DOI: https://doi.org/10.21203/rs.3.rs-1772789/v1
Vanadium, one of the metal family members, is widely applied all over the world duo to the excellent property. However, vanadium exposure attracted more attention recently due to the adverse impacts on human health, especially the lung [1-7]. For instance, the long-term exposure of vanadium dioxide (VO2) caused an inhibition of lung cell lines proliferation. In A549 cells, the exposure of VO2 resulted in the genotoxicity, including micronuclei and DNA damage by decreasing the reduced glutathione generation and improving the reactive oxygen species production [1]. In addition, the cytotoxicity induced by VO2 was dose-dependent in lung cell. The VO2 exposure caused the cell viability loss as well as the proliferation and apoptosis inhibition [2]. Vanadium exposure was in connection with the lower DNA methylation of IFNγ, NOS2A and IL4 genes in lung cells [3]. Chronic exposure of vanadium pentoxide (V2O5) resulted in alveolar/bronchiolar tumors [4]. KRAS mutations have been detected in V2O5-exposed mice [4]. The V2O5 incubation caused the genotoxicity in lung cells and inducted the airway inflammation and fibrosis [5]. The vanadium exposure caused the iron homeostasis disorder in lung cell [6]. After the vanadium oxide (V2O3) exposure, the viability was reduced by almost 90% in the epi- and endothelial lung cells [7]. V2O3 incubation led to an increase in the expression of HO1 gene in ECV304 cells, but a reduction in the HO1 protein level in A549 cells [7].
Few studies have shown that certain genes and signaling pathways were related to vanadium exposure-induced lung injury. Turpin et al reported that V2O5 exposure obviously enhanced the expression of IFNα, CTGF, CXCL10, TGFβ1, CXCL9, PDGFC, IFNβ, and COL1A2 genes in lung [5]. V2O5 induced the phosphorylation of STAT1 protein mediated by IFNβ protein in lung fibroblasts. V2O5 incubation also increased CXCL10 expression which was a STAT1-dependent chemokine [8]. The DNA-binding proteins (STAT1 and GAS1), chemokines (CXCL9, CXCL10, and IL8), growth factors (HBEGF, VEGF, and CTGF), and PIPOX, SOD2, and OXR1 associated with the immune responses and inflammation induced by V2O5 exposure in lung fibrotic [9]. MAPK and NF-κB pathways mediated V2O5-induced mucin production in chronic airway diseases [10]. V2O5 exposure upregulated HB-EGF via ERK and p38-MAPK pathways in human lung fibroblasts [11]. Vanadium exposure caused the pathological outcomes (such as abnormal cytokine expression, cell apoptosis and proliferation) and the generation of hydrogen peroxide by activating the p38, MAPK, and EGF pathways in pulmonary cell types [12]. However, the mechanism underlying vanadium exposure-induced pulmonary toxicity is unclear. Therefore, our study was designed to reveal the core genes and the associated enriched signaling pathways link to the vanadium exposure-induced pulmonary toxicity.
Cell Culture
Human bronchial epithelial cells (HBECs) BEAS-2B were maintained in Dulbecco’s Modified Eagle’s Medium (GIBCO) at the temperature of 37 ℃ in a humidified air with 5% CO2. 10.0 mM soluble sodium metavanadate (SMV, Sigma) was supplemented into the sterilized media. The culture medium was changed every 48 hours [13].
Growth in Soft Agar
HBECs were rinsed with polybutylene succinate to remove SMV from the culture media after 30 days of SMV exposure. Then HBECs were seeded in DIFCO BACTO agar (Sigma) following the standard protocols. HBECs were seeded in soft agar until individual isolated colonies were large enough to collect. After an additional three weeks, three control and five transformed colonies were selected for RNA extraction [13].
RNA Extraction and Microarray Hybridization
Total RNA extraction was implemented with the Trizol reagent according to the manufacturer instructions. The cDNA probes were subjected to hybridization with GeneChip Human Gene 1.0 ST Array [13].
Screening of Differentially Expressed Genes
To identify the differentially expressed genes (DEGs) in HBECs treated with and without SMV, GEO2R (http://www.ncbi.nlm.nih.gov/geo/geo2r) was used to analyze the data from eight HBEC samples, including the five 10.0 mM SMV-treated (GSM898741, GSM898742, GSM898743, GSM898744, and GSM898745) and three control samples (GSM898718, GSM898719, and GSM898720) collected after 30-day cell culture. The DEGs was sorted out by the thresholds of P < 0.05 with |log2Fold Change (FC)| > 1. The significantly up- and down-regulated genes were stored in Excel files.
