Gastrin ameliorates heart failure and suppresses myocardial remodeling via the JAK2/STAT3 and ERK1/2 pathways

Figures

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  Fig 1. Effects of gastrin on ISO-induced heart failure. (A) Gastrin level of ISO and control group. (B) Comparison of heart size in each group of mice; (C)Echocardiography images of five groups; (D) Cardiac Output(CO) of five groups; (E) Left ventricular ejection fraction (EF) of five groups; (F) Left ventricular fraction shortening(FS) of five groups, n = 7.*P < 0.05, **P < 0.01,***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. A student’s t-test was performed for (Panel A), and one-way ANOVA with Tukey’s post-hoc was conducted for (Panel B-F).

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  Fig 2. Effects of gastrin on ISO-induced myocardial hypertrophy in vivo. (A)Heart weight/Body weight(HW/BW) of five groups; (B)Heart weight/Tibial length(HW/TL) of five groups;(C)Posterior wall of left ventricle(LVPWs) of five groups; (D) Cardiomyocyte cross-sectional areas of five groups; n = 7.(E)Histological cross-sectional image of cardiomyocytes stained with hematoxylin and eosin (H&E);(F)(G)(H)Expression of ANP, BNP, and β-MHC in the hearts of mice across Control, Gastrin, ISO, and ISO+Gastrin groups；(I)(J)(K)Expression of ANP, BNP, and β-MHC in the hearts of mice across Control, ISO, ISO+Gastrin and ISO+Gastrin+CI-988 groups.*P < 0.05,** < 0.01,***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. One-way ANOVA with Tukey’s post-hoc was conducted for (Panel A,C,D). Welch’s one-way ANOVA analysis was performed of (Panel B,F-K).

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  Fig 3. The impact of gastrin on ISO-induced myocardial fibrosis in vivo(1). (A)Masson trichrome staining images of each group(200x magnification); (B)Immunohistochemical staining images for collagen I and collagen III of each group(200x magnification); (C)The relative fibrotic area as demonstrated by Masson's trichrome staining; (D) Quantitative analysis of type I collagen areas; (E)Quantitative analysis of type III collagen areas.*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. One-way ANOVA with Tukey’s post-hoc was conducted for (Panel C-E).

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  Fig 4. The impact of gastrin on ISO-induced myocardial fibrosis in vivo(2). (A)Immunofluorescence staining for alpha-smooth muscle actin (α-SMA); (B)(C)(D)The expression level of collagen I, collagen III and α-SMA in myocardial tissue of mice in the control, gastrin, ISO, and ISO+gastrin groups were quantified by qRT-PCR; (E)(F)(G)The expression level of collagen I, collagen III and α-SMA in myocardial tissue of mice in the control, ISO, ISO+gastrin, and ISO+gastrin+CI-988 groups were quantified by qRT-PCR;*P < 0.05, **P < 0.01,***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. Welch’s one-way ANOVA analysis was performed of (Panel B-G).

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  Fig 5. The myocardial protective effect of gastrin through JAK2/STAT3 and ERK1/2 pathways in vivo. In the control, gastrin, ISO, and ISO+gastrin groups:(A)The protein expression levels of JAK2,p-JAK2, STAT3, p-STAT3, ERK1/2, (p-ERK, and GAPDH by Western blot; (B)The level of p-JAK2/JAK2; (C)The level of p-STAT3/STAT3; (D)The level of p-ERK/ERK; In the control, ISO, ISO+gastrin, and ISO+gastrin+CI-988 groups: (E)The protein expression levels of JAK2,p-JAK2, STAT3, p-STAT3, ERK1/2, p-ERK, and GAPDH by Western blot; (F)The level of p-JAK2/JAK2; (G)The level of p-STAT3/STAT3; (H)The level of p-ERK/ERK.*P < 0.05, **P < 0.01,***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. One-way ANOVA with Tukey’s post-hoc was conducted for (Panel B-D,F-H).

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  Fig 6. Effect of gastrin on H9C2 cardiomyocyte hypertrophy. (A)Rhodamine-phalloidin staining of F-actin(400×), DAPI stainded nuclei; (B) Quantificantion analysis of cardiomyocyte surface areas; (C-D) RT-PCR analysis for ANP and BNP mRNA in control, gastrin, ISO, and ISO+gastrin groups; (E-F) RT-PCR analysis for ANP and BNP mRNA in control, ISO, ISO+gastrin, and ISO+gastrin+ CI-988 groups;*P < 0.05, **P < 0.01,***P < 0.001, ****P < 0.0001. The data are presented as the mean±SEM. One-way ANOVA with Tukey’s post-hoc was conducted for (Panel B). Welch’s one-wayANOVA analysis was performed of (Panle C-F).

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  Fig 7. Mechanism of gastrin in alleviating ISO-induced cardiomyocyte hypertrophy. In the control,gastrin, ISO, and ISO+gastrin groups:(A)The protein expression levels of JAK2,p-JAK2, STAT3, p-STAT3, ERK1/2, p-ERK, and GAPDH by Western blot; (B)The level of p-JAK2/ JAK2; (C)The level of p-STAT3/ STAT3;(D)The level of p-ERK/ERK. In the control, ISO, ISO+ gastrin, and ISO+gastrin+ CI-988 groups: (E)The protein expression levels of JAK2,p-JAK2, STAT3, p-STAT3, ERK1/2, p-ERK, and GAPDH by Western blot;(F)The level of p-JAK2/ JAK2;(G)The level of p-STAT3/ STAT3;(H)The level of p-ERK/ERK. The expression of ANP(I),BNP(J), protein ERK1/2, p-ERK(K) and the ratio of p-ERK/ERK (L)after the addition of ERK agonist mSIRK. The expression of ANP(M),BNP(N), protein STAT3,p-STAT3(O) and the ratio of p-STAT3/STAT3 (P)after the addition of STAT3 agonist colevelin.*P < 0.05, **P < 0.01. ***P < 0.001. ****P < 0.0001. The data are presented as the mean±SEM. One-way ANOVA with Tukey’s post-hoc was conducted for (Panel B-D,F–-H,L,P). Welch’s one-wayANOVA analysis was performed of (Panel I,J,M,N).