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. 2021 Jan 28:12:625846.
doi: 10.3389/fgene.2021.625846. eCollection 2021.

Comprehensive Analysis of Differentially Expressed lncRNA, circRNA and mRNA and Their ceRNA Networks in Mice With Severe Acute Pancreatitis

Bing Wang  1   2 , Jun Wu  1   2 , Qilin Huang  1   2 , Xiaohui Yuan  1   2 , Yi Yang  1   2 , Wen Jiang  1   2 , Yi Wen  1 , Lijun Tang  1   2 , Hongyu Sun  1   2   3
Affiliations
Free PMC article

Comprehensive Analysis of Differentially Expressed lncRNA, circRNA and mRNA and Their ceRNA Networks in Mice With Severe Acute Pancreatitis

Bing Wang et al. Front Genet. .
Free PMC article

Abstract

Severe acute pancreatitis (SAP) is an acute digestive system disease with high morbidity mortality and hospitalization rate worldwide, due to various causes and unknown pathogenesis. In recent years, a large number of studies have confirmed that non-coding RNAs (ncRNAs) play an important role in many cellular processes and disease occurrence. However, the underlying mechanisms based on the function of ncRNAs, including long noncoding RNA (lncRNA) and circular RNA (circRNA), in SAP remain unclear. In this study, we performed high-throughput sequencing on the pancreatic tissues of three normal mice and three SAP mice for the first time to describe and analyze the expression profiles of ncRNAs, including lncRNA and circRNA. Our results identified that 49 lncRNAs, 56 circRNAs and 1,194 mRNAs were differentially expressed in the SAP group, compared with the control group. Furthermore, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differentially expressed lncRNAs and circRNAs, and found that the functions of the parental genes are enriched in the calcium-regulated signaling pathway, NF-κB signaling pathway, autophagy and protein digestion and absorption processes, which are closely related to the central events in pathogenesis of SAP. We also constructed lncRNA/circRNA-miRNA-mRNA networks to further explore their underlying mechanism and possible relationships in SAP. We found that in the competitive endogenous RNA (ceRNA) networks, differentially expressed lncRNAs and circRNAs are mainly involved in the apoptosis pathway and calcium signal transduction pathway. In conclusion, we found that lncRNAs and circRNAs play an important role in the pathogenesis of SAP, which may provide new insights in further exploring the pathogenesis of SAP and seek new targets for SAP.

Keywords: circular RNA; competitive endogenous RNA; long noncoding RNA; mRNA; severe acute pancreatitis.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Evaluation of the SAP Model. (A) Compared with the control group (con), the histopathological sections of the pancreas in the SAP group showed focal expansion of the interlobular space of the pancreas, edema of the pancreatic lobules, extensive acinar cell necrosis and granulocyte infiltration (100x). (B) The histological score of pancreatic tissue. (C,D) The lipase and amylase levels were also significantly higher than the control group (p < 0.05). ***p < 0.001, vs. control group, n = 3 per group.
Figure 2
Figure 2
Comparison of the characteristics of lncRNA, circRNA, and mRNA expression profiles in pancreatic tissues between SAP group and control group. (A) The distribution of lncRNAs and circRNAs on chromosomes, (B) LncRNAs and mRNAs on chromosomes. (C,D) Biotypes of lncRNAs and circRNAs.
Figure 3
Figure 3
Differential expression of lncRNA, circRNA and mRNA. (A) The specific lncRNAs, circRNAs, and mRNAs shared between SAP group and control (con) group. The red points showed the up-regulated lncRNAs, circRNAs, mRNAs. The green points showed the down-regulated lncRNAs, circRNAs, and mRNAs. (B) The volcano diagrams showed the DE lncRNAs, circRNAs and mRNAs in the pancreatic head tissue of the three pairs of SAP group and control group, respectively. (C) Heatmap showing DE lncRNAs, circRNAs and mRNAs from pancreatic tissues of SAP mice compared to pancreatic tissues of control mice, respectively. Row and column represent DE lncRNA/DE circRNA/DE mRNA transcripts and tissue samples, red color represents up-regulated DERNAs, green color represents down-regulated DERNAs, and heavier color represents higher fold change.
Figure 4
Figure 4
qRT-PCR validation of DELs and DECs in SAP rats compared with matched tissues of control rats. (A) The expression level of lncRNA AK004187, AK035396, AK162674, NR_040328, uc029sug.1, TCONS_00010866. (B) The expression level of circRNA CircAmy2a5, CircZFP644, CircDTNB, CircCELA3b, CircARHGEF38, and CircSRPK2. *represents p < 0.05, **represents p < 0.01, and ***represents p < 0.001.
Figure 5
Figure 5
Functional analysis for the differentially expressed mRNAs, lncRNAs and circRNAs. (A) GO enrichment analysis for up-regulated mRNAs, up-regulated lncRNAs and the down-regulated circRNAs. Red bars are biological process, green bars are cell component, and blue bars are molecular function. The ordinate is -Log10 p value (-LgP). The larger the -LgP value, the smaller the P value, indicating that the enrichment of differentially expressed mRNA/lncRNA/circRNA in a given pathway is more significant. (B) KEGG pathway enrichment analysis for up-regulated mRNAs, up-regulated lncRNAs, and for down-regulated circRNAs. Size represents the number of enriched genes, and color indicates the degree of enrichment. Higher enrichment scores correlate with lower p-value, indicating that the enrichment of differentially expressed genes in the given pathway is significant.
Figure 6
Figure 6
Construction of the lncRNA/circRNA-miRNA-mRNA Network. (A) Network analysis of lncRNA-miRNA-mRNA. The square nodes represent mRNA. The triangle node represents miRNA. The round node represents lncRNA. (B) Network analysis of circRNA-miRNA-mRNA. The square nodes represent mRNA. The triangle node represents miRNA. The round node represents circRNA. (C) Network analysis of lncRNA-circRNA-miRNA. The purple nodes represent lncRNA. The green node represents miRNA. The red node represents circRNA.
Figure 7
Figure 7
PPI Network and Hub Gene Analysis. (A) Import the up-regulated DEMs into the STRING database to build a PPI network with 433 nodes and 1,137 edges. The darker the red of the circle means that the gene has a higher degree and has more connections with other genes; conversely, the closer the color of the circle is to yellow, the less the degree of the gene is, and the less the connection with other genes. (B) Recalculate the degree of 24 DEMs with degree >20 and get the top five genes as hub genes. These hub genes are Socs3 (degree = 26), Rnf41 (degree = 24), Smurf1 (degree = 23), Mib2 (degree = 21) and Ube2z (degree = 21). (C) GO analysis of DEMs. (D) KEGG analysis of DEMs.
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