Gene Ontology
To understand the selected DEGs better, gene ontology (GO) analysis for DEGs was implemented using DAVID (https://david.ncifcrf.gov/summary.jsp).
KEGG analysis
To reveal the signing pathways associated with the selected DEGs, KOBAS 3.0 (http://kobas.cbi.pku.edu.cn/kobas3/genelist/) was implemented KEGG analysis.
Protein Classification and Reactome Analysis
Protein classification for DEGs was performed with the PANTHER (http://pantherdb.org/). Moreover, Reactome analysis for DEGs was also employed using KOBAS 3.0.
Protein-Protein Interaction Network
Protein-protein interaction network (PPI) and its further visualization for the DEGs were performed with STRING database (https://string-db.org/) and Cytoscape 3.8.0 software (http://www.cytoscape.org/), respectively.
Hub Genes and Their Functions
CytoHubba (http://apps.cytoscape.org/apps/cytohubba) were used for the identification of hub genes, and hub gene functions were summarized with GeneCards (https://www.genecards.org/), STRING, NCBI, and the previous literature.
Ethics Statement
The procedures used in this study were approved by Institutional Animal Care and Use Committee of Anhui Science and Technology University (ECASTU-2015-P08).
Outline of Transcripts and Genes in HBECs
A total of 32,113 transcripts and 18,809 genes were screened out from the HBECs in both SMV-treated and control groups (Supplementary file 1 and 2). Fig. 1A-F indicated the UMAP, expression density, probes distribution, mean-variance trend, moderated T statistic, and the volcano plot of the DEGs, respectively.
Totally 513 differentially expressed transcripts (Supplementary file 3; 232 down-regulated and 281 up-regulated) and 455 DEGs (Supplementary file 4; 201 down-regulated and 254 up-regulated) were identified. The top 25 up- and down-regulated genes in SMV-treated HBECs were shown in Tables 2 and 3, respectively.
GO Enrichment for DEGs
As indicated in Fig. 2A and Supplementary file 5, upregulated genes might be related to cell adhesion, RNA processing, angiogenesis, response to xenobiotic stimulus, the regulation of cell proliferation, and cell proliferation. Also, down-regulated genes were likely to involve signal transduction, response to drug, cell-cell adhesion, cell adhesion, transcription regulation, NF-κB transcription factor activity, and mitochondrion localization (Fig. 2B and Supplementary file 6). In addition, the expression profiles of the DEGs in cell adhesion, cell migration, the positive regulation of cell migration, the negative regulation of cell proliferation, the regulation of cell proliferation, and the positive regulation of cell proliferation were visualized in Fig. 2C-G with heatmaps, respectively.
KEGG Enrichment for DEGs
Upregulated genes in SMV-treated HBECs mainly participated in metabolic pathway, MAPK, Wnt, Rap1, ErbB, FoxO, cAMP, Ras, PI3K-Akt, relaxin, apelin, calcium, phagosome, axon guidance, focal adhesion, non-small cell lung cancer (NSCLC), transcriptional misregulation in cancer, regulation of actin cytoskeleton, drug metabolism-cytochrome P450, complement and coagulation cascades, small cell lung cancer (SCLC), pancreatic secretion, thyroid hormone, vascular smooth muscle contraction, glycerolipid metabolism, and cortisol synthesis and secretion signaling pathways (Fig. 3A and Supplementary file 7).
Also, as shown in Fig. 3B and Supplementary file 8, the downregulated genes in SMV-treated HBECs mainly related to oxytocin, Wnt, relaxin, calcium, PPAR, thyroid hormone, metabolic pathways, NOD-like receptor, axon guidance, NF-κB, CAMs, transcriptional misregulation in cancer, glutathione metabolism, focal adhesion, mucin type O-glycan biosynthesis, cholinergic synapse, glutamatergic synapse, platelet activation, fluid shear stress and atherosclerosis, arginine biosynthesis, lysine degradation, basal cell carcinoma, acute myeloid leukemia, and long-term potentiation.
Moreover, Fig. 3C-I represented the heatmaps of the DEGs in MAPK, metabolic pathways, NSCLC, PI3K-Ak, SCLC, CAMs, and PPAR signaling pathways, respectively.
Reactome enrichment and Protein classification for DEGs
The upregulated DEGs had a vital role in many signaling pathways, including metabolism, GPCR downstream signaling, developmental biology, signaling by receptor tyrosine kinases, extracellular matrix organization, disease, Axon guidance, hemostasis, signaling by GPCR, metabolism of lipids, diseases of signal transduction, GPCR ligand binding, L1CAM interactions, neuronal system, ECM proteoglycans, platelet activation, signaling and aggregation (Fig. 4A).
