Effect of berberine on global modulation of lncRNAs and mRNAs expression profiles in patients with stable coronary heart disease

Abstract

Background: Berberine (BBR) is an isoquinoline alkaloid found in the Berberis species. It was found to have protected effects in cardiovascular diseases. Here, we investigated the effect the regulatory function of long noncoding RNAs (lncRNAs) during the treatment of stable coronary heart disease (CHD) using BBR. We performed microarray analyses to identify differentially expressed (DE) lncRNAs and mRNAs between whole blood samples from 5 patients with stable CHD taking BBR and 5 no BBR volunteers. DE lncRNAs and mRNAs were validated by quantitative real-time PCR.

Results: A total of 1703 DE lncRNAs and 912 DE mRNAs were identified. Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated DE mRNAs might be associated with mammalian target of rapamycin and mitogen-activated protein kinase pathway. These pathways may be involved in the healing process after CHD. To study the relationship between mRNAs encoding transcription factors (DNA damage inducible transcript 3, sal-like protein 4 and estrogen receptor alpha gene) and CHD related de mRNAs, we performed protein and protein interaction analysis on their corresponding proteins. AKT and apoptosis pathway were significant enriched in protein and protein interaction network. BBR may affect downstream apoptosis pathways through DNA damage inducible transcript 3, sal-like protein 4 and estrogen receptor alpha gene. Growth arrest-specific transcript 5 might regulate CHD-related mRNAs through competing endogenous RNA mechanism and may be the downstream target gene regulated by BBR. Verified by the quantitative real-time PCR, we identified 8 DE lncRNAs that may relate to CHD. We performed coding and non-coding co-expression and competing endogenous RNA mechanism analysis of these 8 DE lncRNAs and CHD-related DE mRNA, and predicted their subcellular localization and N⁶-methyladenosine modification sites.

Conclusion: Our research found that BBR may affect mammalian target of rapamycin, mitogen-activated protein kinase, apoptosis pathway and growth arrest-specific transcript 5 in the process of CHD. These pathways may be involved in the healing process after CHD. Our research might provide novel insights for functional research of BBR.

Keywords: Berberine; Stable coronary heart disease; lncRNA; mRNA.

Fig. 1

Identification of DE lncRNAs and mRNAs. Volcano plots presenting differences in the expression of lncRNAs (A) and mRNAs (B) between the BBR and control groups. Heatmaps showing the expression profiles of the top DE lncRNAs (C) and mRNAs (D). Chromosome distribution of DE lncRNAs (E) and mRNAs (F). Up-regulated (G) and down-regulated (H) DE lncRNAs counts and categorizations distribution. DE, differentially expressed; BBR, berberine; Ctrl, control

Fig. 2

GO and KEGG pathway analyses of the DE mRNAs. A Top 10 terms from the GO analysis of DE mRNAs. B KEGG pathway analysis of up-regulated and down-regulated DE mRNAs. Left and right terms represented up-regulated and down-regulated DE mRNAs enriched pathways associated with CHD. GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; DE, differentially expressed

Fig. 3

Construction of protein-protein interaction network. A PPI network of SALL4, DDIT3, ESR1 and their interacting proteins. The red dots and green dots represent represent up-regulated and down-regulated DE mRNAs and TFs, respectively. Solid lines represent positive relationship and dash lines represent negative relationship; Line thickness represents combine score. B Network of Reactome pathways and enriched proteins. Blue diamond represents pathway; Red and green elipse represents up-regulated and down-regulated mRNAs, respectively. PPI, protein-protein interaction; DDIT3, DNA damage inducible transcript 3; SALL4, sal-like protein 4; ESR1, estrogen receptor alpha gene; TF, transcription factor

Fig. 4

Construction of the CNC network. Red and green V represents up-regulated and down-regulated DE lncRNAs, respectively. Red diamond and ellipse represent up-regulated DE mRNAs associate with CHD; Green diamond and ellipse represent down-regulated DE mRNAs associate with CHD; Ellipse represent transcription factor; Solid lines represent positive relationship and dash lines represent negative relationship; Line thickness represents combine score. DE, differentially expressed; CNC, coding and non-coding co-expression; CHD, coronary heart disease

Fig. 5

Construction of the ceRNA network. Red and green V represents up-regulated and down-regulated DE lncRNAs, respectively. Blue diamond represents microRNA; Red and green ellipse represent up-regulated and down-regulated DE mRNAs associate with CHD; Red and green rectangle represent up-regulated and down-regulated transcription factor associate with CHD. DE, differentially expressed; CHD, coronary heart disease

Fig. 6

Construction of the ceRNA network of 8 DE lncRNAs. Red and green diamond represents up-regulated and down-regulated DE lncRNAs, respectively. Blue V represents microRNA. Red and green ellipse represent up-regulated and down-regulated DE mRNAs associate with CHD; Red and green rectangle represent up-regulated and down-regulated transcription factor associate with CHD. Red solid lines represent positive relationship and lncRNA-mRNA coexpression. Green solid represent negative relationship and lncRNA-mRNA coexpression. Line thickness represents combine score. Dash dot represent lncRNA-microRNA-mRNA competitive combination

Fig. 7

Flow chart of the approach utilized in the present study