Principles of Virology, Volume 1. Jane Flint

Principles of Virology, Volume 1 - Jane Flint


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which bipolar neurons migrate. ZIKV infection perturbed the RGP scaffold compared with control slices.

      Glial fibers are visible as parallel tracks in the mouse embryonic brain slice cultures stained with an antibody to vimentin, a protein component of the fibers (image, left panel). When embryonic brain slice cultures were infected with Zika virus, the structure of the glial tracks was altered. Instead of parallel tracks, the fibers assumed a twisted morphology that would not allow neurons to travel from the ventricular zone to the developing neocortex (image, right panel). Disruption of glial fibers was observed after infection with Zika viruses isolated from 1947 to 2016.

      These results suggest that Zika virus-mediated disruption of glial fibers during embryonic development contributes to microcephaly: if neurons cannot migrate to the pial surface, the neocortex will be thinner.

       Rosenfeld AB, Doobin DJ, Warren AL, Racaniello VR, Vallee RB. 2017. Replication of early and recent Zika virus isolates throughout mouse brain development. Proc Natl Acad Sci U S A 114:12273–12278.

      Monolayer and suspension cell cultures do not reproduce the cell type diversity and architecture typical of tissues and organs. One way to overcome this limitation is by the use of organotypic slice cultures, which can be produced from a variety of organs, including brain, liver, and kidney. These cultures are prepared by slicing embryonic or postnatal rodent organs into 100- to 400-micrometer slices. They are placed on substrates, such as porous or semiporous membranes, and bathed in cell culture medium. Such cultures remain viable for 1 to 2 weeks. The effect of Zika virus infection on neuronal migration has been examined in organotypic brain slice cultures derived from embryonic mice (Box 2.3).

      The differentiation of stem cells into organoids depends on growth conditions and nutrients. For example, one type of brain organoid can be established from human pluripotent stem cells by embedding the cells in a gelatinous protein mixture that resembles the extracellular environment of many tissues. In the absence of further cues, the stem cells differentiate into structures typical of many diverse brain regions, including the cortex. In contrast, the production of intestinal organoids requires agonists of a particular signal transduction pathway. Current attempts to improve organoid cultures include the addition of immune cells, vasculature, and commensal microorganisms, to more accurately reflect the details of tissue and organ architectures.

       Evidence of Viral Reproduction in Cultured Cells

      Before quantitative methods for measuring viruses were developed, evidence of viral propagation was obtained by visual inspection of infected cells. Some viruses kill the cells in which they reproduce, and they may eventually detach from the cell culture plate. As more cells are infected, the changes become visible and are called cytopathic effects.

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      TERMINOLOGY

      In vitro and in vivo

      The terms “in vitro” and “in vivo” are common in the virology literature. In vitro means “in glass” and refers to experiments carried out in an artificial environment, such as a glass or plastic test tube. Unfortunately, the phrase “experiments performed in vitro” is used to designate not only work done in the cell-free environment of a test tube but also work done within cultured cells. The use of the phrase in vitro to describe living cultured cells leads to confusion and is inappropriate. In vivo means “in a living organism” but may be used to refer to either cells or animals. Those who work on plants avoid this confusion by using the term “in planta.”

      In this textbook, we use in vitro to designate experiments carried out in the absence of cells, e.g., in vitro translation. Work done in cells in culture is done ex vivo, while research done in animals is carried out in vivo.

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