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. 2021 Feb 2;12:753. doi: 10.1038/s41467-021-21029-2

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© The Author(s) 2021

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 6 — a (i) Schematic of cholesterol modified miR302 (chol-miRNA 302 b/c) delivery to scarred cardiac spheroids for 0, 0–2, 0–4, and 0–7 days. (ii) Contraction amplitude (a.u) and (iii) peak-to-peak time (ms) within scarred spheroids measured after 2, 4, and 7 days for each treatment period (n = 4, 6, 5, 5, 7, 5, 5, 5, 5 biologically independent samples (from left to right), mean ± s.d, one-way ANOVA, (ii) day 4: 0 vs. 0–4 days treatment p = 0.014, (ii) day 7: 0 vs. 0–7 days treatment p = 0.0045, (iii) day 4: 0 vs. 0–4 days treatment p = 1.6 × 10⁻⁷, (iii) day 7: 0 vs. 0–7 days treatment p = 4.1 × 10⁻⁹). b (i) Immunofluorescence staining for cTnT (red; iPSC-CMs), vimentin (green; cardiac fibroblasts), and EdU (proliferation marker) in scarred spheroids at day 7 for each treatment condition. Top panel scalebar 50 µm, and bottom panel scalebar 40 µm. (ii) Quantification of cardiomyocyte proliferation (EdU⁺ and cTnT⁺) and (iii) fibroblast proliferation (EdU⁺ and Vimentin⁺) at day 7 for each treatment condition. (n = 3, 4, 4, 4 biologically independent samples (from left to right), mean ± s.d, one-way ANOVA, 0 vs. 0–4 days treatment p = 0.044, 0 vs. 0–7 days treatment p = 0.026). Scalebar 50 µm. c (i) Experimental outline where scarred microtissues are bioprinted in the support hydrogel as previously described, followed by 4 days treatment with miR302, and analysis compared to non-treated controls. (ii) Calcium mapping in scarred cardiac microtissues at 9 days (5 days culture in support hydrogel; (+) and (−) 4 days miRNA treatment) each frame 40 ms. Scalebar 100 µm. (iii) Representative calcium traces from regions 1 and 2 in treated and non-treated scarred cardiac microtissues. Scalebars 0.5 ΔF/F_o (y), 500 ms (x). (iv) Activation time maps across scarred regions for treated and non-treated scarred cardiac microtissues, and quantification of activation delay (ms) (n = 5 biologically independent samples, mean ± s.d, two-sided student t test, p = 0.019). d Immunofluorescence staining for cTnT (iPSC-CMs), vimentin (CFs), and EdU (proliferation) in scarred cardiac microtissues at day 9 for treated and non-treated scarred cardiac microtissues. Scalebar 50 µm. All experiments from a single iPSC-CM donor (donor B). (n.s. not significant, *p < 0.05, **p < 0.01, **** p < 0.0001).