Exploration of Feasible RNA Biomarkers for Real Clinical Practice

Editors

Xiaoyan Wang

Shanghai Jiao Tong University

Tingxiang Yan

Mayo Clinic Florida

Zhougui Ling

Department of Pulmonary and Critical Care Medicine, the Fourth Affiliated Hospital of Guangxi Medical University,

Impact

Original Research

11 May 2023

Identification of hsa-miR-619-5p and hsa-miR-4454 in plasma-derived exosomes as a potential biomarker for lung adenocarcinoma

Linxiang Feng

, 6 more and

Zuojun Shen

Introduction: Lung cancer has long been at the forefront of all cancers in terms of incidence and mortality. Lung adenocarcinoma is the most common type of lung cancer, accounting for 40% of all lung cancer types. Exosomes can act as biomarkers of tumors and thus play an important role.

Methods: In this article, high-throughput sequencing of miRNAs in plasma exosomes from lung adenocarcinoma patients and healthy individuals was performed to obtain 87 upregulated miRNAs, which were then combined with data from the GSE137140 database uploaded by others for screening. The database included 1566 preoperative lung cancer patients, 180 postoperative patients, and 1774 non-cancerous controls. We overlapped the miRNAs upregulated in the serum of lung cancer patients in the database relative to those of non-cancer controls and post-operative patients with the upregulated miRNAs obtained from our next-generation sequencing to obtain nine miRNAs. Two miRNAs that were not reported as tumor markers in lung cancer, hsa-miR-4454 and hsa-miR-619-5p, were selected from them and then validated by qRT-PCR, and further analysis of miRNAs was performed using bioinformatics.

Results: Real-time quantitative PCR showed that the expression levels of hsa-miR-4454 and hsa-miR-619-5p in plasma exosomes of patients with lung adenocarcinoma were significantly up-regulated. The AUC values of hsa-miR-619-5p and hsa-miR-4454 were 0.906 and 0.975, respectively, both greater than 0.5, showing good performance. The target genes of miRNAs were screened by bioinformatics methods, and the regulatory network between miRNAs and lncRNAs and mRNAs was studied.

Discussion: Our work demonstrated that hsa-miR-4454 and hsa-miR-619-5p have the potential to be used as biomarkers for the early diagnosis of lung adenocarcinoma.

2,237 views

5 citations

Validation of the experimentally confirmed target genes predicted from the bioinformatics analysis. (A) The expression of GCM1 significantly decreased after transfection with t0048674 knock down plasmid. (B) The expression of ACTG1 significantly decreased after transfection with t00846456 knock down plasmid. (C–E) Western blot results demonstrating that after transfection with t0048674 knock down plasmid, the corresponding ACTG1 protein was significantly decreased, and after transfection with t00846456 knock down plasmid, the corresponding GCM1 protein was significantly decreased. (F) A bar graph showing that the migration ability of cells transfected with over-expression of t00846456 and t0048674 significantly increased. (G) A trans well graph showing that the migration ability of cells significantly increased after transfection with over-expression of t00846456 and t0048674.

Original Research

23 March 2023

Mitochondrial non-coding RNA in nasopharyngeal carcinoma: Clinical diagnosis and functional analysis

Feng Wang

, 1 more and

Cong Li

1,624 views

0 citations

Original Research

27 February 2023

Identification and validation of metabolism-related hub genes in idiopathic pulmonary fibrosis

Youjie Zeng

, 4 more and

Wen Ouyang

Functional enrichment analysis of DEMRGs. The dot size indicates the number of DEMRGs enriched to the corresponding term, and the dot color indicates the enrichment significance of the corresponding term. (A) The top 10 significantly enriched terms for Gene ontology biological process. (B) The top 10 significantly enriched terms for Gene ontology cellular component (C) The top 10 significantly enriched terms for Gene ontology molecular function. (D) The top 10 significantly enriched terms for the KEGG pathway. (E) The top 10 significantly enriched terms for the Reactome pathway.

Background: Idiopathic pulmonary fibrosis (IPF) is a fatal and irreversible interstitial lung disease. The specific mechanisms involved in the pathogenesis of IPF are not fully understood, while metabolic dysregulation has recently been demonstrated to contribute to IPF. This study aims to identify key metabolism-related genes involved in the progression of IPF, providing new insights into the pathogenesis of IPF.

