Exosome-Mediated Delivery of Inducible miR-423-5p Enhances Resistance of MRC-5 Cells to Rabies Virus Infection
Abstract
:1. Introduction
2. Results
2.1. Blocking Exosomal Release Promotes RABV Infection in MRC-5 Cells
2.2. RABV Infection Alters miRNA Expression Patterns in Exosomes
2.3. miR-423-5p Targets Suppressor of Cytokine Signaling 3 (SOCS3)
2.4. miR-423-5p Feedback Inhibits RABV Replication in MRC-5 Cells
2.5. RABV Infection Induces Up-Regulation of IFN-β in MRC-5 Cells
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Exosome Isolation
4.3. Exosome Labeling and Uptake Analysis
4.4. Small RNA Deep Sequencing Analysis
4.5. Transient Transfection of siRNAs and miRNAs
4.6. Luciferase Reporter Assays
4.7. RNA Extraction and Quantitative RT-PCR
4.8. Western Blotting
4.9. Transmission Electron Microscopy
4.10. Nanoparticle Tracking Analysis
4.11. Immunofluorescence Microscopy
4.12. Statistical Analysis
Supplementary Materials
Author Contributions
Acknowledgments
Conflicts of Interest
Abbreviations
DAPI | 4′,6-diamidino-2-phenylindole |
DiO | dioctadecyloxacarbocyanine perchlorate |
Exo | exosome |
HCV | hepatitis C virus |
HDCs | human diploid cells |
IFN | interferon |
JAK-SATA | Janus Kinase-Signal Transducers and Activators of Transcription |
KSHV | Kaposi’s sarcoma-associated herpesvirus |
miRNA | microRNA |
MRC-5 cells | human embryonic lung fibroblast cells |
Mut | mutant |
MVB | multivesicular body |
NTA | nanoparticle tracking analysis |
RABV | rabies virus |
siRNA | small interfering RNA |
SOCS3 | suppressor of cytokine signaling 3 |
SSI | STAT-induced STAT inhibitor |
TEM | transmission electron microscopy |
WT | wild-type |
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Gene | Forward Primer (5′-3′) | Reverse Primer (5′-3′) |
---|---|---|
Rabies virus N protein | CAAGATGTGTGCYAAYTGGAG | AGCCCTGGTTCGAACATTCT |
IFN-β | GTCTCCTCCAAATTGCTCTC | ACAGGAGCTTCTGACACTGA |
STAT1 | TTCTGTGTCTGAAGTGTAAGTGAA | TAACACGGGGATCTCAACAAGTTC |
OAS1 | AGAAGGCAGCTCACGAAACC | CCACCACCCAAGTTTCCTGTA |
IRF7 | GAGCCCTTACCTCCCCTGTTAT | CCACTGCAGCCCCTCATAG |
18S | CTTAGAGGGACAAGTGGCG | ACGCTGAGCCAGTCAGTGTA |
miR-423-5p | GGGGTGAGGGGCAGAGAG | universal reverse primers |
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Wang, J.; Teng, Y.; Zhao, G.; Li, F.; Hou, A.; Sun, B.; Kong, W.; Gao, F.; Cai, L.; Jiang, C. Exosome-Mediated Delivery of Inducible miR-423-5p Enhances Resistance of MRC-5 Cells to Rabies Virus Infection. Int. J. Mol. Sci. 2019, 20, 1537. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms20071537
Wang J, Teng Y, Zhao G, Li F, Hou A, Sun B, Kong W, Gao F, Cai L, Jiang C. Exosome-Mediated Delivery of Inducible miR-423-5p Enhances Resistance of MRC-5 Cells to Rabies Virus Infection. International Journal of Molecular Sciences. 2019; 20(7):1537. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms20071537
Chicago/Turabian StyleWang, Jingyu, Yawei Teng, Guanshu Zhao, Fang Li, Ali Hou, Bo Sun, Wei Kong, Feng Gao, Linjun Cai, and Chunlai Jiang. 2019. "Exosome-Mediated Delivery of Inducible miR-423-5p Enhances Resistance of MRC-5 Cells to Rabies Virus Infection" International Journal of Molecular Sciences 20, no. 7: 1537. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms20071537