Clin Exp Vaccine Res.  2019 Jan;8(1):54-63. 10.7774/cevr.2019.8.1.54.

Nucleoprotein vaccine induces cross-protective cytotoxic T lymphocytes against both lineages of influenza B virus

• Lee SY
• Kang JO
• Chang J

• KMID: 2438964
• DOI: http://doi.org/10.7774/cevr.2019.8.1.54

Abstract

PURPOSE
The influenza B virus diverges into two antigenically distinct lineages: B/Yamagata and B/Victoria. Influenza B is the dominant circulating virus during some influenza seasons, and recent data demonstrated that influenza A and B infection similarly cause severe clinical symptoms in hospitalized patients. Nucleoprotein (NP) is a good target for a universal influenza vaccine. This study investigated whether NP epitope variation within two lineages affects the dominant cytotoxic T lymphocyte (CTL) responses induced by vaccination and the resultant protective immunity.
MATERIALS AND METHODS
The NP of B/Yamagata/16/1988, the representative strain of the Yamagata lineage, includes a dominant CTL epitope, FSPIRITFL, while B/Shangdong/7/1997 from the Victoria lineage has one amino acid difference in this sequence, FSPIRVTFL. Two recombinant replication-deficient adenovirus (rAd)-vectored vaccines expressing either NP were prepared (rAd/B-NP(I) and rAd/B-NP(V), respectively) and administered to BALB/c mice intranasally. To examine the efficacy of vaccination, antibody responses, CTL responses, and morbidity/mortality after challenge were measured.
RESULTS
Both vaccines induce similar antibody and CD8 T-cell responses cross-reacting to both epitopes, and also confer cross-protection against both lineages regardless of amino acid difference.
CONCLUSION
The rAd-vectored vaccine expressing the NP could be developed as universal influenza B vaccine which provides broader protection.

Keyword

Influenza B virus; Cross protective immunity; Nucleoproteins; Cytotoxic T lymphocytes; Recombinant adenovirus; Epitope

MeSH Terms

Adenoviridae
Animals
Antibody Formation
Epitopes
Humans
Influenza B virus*
Influenza Vaccines
Influenza, Human*
Lymphocytes
Mice
Nucleoproteins*
Seasons
T-Lymphocytes
T-Lymphocytes, Cytotoxic*
Vaccination
Vaccines
Victoria
Epitopes
Influenza Vaccines
Nucleoproteins
Vaccines

Figure

• 

  Fig. 1 Construction of replication-defective adenoviruses expressing nucleoprotein (NP) of influenza B virus and expression in HEK293 cells. (A) Cytotoxic T lymphocyte epitope sequences of NP restricted to MHC class I H-2Dd. (B) The recombinant adenovirus encoding NP is generated following homologous recombination with the shuttle vector and adenoviral genome. (C) Expression of NP antigens in HEK293 cells (arrow indicated) infected with each recombinant adenovirus is confirmed by Western blotting as described in the Materials and Methods.

• 

  Fig. 2 Characterization of humoral immune responses induced by immunization of the recombinant adenoviruses. BALB/c mice were immunized once with 1×108 PFU rAd/mock, rAd/B-NP(V), or rAd/B-NP(I) via intranasal route. Two weeks later, average anti-nucleoprotein (NP) IgG antibody responses in the sera were measured by enzyme-linked immunosorbent assay. a)p<0.001.

• 

  Fig. 3 Nucleoprotein (NP)-specific CD8 T-cell responses measured by the tetramers. rAd/B-NP(I) (A, B) and rAd/B-NP(V) (C, D) vaccinated mice were challenged with 5LD50 doses of Influenza B/Shangdong or B/Yamagata. Lung cells were harvested 5 days after challenge, and stained with two types of Dd/NP166–174 tetramer (V or I), anti-CD8, and anti-CD44 antibodies. (A, C) Representative dot plots of tetramer staining. The upper right quadrant area shows tetramer+CD44+ cells. (B, D) The percentages of tetramer-positive cells calculated among each group. a)p<0.01.

• 

  Fig. 4 Nucleoprotein (NP)-specific CD8 T cells in the lungs measured by interferon-γ (IFN-γ) intracellular cytokine staining. Mice were immunized with rAd/B-NP(I) (A, B) or rAd/B-NP(V) (C, D), and lung cells were prepared and stained with anti-CD8, CD44, and IFN-γ after stimulation with two types of peptide (I or V). (A, C) Representative dot plots of CD44+IFN-γ+ cells. (B, D) The percentages of CD44+IFN-γ+ population. ns, not significant. a)p<0.01, b)p<0.001.

• 

  Fig. 5 Protective efficacy of the rAd/B-NP vaccines against influenza B infection. Mice were immunized with 1×108 plaque-forming units of each vaccine intranasally, and were challenged with 5LD50 of B/Shangdong or B/Yamagata. Body weight changes (A) and survival rates (B) were recorded for 10 days.

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