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. 2020 Oct 5;10:16459. doi: 10.1038/s41598-020-73177-y

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Nanostring mRNA analysis reveals synaptic changes in response to hyaluronan manipulations. (A) Experimental timeline for HA manipulation. Human IPSCs (Day 0) are used to make cortical spheroids (Day 90). Middle image shows intermediate stage embryoid bodies around day 9. 90-day-old spheroids are treated with purified HA or streptomyces hyaluronidase to digest HA. After 24 h of treatment, spheroids are harvested for subsequent analysis. (B) Average pathway scores were plotted for + HA (hyaluronan addition) and − HA (hyaluronidase treatment), revealing three pathways that were preferentially increased by hyaluronidase treatment: Transcription and Splicing (purple), Growth Factor Signaling (green), and Neuronal Cytoskeleton (pink). (C) Volcano plot of the differential expression of all genes analyzed. − HA differential expression is plotted against + HA differential expression, such that mRNAs to the right of 0.0 on the x-axis are upregulated in response to hyaluronidase treatment. The top four differentially-expressed mRNAs (as indicated by blue dots above the p < 0.01 line) are all synapse-associated proteins. (D) Representative image of a synapse showing the pre- or post-synaptic protein localization corresponding to the differentially-expressed mRNAs.