A time-series minimally invasive transverse aortic constriction mouse model for pressure overload-induced cardiac remodeling and heart failure

Figures

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  Figure 1. Homemade surgical instruments. (A) Creating the ligation spacer and curved needle with suture: bend the blunt 25-gauge needle 30°~45° with a needle holder. (B) Making a simple surgical retractor with the paper clip, as illustrated previously.

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  Figure 2. The procedure of minimally invasive TAC. (A) Open a 1.5 ~ 2 cm incision at the midline of the neck and chest. (B) Cut the sternum stem about 3~5 mm. (C) Separate the aortic arch from the nearby connective tissue by using the above-mentioned simple surgical retractor. (D) Guide the suture under the aortic arch by using the above-mentioned curved needle with suture. (E) Pull out of the suture from the curved needle. (F) Binding a constriction of the desired diameter with the above-mentioned 25-gauge curved needle. (G) Completed constriction.

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  Figure 3. Color and pulsed-wave Doppler ultrasound. (A) Representative color Doppler imaging of an aortic flow velocity peak of the non-ligated (sham) mouse (top) and the ligated (TAC) mouse (bottom). The constriction between the two carotid arteries on the aortic arch is visible (yellow arrowheads). (B) Quantification of the AV peak velocity after surgery. (C) AV peak pressure was calculated by doppler velocities. n = 25~50. Unpaired t-test for two-group comparisons, ****p < 0.0001 vs. Sham.

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  Figure 4. Echocardiography at various time points after surgery. (A) Representative two-dimensional guided M-mode echocardiogram of the various groups. (B) The thickness of the diastolic left ventricular anterior wall. (C) Systolic left ventricular anterior wall. (D) Diastolic left ventricular posterior wall. (E) Systolic left ventricular posterior wall. (F) Left ventricular mass AW. (G) Quantification of ejection fraction (EF). (H) Quantification of fractional shortening (FS). (I) Diastolic left ventricular internal dimensions. (J) Systolic left ventricular internal dimensions. (K) Diastolic left ventricular volume. (L) Systolic left ventricular volume. (M) Heart rate. n = 12~34. Unpaired t-test for two-group comparisons, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs. corresponding Sham.

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  Figure 5. Survival of mice after TAC surgery. C57BL/J mice were randomized to TAC with a 25-gauge needle or sham surgery. The survival of mice was monitored for 12 weeks after surgery. Log-rank (Mantel–Cox) test and Gehan–Breslow–Wilcoxon test were performed for the survival rate of mice after TAC. **p < 0.01 vs. Sham.

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  Figure 6. Mouse heart weight and size had a time-series increase after TAC. (A) Representative gross heart of mice at various times upon surgery (top), representative images of hematoxylin and eosin (H&E) staining of heart section across the heart’s short axis (transverse section) at various times upon surgery (middle), and representative images of H&E staining of heart section along the heart’s long axis (coronal section) at various times upon surgery (bottom). (B) HW/BW ratios at various times upon surgery. (C) HW/TL ratios at various times upon surgery. n = 10~27. Unpaired t-test for two-group comparisons, ****p < 0.0001 vs. the corresponding sham.

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  Figure 7. Cardiomyocyte area during TAC-induced time-series cardiac remodeling. Representative images of wheat germ agglutinin staining of heart sections (top). Quantification of cross-sectional areas of cardiomyocytes (bottom), n = 8~15. Unpaired t-test for two-group comparisons, ***p < 0.001, ****p < 0.0001 vs. corresponding sham.

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  Figure 8. Expression of fetal genes during TAC-induced time-series cardiac remodeling. Expression of the Anf (A), Bnp (B), and Myh7 (C) in mouse hearts was quantified by qPCR at various times upon surgery. n = 8~20. Unpaired t-test for two-group comparisons, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs. the corresponding sham.

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  Figure 9. Time-series cardiac fibrosis induced by TAC. (A) Representative images of Masson’s trichrome staining of heart sections (top). (B) Quantification of the fibrotic area (bottom), n = 8~15. (C) Expression of Col1a2 and Col3a1 in mouse heart was detected by qPCR at various times upon surgery, n = 8~20. Unpaired t-test for two-group comparisons, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs. corresponding sham.