Idiopathic pulmonary arterial hypertension (IPAH), a rare but life-threatening cardiopulmonary disease without any known associated disease or genetic cause, is characterized by progressively increased pulmonary artery pressure (PAP) and pulmonary vascular resistance (PVR) [
microRNA (miRNA) is a type of small, noncoding RNA, which negatively regulates the expression of targeted genes via posttranscriptional regulation [
Bioinformatic analysis has been widely used to investigate potential mechanisms in the pathology of disease since it was developed [
In this study, microarray datasets of IPAH were searched and downloaded from Gene Expression Omnibus (GEO). miRNA-mRNA network was established accompanied with associated comprehensive analyses in order to better understand the mechanisms of IPAH. Additionally, potential drugs targeting hub genes were investigated to contribute to future therapies of IPAH.
The mRNA and miRNA expression profiles of IPAH patients from Gene Expression Omnibus (
R-platform (
R package clusterProfiler [
The online tool, STRING (
To better understand the function of DEMI, miRDB (
L1000 platform (
The basic information of the datasets related to IPAH is shown in Table
Details of datasets related to IPAH patients.
GEO ID | Platform | Organism | Experiment type | Samples (case vs. control) | Country | Year | |
---|---|---|---|---|---|---|---|
mRNA | GSE117261 | GPL6244 | Homo sapiens | Expression profiling by array | 32 vs. 25 | United States of America | 2018 |
GSE113439 | GPL6244 | Homo sapiens | Expression profiling by array | 6 vs. 11 | Canada | 2018 | |
miRNA | GSE67597 | GPL18402 | Homo sapiens | Noncoding RNA profiling by array | 7 vs. 8 | United States of America | 2015 |
IPAH: idiopathic pulmonary arterial hypertension; GEO; Gene Expression Omnibus.
(a) Volcano plot of DEGs in IPAH. Green represents downregulated DEGs; red represents upregulated DEGs; and black represents no difference. (b) Heat map of the DEGs in IPAH compared with normal controls. Red represents greater expression and green represents less expression. (c) Volcano plot of DEMIs in IPAH. DEG: differentially expressed gene; IPAH: idiopathic pulmonary arterial hypertension; DEMI: differentially expressed miRNA.
Subsequently, functional enrichment analyses were performed to reveal the function of DEGs and the results are shown in Figure
The top 10 enriched Gene Ontology terms in biological process (a), cellular component (b), molecular function (c) and enriched Kyoto Encyclopedia of Genes and Genomes pathway (d) of DEGs. DEG: differentially expressed gene.
Go enrichment analysis revealed that DEG-related biological processes (BP) were mostly enriched in antimicrobial humoral response (
To investigate the crucial pathways of these DEGs, KEGG pathways analysis was performed and the significant pathways are shown in Figure
Using the STRING database, we performed PPI network analysis for DEGs. As we have shown in Figure
Protein-protein interaction network analysis (a) and hub genes (b) identified by Cytoscape.
In order to further identify important miRNA-mRNA-regulated axes in IPAH, four databases (miRDB, TargetScan, TargetMiner, and miRWalk) were employed to predict the target genes of DEMIs. 9095 genes were predicted as target genes of 6 DEMIs. GO and KEGG analyses of those genes were performed with results shown in Supplement Figure
DEMI-DEG network. Rhombus represents miRNA while rectangle represents mRNA. DEG: differentially expressed gene; DEMI: differentially expressed miRNA.
Details of the 16 DEGs included in DEMI-DEG network.
Gene | Log2FC | Adjusted | |
---|---|---|---|
HBB | ENSG00000244734 | 2.1829 | 3.26 |
COL14A1 | ENSG00000187955 | 1.1296 | 3.55 |
WIF1 | ENSG00000156076 | 1.3922 | 3.82 |
OGN | ENSG00000106809 | 1.0880 | 1.34 |
RGS1 | ENSG00000090104 | 1.2087 | 1.95 |
AQP9 | ENSG00000103569 | -1.3423 | 2.29 |
ASPN | ENSG00000106819 | 1.3752 | 2.96 |
ESM1 | ENSG00000164283 | 1.1911 | 3.45 |
SFRP2 | ENSG00000145423 | 1.3301 | 3.83 |
ENPP2 | ENSG00000136960 | 1.1842 | 9.72 |
SAA1 | ENSG00000173432 | -1.1547 | 0.0002 |
EDN1 | ENSG00000078401 | 1.0631 | 0.0006 |
VCAM1 | ENSG00000162692 | 1.0820 | 0.0006 |
SOSTDC1 | ENSG00000171243 | -1.1114 | 0.0014 |
CCDC80 | ENSG00000091986 | 1.0089 | 0.0024 |
SPP1 | ENSG00000118785 | -1.2404 | 0.0026 |
DEG: differentially expressed gene; DEMI: differentially expressed miRNA; FC: fold change.
Details of the 4 DEMIs included in the DEMI-DEG network.
Gene | miRBase | Log2FC | |
---|---|---|---|
hsa-miR-205-5p | MIMAT0000266 | 1.2088 | 0.0038 |
hsa-miR-199a-3p | MIMAT0000232 | 1.2975 | 0.0109 |
hsa-miR-34b-5p | MIMAT0000685 | 1.2421 | 0.0270 |
hsa-miR-26b-5p | MIMAT0000083 | 1.1240 | 0.0418 |
DEG: differentially expressed gene; DEMI: differentially expressed miRNA; FC: fold change.
The 12 upregulated DEGs in the DEMI-DEG network were uploaded to the L1000 platform to search for potential drugs, and the top 20 ranked by CMap connectivity score are listed in Table
The top 10 and bottom 10 chemical compounds identified by L1000 platform.
