Genetic deletion of MrgD receptor disrupts cardiac protein homeostasis in mice

Figures

• 

  Fig. 1. Evaluation of cardiac morphology. (A) Macroscopic view of the whole heart (bar = 1 cm). (B, C) Representative photomicrographs of hematoxylin and eosin-stained histological sections of the heart in low (B, bar = 1 mm) and high (C, bar = 50 μm, with cross-sectional cardiomyocytes) magnifications. (D) LV wall thickness. (E) LV chamber area. (F) Cardiomyocyte cross-sectional area. (G) Cardiomyocyte profile by area. Data presented as mean ± SD (n = 5/group). Significant differences between groups are indicated with the symbols: ** ≠ WT (p < 0.01), as determined by unpaired Student’s t test with Welch’s correction

• 

  Fig. 2. Evaluation of cardiac collagen deposition. (A) Representative photomicrographs of Picrosirius Red-stained histological sections of the heart in the same magnification (bar = 50 μm), and collagen deposition. (B) Representative photomicrographs of collagen type 1 immunostaining counterstained with hematoxylin in cardiac tissue at the same magnification in all groups (bar = 60 μm). (C) Representative photomicrographs of collagen type 3 immunostaining counterstained with hematoxylin in cardiac tissue at the same magnification in all groups (bar = 60 μm). (D) Col1a1 and (E) Col3a1 mRNA expression. (F) MMP-2 protein expression and representative Western blot analysis of proteins in LV. Data presented as mean ± SD (n = 5/group). Significant differences between groups are indicated with the symbols: * ≠ WT (p < 0.05), *** ≠ WT (p < 0.001), **** ≠ WT (p < 0.0001), as determined by unpaired Student’s t test with Welch’s correction

• 

  Fig. 3. Evaluation of pro-oxidative response. NOX2 (A) mRNA and (B) protein expression. NOX4 (C) mRNA and (D) protein expression. (E) ERO1α protein expression. (F) MDA levels. (G) Protein carbonyl content. (H) Representative Western blot analysis of proteins in LV. Data presented as mean ± SD (n = 5/group). Significant differences between groups are indicated with the symbols: * ≠ WT (p < 0.05), ** ≠ WT (p < 0.01), as determined by unpaired Student’s t test with Welch’s correction

• 

  Fig. 4. Evaluation of ER stress. GRP78 (A) mRNA and (B) protein expression. CHOP (C) mRNA and (D) protein expression. (E) Representative Western blot analysis of proteins in LV. Data presented as mean ± SD (n = 5/group). Significant differences between groups are indicated with the symbols: * ≠ WT (p < 0.05), ** ≠ WT (p < 0.01), as determined by unpaired Student’s t test with Welch’s correction

• 

  Fig. 5. Evaluation of protein homeostasis. (A) MuRF-1, (B) Atrogin-1, and (C) Polyubiquitinated proteins expression. (D) Representative Western blot analysis of proteins in LV. Data presented as mean ± SD (n = 5/group). Significant differences between groups are indicated with the symbols: * ≠ WT (p < 0.05), ** ≠ WT (p < 0.01), as determined by unpaired Student’s t test with Welch’s correction

• 

  Fig. 6. MrgD deficiency disrupts cardiac protein homeostasis. MrgD deficiency led to a pattern of dilated cardiomyopathy with a remodeling process characterized by cardiac atrophy and fibrosis. Changes in cardiac extracellular matrix were evidenced by increased collagen types 1 and 3 deposition. The atrophic response was associated with increased protein degradation, which occurred as a result of a disruption in protein homeostasis. MrgD deletion increased ROS by augmenting both NOX and ERO1α protein expression, which increased ER stress and, consequently, the ubiquitin-proteasome pathway