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. 2023 Jan 6:9:1050242.
doi: 10.3389/fsurg.2022.1050242. eCollection 2022.

Exosomal miRNA-profiling of pleural effusion in lung adenocarcinoma and tuberculosis

Xuede Zhang  1 , Lingling Bao  2 , Guohua Yu  1 , Haifeng Wang  2
Affiliations

Exosomal miRNA-profiling of pleural effusion in lung adenocarcinoma and tuberculosis

Xuede Zhang et al. Front Surg. .

Abstract

Background: Pleural effusion (PE) caused by lung cancer is prevalent, and it is difficult to differentiate it from PE caused by tuberculosis. Exosome-based liquid biopsy offers a non-invasive technique to diagnose benign and malignant PE. Exosomal miRNAs are potential diagnostic markers and play an essential role in signal transduction and biological processes in tumor development. We hypothesized that exosomal miRNA expression profiles in PE would contribute to identifying its diagnostic markers and elucidating the molecular basis of PE formation in lung cancer.

Methods: The exosomes from PE caused by lung adenocarcinoma (LUAD) and pulmonary tuberculosis were isolated and verified by transmission electron microscopy. The exosomal miRNA profiles were identified using deep sequencing and validated with quantitative real-time PCR (qRT-PCR). We performed bioinformatic analysis for differentially expressed miRNAs to explore how exosomal miRNAs regulate pleural effusion.

Results: We identified 99 upregulated and 91 downregulated miRNAs in malignant pleural effusion (MPE) compared to tuberculous pleural effusion (TPE). Seven differentially expressed miRNAs (DEmiRNAs) were validated by qRT-PCR, out of which 5 (71.4%) were confirmed through sequencing. Gene Ontology (GO) analysis revealed that most exosomal miRNAs target genes were involved in regulating cellular processes and nitrogen compound metabolism. According to the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, the exosomal miRNAs target genes were mainly involved in Fc gamma R-mediated phagocytosis, Rap1 signaling pathway, and breast cancer. The hub genes, including ITGAM, FOXO1, MAPK14, YWHAB, GRIN1, and PRF1, were screened through plug-in cytoHubba. The PFR1 was identified as a critical gene in MPE formation using single-cell sequencing analysis. Additionally, we hypothesized that tumor cells affected natural killer cells and promoted the generation of PE in LUAD via the exosomal hsa-miR-3120-5p-PRF1 axis.

Conclusions: We identified exosomal miRNA profiles in LUAD-MPE and TPE, which may help in the differential diagnosis of MPE and TPE. Bioinformatic analysis revealed that these miRNAs might affect PE generation through tumor immune response in LUAD. Our results provided a new theoretical basis for understanding the function of exosomal miRNAs in LUAD-MPE.

Keywords: bioinformatic analysis; exosomes; lung adenocarcinoma; miRNAs; pleural effusions.

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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
Characterization of exosomes from PE. (A) The exosomes from MPE. (B) The exosomes from TPE.
Figure 2
Figure 2
The distribution, heatmap, and volcano plot of DEmiRNAs. (A) The distribution of DEmiRNAs. (B) Heatmap of DEmiRNAs. (C) Volcano plot of DEmiRNAs.
Figure 3
Figure 3
Qrt–PCR validation of miRNA sequencing results.
Figure 4
Figure 4
Go and KEGG analysis for target genes of DEmiRNAs. (A) BP group for upregulated DEmiRNAs. (B) CC group for upregulated DEmiRNAs. (C) MF group for upregulated DEmiRNAs. (D) KEGG for upregulated DEmiRNAs. (E) BP group downregulated DEmiRNAs. (F) CC group for downregulated DEmiRNAs. (G) MF group for downregulated DEmiRNAs. (H) KEGG for downregulated DEmiRNAs.
Figure 5
Figure 5
Hub genes and modules analysis. (A) PPI for the top 20 ranked hub genes for upregulated DEmiRNAs. (B) Module 1, (C) Module 2, and (D) Module 3 for target genes of upregulated DEmiRNAs. (E) PPI for the top 20 ranked hub genes for downregulated DEmiRNAs. (F) Module 1, (G) Module 2, and (H) Module 3 for target genes of downregulated DEmiRNAs.
Figure 6
Figure 6
The natural killer cell-mediated cytotoxicity pathway.
See this image and copyright information in PMC

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