Name | CMap connectivity score | Description |
---|---|---|
Nimodipine | 99.89 | Calcium channel blocker |
GW-501516 | 99.82 | PPAR receptor agonist |
Icilin | 99.82 | TRPV agonist |
Fexofenadine | 99.82 | Histamine receptor antagonist |
Modafinil | 99.79 | Adrenergic receptor agonist |
Guanabenz | 99.79 | Alpha-2 selective adrenergic agonist |
Mianserin | 99.75 | Serotonin receptor antagonist |
CGP-53353 | 99.75 | EGFR inhibitor |
TUL-XXI039 | 99.72 | Serine/threonine kinase inhibitor |
Diphenoxylate | 99.68 | Opioid receptor agonist |
Huperzine-a | -99.46 | Acetylcholinesterase inhibitor |
Perindopril | -99.47 | ACE inhibitor |
Solanine | -99.47 | Acetylcholinesterase inhibitor |
Phenothiazine | -99.51 | Dopamine receptor antagonist |
Tyrphostin-AG-1295 | -99.68 | PDGFR receptor inhibitor |
Y-27152 | -99.72 | Potassium channel activator |
Lenalidomide | -99.79 | Antineoplastic |
Androstenedione | -99.89 | Cytochrome P450 inhibitor |
Repaglinide | -99.89 | Insulin secretagogue |
Neurodazine | -99.89 | Neurogenesis of nonpluripotent C2C12 myoblast inducer |
Though new therapies have been developed in recent years, IPAH remains a severe disease with continuous suffering to patients and society. Therefore, better understanding of potential mechanisms in the pathogenesis of IPAH may shed a light for future studies.
In the present study, 30 DEGs, mostly related to immune and inflammatory response such as neutrophil chemotaxis and migration, integrin binding and Toll-like receptor binding, and significantly enriched in inflammation pathways like IL-17 signaling pathway, and 6 DEMIs were found in IPAH. The DEMI-DEG network was conducted subsequently with 4 DEMIs (hsa-miR-34b-5p, hsa-miR-26b-5p, hsa-miR-205-5p, and hsa-miR-199a-3p) and 16 DEGs. We found that 5 DEGs (AQP9, SPP1, END1, VCAM1, and SAA1) were included in the top 10 hub genes of the PPI network, indicating those factors may play important roles in the pathogenesis of IPAH. Finally, drugs including nimodipine were identified. To our knowledge, this is the first study that conducted the network of mRNA and miRNA and predicted potential drugs in IPAH, which provides a foundation for further research and development of therapy.
Previous studies have revealed the important role of the miRNAs and mRNAs that is identified in our DEMI-DEG network in the pathogenesis of IPAH. Wu et al. identified dysregulated miRNAs in IPAH via miRNA profiling and qRT-PCR. miR-26b-5p and miR-199a-3p were detected as two of the top 5 most significantly increased miRNAs targeting major PAH related pathways including Wnt/
However, some of the factors in our DEMI-DEG network were poorly studied. The role of hsa-miR-34b-5p, which was connected with most of the predicted target DEGs, has not been studied in IPAH before. miR-34b was known to affect cell proliferation and adhesion-independent growth in several types of cancer [
The calcium channel blocker is an effective treatment for IPAH patients with a positive vasodilator response, which dramatically improves the survival of those patients and has been widely applied recent years [
There are some limitations in our study. Firstly, only two mRNA expression profiles and one miRNA expression profile were found in GEO; the relatively small number of samples may make the results less convincing. Secondly, since the expression profiles of miRNA and mRNA were obtained from different datasets in this study, coexpression analysis cannot be performed currently, which prevents us from obtaining a more accurate miRNA-mRNA regulation network. In addition, PCR or WB should be done to further confirm our results, and the relationship between miRNAs and mRNAs in our DEMI-DEG network should be validated via
Our study initially conducted a miRNA-mRNA network including 4 miRNAs and 5 mRNAs to systematically analyze the pathogenesis of IPAH and provided a new insight for future therapies by predicting potential drugs acting with hub genes.
Idiopathic pulmonary arterial hypertension
microRNA
Differentially expressed genes
Differentially expressed miRNAs
Gene Expression Omnibus
Gene Ontology
Kyoto Encyclopedia of Genes and Genomes
Protein-protein interaction
Biological processes
Cellular components
Molecular function
Pulmonary artery pressure
Pulmonary vascular resistance
Phosphodiesterase type 5
Pulmonary vascular smooth muscle cell
Endothelin-1
Vascular cell adhesion molecule 1
Aquaporin 9
Secreted phosphoprotein 1
Serum amyloid A1.
All data generated or analysed during this study are included in this published article [and its supplementary information files].
The authors declare that they have no competing interests.
All authors made substantive intellectual contributions to this study to qualify as authors. Ruizheng Shi conceived of the design of the study. Chan Li performed the study, collected the data, and contributed to the design of the study. Chan Li and Zeyu Zhang drafted the manuscript. Qian Xu revised the manuscript. All authors read and approved the final manuscript.
The work was supported by grants from the National Nature Science Foundation of China (No. 81570453 to S.R.Z.) and the Chinese Cardiovascular Association V.G foundation (No. 2017-CCA-VG-005 to S.R.Z).
Supplement Table 1: the 30 DEGs in the lung tissue of patients with IPAH. Supplement Table 2: the 6 DEMIs in the lung tissue of patients with IPAH. Supplement Figure 1: the top 10 enriched Gene Ontology terms in biological process (A), cellular component (B), molecular function (C), and enriched Kyoto Encyclopedia of Genes and Genomes pathway (D) of the targets of DEMIs. DEMIs: differential expressed miRNAs.