Related Experiment Video
Updated: Jul 18, 2025
Polarization of M1 and M2 Human Monocyte-Derived Cells and Analysis with Flow Cytometry upon Mycobacterium tuberculosis Infection
Published on: September 18, 2020
Differential expression analysis of miRNAs in macrophage-derived exosomes in the tuberculosis-infected bone
Zhicheng Sun1, Xiaoyang Pang1, Xiyang Wang1
1Department of Spinal Surgery, Xiangya Hospital of Central South University, Changsha, Hunan, China.
Background:
Macrophages play an important role in regulating the course of spinal tuberculosis within the bone microenvironment. This study aimed to investigate the differential expression of miRNA in macrophage-derived exosomes within the tuberculosis-infected bone microenvironment, to identify miRNAs that hold potential as diagnostic markers and therapeutic targets.
Methods:
We established study cohorts for spinal tuberculosis, collected bone marrow blood samples, isolated macrophage exosomes, and performed exosome miRNA sequencing. A miRNA-mRNA co-expression network was constructed using WGCNA analysis. Gene GO analysis and KEGG pathway enrichment analysis were performed using KOBAS software. Target miRNAs were selected based on fold change, P-value, and false discovery rate, and their validation was carried out using qRT-PCR and ROC curve studies. Subsequently, we constructed a target gene network for these miRNAs and performed KEGG pathway enrichment analysis to explore the potential signaling mechanisms involved in regulating the disease course of spinal tuberculosis.
Results:
Our findings revealed that macrophages from the tuberculosis-infected bone microenvironment exhibited an M1 phenotype. The successful extraction of exosomes from macrophage supernatants was confirmed through electron microscopy, particle size analysis, and protein blot analysis. Exosome miRNA-seq demonstrated that 28 miRNAs were up-regulated, while 34 miRNAs were down-regulated in individuals with spinal tuberculosis. GO analysis and KEGG pathway enrichment analysis indicated that the differentially expressed miRNAs were involved in various biological processes, cell components, molecular functions, and signaling pathways, which collectively contribute to the regulation of the disease course of spinal tuberculosis. Notably, miRNA-125b-5p was successfully selected based on fold change, p-value, and false discovery rate. qRT-PCR validation further confirmed the significant up-regulation of miRNA-125b-5p in spinal tuberculosis. The ROC curve revealed that miR-125b-5p is a potential diagnostic biomarker for spinal tuberculosis. Moreover, construction of the miRNA-125b-5p target gene network and subsequent KEGG enrichment analysis highlighted the importance of MAPK, TNF, Ras, Rap1, and the PI3K-Akt signaling pathways in the regulation of the disease course of spinal tuberculosis.
Conclusion:
Our study demonstrates differential expression of miRNAs in macrophage-derived exosomes in the tuberculosis-infected bone microenvironment. Specifically, MiRNA-125b-5p is significantly up-regulated in spinal tuberculosis and shows potential as a diagnostic biomarker for spinal tuberculosis.
Insights
Researchers identified specific microRNAs (miRNAs) in exosomes from macrophages within the bone microenvironment of spinal tuberculosis patients. MiRNA-125b-5p was found to be significantly elevated, showing potential as a diagnostic marker for spinal tuberculosis.
Area of Science:
- Molecular Biology
- Immunology
- Microbiology
Background:
- Macrophages are key regulators in the bone microenvironment during spinal tuberculosis.
- Investigating macrophage-derived exosomal microRNAs (miRNAs) is crucial for understanding disease pathogenesis.
- This study focuses on identifying potential diagnostic markers and therapeutic targets within these exosomes.
Purpose of the Study:
- To investigate differential miRNA expression in macrophage-derived exosomes from the spinal tuberculosis bone microenvironment.
- To identify specific miRNAs with potential as diagnostic biomarkers and therapeutic targets for spinal tuberculosis.
Main Methods:
- Established spinal tuberculosis cohorts and collected bone marrow blood samples.
- Isolated macrophage exosomes and performed exosome miRNA sequencing.
- Utilized WGCNA for miRNA-mRNA co-expression network construction, GO and KEGG analyses for functional enrichment, and qRT-PCR and ROC curves for miRNA validation.
Main Results:
- Macrophages in the infected bone microenvironment exhibited an M1 phenotype.
- Exosome miRNA sequencing revealed 28 up-regulated and 34 down-regulated miRNAs in spinal tuberculosis patients.
- MiRNA-125b-5p was significantly up-regulated, validated by qRT-PCR, and demonstrated potential as a diagnostic biomarker via ROC curve analysis.
Conclusions:
- Differential miRNA expression exists in macrophage-derived exosomes within the spinal tuberculosis bone microenvironment.
- MiRNA-125b-5p is significantly up-regulated in spinal tuberculosis.
- MiRNA-125b-5p holds promise as a diagnostic biomarker for spinal tuberculosis.
