A High-Fat and High-Cholesterol Diet Induces Cardiac Fibrosis, Vascular Endothelial, and Left Ventricular Diastolic Dysfunction in SHRSP5/Dmcr Rats

Fig. 1.. Changes in body weight (A), food intake (B), systolic blood pressure (SBP; C), and heart rate (HR; D) at 916 weeks of age The following 4 groups were used: CONT group; WKY rats were fed a stroke-prone (SP) diet, CONT + HFC group; WKY rats were fed a high-fat and high-cholesterol (HFC) diet, SHRSP5 group; SHRSP5/Dmcr rats were fed an SP diet, and SHRSP5 + HFC group; SHRSP5/Dmcr rats were fed an HFC diet, which induce the non-alcoholic steatohepatitis (NASH). Data are shown as means ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Supplemental Fig. 1.. Change in mean aortic pressure (MAP) from 9 to 17 weeks of age All data are shown as mean ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Fig. 2.. Pathological liver conditions in the 4 groups at 18 weeks of age A: Macroscopic findings of the liver in the abdominal cavity. B: Masson trichrome staining for hepatic fibrosis. Scale bars = 100 µm. C and D: Hepatic function in blood biochemistry assays. The serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were measured. E: Liver weight corrected by body weight. All data are shown as means ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Fig. 3.. Evaluation of left-ventricle (LV) hypertrophy in the 4 groups at 18 weeks of age AF: Organ weights corrected by body weight (BW) for heart (A), atrium (B), ventricle (C), LV (D), right-ventricle (RV) (E), and lung (F). G: Hematoxylin and eosin (HE) staining of cross sections of the LV myocardium. Scale bars = 100 µm. H: Cross-sectional areas of cardiac myocytes calculated by those in (G). I: Quantitative RT-PCR analysis of atrial natriuretic peptide (ANP). The amount of mRNA was normalized to that of GAPDH mRNA and then expressed relative to the mean value of the CONT group. All data are shown as means ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Supplemental Fig. 2.. Evaluation of left ventricular (LV) hypertrophy in the 4 groups at 18 weeks of age AF: Organ weights (heart (A), atrium (B), ventricle (C), LV (D), right ventricle (RV) (E), and lung (F)) corrected by tibial length (G). All data are shown as mean ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Fig. 4.. Evaluation of cardiac fibrosis in the 4 groups at 18 weeks of age AD: Collagen deposition as revealed by picro-sirius red (PSR) staining in perivascular (A) and interstitial (C) regions of the LV myocardium. The perivascular fibrosis area was corrected by the area of the perimeter (B), and interstitial fibrosis was expressed relative to the CONT group (D). Scale bars = 100 µm. All data are shown as means ± SE; n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Fig. 5.. Macrophage infiltration and inflammation-related gene expression in the LV of rats from the 4 groups at 18 weeks of age A: Immunohistochemical staining with antibodies of the monocyte-macrophage marker CD68. Scale bars = 20 µm. B: Density of CD68-positive cells per unit area (mm2) determined from sections similar to those shown in A. C, D: Quantitative RT-PCR analysis of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) in the LV. The amount of mRNA was normalized to that of GAPDH mRNA and then expressed relative to the mean value for the CONT group. All data are shown as mean ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Fig. 6.. Lipid deposition in the branches of the mesenteric artery in the 4 groups at 18 weeks of age Isolated mesenteric arteries were stained with oil red O to visualize ring-shaped lipid deposition. Scale bar = 2 mm.

Fig. 7.. Endothelium-dependent relaxation curve induced by acetylcholine (ACh) of aortic rings Statistical analysis for the 4 groups was performed at the ACh 10−4M. All data are shown as means ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.

Supplemental Fig. 3.. A quantitative reverse transcription–polymerase chain reaction analysis of RAS-related C3 botulinum toxin substrate 1 (Rac-1) (A) and Toll-like receptor 4 (TLR4) (B) in the left ventricle of the 4 groups at 18 weeks of age The amount of mRNA was normalized to that of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA and then expressed relative to the mean value of the CONT group. All data are shown as mean ± standard error (SE); n = 5 in each group. *P < 0.05 vs. the CONT group, †P < 0.05 vs. the CONT + HFC group, ‡P < 0.05 vs. the SHRSP5 group.