Pelargonidin inhibits isoproterenol induced myocardial fibrosis via regulating transforming growth factor-β/Smad2/3 signaling and Th2 cytokines in mice

Figures

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  Fig. 1. Pelargonidin reduces ISO-induced cardiac hypertrophy in mice. HW/BW ratios were measured after 47 days of treatment. ISO significantly increased HW/BW compared to controls (P=0.02), indicating hypertrophy. Pelargonidin (low: 20 mg/kg/day; high: 40 mg/kg/day) and Captopril (15 mg/kg/day) significantly reduced HW/BW compared to ISO (P=0.03). The ISO + high-dose Pelargonidin (P=0.01) and ISO + Captopril (P=0.007) groups showed greater reduction than ISO + low-dose Pelargonidin. Data are mean ± SEM (n=3). Statistical significance: # P<0.05 vs. control, * P<0.05, ** P<0.01 vs. ISO.

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  Fig. 2. Pelargonidin reduces myocar-dial injury and fibrosis in ISO-induced heart failure. (A, B) CK and LDH acti-vity levels were measured in serum to assess myocardial injury. ISO treatment significantly elevated CK and LDH levels compared to controls, indicating cardiac damage. Pelargo-nidin treatment at both low (20 mg/kg/day) and high (40 mg/kg/day) doses significantly reduced these enzyme levels, suggesting cardioprotective effects. (C, D) Representative Western blot images and corresponding densito-metric quantification show the expression of fibrosis-related proteins FN1, α-SMA, and COL3A1 in myocardial tissue. ISO treatment markedly increased these proteins, reflecting cardiac fibrosis, while Pelargonidin treatment suppressed their expression in a dose-dependent manner. Data are expressed as mean ± SEM (n=3 per group). Statistical significance was determined by one-way ANOVA followed by Tukey’s post hoc test. # P<0.05, ## P<0.01, vs. control group, * P<0.01, ** P<0.001 vs. ISO group.

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  Fig. 3. Pelargonidin attenuates collagen deposition in ISO-induced heart failure. (A) Representative immunohistochemistry images (40×) of left ventricular tissue illustrating collagen I expression across experimental groups: (I) Control, showing minimal collagen deposition; (II) ISO-induced CHF, exhibiting marked collagen accumulation indicative of myocardial fibrosis; (III) Low-dose Pelargonidin (20 mg/kg/day), showing partial reduction of collagen deposition; (IV) High-dose Pelargonidin (40 mg/kg/day), showing greater reduction; (V) Captopril (15 mg/kg/day) as a positive control, demonstrating reduced collagen levels. (B) Quantitative assessment of total collagen content was performed using a hydroxyproline assay, confirming histological findings. Data are presented as mean ± SEM (n=3). Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test: # P<0.05 vs. control group, * P<0.05, vs. ISO group.

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  Fig. 4. Pelargonidin suppresses TGF-β1/Smad signaling in ISO-induced heart failure. (A) Representative Western blot images showing expression of TGF-β1, Smad2, Smad3, p-Smad2, and p-Smad3 in left ventricular tissue across experimental groups. (B) Densitometric quantification of the blots confirming the effects of Pelargonidin on the Smad signaling pathway. Data are expressed as mean ± SEM (n=6 per group). Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test: # P<0.05, vs. control group, * P<0.01, ** P<0.001 vs. ISO group.

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  Fig. 5. Pelargonidin regulates mRNA expression of ECM-associated genes in ISO-induced heart failure. Quantitative PCR analysis of left ventricular tissue showing relative mRNA expression of MMP-9 (A), TIMP1 (B), and elastin (C) across experimental groups. ISO treatment significantly increased expression of these genes compared to controls, while Pelargonidin treatment dose-dependently suppressed their expression. Data are expressed as mean ± SEM (n=3). Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test: # P<0.05, ## P<0.01, vs. control group, * P<0.01, ** P<0.001 vs. ISO group.

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  Fig. 6. Pelargonidin reduces Th2 inflammatory cytokines in ISO-induced heart failure. Quantification of myocardial IL-4 (A) and IL-13 (B) levels in different experimental groups. ISO treatment significantly increased cytokine expression compared to controls, while Pelargonidin at both low (20 mg/kg/day) and high (40 mg/kg/day) doses dose-dependently suppressed IL-4 and IL-13 levels. Data are presented as mean ± SEM (n=3). Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test: # P<0.05, ## P<0.01, vs. control group, * P<0.01, ** P<0.001 vs. ISO group.