Salvianolic acid B plays an anti-obesity role in high fat diet-induced obese mice by regulating the expression of mRNA, circRNA, and lncRNA

Abstract

Background: Adipose tissue plays a central role in obesity-related metabolic diseases such as type 2 diabetes. Salvianolic acid B (Sal B), a water-soluble ingredient derived from Salvia miltiorrhiza, has been shown to reduce obesity and obesity-related metabolic diseases by suppressing adipogenesis. However, the role of Sal B in white adipose tissue (WAT) is not yet clear.

Methods: Illumina Hiseq 4000 was used to study the effects of Sal B on the expression of long non-coding RNA (lncRNA) and circular RNA (circRNA) in epididymal white adipose tissue induced by a high fat diet in obese mice.

Results: RNA-Seq data showed that 234 lncRNAs, 19 circRNAs, and 132 mRNAs were differentially expressed in WAT under Sal B treatment. The up-regulated protein-coding genes in WAT of the Sal B-treated group were involved in the insulin resistance pathway, while the down-regulated genes mainly participated in the IL-17 signaling pathway. Other pathways may play an important role in the formation and differentiation of adipose tissue, such as B cell receptor signaling. Analysis of the lncRNA-mRNA network provides potential targets for lncRNAs in energy metabolism. We speculate that Sal B may serve as a potential therapeutic approach for obesity.

Keywords: Adipose tissue; Obesity; Sal B; circRNA; lncRNAs.

Figure 1. Analysis of DEmRNAs (A, C) and DEcircRNAs (B, D).

Hierarchical clustering. Each row represents an mRNA and each column represents a sample. Green and red represent down-and up-regulated mRNAs or circRNAs, respectively. Genes in the volcano-Plot above the green parallel line (p < 0.05) and outside the two longitudinal green lines indicated DEmRNAs and DEcircRNAs between the two compared samples.

Figure 2. GSEA Cluster Heat Map of top 10 DElncRNAs, in up-regulation and down-regulation, respectively.

(A) Biological process; (B) cellular components; (C) molecular functions, and (D) KEGG pathway, each row represents a functional entry, and each column represents an lncRNA. GSEA is a method used to determine whether a given gene set has significant differences among different groups. Genes in these sets have some degree of correlation. Therefore, enrichment analysis of gene sets can make up for the shortcomings of single gene in the analysis.

Figure 3. Sequencing and quantitative PCR.

Sequencing and quantitative PCR for mRNAs (Wbscr27, Sfrp5, Adig, and Saa3), circRNAs- chr7:67264864–67268400:- and lncRNA-ENSMUST00000169194. The quantitative PCR results were consistent with the sequencing data. n = 5.

Figure 4. LncRNA–mRNA regulatory network (ENSMUST0000140351 and ENSMUST00000169194).

Squares represent lncRNAs, circles represent mRNAs; red indicates up-regulated expression and blue indicates down-regulated expression. The solid line is positively correlated and the dotted line is negatively correlated. LncRNA–mRNA regulatory network was constructed using Cytoscape v2.8.2 software.

Figure 5. GO analysis.

(A) up-regulated and (B) down-regulated of DEmRNAs. Using GO database (http://www.geneontology.org) analysis the GO enrichment of the DEmRNAs, based on three aspects: biological processes (BP), cellular components (CC), and molecular functions (MF). The log 10 values (p-value) denote enrichment scores and represent the significance of the GO term enrichment among the DEmRNAs.