Extended Data Fig. 8: Rarβ is indispensable for 4-oxo-RA’s beneficial effects.
a, Experimental design to characterize Rarβ KO HSC response after 4-oxo-RA in vivo treatment following MI. b, UMAP plot of scRNA-seq on BM HSPC cells isolated from Rarβ KO mice that received in vivo treatment with vehicle or 4-oxo-RA post MI, as well as from Rarβ KO mice undergoing sham surgery. n = 2. c, UMAP density plots of each condition in BM Rarβ KO HSPC scRNA-seq depicting relative cell abundance. d, Bar plot of quantified relative cluster abundance in each condition based on scRNA-seq data of BM Rarβ KO HSPC cells. e, GSEA of published mouse HSPC signatures in the pairwise comparisons between MI+vehicle vs. sham HSC cluster and MI + 4-oxo-RA vs. MI+vehicle HSC cluster based on Rarβ KO scRNA-seq. f, ScHSC division assay after 48 h in sham, MI+vehicle and MI + 4-oxo-RA Rarβ KO HSCs. The percentage of cells is shown. Depicted p values correspond to the percentage of non-divided cells. Statistics denote comparisons between vehicle or 4-oxo-RA condition and the sham condition. Ordinary two-way ANOVA. n = 5–6. g, 1st, 2nd and 3rd plating of Rarβ KO HSC CFU assay comparing sham, MI+vehicle and MI + 4-oxo-RA condition. Ordinary two-way ANOVA. n = 6. h and i, Flow cytometry-based analysis of emergency hematopoiesis in Rarβ KO mice during the acute phase at day three post MI. Percentage of leukocyte cell frequencies is depicted in BM and myocardium. Ordinary two-way ANOVA. BM and myocardium, n = 4–6. j, Left, Representative echocardiographic images showing left ventricular (LV) morphology during diastole in sham, MI with vehicle, and MI with 4-oxo-RA treatment groups. Ao, ascending aorta; LA, left atrium; LV, left ventricle. Right, Analysis of ejection fraction and endiastolic volume at day one post MI (as quality control) and in the chronic phase at day 28 post MI. Ordinary one-way ANOVA. n = 7-10. k, Left, Schematic design of chimera experiments to characterize the transplantation of BM Rarβ-KO cells into wildtype mice followed by MI and 4-oxo-RA in vivo treatment. Right, Analysis of ejection fraction, stroke volume and endiastolic volume in sham, MI with vehicle, and MI with 4-oxo-RA treatment groups at day one post MI (as quality control) and in the chronic phase at day 28 post MI. Ordinary one-way ANOVA. n = 8-11. In c, cardiac fibroblast were isolated 14 day after MI. In (a-e), cells were isolated in the acute phase at day three after MI. Data are presented as mean ± standard deviation. n indicates the number of biological replicates per condition or total number of human BM donors. For (b-e), two or more independent experiments were performed.
