iPSC Platform

Our knowledge on virus-associated alterations during human embryonal development is limited. Human induced pluripotent stem cells (iPSCs) are a highly suitable cell culture model not only for human embryonal development, but also for the analysis of congenital virus infections. As a pluripotent stem cell type iPSCs are similar to human embryonic stem cells (ESCs), which are derived from the inner cell mass of human blastocysts. Both cell types can be differentiated into the three embryonic germ layers, ectoderm, mesoderm, and endoderm. Thus, they can recapitulate blastocyst- and early gastrulation-like stages and as such the viral impact on early human development (Figure 1). The combination of developmental biology with congenital virology opens up new and exciting perspectives (Claus et al., 2020).
Figure 1. Overview of induced pluripotent stem cells and iPSC-derived ectodermal, mesodermal, and endodermal cells as a cell culture model for early human development.
Rubella virus can infect iPSCs and be maintained in culture for several passages (Hübner et al., 2017). In contrast to recombinant coxsackievirus B3 expressing green fluorescent protein (EGFP) and measles virus (Edmonston strain) RV does not display any notable cytopathogenicity on iPSCs and as such enables their differentiation into derivatives of the three embryonic germ layers, ectoderm, mesoderm, and endoderm (Figure 2).
Figure 2. The pattern of infection of rubella virus (RV) on iPSCs as compared to recombinant coxsackievirus B3 expressing green fluorescent protein (CVB3-EGFP) and measles virus (MV, Edmonston strain). Infection by CVB3-EGFP, MV, and RV is shown through expression of EGFP and the viral proteins E1 envelope glycoprotein and phosphoprotein (P), respectively. The course of infection was assessed with regard to alterations of cellular metabolism, induction of cytopathogenic effects and the effect on differentiation capacity.
In a follow-up study we addressed whether alterations of developmental pathways during RV infection of iPSCs can be correlated to the clinical symptoms of congenital rubella syndrome (CRS). Figure 3 illustratively summarizes our findings on rubella virus-associated alterations of the differentiation capacity of iPSCs (Bilz et al., 2020).
Figure 3. Summary of rubella virus-associated alterations during undirected and directed differentiation of iPSCs. Adopted from “Teratogenic rubella virus alters the endodermal differentiation capacity of human induced pluripotent stem cells” by Bilz NC, Willscher E.,…, Claus C. 2019, Cells, 8(8). pii: E870. doi: 10.3390/cells8080870.
A comparable transcriptomic profile of rubella virus-infected and uninfected iPSCs after initiation of undirected differentiation indicates that RV infection does not redirect differentiation of iPSC s into a distinct developmental pathway. This was followed by the examination of its impact on directed differentiation.
The ectoderm is involved in eye development, which is altered during congenital rubella virus infection. During ectodermal differentiation of rubella virus-infected cells eye field transcription factors such as SIX homeobox 3 (SIX3) and retina and anterior neural fold homeobox (RAX) are reduced in comparison to the uninfected population. During directed differentiation of RV-infected iPSCs the endoderm appeared to be the most affected cell population. The increased expression of markers for definitive endoderm such as Eomesodermin (EOMES) and Cerberus (CER) could affect its crosstalk with the ectoderm and especially the mesoderm during embryonal development. The mesoderm is involved in cardiac development, which is affected during congenital rubella virus infection. For more details please refer to our publication (Bilz et al., 2019).

Bilz NC*, Willscher E*, Binder H, Böhnke J, Stanifer ML, Hübner D, Boulant S, Liebert UG, Claus C. Teratogenic rubella virus alters the endodermal differentiation capacity of human induced pluripotent stem cells. Cells. 2019; 8(8). pii: E870. doi: 10.3390/cells8080870.

Claus C, Jung M, Hübschen JM. Pluripotent stem cell-based models: a peephole into virus infections during early pregnancy. Cells. 2020, 9, 542. doi: 10.3390/cells9030542.

Hübner D, Jahn K, Pinkert S, Böhnke J, Jung M, Fechner H, Rujescu D, Liebert UG, Claus C. Infection of iPSC Lines with Miscarriage-Associated Coxsackievirus and Measles Virus and Teratogenic Rubella Virus as a Model for Viral Impairment of Early Human Embryogenesis. ACS Infect Dis. 2017 Dec 8;3(12):886-897. doi: 10.1021/acsinfecdis.7b00103. Epub 2017 Oct 26.