The down-regulated DEGs played an important role in gene expression, immune system, signal transduction, metabolism, metabolism of proteins, innate immune system, developmental biology, neuronal system, neutrophil degranulation, adaptive immune system, hemostasis, interferon signaling, transmission across chemical synapses, extracellular matrix organization, G alpha (i) signalling events, inositol phosphate metabolism, G-protein mediated events, peptide hormone metabolism, opioid signaling, and stimuli-sensing channels (Fig. 4B).
The upregulated DEGs in SMV-treated HBECs were mainly involved in integrin, metalloprotease, intercellular signal molecule, g-protein coupled receptor, extracellular matrix protein, cell adhesion molecule, protease, growth factor, membrane-bound signaling molecule, histone modifying enzyme, lambda repressor-like transcription factor, and oxidase (Fig. 4C).
Down-regulated DEGs in SMV-treated HBECs played a significant role in cell adhesion molecule, cadherin, DNA-binding transcription factor, C2H2 zinc finger transcription factor, gene-specific transcriptional regulator, glycosyltransferase, Zinc finger transcription factor, immunoglobulin superfamily cell adhesion molecule, actin binding motor protein, general transcription factor, ion channel, and membrane-bound signaling molecule (Fig. 4D).
Integration of the PPI Network
To further discover the key genes linked to SMV-induced pulmonary toxicity, PPI networks of the DEGs were constructed. as shown in Fig. 5A and B, a set of upregulated genes (including LAMC2, ITGA5, LAMB1, LOX, ITGB3, ITGA2, HSPG2, COL5A2, MMP2, and LAMB3, et al) and downregulated genes (including GLUL, BIRC3, IL7R, BRCA1, PAX6, WT1, GPX7, and KAT2A, et al) might play a vital role in SMV-induced pulmonary toxicity.
Hub genes and their functions
The hub genes associated with SMV-induced pulmonary toxicity were obtained based on the MCC method. As shown in Fig. 6A and Table 4, the top 20 hub genes upregulated in SMV-treated HBECs included ITGA5, ITGB3, ITGA2, LAMB1, LAMC2, HSPG2, COL5A2, MMP2, LOX, LAMB3, SNAI2, ACTA2, LOXL2, ITGBL1, TGFBR2, LOXL1, JAG1, WNT5A, LTBP1, and EREG. In addition, the top 20 hub genes downregulated in SMV-treated HBECs included ICAM1, ITGA4, ASS1, COL1A1, NCAM1, IL7R, BRCA1, PAX6, GLUL, BIRC3, WT1, GPX7, KAT2A, NPPB, ALDH2, NRXN3, E2F5, SYDE2, SULF1, and ACTC1 (Fig. 6B and Table 5).
GO enrichment suggested that the top 20 upregulated hub genes played a key role in animal organ morphogenesis, endodermal cell differentiation, peptidyl-lysine oxidation, collagen fibril organization, angiogenesis, cell migration, cell-substrate adhesion, substrate adhesion-dependent cell spreading, keratinocyte differentiation, and the response to xenobiotic stimulus, et al (Fig. 6C).
Similarly, the top 20 downregulated hub genes participated in the positive regulation of gene expression, response to drug, regulation of cell cycle, response to ionizing radiation, kidney development, negative regulation of centriole replication, diaphragm development, glomerular basement membrane development, cell adhesion, leukocyte migration, positive regulation of vascular endothelial growth factor production, leukocyte cell-cell adhesion, embryonic skeletal system development, and blood vessel development (Fig. 6D).
The top 20 upregulated hub genes participated in various signing pathways, such as PI3K-Akt, TGF-beta, Hippo, apelin, MAPK, toxoplasmosis, focal adhesion, endocrine resistance, platelet activation, SCLC, ECM-receptor interaction, relaxin, phagosome, regulation of actin cytoskeleton, osteoclast differentiation signaling pathways (Fig. 8E).
Meanwhile, the top 20 down-regulated Hub genes had a major role in PI3K-Akt, NF-κB, TNF, metabolic pathways, focal adhesion, arginine biosynthesis, alanine, aspartate and glutamate metabolism, platinum drug resistance, biosynthesis of amino acids, ECM-receptor interaction, fluid shear stress and atherosclerosis, and necroptosis signaling pathways (Fig. 8F).
