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. 2021 May 19;12:2951. doi: 10.1038/s41467-021-23217-6

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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. 2 — a–b” Fibronectin and collagen immunochemistry. Whole-mount staining and confocal imaging of zebrafish myotomes centred on myotome 10 (1 dpf) and myotome 12 (5 dpf). Images are z projections of the mediolateral extent of the myotome stained for f-actin (red) to mark muscle fibres and DAPI (blue) for nuclei. a Fibronectin (Fn) staining (green) at the vertical myosepta at 1 dpf, images representative of n = 9, three larvae from three separate clutches on different weeks, most severe and weakest phenotypes excluded (scale bar = 30 μm). a’ dag1^−/−, a” fkrp^−/−. b Collagen-1 (Col1a, green) staining of the muscle basement membrane in wild-type sibling fish. b’ dag1^−/− and b” fkrp^−/−, white arrow: absent collagen, scale bar = 40 μm. a–b” Lateral views anterior to the left. c Transmission electron micrographs (TEM) of longitudinal sections of 7 dpf zebrafish myotome centred on the muscle basement membrane (red arrowheads). Red arrows: absent collagen fibrils, red star: fibre detachment from vertical myosepta, scale bar = 0.5 μm. Wild-type siblings, c’, dag1^−/− c”, fkrp^−/−, micrographs representative of (n = 9), three larvae from three separate clutches on different weeks, most severe and weakest phenotypes excluded. d Schematic of fibre-crossing and detachment model at the Myotendinous Junction (MTJ) and disease progression, d’ canonical DGC axis in dag1^−/− and, d” combined detachment crossing in fkrp^−/−. e, f Measurement of max intensity from fibronectin (e) and collagen (f) staining analysed by two-way ANOVA, ***(P = <0.0001), plotted points outside 95% confidence interval, box represents 5–95%, median centre line, whiskers = SEM, Tukey’s multiple comparison analysis, three independent experiments. g Passive force measurement at 6 dpf, calculated by plotting the maximum active force at given loads, as passive tension (mN) against external stretch (Lo%), with representative linear regression analysis of the plotted points of fkrp^+/+ sibling (sib); black line (n = 8), fkrp^−/−, red line (n = 6) and dag1^−/− blue line (n = 5) was used to test the significance of the differences (***P = <0.0001), error bars = SEM.