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. 2023 Aug 28:11:e15939.
doi: 10.7717/peerj.15939. eCollection 2023.

Identification of Zip8-correlated hub genes in pulmonary hypertension by informatic analysis

FanRong Zhao #  1   2   3 , Yujing Chen #  1   2   3 , Yuliang Xie  1   2   3 , Shuang Kong  1   2   3 , LiaoFan Song  1   2   3 , Hanfei Li  1   2   3 , Chao Guo  1   2   3 , Yanyan Yin  1 , Weifang Zhang  1   4 , Tiantian Zhu  1   2   3
Affiliations

Identification of Zip8-correlated hub genes in pulmonary hypertension by informatic analysis

FanRong Zhao et al. PeerJ. .

Abstract

Background: Pulmonary hypertension (PH) is a syndrome characterized by marked remodeling of the pulmonary vasculature and increased pulmonary vascular resistance, ultimately leading to right heart failure and even death. The localization of Zrt/Irt-like Protein 8 (ZIP8, a metal ion transporter, encoded by SLC39A8) was abundantly in microvasculature endothelium and its pivotal role in the lung has been demonstrated. However, the role of Zip8 in PH remains unclear.

Methods: Bioinformatics analysis was employed to identify SLC39A8 expression patterns and differentially expressed genes (DEGs) between PH patients and normal controls (NC), based on four datasets (GSE24988, GSE113439, GSE117261, and GSE15197) from the Biotechnology Gene Expression Omnibus (NCBI GEO) database. Gene set enrichment analysis (GSEA) was performed to analyze signaling pathways enriched for DEGs. Hub genes were identified by cytoHubba analysis in Cytoscape. Reverse transcriptase-polymerase chain reaction was used to validate SLC39A8 and its correlated metabolic DEGs expression in PH (SU5416/Hypoxia) mice.

Results: SLC39A8 expression was downregulated in PH patients, and this expression pattern was validated in PH (SU5416/Hypoxia) mouse lung tissue. SLC39A8-correlated genes were mainly enriched in the metabolic pathways. Within these SLC39A8-correlated genes, 202 SLC39A8-correlated metabolic genes were screened out, and seven genes were identified as SLC39A8-correlated metabolic hub genes. The expression patterns of hub genes were analyzed between PH patients and controls and further validated in PH mice. Finally, four genes (Fasn, Nsdhl, Acat2, and Acly) were downregulated in PH mice. However, there were no significant differences in the expression of the other three hub genes between PH mice and controls. Of the four genes, Fasn and Acly are key enzymes in fatty acids synthesis, Nsdhl is involved in cholesterol synthesis, and Acat2 is implicated in cholesterol metabolic transformation. Taken together, these results provide novel insight into the role of Zip8 in PH.

Keywords: Differentially expressed genes; Fatty acids; Informatic analysis; Pulmonary hypertension; Zip8.

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Conflict of interest statement

The authors declare there are no competing interests.

Figures

Figure 1
Figure 1. SLC39A8 expression was downregulated in PH patients and mouse PH models.
(A) The volcano plots of DEGs in the merged datasets (153 PH and 71 NC, merged from these four datasets), in which 3,031 DEGs were downregulated (log2FC > 0, FDR < 0.05) and 2,197 DEGs upregulated (log2FC > 0, FDR < 0.05). (B) Heatmap of DEGs in the merged datasets (C) GSEA analysis shows enriched KEGG pathways. (D) GSEA analysis shows enriched Reactome pathways. (E) Expression levels of SLC39A8 in PH and NC groups in the merged datasets. (F) Expression levels of Slc39a8 in the lungs of normoxia and Su/Hx treated PH mice (n = 10). Data were presented as mean ± SEM. ***P < 0.001.
Figure 2
Figure 2. Correlation analysis of all DEGs and SLC39A8.
(A) Heatmap of the top 20 genes positively or negatively correlated with SLC39A8. Red represents positive correlation and blue represents negative correlation. (B) Top five genes positively correlated with SLC39A8 were displayed. (C) Top five genes negatively correlated with SLC39A8 were displayed.
Figure 3
Figure 3. The GSEA analysis of SLC39A8-correlated DEGs between PH and NC.
(A) GSEA classical plots generated based on NES score in canonical Wikipathways. (B) The top three WikiPathways are listed respectively. (C) GSEA classical plots generated based on NES score in canonical Reactome pathways. (D) GSEA classical plots generated based on NES score in canonical KEGG pathways. P.adj < 0.05 and false discovery rate (FDR, qvalue) <0.25 were used to indicate significant enrichment score.
Figure 4
Figure 4. Identification of hub genes SLC39A8-correlated metabolic DEGS between PH and NC.
(A) Venn diagram of common genes in three groups (1,600 transcripts of metabolic genes, 6083 SLC39A8-correlated DEGs, and 5,228 DEGs). (B) PPI network was constructed by the STRING database and visualized by cytoscape software (v3.9.1), and each blue filled node represents a SLC39A8-related gene; (C–E) The top 15 Hub genes were identified via cytoscape software (cytohubba) using MCC (C), degree (D), and closeness (E). (F) Venn diagram of common genes in these three hub gene sets.
Figure 5
Figure 5. Verification of hub genes expression at the mRNA level.
(A) The expression of 7 key SLC39A8-correlated metabolic DEGs in the merged dataset. (B) The correlations between 7 key SLC39A8-correlated metabolic DEGs and SLC39A8 were presented independently. (C) RT-PCR analysis of the expression of Acat2, Nsdhl, Acly and Fasn in lungs of normoxia and Su/Hx treated PH mice. n = 10 for each group. Data are shown as mean ± SEM; ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; ns, no significance.
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