Hub Genes associated with SMV-induced Pulmonary Toxicity
Lung is an important respiratory organ for humans and animals, and its health is directly related to many life activities. In recent years, vanadium exposure inducted-pulmonary toxicity, especially lung cancer, has attracted an increasing attention [14, 15]. However, the mechanism remains unclear. In our study, a set of genes, such as ITGA5, ITGB3, ITGA2, LAMC2, MMP2, ITGA4, were proven to be involved in SMV-induced pulmonary toxicity. For example, ITGA5 was a prognostic biomarker for non-small-cell and squamous cell lung cancer [16, 17]. ITGB3 could improve the cell migration, proliferation, and invasion in lung cancer [18, 19].
ITGA2, named integrin alpha-2, has a vital role in the proliferation and apoptosis of lung cancer cells. In this study, ITGA2 expression was increased in SMV-treated HBECs, which was consistent with the previous report that ITGA2 was closely with NSCLC cell proliferation and apoptosis, and the suppressed expression of ITGA2 gene could impede the proliferation and triggered the apoptosis of NSCLC cell [20]. Ren et al also reported that the methylation-associated genes (including ITGA2) and CD8 T cell-related genes might be the prognostic factors for lung adenocarcinoma [21].
MMP2 (matrix metalloproteinase-2) might be link to lung cancer proliferation and metastasis, and lung injury. In our study, we found that MMP2 was significantly up-regulated in SMV-treated HBECs and it was involved in many functions such as tissue repair, inflammation, vasculature remodeling, angiogenesis, and tumor invasion (Table 4). The finding was consistent with the previous reports [22-24]. MMP2 might be closely associated with the lung cancer cell proliferation and metastasis [22]. MMP2 also played a main regulatory role in the cytotoxicity induced by amyloid-beta in lung cancer cells [23]. Moreover, MMP2 blockade contribute to the protection against transient receptor potential vanilloid 4-induced lung injury [24].
COL1A1, a fibril-forming collagen found in most connective tissues (Table 4), was an important biomarker lung injury and diseases. Geng et al [25] reported that COL1A1 was closely related to the overall survival, and the cell infiltration level in the lung cancer patients. The cell type included CD4+ T, dendritic, neutrophil, and macrophage cell [25]. In NSCLC cell and tissue, COL1A1 was upregulated. COL1A1 overexpression could increase the cell proliferation and viability, decrease apoptosis rate and the expression of apoptosis-associated protein BAX and PTEN in NSCLC cell [26].
Signaling Pathways related to SMV-induced Pulmonary Toxicity
Previous studies have shown that diverse signaling pathways were related to SMV-induced pulmonary toxicity [23, 27-30]. In our study, we found that MAPK, NF-κB, PI3K-Akt, Wnt, Rap1, and PPAR signaling pathways associated with SMV-induced pulmonary toxicity. As mentioned in the previous study, PI3K-Akt and NF-κB signaling pathways might be involved in lung cancer and cell injury [23]. In A549 and H1299, the up-regulation of phosphorylation levels of core proteins in the PI3K/Akt/mTOR pathway obviously enhanced the cell migration, invasion, and proliferation, whereas down-regulation of phosphorylation levels inhibited the above capacities [27]. LY294002, a PI3K inhibitor, could block the PI3K/Akt and aggravate the fine particulate matter-induced lung injury, increase inflammation, pulmonary edema and the TNFα and IL-1β levels in the bronchoalveolar lavage fluid, upregulate the expression of 4-hydroxynonenal, and downregulate the ferroptosis-associated proteins (GPX4, NRF2, and SLC7A11) [28]. PI3K/Akt/GSK3 beta/Snail pathway mediated the miR-4732-5p effects on lung cancer cells, such as an inhibition of the invasion, migration, and metastasis in vivo and in vitro [29]. PI3K/AKT pathway played a vital role in the process of Agrin promoting Treg infiltration and cell growth, increasing IL6 protein expression and secretion in NSCLC [30].
NF-κB signaling pathways mediated the lung cell metabolism and the injury caused by the multiple harmful exposures and. For instance, NF-κB was found significantly up-regulated in the lung exposed to radiation [31]. Suppression of NF-κB relieved the lung injury by inhibiting the inflammatory and pro-fibrotic cytokines, including IL6, TNFα, TGFβ1, IL18, and IL1β [31]. NF-κB inhibition prevented the perivascular infiltration and prolonged the inflammatory cascade which could resulted in the chronic radiation fibrosis [31]. NF-κB pathway inhibition relieved the acute lung injury (ALI) and reduced the lung inflammatory response, such as a decrease in the number of neutrophils and total cells in the bronchoalveolar lavage fluid (BALF), the BALF protein concentration, and the MPO activity [32]. NF-κB signaling could negatively regulate the expression of OVOL2 which was downregulated in NSCLC cells. In addition, OVOL2 overexpression significantly compromise the GLUT1 translocation and aerobic glycolysis induced by NF-κB signaling in NSCLC cells [33]. Inhibition of NF-κB and TGF-beta/pSmad2 pathways could effectively relieve the injury and restrain the inflammation in the irradiated lung [34].
