Simulated Microgravity Impairs Cardiac Autonomic Neurogenesis from Neural Crest Cells

FIG. 8. RCCS compromises NC differentiation into cardiac ANS. (A–F) Confocal immunofluorescence of TH, Nkx2.5, and tdTomato on day 19 iPSCWnt1-Cre EBs, transitioned from RCCS on day 10 (A–C) or continuously grown in SC (D–F). Exposure to RCCS supports differentiation of EBs into tdTomato+/Nkx2.5+ myocardium, which however lacks sympathetic innervation, as indicated by the complete lack of TH+ neurons (A–C). In contrast, SC-differentiated Nkx2.5+ myocardium is innervated with tdTomato+/TH+ sympathetic neurons (D–F). (C, F) Are blownup images of the areas depicted in insets in (A, B, D, E), respectively. (G, H) Confocal immunofluorescence of vesicular acetylcholine transporter (VAChT) and tdTomato on day 19 iPSCWnt1-Cre EBs, transitioned from RCCS on day 10 (G–I) or continuously grown in SC (J–L). Exposure to RCCS results in EB differentiation into VAChT+/tdTomato-negative parasympathetic neurons (G–I). In contrast, parasympathetic neurons in SC-differentiated EBs are derived from NCs, as indicated by colocalization of tdTomato+/VAChT+ (J–L). (I, L) Are higher magnification images of the areas depicted in insets in (G, H, J, K), respectively. Scale bars 100 μm.

Simulated Microgravity Impairs Cardiac Autonomic Neurogenesis from Neural Crest…

Microgravity-induced alterations in the autonomic nervous system (ANS) contribute to derangements in both the mechanical and electrophysiological function of the cardiovascular system, leading to severe symptoms in humans following space travel. Because the ANS forms embryonically from neural crest (NC) progenitors, we hypothesized that microgravity can impair NC-derived cardiac structures. Accordingly, we conducted in vitro simulated microgravity experiments employing NC genetic lineage tracing in mice with cKitCreERT2/+, Isl1nLacZ, and Wnt1-Cre reporter alleles. Inducible fate mapping in adult mouse hearts and pluripotent stem cells (iPSCs) demonstrated reduced cKitCreERT2/+-mediated labeling of both NC-derived cardiomyocytes and autonomic neurons (P < 0.0005 vs. controls). Whole transcriptome analysis, suggested that this effect was associated with repressed cardiac NC- and upregulated mesoderm-related gene expression profiles, coupled with abnormal bone morphogenetic protein (BMP)/transforming growth factor beta (TGF-β) and Wnt/β-catenin signaling. To separate the manifestations of simulated microgravity on NC versus mesodermal-cardiac derivatives, we conducted Isl1nLacZ lineage analyses, which indicated an approximately 3-fold expansion (P < 0.05) in mesoderm-derived Isl-1+ pacemaker sinoatrial nodal cells; and an approximately 3-fold reduction (P < 0.05) in cardiac NC-derived ANS cells, including sympathetic nerves and Isl-1+ cardiac ganglia. Finally, NC-specific fate mapping with a Wnt1-Cre reporter iPSC model of murine NC development confirmed that simulated microgravity directly impacted the in vitro development of cardiac NC progenitors and their contribution to the sympathetic and parasympathetic innervation of the iPSC-derived myocardium. Altogether, these findings reveal an important role for gravity in the development of NCs and their postnatal derivatives, and have important therapeutic implications for human space exploration, providing insights into cellular and molecular mechanisms of microgravity-induced cardiomyopathies/channelopathies.

Hatzistergos KE, Jiang Z, Valasaki K, Takeuchi LM, Balkan W, Atluri P, Saur D, Seidler B, Tsinoremas N, DiFede DL, Hare JM. Simulated Microgravity Impairs Cardiac Autonomic Neurogenesis from Neural Crest Cells. Stem Cells Dev. 2018 Jun 15;27(12):819-830. doi: 10.1089/scd.2017.0265. Epub 2018 Mar 20.