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  • Küçük Resim
    Öğe
    Differentiation of inflammation-responsive astrocytes from glial progenitors generated from human induced pluripotent stem cells
    (Cell Press, 2017) Santos, Renata; Vadodaria, Krishna C.; Jaeger, Baptiste N.; Mei, Arianna; Lefcochilos-Fogelquist, Sabrina; Mendes, Ana P. D.; Erikson, Galina; Shokhirev, Maxim; Randolph-Moore, Lynne; Fredlender, Callie; Dave, Sonia; Oefner, Ruth; Fitzpatrick, Conor; Pena, Monique; Barron, Jerika J.; Ku, Manching; Denli, Ahmet M.; Kerman, Bilal Ersen; Charnay, Patrick; Kelsoe, John R.; Marchetto, Maria C.; Gage, Fred H.
    Astrocyte dysfunction and neuroinflammation are detrimental features in multiple pathologies of the CNS. Therefore, the development of methods that produce functional human astrocytes represents an advance in the study of neurological diseases. Here we report an efficient method for inflammation-responsive astrocyte generation from induced pluripotent stem cells (iPSCs) and embryonic stem cells. This protocol uses an intermediate glial progenitor stage and generates functional astrocytes that show levels of glutamate uptake and calcium activation comparable with those observed in human primary astrocytes. Stimulation of stem cell-derived astrocytes with interleukin-1 beta or tumor necrosis factor a elicits a strong and rapid pro-inflammatory response. RNA-sequencing transcriptome profiling confirmed that similar gene expression changes occurred in iPSC-derived and primary astrocytes upon stimulation with interleukin-1 beta. This protocol represents an important tool for modeling in-a-dish neurological diseases with an inflammatory component, allowing for the investigation of the role of diseased astrocytes in neuronal degeneration.
  • Küçük Resim
    Öğe
    In vitro myelin formation using embryonic stem cells
    (Company of Biologists, 2015) Kerman, Bilal Ersen; Kim, Hyung-joon; Padmanabhan, Krishnan; Mei, Arianna; Georges, Shereen; Joens, Matthew; Fitzpatrick, James; Jappelli, Roberto; Chandross, Karen; August, Paul; Gage, Fred
    Myelination in the central nervous system is the process by which oligodendrocytes form myelin sheaths around the axons of neurons. Myelination enables neurons to transmit information more quickly and more efficiently and allows for more complex brain functions; yet, remarkably, the underlying mechanism by which myelination occurs is still not fully understood. A reliable in vitro assay is essential to dissect oligodendrocyte and myelin biology. Hence, we developed a protocol to generate myelinating oligodendrocytes from mouse embryonic stem cells and established a myelin formation assay with embryonic stem cell-derived neurons in microfluidic devices. Myelin formation was quantified using a custom semi-automated method that is suitable for larger scale analysis. Finally, early myelination was followed in real time over several days and the results have led us to propose a new model for myelin formation.