MAPK signaling pathway closely linked to lung injury, and the subsequent oxidative stress and inflammation in lung cells. MAPK/ NF-κB signaling pathway mediated the protective effects of Hydnocarpin D on ALI, including ameliorating the lung tissues histological alterations, and decreasing inflammatory cell infiltration, lung edema, cytokines secretion, and the content of protein in the bronchoalveolar lavage fluid [35]. In lipopolysaccharide (LPS)-induced ALI, MAPK/NF-κB signaling pathway and IL-17 activation played a vital role in lipopolysaccharide and inflammatory cells infiltration induced by LPS [36]. MAPK pathway was associated with the oxygen deprivation/reoxygenation-induced lung epithelial cells injury characterized by an increase of cell apoptosis and the inflammatory cytokines release [37].
Wnt pathway linked to cell proliferation and invasion in NSCLC. Wnt/β-catenin pathway activation partially reversed the suppression of cell proliferation, migration, and invasion elicited by the LRP8 depletion [38]. In ALI, the expression of three key proteins (β-catenin, Wnt5a, and APC) in Wnt/β-catenin pathway, was positively correlated with the number of inflammatory factors in bronchoalveolar lavage fluid. In addition, the expression of three key protein was obviously decreased following the reducing degree of lung injury [40]. Wnt/β-catenin suppression and NF-κB activation mediated the alleviated the ALI, cell apoptosis and inflammation [41]. Wnt/β-catenin pathway activation obviously relieved the lung inflammation induced by LPS, promoted Th17 response, and increased pro-inflammatory cytokines production and neutrophils infiltration [42].
In conclusion, multiple signaling pathways (MAPK, NF-κB, Wnt, and PI3K-Akt signaling pathways) and the core genes, including ITGA5, ITGB3, ITGA2, LAMC2, MMP2, and ITGA4, might contribute to SMV-induced pulmonary toxicity. Our findings provided the valuable insights for the identification of novel biomarkers in SMV-induced pulmonary toxicity.
Authors’ contributions
B Yang designed the study and analyzed the results. XF Li prepared the manuscript. AE Abdel-Moneim visualized the results. The authors reviewed the manuscript.
Funding
This work was funded by the Talent Introduction Program of Anhui Science and Technology University (No. DKYJ202003).
Availability of Data and Materials
All data of the present article are included in Supplementary Information files or in the main text.
Interest conflict
There are no competing interests.
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Table 1. The transcripts and genes differentially expressed in SMV-treated HBECs
|
Items |
Total |
Down-regulated |
Up-regulated |
|
Transcripts |
513 |
281 |
232 |
|
Genes |
455 |
254 |
201 |
Table 2. The top 25 significantly up-regulated genes in SMV-treated HBECs
|
Gene symbol |
Log2FC |
P Value |
Description |
|
TMEM47 |
5.87 |
5.54E-07 |
Transmembrane protein 47 |
|
RPS4Y1 |
5.63 |
8.90E-10 |
Ribosomal protein S4, Y-linked 1 |
|
EYA4 |
5.14 |
5.26E-11 |
EYA transcriptional coactivator and phosphatase 4 |
|
SPTLC3 |
5.09 |
3.78E-07 |
Serine palmitoyltransferase long chain base subunit 3 |
|
EIF1AY |
4.56 |
1.56E-07 |
Eukaryotic translation initiation factor 1A, Y-linked |
|
TMPRSS15 |
4.28 |
1.31E-03 |
Transmembrane protease, serine 15 |
|
DDX3Y |
4.22 |
6.49E-10 |
DEAD-box helicase 3, Y-linked |
|
EDIL3 |
3.91 |
8.92E-05 |
EGF like repeats and discoidin domains 3 |
|
CD24 |
3.81 |
5.03E-05 |
CD24 molecule |
|
UTY |
3.77 |
8.10E-09 |
Ubiquitously transcribed tetratricopeptide repeat containing, Y-linked |
|
GABRA3 |
3.65 |
1.30E-04 |
Gamma-aminobutyric acid type A receptor alpha3 subunit |
|
ZNF730 |
3.48 |
1.12E-06 |
Zinc finger protein 730 |
|
RARB |
3.34 |
4.56E-05 |
Retinoic acid receptor beta |
|
USP9Y |
3.29 |
6.79E-09 |
Ubiquitin specific peptidase 9, Y-linked |
