Mst1/Hippo signaling pathway drives isoproterenol-induced inflammatory heart remodeling

Figure 1. Mst1 ablation mitigates isoproterenol-induced myocardial dysfunction and preserves cardiomyocyte viability. Cardiac-specific Mst1 knockout (Mst1Cko) mice and Mst1f/f mice were treated with isoproterenol (ISO, 100 mg/kg) for two consecutive days. Cardiac tissues were harvested four weeks post-treatment for functional analyses. A-F. Echocardiography was used to evaluate the heart function. G-L. Cardiomyocytes were isolated from heart and the cardiomyocyte contractility was assessed under 1 Hz field stimulation using an IonOptix system. Briefly, contractions were elicited via platinum electrodes delivering rectangular depolarizing pulses. *p<0.05 vs. PBS+ Mst1f/f mice, #p<0.05 vs. ISO+ Mst1f/f mice.

Figure 2. Mst1 ablation protects against isoproterenol-induced cardiomyocyte apoptosis, oxidative stress, and calcium overload. Cardiac-specific Mst1 knockout (Mst1Cko) mice and Mst1f/f mice were treated with isoproterenol (ISO, 100 mg/kg) for two consecutive days. Cardiac tissues were harvested four weeks post-treatment for functional analyses. In vitro, HL-1 cells were transfected with Mst1-targeting siRNA (si-Mst1) or control siRNA (si-Ctrl) and then treated with 20 µM isoproterenol for 24 hrs. A. ELISA was used to evaluate the caspase-3 activity. B. Cell viability was measured by MTT assay. C. ROS production was determined by immunofluorescence. D-F. ELISA kits were used to evaluate the concentration of GSH, SOD, and GPX. G. Calcium content was measured by immunofluorescence in HL-1 cells. *p<0.05 vs. PBS+ Mst1f/f mice, #p<0.05 vs. ISO+ Mst1f/f mice.

Figure 3. Endoplasmic reticulum stress and mitochondrial fission are inhibited by Mst1 deletion in isoproterenol-mediated myocardial injury. Cardiac-specific Mst1 knockout (Mst1Cko) mice and Mst1f/f mice were treated with isoproterenol (ISO, 100 mg/kg) for two consecutive days. Cardiac tissues were harvested four weeks post-treatment for functional analyses. A-C. qPCR analysis of the markers of ER stress. D-E. RNA was isolated from heart tissues and the transcription of mitochondrial fission-related genes were determined by qPCR. *p<0.05 vs. PBS+ Mst1f/f mice, #p<0.05 vs. ISO+ Mst1f/f mice.

Figure 4. Inflammation response is controlled by Mst1 in in isoproterenol-mediated myocardial injury. Cardiac-specific Mst1 knockout (Mst1Cko) mice and Mst1f/f mice were treated with isoproterenol (ISO, 100 mg/kg) for two consecutive days. Cardiac tissues were harvested four weeks post-treatment for functional analyses. A-C. RNA was isolated from heart tissues and the transcription of inflammation-related genes were determined by qPCR. D. Immunofluorescence was used to measure the levels of GR-1 in myocardium. *p<0.05 vs. PBS+ Mst1f/f mice, #p<0.05 vs. ISO+ Mst1f/f mice.

Figure 5. GO and KEGG pathway enrichment analysis of differentially expressed genes (DEGs) in isoproterenol-treated mice. A. GO analysis of DEGs. B. Chord diagram illustrating the connections between significantly enriched GO pathways and their associated genes. C. KEGG pathway classification of DEGs, showing the percentage of genes in each category compared to the entire set of genes. D. Chord diagram illustrating the connections between significantly enriched KEGG pathways and their associated genes.