Some children with proven intrauterine Zika virus (ZIKV) infection who were born asymptomatic subsequently manifested neurodevelopmental delays, pointing to impairment of development perinatally and postnatally. To model this, we infected postnatal day (P) 5-6 (equivalent to the perinatal period in humans) susceptible mice with a mammalian cell-propagated ZIKV clinical isolate from the Brazilian outbreak in 2015. All infected mice appeared normal up to 4 days post-intraperitoneal inoculation (dpi), but rapidly developed severe clinical signs at 5-6 dpi. All nervous tissue examined at 5/6 dpi appeared grossly normal. However, anti-ZIKV positive cells were observed in the optic nerve, brain, and spinal cord; predominantly in white matter. Co-labeling with cell type specific markers demonstrated oligodendrocytes and astrocytes support productive infection. Rarely, ZIKV positive neurons were observed. In spinal cord white matter, which we examined in detail, apoptotic cells were evident; the density of oligodendrocytes was significantly reduced; and there was localized microglial reactivity including expression of the NLRP3 inflammasome. Together, our observations demonstrate that a clinically relevant ZIKV isolate can directly impact oligodendrocytes. As primary oligodendrocyte cell death can lead later to secondary autoimmune demyelination, our observations may help explain neurodevelopmental delays in infants appearing asymptomatic at birth and commend lifetime surveillance.

Oligodendrocytes are susceptible to Zika virus infection in a mouse model of perinatal exposure: Implications for CNS complications.

A century ago this year, Pio del Rio-Hortega (1921) coined the term ‘oligodendroglia’ for the ‘interfascicular glia’ with very few processes, launching an extensive discovery effort on his new cell type. One hundred years later, we review his original contributions to our understanding of the system of cytoplasmic channels within myelin in the context of what we observe today using light and electron microscopy of genetically encoded fluorescent reporters and immunostaining. We use the term myelinic channel system to describe the cytoplasm-delimited spaces associated with myelin; being the paranodal loops, inner and outer tongues, cytoplasm-filled spaces through compact myelin and further complex motifs associated to the sheath. Using a central nervous system myelinating cell culture model that contains all major neural cell types and produces compact myelin, we find that td-tomato fluorescent protein delineates the myelinic channel system in a manner reminiscent of the drawings of adult white matter by Rio-Hortega, despite that he questioned whether some cytoplasmic figures he observed represented artefact. Together, these data lead us to propose a slightly revised model of the ‘unrolled’ sheath. Further, we show that the myelinic channel system, while relatively stable, can undergo subtle dynamic shape changes over days. Importantly, we capture an under-appreciated complexity of the myelinic channel system in mature myelin sheaths.

Río-Hortega's drawings revisited with fluorescent protein defines a cytoplasm-filled channel system of CNS myelin.

The link between gliomas infiltration and white matter architecture investigated with electron microscopy and diffusion tensor imaging.

Demyelinating disorders of the central nervous system (CNS) occur when myelin and oligodendrocytes are damaged or lost. Remyelination and regeneration of oligodendrocytes can be achieved from endogenous oligodendrocyte precursor cells (OPCs) that reside in the adult CNS tissue. Using a cuprizone mouse model of demyelination, we show that infusion of fractalkine (CX3CL1) into the demyelinated murine brain increases de novo oligodendrocyte formation and enhances remyelination in the corpus callosum and cortical gray matter. This is achieved by increased OPC proliferation in the cortical gray matter as well as OPC differentiation and attenuation of microglia/macrophage activation both in corpus callosum and cortical gray matter. Finally, we show that activated OPCs and microglia/macrophages express fractalkine receptor CX3CR1 in vivo, and that in OPC-microglia co-cultures fractalkine increases in vitro oligodendrocyte differentiation by modulating both OPC and microglia biology. Our results demonstrate a novel pro-regenerative role of fractalkine in a demyelinating mouse model. 

http://www.cell.com/article/S2213671122005902/pdf

Fractalkine enhances oligodendrocyte regeneration and remyelination in a demyelination mouse model

Culture Protocol and Transcriptomic Analysis of Murine SVZ NPCs and OPCs

Transmission electron microscopy of central nervous system white matter has provided unparalleled access to the ultrastructural features of axons, their myelin sheaths, and the major cells of white matter; namely, oligodendrocytes, oligodendrocyte precursors, astrocytes, and microglia. In particular, it has been invaluable in elucidating pathological changes in axons and myelin following experimentally induced injury or genetic alteration, in animal models. While also of value in the examination of human white matter, the tissue is rarely fixed adequately for the types of detailed analyses that can be performed on well-preserved samples from animal models, perfusion fixed at the time of death. In this chapter we describe methods for obtaining, processing, and visualizing white matter samples using transmission electron microscopy of perfusion fixed tissue and for unbiased morphometry of white matter, with particular emphasis on axon and myelin pathology. Several advanced electron microscopy techniques are now available, but this method remains the most expedient and accessible for routine ultrastructural examination and morphometry.

Transmission Electron Microscopy and Morphometry of the CNS White Matter.

Generation of Rat Neural Stem Cells to Produce Different Astrocyte Phenotypes.

There is an urgent need for therapies that target the multicellular pathology of central nervous system (CNS) disease. Modified, nonanticoagulant heparins mimic the heparan sulfate glycan family and are known regulators of multiple cellular processes. In vitro studies have demonstrated that low sulfated modified heparin mimetics (LS‐mHeps) drive repair after CNS demyelination. Herein, we test LS‐mHep7 (an in vitro lead compound) in experimental autoimmune encephalomyelitis (EAE) and cuprizone‐induced demyelination. In EAE, LS‐mHep7 treatment resulted in faster recovery and rapidly reduced inflammation which was accompanied by restoration of animal weight. LS‐mHep7 treatment had no effect on remyelination or on OLIG2 positive oligodendrocyte numbers within the corpus callosum in the cuprizone model. Further in vitro investigation confirmed that LS‐mHep7 likely mediates its pro‐repair effect in the EAE model by sequestering inflammatory cytokines, such as CCL5 which are upregulated during immune‐mediated inflammatory attacks. These data support the future clinical translation of this next generation modified heparin as a treatment for CNS diseases with active immune system involvement.

https://onlinelibrary.wiley.com/doi/pdfdirect/10.1002/glia.24363

Rebecca Sherrard Smith

Papers

Transmission Electron Microscopy and Morphometry of the CNS White Matter.

Generation of Rat Neural Stem Cells to Produce Different Astrocyte Phenotypes.

Low sulfated heparan sulfate mimetic differentially affects repair in immune‐mediated and toxin‐induced experimental models of demyelination