hsd icr cd1 mice  (Envigo)

Journal: Cell Systems

Article Title: Cold and hot fibrosis define clinically distinct cardiac pathologies

doi: 10.1016/j.cels.2025.101198

Figure Lengend Snippet: Acute myocardial infarction results in cold fibrosis (A) Mathematical model of the myofibroblast-macrophage cell circuit predicts three possible outcomes: hot fibrosis, cold fibrosis, and healing based on the abundance of these two populations (axes). The basin of attraction for the healing state is bounded by a separatrix. The scheme also denotes the growth factor signaling as either ON (black lines) or OFF (gray lines) at each specific fixed point. (B) Single-cell RNA sequencing (scRNA-seq) dynamic analysis of total macrophage and myofibroblast populations (% of total interstitial cells) in an adult mouse heart following-MI data obtained from Forte et al. Data are represented as cell composition fold change (FC) to day 0. These data are further described in Figures S1 A and S1B. UMAP of clusters highlighting macrophages (green, full line) and myofibroblasts (red, straited line). Cell-type clusters also include smooth muscle cells, fibroblasts, endothelial cells, granulocytes, NK/T cells, and B cells. (C) Breeding strategy used to create the macrophage-myofibroblast double-reporter (MAMY) mice. First, we crossed Postn MCM/MCM mice with Rosa26 tdTom a to/tdTom a to mice to obtain an established tamoxifen-inducible cardiac myofibroblasts lineage-tracer , . We then crossed these mice with a global monocyte/macrophage reporter-Cx3cr1 GFP/GFP . Triple-heterozygote mice were used as experimental animals (Postn MCM/+ Rosa26 TdTomto/+ Cx3cr1 GFP/+ ). (D) Experimental scheme whereby MAMY mice (Postn MCM/+ Rosa26 TdTomto/+ Cx3cr1 GFP/+ ) underwent sham or MI by permanent left anterior descending artery (LAD) ligation. To label myofibroblasts with tdTomato, mice were given daily tamoxifen injections following MI (red line). Cardiac function was tracked using 2D echocardiography at multiple time points following MI: 7 days prior to MI (baseline), and 7, 14, and 28 days post-MI (black arrows). Hearts were collected for histological fibrosis and immunofluorescence (IF) analysis on days: 2, 4, 7, 14, and 28 post-MI/sham (red arrows). (E) Ejection fraction (EF; %) analysis was performed on MI/Sham operated MAMY mice (Sham, n = 4; MI, n = 7). To extract the direct % change in EF between days 28 post-MI and baseline, we calculated the ΔEF value per animal . Red, MI; blue, sham. Results are represented as mean ± SEM. Statistical analysis performed using two-way ANOVA with Sidak’s adjusted p values (left) or two-tailed unpaired t test (right). Individual points represent individual biological replicates. (F) Representative histology images of MAMY hearts stained with sirius red on days 2, 4, 7, 14, and 28 post-MI/sham. Scale bar, 1 mm. Collagen, red; healthy myocardium, yellow. (G) MAMY mice fibrosis area/left ventricle (LV) in % was calculated per timepoint: Sham ( n = 12), day 2 ( n = 7), day 4 ( n = 4), day 7 ( n = 9), day 14 ( n = 4), and 28 ( n = 13) post-MI. Mix-max violin plot represent the median (middle full line) and quartiles (striated lines). Statistical analysis performed using one way ANOVA with Tukey’s multiple comparisons test. (H–K) Quantification of spatiotemporal distribution of cardiac-troponin-T cells (cardiomyocytes, cTnT+; purple), the lineage-traced (tdTomato+; red) myofibroblasts, and macrophages (GFP+, green) in Sham hearts and infarct zone of MI operated hearts. cTnT+, tdTomato+, and GFP+ cells were also measured at the suture area of 28 days post-MI hearts ( , Figure S1 C). (H) Representative immunofluorescence (IF) images of Sham ( n = 5; 1,487 ± 118.5 SEM cells per replicate) and MI operated hearts on days 2 ( n = 4; 1,088 ± 341.8 SEM cells per replicate), 4 ( n = 4; 1,027 ± 352.6 SEM cells per replicate), 7 ( n = 4; 1,792 ± 153.3 SEM cells per replicate), 14 ( n = 3; an average of 1,471 ± 275.2 SEM cells per replicate), and 28 ( n = 5; 1,627 ± 248.7 and 1,049 ± 315.9 SEM cells per replicate for infarct zone and suture, respectively) post-MI. Images are represented as either single channel for GFP, tdTomato, and cTnT, as a merged IF image and as a cell-type map that illustrates the identified cells as representative points. White scale bar, 100 μm. cTnT+ cells (I), GFP+ cells (J), and tdTomato+ cells (K) were quantified as count per 100 μm neighborhood or as % of cells per 100 μm neighborhood ( , Figure S1 C). Results are represented as mean ± SEM. In (I)–(K), infarct zone is denoted as a full circle, day 28 suture is denoted as an upside-down triangle. Statistical analysis performed using the Mann-Whitney test with Bonferroni adjusted p values (I–K). (L) Experimental scheme whereby adult Hsd:ICR (CD1) mice hearts underwent MI by permanent LAD ligation and further processed by flow cytometry . LV samples below the suture of MI or sham operated mice were collected at 6 different time points following injury at days: 0/Sham ( n = 11), 2 ( n = 5), 4 ( n = 5), 7 ( n = 7), 14 ( n = 5), and 65 ( n = 5). (M–Q) (M) Representative flow cytometry plots of the gating scheme used to identify (N) total cardiac macrophages (out of total immune cells- CD45 + ), (O) infiltrating cells (monocytes and neutrophils out of total immune cells-CD45 + ), (P) cardiac activated macrophages, measured by CD11b mean fluorescence intensity (MFI), and (Q) Resident macrophages (TIM4 + macrophages out of total macrophages. Data presented as mean ± SEM. Statistical analysis performed using one way ANOVA with Dunnett’s multiple comparisons procedure.

Article Snippet: Hsd: ICR (CD1) mice , Envigo , N/A.

Techniques: RNA Sequencing, Ligation, Immunofluorescence, Two Tailed Test, Staining, MANN-WHITNEY, Flow Cytometry, Fluorescence

Journal: Cell Systems

Article Title: Cold and hot fibrosis define clinically distinct cardiac pathologies

doi: 10.1016/j.cels.2025.101198

Figure Lengend Snippet:

Article Snippet: Hsd: ICR (CD1) mice , Envigo , N/A.

Techniques: Control, Recombinant, Synthesized, Sequencing, Mutagenesis, Software, Flow Cytometry, Electron Microscopy, Staining, Gene Expression