Extended Data Fig. 5: Single cell- and single nucleus RNA-seq analysis of cardiac cells from TCRM mice treated 4 for weeks with isotype control or GREM1/2-neutralizing antibody 14-D10-2 and further validation experiments.
(a) Schematic representation of the experimental set up. (b) UMAP representation showing marker gene assigned cell populations in the murine heart. (c) Pie charts depicting the proportions of the different cell populations based on snRNA-seq analysis of TCRM hearts; isotype, 13,498 nuclei; 14-D10-2, 13,662 nuclei. (d) Enumeration of the indicated heart-infiltrating immune cell populations in 12-week-old TCRM mice treated with Ig2b isotype control or GREM1/2-neutralizing antibody (14-D10-2) using flow cytometry (N = 7 mice per group, pooled data from 2 independent experiments). Box and whiskers show min to max, mean ± interquartile range; dots indicate individual mice. (e) Heatmap showing changes in the expression of differentially expressed genes grouped by function from the fibroblast population in TCRM mice treated with isotype or 14-D10-2 antibodies. (f) Representative images showing immune cell infiltration and collagen deposition in hearts from 12-week-old TCRM mice treated with the indicated antibodies; hematoxylin-eosin (left panels) and confocal microscopy with indicated antibody staining (right panels). (g) Assessment of blood cell composition and the concentration of key serum enzymes in 16-week-old Balb/c mice according to the indicated treatment scheme; WBC, white blood cells; RBC, red blood cells; PLT, platelets; AST, aspartate aminotransferase; ALT, alanine transaminase; CK, creatine kinase (N = 6 mice per group, pooled data from two independent experiments; Bars indicate mean ± SEM). Statistical analysis was performed using Student’s t test (d, g, h) and 2-way Anova with Sidak’s multiple comparisons test (disease activity score in h), with *, p < 0.05; **, p < 0.01; ***, p < 0.001.