|
CYP1B1 |
3.16 |
1.19E-04 |
Cytochrome P450 family 1 subfamily B member 1 |
|
SEMA3A |
3.08 |
5.35E-04 |
Semaphorin 3A |
|
ZFY |
3.03 |
5.11E-07 |
Zinc finger protein, Y-linked |
|
SNORD116-14 |
3.03 |
1.49E-02 |
Small nucleolar RNA, C/D box 116-14 |
|
ADGRL2 |
2.94 |
7.44E-03 |
Adhesion G protein-coupled receptor L2 |
|
COL5A2 |
2.88 |
1.14E-06 |
Collagen type V alpha 2 chain |
|
TXLNGY |
2.85 |
7.90E-07 |
Taxilin gamma pseudogene, Y-linked |
|
SNORD116-20 |
2.76 |
1.27E-02 |
Small nucleolar RNA, C/D box 116-20 |
|
SNORD116-15 |
2.74 |
1.71E-02 |
Small nucleolar RNA, C/D box 116-15 |
|
DNER |
2.70 |
1.61E-03 |
Delta/notch like EGF repeat containing |
|
WNT5A |
2.66 |
4.46E-05 |
Wnt family member 5A |
Table 3. The top 25 significantly down-regulated genes in SMV-treated HBECs
|
Gene symbol |
Log2FC |
P Value |
Description |
|
DSC3 |
-5.34 |
2.17E-08 |
Desmocollin 3 |
|
ZNF93 |
-3.68 |
6.62E-06 |
Zinc finger protein 93 |
|
ARHGAP28 |
-3.33 |
1.97E-05 |
Rho gtpase activating protein 28 |
|
ZNF788 |
-3.26 |
2.18E-11 |
Zinc finger family member 788 |
|
TNFRSF11B |
-3.17 |
9.39E-05 |
TNF receptor superfamily member 11b |
|
ZNF542P |
-3.15 |
5.44E-09 |
Zinc finger protein 542, pseudogene |
|
GCA |
-3.06 |
3.63E-05 |
Grancalcin |
|
KCNQ5 |
-3.06 |
9.00E-10 |
Potassium voltage-gated channel subfamily Q member 5 |
|
GLUL |
-3.04 |
1.78E-04 |
Glutamate-ammonia ligase |
|
ZC4H2 |
-3.01 |
1.01E-07 |
Zinc finger C4H2-type containing |
|
PAX6 |
-2.96 |
1.89E-08 |
Paired box 6 |
|
MAL2 |
-2.89 |
1.37E-03 |
Mal, T-cell differentiation protein 2 |
|
NFIA |
-2.86 |
2.33E-08 |
Nuclear factor I A |
|
RBP7 |
-2.83 |
3.36E-06 |
Retinol binding protein 7 |
|
MGAT4A |
-2.81 |
2.99E-08 |
Mannosyl (alpha-1,3-)-glycoprotein beta-1,4-N-acetylglucosaminyltransferase, isozyme A |
|
GULP1 |
-2.72 |
1.38E-02 |
GULP, engulfment adaptor PTB domain containing 1 |
|
RRAGD |
-2.67 |
2.37E-04 |
Ras related GTP binding D |
|
RAVER2 |
-2.66 |
1.98E-04 |
Ribonucleoprotein, PTB binding 2 |
|
DIO2 |
-2.65 |
2.45E-04 |
Deiodinase, iodothyronine, type II |
|
ZNF506 |
-2.64 |
3.10E-03 |
Zinc finger protein 506 |
|
SKAP2 |
-2.63 |
7.55E-07 |
Src kinase associated phosphoprotein 2 |
|
OLR1 |
-2.61 |
7.24E-03 |
Oxidized low density lipoprotein receptor 1 |
|
MARK1 |
-2.60 |
1.35E-05 |
Microtubule affinity regulating kinase 1 |
|
TMEM98 |
-2.49 |
1.12E-04 |
Transmembrane protein 98 |
|
TMEM52B |
-2.47 |
1.31E-03 |
Transmembrane protein 52B |
Table 4. The top 20 hub genes up-regulated in SMV-treated HBECs
|
Gene symbol |
Full name |
Functions |
|
ITGA5 |
Integrin alpha-5 |
ITGA5, a receptor for fibronectin and fibrinogen, recognizes the sequence R-G-D in its ligands. ITGA5:ITGB1 acts as a receptor for fibrillin-1 (FBN1) and mediates R-G-D-dependent cell adhesion to FBN1 |
|
ITGB3 |
Integrin beta-3 |
ITGB3 is a receptor for cytotactin fibronectin, laminin, matrix metalloproteinase-2, osteopontin, osteomodulin, prothrombin, thrombospondin, vitronectin and von Willebrand factor. |
|
ITGA2 |
Integrin alpha-2 |
ITGA2 is a receptor for laminin, collagen, collagen C-propeptides, fibronectin and E-cadherin. It is responsible for adhesion of platelets and other cells to collagens, modulation of collagen and collagenase gene expression, force generation and organization of newly synthesized extracellular matrix. |
|
LAMB1 |
Laminin subunit beta-1 |
LAMB1 mediates the attachment, migration and organization of cells into tissues during embryonic development. It is probably required for the integrity of the basement membrane/glia limitans that serves as an anchor point for the endfeet of radial glial cells and as a physical barrier to migrating neurons. |
|
LAMC2 |
Laminin subunit gamma-2 |
LAMC2 can bind to cells via a high affinity receptor. LAMC2 exerts cell- scattering activity toward a wide variety of cells, including epithelial, endothelial, and fibroblastic cells. |
