which HCMV disturbs neural cell development in vitro. Similar events in vivo may be associated with HCMV-mediated neuropathogenesis during congenital infection in the fetal brain. IMPORTANCE Congenital human cytomegalovirus (HCMV) infection is the leading cause of birth defects that primarily manifest as neurological disabilities. Neural progenitor cells (NPCs), key players in fetal brain development, are the most susceptible cell type for HCMV infection in the fetal brain. Studies have shown that NPCs are fully permissive for HCMV infection, which causes neural cell loss and premature differentiation, thereby perturbing NPC fate. Elucidation of virus-host interactions that govern NPC proliferation and differentiation is critical to understanding neuropathogenesis. The Notch signaling pathway is critical for maintaining stem cell status and functions as a switch for differentiation of NPCs. Our investigation into the impact of HCMV infection on this pathway revealed that HCMV dysregulates Notch signaling by altering expression of the Notch ligand Jag1, Notch1, and its active effector in NPCs. These results suggest a mechanism for the neuropathogenesis induced by HCMV infection that includes altered NPC differentiation and proliferation.

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“…The ability of the virus to infect the developing fetal brain is a key factor in its neuropathogenesis (3)(4)(5)(6)(7)(8). While considerable experimental data were obtained from newborn and embryonic mouse models (6,(9)(10)(11)(12)(13)(14)(15), the strict species specificity precludes animal models of HCMV. Recent studies in human neuronal progenitor cells (NPC) derived from fetal and neonatal brains have revealed productive HCMV infection of NPC, with resultant functional alterations (16)(17)(18)(19)(20).…”

mentioning

confidence: 99%

Transition toward Human Cytomegalovirus Susceptibility in Early Human Embryonic Stem Cell-Derived Neural Precursors

Berger

Gil

Panet

et al. 2015

J Virol

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Congenital human cytomegalovirus (HCMV) infection is associated with neurodevelopmental disabilities. To dissect the earliest events of infection in the developing human brain, we studied HCMV infection during controlled differentiation of human embryonic stem cells (hESC) into neural precursors. We traced a transition from viral restriction in hESC, mediated by a block in viral binding, toward HCMV susceptibility in early hESC-derived neural precursors. We further revealed the role of plateletderived growth factor receptor alpha (PDGFR␣) as a determinant of the developmentally acquired HCMV susceptibility. Human cytomegalovirus (HCMV) is a leading cause of congenital infection (1), associated with neurodevelopmental disabilities (2, 3). The ability of the virus to infect the developing fetal brain is a key factor in its neuropathogenesis (3)(4)(5)(6)(7)(8). While considerable experimental data were obtained from newborn and embryonic mouse models (6, 9-15), the strict species specificity precludes animal models of HCMV. Recent studies in human neuronal progenitor cells (NPC) derived from fetal and neonatal brains have revealed productive HCMV infection of NPC, with resultant functional alterations (16)(17)(18)(19)(20). Notably, NPC do not represent early neural embryonic development but rather a more advanced stage along the neuronal differentiation route. Hence, the earliest events defining the susceptibility of the developing human nervous system to HCMV have remained largely unknown.Human embryonic stem cells (hESC) provide an opportunity to study early human neural development (21-23). We have developed highly reproducible protocols for controlled induction of differentiation of hESC toward a neural lineage, giving rise to enriched populations of proliferating, developmentally multipotent early NPC (24,25).Here, we have employed experimental HCMV infection in a dynamic model of controlled differentiation of hESC into neural precursors, to gain insight into the molecular events mediating HCMV infection during early human neurodevelopment.We first sought to track the earliest stage of neural differentiation at which the cells become susceptible to HCMV. To this end, differentiation of hESC was induced in floating cell clusters toward the formation of neural spheres (Fig. 1A) as detailed previously (24, 25). The emerging neural spheres were infected at sequential time points along the neural differentiation process with the broadly tropic HCMV TB40/E strain expressing green fluorescent protein (GFP) (26,27) (Fig. 1B). In accordance with previous studies (25), immunostaining analysis for the pluripotent stem cell marker TRA 1-81 and the neural differentiation marker PSA-NCAM showed that the majority of the cells underwent early neural differentiation within 11 days (Fig. 1C). Importantly, we have identified a transition toward HCMV susceptibility occurring between 4 and 11 days post-differentiation induction (Fig. 1B). A similar susceptibility pattern was observed for low-passage-number, cell-associated, clinic...

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Mouse Embryonic Stem Cells Inhibit Murine Cytomegalovirus Infection through a Multi-Step Process

Cited by 20 publications

References 78 publications

Effects of cytomegalovirus infection in human neural precursor cells depend on their differentiation state

Effects of cytomegalovirus infection in human neural precursor cells depend on their differentiation state

Human Cytomegalovirus Infection Dysregulates the Localization and Stability of NICD1 and Jag1 in Neural Progenitor Cells

Transition toward Human Cytomegalovirus Susceptibility in Early Human Embryonic Stem Cell-Derived Neural Precursors

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