MiRNA Targeted NP Genome of Live Attenuated Influenza Vaccines Provide Cross-Protection against a Lethal Influenza Virus Infection
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cells, Viruses, and Mice
2.2. Plasmid Construction, Virus Rescue and Multicycle Growth Curve Analysis
2.3. MDCK Cells Transfection
2.4. RT-qPCR
2.5. Immunization and Viral Challenge
2.6. Enzyme-Linked Immunosorbent Assay (ELISA)
2.7. Enzyme-Linked Immune Spot Test (ELISpot)
2.8. Statistics
3. Results
3.1. Construction of Vaccines by Inserted with Different Perfect miRNA Target Sites
3.2. Viruses with Different Perfect miRNA Target Sites Inserted into NP Genome Segment Demonstrate Variable Attenuations in Cells
3.3. Viruses with Different Perfect miRNA Target Sites Inserted into NP Genome Segment Demonstrate Variable Attenuations in Mice
3.4. MiRNA Targeted Viruses of 192t-6 and 192t-9 Provide Protection from Homologous Virus Infection
3.5. MiRNA Targeted Viruses of 192t-6 and 192t-9 Provide Protective Immunity during Lethal Challenge of Homologous Virus
3.6. MiRNA Targeted Viruses of 192t-6 and 192t-9 Provide Protection Against Heterosubtypic Virus Challenge
4. Discussion
Author Contributions
Funding
Conflicts of Interest
References
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Group | Dose (pfu/20 μL) | Lung Virus Titer a (Log10TCID50/mL) | Survival Rate (No. of Survivors/No. Tested) |
---|---|---|---|
192t | 103 | 5.67 ± 0.89 | 0/10 |
104 | 6.60 ± 0.28 | 0/10 | |
192t-3 | 103 | 5.13 ± 0.46 | 0/10 |
104 | 6.33 ± 0.34 | 0/10 | |
192t-6 | 103 | ND | 10/10 |
104 | 2.42 ± 0.49 | 5/10 | |
192t-9 | 103 | ND | 10/10 |
104 | 2.44 ± 0.16 | 5/10 | |
TS IAV | 103 | ND | 10/10 |
104 | 2.39 ± 0.08 | 5/10 | |
ctrl | 103 | 6.10 ± 0.42 | 0/10 |
ctrl-3 | 103 | 5.80 ± 0.76 | 0/10 |
ctrl-6 | 103 | 5.83 ± 0.24 | 0/10 |
ctrl-9 | 103 | 5.47 ± 0.49 | 0/10 |
WT | 103 | 5.90 ± 0.36 | 0/10 |
Group | Virus-Specific Antibody Titer (ELISA, 2n) a | ||||
---|---|---|---|---|---|
Serum IgG | Lung Homogenate IgA | Serum IgG1 | Serum IgG2a | Serum IgG2b | |
10 pfu 192t-6 | 16.8 ± 2.17 | 7.6 ± 0.89 | 12.8 ± 1.79 | 14.4 ± 0.89 | 13.0 ± 1.23 |
100 pfu 192t-6 | 17.6 ± 0.89 | 10.2 ± 0.84 | 16.0 ± 0.71 | 17.6 ± 0.89 | 16.4 ± 0.89 |
1000 pfu 192t-6 | 20.8 ± 0.45 | 13.8 ± 0.96 | 17.2 ± 1.10 | 18.2 ± 0.45 | 17.2 ± 2.17 |
10 pfu 192t-9 | 15.4 ± 4.10 | 7.2 ± 0.84 | 12.0 ± 3.00 | 13.4 ± 1.14 | 12.0 ± 2.12 |
100 pfu 192t-9 | 17.4 ± 1.14 | 10.4 ± 0.55 | 16.8 ± 1.10 | 17.2± 1.10 | 15.4 ± 1.14 |
1000 pfu 192t-9 | 20.4 ± 0.55 | 13.0 ± 1.58 | 17.4± 0.55 | 19.0 ± 0.71 | 16.8 ± 1.79 |
10 pfu TS IAV | 9.2 ± 3.27 | 5.2 ± 0.84 | 8.8 ± 4.44 | 11.8 ± 1.79 | 9.2 ± 4.15 |
100 pfu TS IAV | 11.6 ± 3.91 | 8.6 ± 1.52 | 10.8 ± 2.95 | 13.2 ± 1.10 | 11.2 ± 1.30 |
1000 pfu TS IAV | 19.3 ± 0.58 | 12.2 ± 1.30 | 14.2 ± 1.10 | 14.2 ± 1.10 | 13.4 ± 1.14 |
UV 192t-6 | 2.4 ± 0.55 | 3.0 ± 0.71 | 2.8 ± 0.84 | 2.6 ± 0.89 | 2.8 ± 0.84 |
UV 192t-9 | 2.4 ± 0.55 | 2.8 ± 0.45 | 3.0 ± 0.71 | 3.2 ± 0.84 | 3.4 ± 0.89 |
Naive | — b | — | — | — | — |
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Gao, F.; Yang, T.; Liu, X.; Xiong, F.; Luo, J.; Yi, Y.; Fan, J.; Chen, Z.; Tan, W.-S. MiRNA Targeted NP Genome of Live Attenuated Influenza Vaccines Provide Cross-Protection against a Lethal Influenza Virus Infection. Vaccines 2020, 8, 65. https://doi.org/10.3390/vaccines8010065
Gao F, Yang T, Liu X, Xiong F, Luo J, Yi Y, Fan J, Chen Z, Tan W-S. MiRNA Targeted NP Genome of Live Attenuated Influenza Vaccines Provide Cross-Protection against a Lethal Influenza Virus Infection. Vaccines. 2020; 8(1):65. https://doi.org/10.3390/vaccines8010065
Chicago/Turabian StyleGao, Feixia, Tianhan Yang, Xueying Liu, Feifei Xiong, Jian Luo, Yinglei Yi, Jiangfeng Fan, Ze Chen, and Wen-Song Tan. 2020. "MiRNA Targeted NP Genome of Live Attenuated Influenza Vaccines Provide Cross-Protection against a Lethal Influenza Virus Infection" Vaccines 8, no. 1: 65. https://doi.org/10.3390/vaccines8010065