|
HSPG2 |
Heparan sulfate proteoglycan 2 |
HSPG2 is responsible for the fixed negative electrostatic membrane charge, and which provides a barrier which is both size- and charge-selective. It serves as an attachment substrate for cells and plays essential roles in vascularization. |
|
COL5A2 |
Collagen Type V Alpha 2 Chain |
COL5A2, a member of group I collagen, is a minor connective tissue component. Type V collagen binds to DNA, heparan sulfate, thrombospondin, heparin, and insulin. COL5A2 is a key determinant in the assembly of tissue- specific matrices. |
|
MMP2 |
Matrix metalloproteinase-2 |
MMP2, a member of ubiquitinous metalloproteinase, is involved in diverse functions such as remodeling of the vasculature, angiogenesis, tissue repair, tumor invasion, inflammation, and atherosclerotic plaque rupture. |
|
LOX |
Protein-lysine 6-oxidase |
LOX is responsible for the post-translational oxidative deamination of peptidyl lysine residues in precursors to fibrous collagen and elastin. It may play a role in tumor suppression and the aortic wall architecture. |
|
LAMB3 |
Laminin subunit beta-3 |
It mediates the attachment, migration and organization of cells into tissues during embryonic development by interacting with other extracellular matrix components. |
|
SNAI2 |
Zinc finger protein SNAI2 |
SNAI2 is a transcriptional repressor that modulates both activator- dependent and basal transcription. It plays a role in mediating RAF1- induced transcriptional repression of the TJ protein, and subsequent oncogenic transformation of epithelial cells. |
|
ACTA2 |
Actin, aortic smooth muscle |
ACTA2, a member of the actin family, is involved in various types of cell motility and are ubiquitously expressed in all eukaryotic cells. |
|
LOXL2 |
Lysyl oxidase homolog 2 |
LOXL2 mediates the post-translational oxidative deamination of lysine residues on target proteins leading to the formation of deaminated lysine. It also acts as a regulator of sprouting angiogenesis, probably via collagen IV scaffolding. |
|
ITGBL1 |
Integrin subunit beta like 1 |
ITGBL1 encodes a beta integrin-related protein that is a member of the EGF-like protein family. The encoded protein contains integrin-like cysteine-rich repeats. |
|
TGFBR2 |
Transforming growth factor beta receptor 2 |
It regulates a plethora of physiological and pathological processes including cell cycle arrest in epithelial and hematopoietic cells, control of mesenchymal cell proliferation and differentiation, wound healing, extracellular matrix. |
|
LOXL1 |
Lysyl oxidase-like protein 1 |
LOXL1 is essential for the biogenesis of connective tissue, and encoding an extracellular copper-dependent amine oxidase. LOXL1 is proteolytically processed to generate the mature enzyme. |
|
JAG1 |
Protein jagged-1 |
JAG1 is involved in the mediation of Notch signaling, and the cell-fate decisions during hematopoiesis. |
|
WNT5A |
Wnt family member 5a |
WNT5A can activate or inhibit canonical Wnt signaling, depending on receptor context. In the presence of FZD4, it activates beta-catenin signaling. |
|
LTBP1 |
Latent transforming growth factor beta binding protein 1 |
LTBP1 may be involved in the assembly, secretion and targeting of TGFB1 to sites at which it is stored and/or activated. It plays critical roles in controlling and directing the activity of TGFB1 as well as extracellular matrix. |
|
EREG |
Epiregulin |
EREG stimulates EGFR and ERBB4 tyrosine phosphorylation. It contributes to inflammation, wound healing, tissue repair, and oocyte maturation by regulating angiogenesis and vascular remodeling and by stimulating cell proliferation |
Table 5. The top 20 hub genes down-regulated in SMV-treated HBECs
|
Gene symbol |
Full name |
Functions |
|
ICAM1 |
Intercellular adhesion molecule 1 |
