Ganoderma lucidum polysaccharides attenuates pressure-overload-induced pathological cardiac hypertrophy

Figures

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  FIGURE 1. GLPs alleviate Ang II-induced hypertrophy in cardiomyocytes. (A) Representative images of cardiomyocytes treated with Ang II (1 µM) and GLPs (50 mg/mL) (green: α-actinin; blue: DAPI). Original magnification ×200, scale bars, 50 μM. (B) Quantification of cell surface areas (n = 50 + cells in each group). (C) Real-time PCR analysis of ANP gene expression in H9C2 cells treated with Ang II and GLPs (n = 3). (D) Real-time PCR analysis of BNP gene expression in H9C2 cells treated with Ang II and GLPs (n = 3). (E) Real-time PCR analysis of β-MHC gene expression in H9C2 cells treated with Ang II and GLPs (n = 3). All values represent the means ± SEM, *p < 0.05, **p < 0.01. Ctrl: control; Ang II: angiotensin II; ANP: atrial natriuretic peptide; BNP: B-type natriuretic peptide; β-MHC: β-myosin heavy chain; GLPs: Ganoderma lucidum polysaccharides.

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  FIGURE 2. GLPs improve cardiac function and attenuate cardiac hypertrophy induced by TAC in vivo. (A) Representative M-mode echocardiographic tracings of the left ventricle for mice after sham operation or TAC surgery. (B) Quantification of the recovery of EF% and FS% in (A) (n = 4 per group). (C) Quantification of the LVID; d in (A). (D) Quantification of the LVID; s in (A). (E) Changes in the HW/BW ratio for WT and GLP mice after sham operation or TAC surgery (n = 4 per group). (F) Histological sections of hearts from WT and GLP mice after sham operation or TAC surgery were stained with H&E to analyze cardiomyocyte size. Original magnification, ×400, scale bars, 100 μM. (G) Quantification of cell surface areas (n = 50 + cells in each group). (H–J) Real-time PCR analysis of hypertrophic markers (ANP, BNP, and β-MHC) in WT and GLP mouse hearts after sham operation or TAC surgery (n = 4 per group). All values represent the means ± SEM, *p < 0.05, **p < 0.01. TAC: transverse aortic constriction; EF: ejection fraction; FS: fractional shortening; LVID; d: end-diastolic left ventricular internal dimension; LVID; s: end-systolic left ventricular internal dimension; HW/BW: heart weight/body…

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  FIGURE 3. GLPs alleviate cardiac fibrosis induced by TAC in vivo. (A) Histological sections of hearts from WT and GLP mice after sham operation or TAC surgery were stained with Masson’s trichrome to analyze collagen deposition. Original magnification, ×400, scale bars, 100 μM. (B) Statistical results of cardiac interstitial fibrosis (n = 4 per group). (C–F) Real-time PCR analysis of fibrotic markers (Col1a1, Col3a1, fibronectin, and α-SMA) in WT and GLP mouse hearts after sham operation or TAC surgery (n = 4 per group). All values represent the means ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001. WT: wild-type; Col1a1: collagen I; Col3a1: collagen III; α-SMA: α-smooth muscle actin; GLPs: Ganoderma lucidum polysaccharides.

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  FIGURE 4. PPARγ and PGC-1α are upregulated by GLPs in both cardiomyocytes and mouse hearts. (A) Representative Western blot results for PPARγ and PGC-1α protein levels in extracts from H9C2 cells treated with Ang II (1 µM) and GLPs (50 mg/mL). (B) Quantification of the PPARγ protein levels shown in (A) (n = 3). (C) Quantification of the PGC-1α protein levels shown in (A) (n = 3). (D) Representative Western blot results for PPARγ and PGC-1α protein levels in WT and GLP mouse hearts after sham operation or TAC surgery. (E) Quantification of the PPARγ protein levels shown in (D) (n = 4). (F) Quantification of the PGC-1α protein levels shown in (D) (n = 4). All values represent the means ± SEM, *p < 0.05, **p < 0.01. TAC: transverse aortic constriction; Ang II: angiotensin II; PPAR γ: peroxisome proliferator-activated receptor gamma; PGC-1α: peroxisome proliferator-activated receptor gamma coactivator-1α.; GLPs: Ganoderma lucidum polysaccharides.

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  FIGURE 5. GLPs alleviate Ang II-induced cardiomyocyte hypertrophy via PPARγ. (A) Representative Western blot results for PPARγ and PGC-1α protein levels in extracts from H9C2 cell treated with Ang II (1 µM) and GLPs (50 mg/mL) and pre-treatment with T0070907 (1 μM-4 h). (B) Quantification of the PPARγ protein levels shown in (B) (n = 3). (C) Quantification of the PGC-1α protein levels shown in (A) (n = 3). (D) Representative images of cardiomyocytes treated with Ang II and GLPs and pre-treated with T0070907 (green: α-actinin; blue: DAPI). Original magnification, ×200, scale bars, 50 μM. (E) Quantification of cell surface areas (n = 50 + cells in each group). (F) Real-time PCR analysis of ANP gene expression in H9C2 cells treated with Ang II and GLPs and pre-treated with T0070907. (G) Real-time PCR analysis of BNP gene expression in H9C2 cells treated with Ang II and GLPs and pre-treated with T0070907 (n = 3). (H) Real-time PCR analysis of β-MHC gene expression in H9C2 cells treated with Ang II and GLPs and pre-treated with T0070907 (n = 3). All values represent the means ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001. Ang II: angiotensin II; PPAR γ: peroxisome proliferator-activated receptor…

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  FIGURE 6. GLPs increase cardiac function and ameliorate cardiac hypertrophy via PPARγ. (A) Representative M-mode echocardiographic tracings of the left ventricle in mice after sham operation or TAC surgery. (B) Quantification of the recovery of EF% and FS% in (A) (n = 4 per group). (C) Quantification of the LVID; d in (A). (D) Quantification of the LVID; s in (A). (E) Changes in the HW/BW ratio for WT and GLP mice after sham operation or TAC surgery (n = 4 per group). (F) Histological sections of hearts from WT and GLP mice after sham operation or TAC surgery were stained with H&E to analyze cardiomyocyte size. Original magnification, ×400, scale bars, 100 μM. (G) Quantification of cell surface areas (n = 50 + cells in each group). (H–J) Real-time PCR analysis of hypertrophic markers (ANP, BNP, and β-MHC) in WT and GLP mouse hearts after sham operation or TAC surgery (n = 4 per group). All values represent the means ± SEM, *p < 0.05, **p < 0.01. TAC: transverse aortic constriction; EF: ejection fraction; FS: fractional shortening; LVID; d: end-diastolic left ventricular internal dimension; LVID; s: end-systolic left ventricular internal dimension; HW/BW: heart weight/body weight; H&E:…

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  FIGURE 7. GLPs attenuate cardiac fibrosis induced by TAC via PPARγ in vivo. (A) Histological sections of hearts from WT and GLP mice after sham operation or TAC surgery were stained with Masson’s trichrome to analyze collagen deposition. Original magnification, ×400, scale bars, 100 μM. (B) Statistical results for cardiac interstitial fibrosis (n = 4 per group). (C–F) Real-time PCR analysis of fibrotic markers (Col1a1, Col3a1, fibronectin, and α-SMA) in WT and GLP mouse hearts after sham operation or TAC surgery (n = 4 per group). All values represent the means ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001. WT: wild-type; Col1a1: collagen I; Col3a1: collagen III; α-SMA: α-smooth muscle actin; T007: T0070907; GLPs: Ganoderma lucidum polysaccharides.