Journal: bioRxiv
Article Title: Satellite stem cell dysfunction in Duchenne muscular dystrophy
doi: 10.1101/2024.07.24.604963
Figure Lengend Snippet: DMD satellite cells have impaired autophagy and senescence. A) Feature plot showing cells positive for the senescence GO term, showing higher amounts in the regions where cycling progenitors and DMD enriched cells cluster. B-D) ddPCR quantification of gene expression in satellite cells in B) senescence-associated genes Cdkn1a and Cdkn2a from non-injured (NI), one day post injury (1DPI) and three days post injury (3DPI) B10 and mdx mice showing dysregulation in mdx . (F = 101.1, P < 0.0001; F = 89.58, P < 0.0001) C) Strain-comparison of the expression of autophagy-associated genes showing reduced expression of autophagy genes in both DMD models compared to healthy controls (t = 5.260, 1.917; 8.707, 1.513; 3.603, 3.130) and D) changes in autophagy-associated genes in NI, 1DPI and 3DPI B10 and mdx satellite cells demonstrating impaired autophagy dynamics in mdx cells during regeneration ([F,P] = [15.51, 0.0004], [62.45, 0.0001], [6.518, 0.0109], [24.28, <0.0001], [20.22, 0.0001]. E) Representative images of IF staining against PAX7, MYOG and MyHC from mdx primary myoblasts treated with an autophagy inhibitor (3-MA, 5 mM and H 2 O control) or autophagy inducer (Tat-D11, 10 µM and Scramble control) for two hours prior to differentiation for two days. F) Fusion index of each condition was determined and shows increased differentiation after Tat-D11 treatment (t = 0.001 (3MA), 2.894 (Tat-D11)). Scalebars = 25 µM, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (two-tailed unpaired t-test ( C, F ) and two-way analysis of variance ( B, D )). Data are expressed as mean ± SD.
Article Snippet: When cells reached 80-90% confluence, cells were treated for two hours with either 10 µM Tat-Beclin 1 D11 peptide (Novus Biologicals) or Tat-Beclin 1 L11S peptide scramble control (vehicle, Novus Biologicals); 5 mM 3-methyladenine (3-MA, Sigma) or water (vehicle) , .
Techniques: Expressing, Comparison, Staining, Control, Two Tailed Test
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![DMD satellite cells have impaired autophagy and senescence. A) Feature plot showing cells positive for the senescence GO term, showing higher amounts in the regions where cycling progenitors and DMD enriched cells cluster. B-D) ddPCR quantification of gene expression in satellite cells in B) senescence-associated genes Cdkn1a and Cdkn2a from non-injured (NI), one day post injury (1DPI) and three days post injury (3DPI) B10 and mdx mice showing dysregulation in mdx . (F = 101.1, P < 0.0001; F = 89.58, P < 0.0001) C) Strain-comparison of the expression of autophagy-associated genes showing reduced expression of autophagy genes in both DMD models compared to healthy controls (t = 5.260, 1.917; 8.707, 1.513; 3.603, 3.130) and D) changes in autophagy-associated genes in NI, 1DPI and 3DPI B10 and mdx satellite cells demonstrating impaired autophagy dynamics in mdx cells during regeneration ([F,P] = [15.51, 0.0004], [62.45, 0.0001], [6.518, 0.0109], [24.28, <0.0001], [20.22, 0.0001]. E) Representative images of IF staining against PAX7, MYOG and MyHC from mdx primary myoblasts treated with an autophagy inhibitor (3-MA, 5 mM and H 2 O control) or autophagy inducer <t>(Tat-D11,</t> 10 µM and Scramble control) for two hours prior to differentiation for two days. F) Fusion index of each condition was determined and shows increased differentiation after Tat-D11 treatment (t = 0.001 (3MA), 2.894 (Tat-D11)). Scalebars = 25 µM, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (two-tailed unpaired t-test ( C, F ) and two-way analysis of variance ( B, D )). Data are expressed as mean ± SD.](https://bio-rxiv-images-cdn.bioz.com/dois_ending_with_63/10__1101_slash_2024__07__24__604963/10__1101_slash_2024__07__24__604963___F7.large.jpg)