ICAM1 is a ligand for the leukocyte adhesion protein LFA-1. During leukocyte trans- endothelial migration, ICAM1 engagement promotes the assembly of endothelial apical cups through ARHGEF26/SGEF and RHOG activation. |
|
ITGA4 |
Integrin alpha-4 |
ITGA4, a receptor for fibronectin, recognizes one or more domains within the alternatively spliced CS-1 and CS-5 regions of fibronectin. |
|
ASS1 |
Argininosuccinate synthase |
ASS1, an enzyme inthe urea cycle, catalyzes the formation of arginosuccinate from aspartate. |
|
COL1A1 |
Collagen type I alpha 1 chain |
COL1A1 is a fibril-forming collagen found in most connective tissues. |
|
NCAM1 |
Neural cell adhesion molecule 1 |
NCAM1 protein is a cell adhesion molecule involved in neuron-neuron adhesion, neurite fasciculation, outgrowth of neurites, etc. |
|
IL7R |
Interleukin 7 Receptor |
IL7R acts as a receptor for thymic stromal lymphopoietin, and it belongs to the type I cytokine receptor family. |
|
BRCA1 |
Breast cancer type 1 susceptibility protein |
BRCA1 specifically mediates the formation of 'Lys-6'-linked polyubiquitin chains and plays a central role in DNA repair by facilitating cellular responses to DNA damage. |
|
PAX6 |
Paired box protein Pax-6 |
PAX6 is a transcription factor with important functions in the development of the eye, nose, central nervous system and pancreas. It regulates specification of the ventral neuron subtypes. |
|
GLUL |
Glutamate-ammonia ligase; |
GLUL belongs to the glutamine synthetase family. It has 2 functions: it catalyzes the production of glutamine and 4-aminobutanoate, the latter in a pyridoxal phosphate-independent manner. |
|
BIRC3 |
Baculoviral IAP repeat-containing protein 3 |
BIRC3 has multi-functional protein which regulates not only caspases and apoptosis, but also modulates inflammatory signaling and immunity, mitogenic kinase signaling and cell proliferation, as well as cell invasion and metastasis. |
|
WT1 |
Wilms tumor protein |
WT1 plays an important role in cellular development and cell survival. It has a tumor suppressor as well as an oncogenic role in tumor formation. |
|
GPX7 |
Glutathione peroxidase 7 |
It protects esophageal epithelia from hydrogen peroxide- induced oxidative stress. It suppresses acidic bile acid-induced reactive oxygen species and protects against oxidative DNA damage and double-strand breaks. |
|
KAT2A |
Histone acetyltransferase KAT2A |
KAT2A functions as a histone acetyltransferase to promote transcriptional activation. |
|
NPPB |
Natriuretic peptides B |
NPPB plays a key role in cardiovascular homeostasis through natriuresis, diuresis, vasorelaxation, and inhibition of renin and aldosterone secretion. |
|
ALDH2 |
Aldehyde dehydrogenase 2 family member |
ALDH2 encodes a mitochondrial isoform, which has a low Km for acetaldehydes, and is localized in mitochondrial matrix. |
|
NRXN3 |
Neurexin-3 |
NRXN3, a kind of neuronal cell surface protein that may be involved in cell recognition and cell adhesion. It may mediate intracellular signaling |
|
E2F5 |
Transcription factor E2F5 |
E2F5 is the transcriptional activator that binds to E2F sites. These sites are present in the promoter of many genes whose products are involved in cell proliferation. E2F5 may mediate growth factor- initiated signal transduction. |
|
SYDE2 |
Rho GTPase-activating protein SYDE2 |
SYDE2 is a GTPase activator for the Rho-type GTPases by converting them to an inactive GDP-bound state. |
|
SULF1 |
Extracellular sulfatase sulf-1 |
SULF1 exhibits arylsulfatase activity and highly specific endoglucosamine-6-sulfatase activity. It can remove sulfate from the C-6 position of glucosamine. |
|
ACTC1 |
Actin, alpha cardiac muscle 1 |
ACTC1 protein, belongs to the actin family, is involved in various types of cell motility and is ubiquitously expressed in all eukaryotic cells. |