Analysis of Therapeutic Targets of A Novel Peptide Athycaltide-1 in the Treatment of Isoproterenol-Induced Pathological Myocardial Hypertrophy

Abstract

Myocardial hypertrophy is a pathological feature of many heart diseases. This is a complex process involving all types of cells in the heart and interactions with circulating cells. This study is aimed at identifying the differentially expressed proteins (DEPs) in myocardial hypertrophy rats induced by isoprenaline (ISO) and treated with novel peptide Athycaltide-1 (ATH-1) and exploring the mechanism of its improvement. ITRAQ was performed to compare the three different heart states in control group, ISO group, and ATH-1 group. Pairwise comparison showed that there were 121 DEPs in ISO/control (96 upregulated and 25 downregulated), 47 DEPs in ATH-1/ISO (27 upregulated and 20 downregulated), and 116 DEPs in ATH-1/control (77 upregulated and 39 downregulated). Protein network analysis was then performed using the STRING software. Functional analysis revealed that Hspa1 protein, oxidative stress, and MAPK signaling pathway were significantly involved in the occurrence and development of myocardial hypertrophy, which was further validated by vivo model. It is proved that ATH-1 can reduce the expression of Hspa1 protein and the level of oxidative stress in hypertrophic myocardium and further inhibit the phosphorylation of p38 MAPK, JNK, and ERK1/2.

Figure 1

HE staining, Masson staining, and protein expression of ANP and BNP. (a) The cross-section of the cardiac tissue (the magnification is 20.0x) by HE staining in each group (n = 3). (b) The cross-section of the cardiac tissue (the magnification is 20.0x) by Masson staining in each group (n = 3). (c) The protein expression of ANP and BNP in each group (n = 3). ^∗P < 0.05 and ^∗∗P < 0.01.

Figure 2

(a) Histogram of upregulated and downregulated DEPs. Upregulated DEPs were in grey, and downregulated DEPs were in yellow. (b) The heatmap and cluster analysis of DEPs. Upregulated DEPs were in red, and downregulated DEPs were in green. (c) Volcano plot representation of DEPs. A: upregulated proteins; B: downregulated proteins; and C: unchanged proteins. (d) GO enrichment and KEGG analysis of DEPs in the ISO/control and (e) ATH-1/ISO comparisons. The top 10 terms of BP, MF, CC, and KEGG analysis were displayed with the parameter protein count and P value. (f) Mapping of DEPs onto a composite network based on predicted protein-protein interaction. A predicted PPI network has been visualized for the identified DEPs.

Figure 3

ATH-1 decreased the protein expression of Hspa1, p-ERK, p-JNK, and p-p38 in hypertrophic myocardium and enhanced the activity of T-SOD. (a) Differentially abundant proteins identified in the iTRAQ experiment. (b) The protein expression of Hspa1 (n = 3). ^∗P < 0.05 and ^∗∗P < 0.01. (c) The activity of T-SOD in each group (n = 3). ^∗∗P < 0.01 and ^∗∗∗P < 0.001. (d) The protein expression of ERK1/2 and p-ERK1/2 (n = 3). ns indicates none significance, ^∗P < 0.05, ^∗∗P < 0.01, and ^∗∗∗P < 0.001. (e) The protein expression of JNK and p-JNK (n = 3). ns indicates none significance, ^∗P < 0.05. (f) The protein expression of p38 MAPK and p-p38 MAPK (n = 3). ns indicates none significance, ^∗P < 0.05 and ^∗∗∗P < 0.001.