Methods: We downloaded four datasets (GSE32537, GSE110147, GSE150910, and GSE92592) from the Gene Expression Omnibus (GEO) database and identified differentially expressed metabolism-related genes (DEMRGs) in lung tissues of IPF by comprehensive analysis. Then, we performed GO, KEGG, and Reactome enrichment analyses of the DEMRGs. Subsequently, key DEMRGs were identified by machine-learning algorithms. Next, miRNAs regulating these key DEMRGs were predicted by integrating the GSE32538 (IPF miRNA dataset) and the miRWalk database. The Cytoscape software was used to visualize miRNA-mRNA regulatory networks. In addition, the relative levels of immune cells were assessed by the CIBERSORT algorithm, and the correlation of key DEMRGs with immune cells was calculated. Finally, the mRNA expression of the key DEMRGs was validated in two external independent datasets and an in vivo experiment.

Results: A total of 101 DEMRGs (51 upregulated and 50 downregulated) were identified. Six key DEMRGs (ENPP3, ENTPD1, GPX3, PDE7B, PNMT, and POLR3H) were further identified using two machine-learning algorithms (LASSO and SVM-RFE). In the lung tissue of IPF patients, the expression levels of ENPP3, ENTPD1, and PDE7B were upregulated, and the expression levels of GPX3, PNMT, and POLR3H were downregulated. In addition, the miRNA-mRNA regulatory network of key DEMRGs was constructed. Then, the expression levels of key DEMRGs were validated in two independent external datasets (GSE53845 and GSE213001). Finally, we verified the key DEMRGs in the lung tissue of bleomycin-induced pulmonary fibrosis mice by qRT-PCR.

Conclusion: Our study identified key metabolism-related genes that are differentially expressed in the lung tissue of IPF patients. Our study emphasizes the critical role of metabolic dysregulation in IPF, offers potential therapeutic targets, and provides new insights for future studies.

5,786 views

3 citations

Differential gene expression in Pediatric Crohn’s Disease and controls. (A) Enhanced volcano plot showing the log2 fold change and -log10 p-value for each gene in the dataset. Genes with a significant difference in expression (p < 0.05) are highlighted in red, with those showing an increased expression in Pediatric Crohn’s Disease in blue and decreased expression in green. (B) Principal component analysis (PCA) plot illustrating the separation of Pediatric Crohn’s Disease and control samples based on gene expression levels. Each point represents a sample, with Pediatric Crohn’s Disease samples shown in red and controls in blue. The first two principal components (PC1 and PC2) are plotted, representing the majority of the variation in the data.

Original Research

01 March 2023

Developing a non-invasive diagnostic model for pediatric Crohn’s disease using RNA-seq analysis

Bin He

, 4 more and

Tao Huang

1,802 views

1 citations

Original Research

29 November 2022

Novel targets in renal fibrosis based on bioinformatic analysis

Yuan Yuan

, 2 more and

Pengcheng Luo

The expression of the five hub genes in three datasets. (A) The expression of the five hub genes in dataset 1. (B) The expression of the five hub genes in dataset 2. (C) The mRNA expression level of the hub genes in dataset GSE36496. *p < 0.05, **p < 0.01, ***p < 0.001.

Background: Renal fibrosis is a widely used pathological indicator of progressive chronic kidney disease (CKD), and renal fibrosis mediates most progressive renal diseases as a final pathway. Nevertheless, the key genes related to the host response are still unclear. In this study, the potential gene network, signaling pathways, and key genes under unilateral ureteral obstruction (UUO) model in mouse kidneys were investigated by integrating two transcriptional data profiles.

Methods: The mice were exposed to UUO surgery in two independent experiments. After 7 days, two datasets were sequenced from mice kidney tissues, respectively, and the transcriptome data were analyzed to identify the differentially expressed genes (DEGs). Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were executed. A Protein-Protein Interaction (PPI) network was constructed based on an online database STRING. Additionally, hub genes were identified and shown, and their expression levels were investigated in a public dataset and confirmed by quantitative real time-PCR (qRT-PCR) in vivo.

Results: A total of 537 DEGs were shared by the two datasets. GO and the KEGG analysis showed that DEGs were typically enriched in seven pathways. Specifically, five hub genes (Bmp1, CD74, Fcer1g, Icam1, H2-Eb1) were identified by performing the 12 scoring methods in cytoHubba, and the receiver operating characteristic (ROC) curve indicated that the hub genes could be served as biomarkers.

Conclusion: A gene network reflecting the transcriptome signature in CKD was established. The five hub genes identified in this study are potentially useful for the treatment and/or diagnosis CKD as biomarkers.

2,800 views

6 citations