Human ex vivo spinal cord slice culture as a useful model of neural development, lesion, and allogeneic neural cell therapy

Abstract

Background: There are multiple promising treatment strategies for central nervous system trauma and disease. However, to develop clinically potent and safe treatments, models of human-specific conditions are needed to complement in vitro and in vivo animal model-based studies. Methods: We established human brain stem and spinal cord (cross-and longitudinal sections) organotypic cultures (hOCs) from first trimester tissues after informed consent by donor and ethical approval by the Regional Human Ethics Committee, Stockholm (lately referred to as Swedish Ethical Review Authority), and The National Board of Health and Welfare, Sweden. We evaluated the stability of hOCs with a semi-quantitative hOC score, immunohistochemistry, flow cytometry, Ca 2+ signaling, and electrophysiological analysis. We also applied experimental allogeneic human neural cell therapy after injury in the ex vivo spinal cord slices. Results: The spinal cord hOCs presented relatively stable features during 7-21 days in vitro (DIV) (except a slightly increased cell proliferation and activated glial response). After contusion injury performed at 7 DIV, a significant reduction of the hOC score, increase of the activated caspase-3 + cell population, and activated microglial populations at 14 days postinjury compared to sham controls were observed. Such elevation in the activated caspase-3 + population and activated microglial population was not observed after allogeneic human neural cell therapy. Conclusions: We conclude that human spinal cord slice cultures have potential for future structural and functional studies of human spinal cord development, injury, and treatment strategies.

Impact of monomeric and aggregated wild‐type and A30P/A53T double‐mutant α‐synuclein on antioxidant mechanism and glutamate metabolic profile of cultured astrocytes

“…These slices were incubated with the respective peptides for 24 hr at 37°C with 5% CO 2 (Loria et al, 2017;Trudler et al, 2021;Xu et al, 2013). After fixing with 2% PFA, slices were minced and then subjected to enzymatic digestion using accutase for tissue dissociation into single cell-suspension and processed for immunostaining and measurement using flow cytometry (Delbridge et al, 2020;Li et al, 2016;Lin et al, 2020;Xu et al, 2015). As per the manufacturer's instruction of primary antibodies (Abcam), for intracellular antigen staining, cells were permeabilized with 0.5% Tween 20 in PBS and nonspecific antigens were blocked using 3% BSA.…”