Journal: bioRxiv

Article Title: KLF4 in smooth muscle cell-derived progenitor cells is essential for angiotensin II-induced cardiac inflammation and fibrosis

doi: 10.1101/2024.06.04.597485

Figure Lengend Snippet: Gli1 -Cre ERT -YFP mice were subjected to Angiotensin II (AngII)-induced cardiac hypertrophy and fibrosis, as described in the Methods section. (A) Masson’s trichrome staining of 4-week AngII or saline treated cardiac tissue; ECM/collagen deposition shown in blue. Representative 20x images of N=4 in each group are shown. (B) Cardiac tissue sections were immunofluorescently stained with anti-GFP antibody and subjected to label-free SHG imaging to visualize the collagen deposition (Red). YFP + cells were imaged (Green) and overlayed to examine their association with ECM/collagen deposition. Representative 40x images of saline (N=10) and AngII-treated (N=12) tissues are shown. ( C ) Cardiac tissue sections were immunofluorescently stained with anti-GFP (Green), anti-αSMA (Red) and DAPI. Representative 60x images of N=6 in each group are shown. (D-I) Cardiac tissue from 2-week AngII- or saline-treated mice were harvested and prepared for scRNA-seq, as described in the Methods section. (D) Uniform Manifold Approximation and Projection (UMAP) visualization is shown. Blue circle highlights the major cluster that is annotated as AdvSca1-SM and fibroblast (SM-Fib) clusters. (E) scRNA-seq data UMAP plot colored by treatment (Saline - blue; AngII - orange). Red circle highlights the AngII-induced shift. (F) YFP transcript positive cells were highlighted in green in the UMAP plot. (G&H) Composition of Saline and AngII-treated samples were visualized for all cells (G) and YFP + cells (H) of the scRNA-seq data. (I) Pathway analysis for genes up-regulated (top) and down-regulated (bottom) by AngII treatment in YFP + AdvSca1-SM cells.

Article Snippet: Sorted cells were plated in gelatin-coated plates with AdvSca1-SM media (α MEM [Gibco, Cat# 32571036]), 10% MSC qualified fetal bovine serum (FBS)(Thermofisher Cat #12662029), 1x Penicillin Streptomycin, 1ng/mL murine basic fibroblast growth factor (R&D systems 3139-FB), and 5ng/mL murine epidermal growth factor (R&D systems 2028-EG) at the density of 20,000/cm 2 .

Techniques: Staining, Saline, Imaging