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Universal vaccine against respiratory syncytial virus A and B subtypes

View Article: PubMed Central - PubMed

ABSTRACT

Respiratory syncytial virus (RSV) is a major cause of acute lower respiratory tract infection in infants, young children, and the elderly. Two subtypes of RSV, A and B, circulate alternately at 1-2-year intervals during epidemics. The attachment glycoprotein (G protein) of RSV is one of the major targets for immune responses. In this study, we generated a recombinant fusion protein, GcfAB, which consists of the central regions (a.a. residues 131–230) of the G proteins of both RSV A (A2 strain) and B (B1 strain) subtypes, and investigated immunogenicity, protective efficacy, and immunopathology. We immunized mice with GcfAB plus cholera toxin as a mucosal adjuvant via intranasal (IN) or sublingual (SL) routes. The IN group showed higher levels of RSV G-specific antibody responses, including serum IgG and mucosal IgA, compared with the SL group. On the contrary, more vigorous RSV G-specific CD4+ T-cell responses were elicited in the SL group than in the IN group after RSV-A but not RSV-B viral challenge. Furthermore, the SL group showed more pulmonary eosinophil recruitment and body weight loss than did the IN group after RSV-A challenge. Both IN and SL immunization with GcfAB provided potential protection against both subtypes of infections. Together, these results suggest that vaccination with GcfAB via an IN route could be a universal vaccine regimen preventing both RSV A and B infections.

No MeSH data available.


Expression and purification of recombinant GcfAB protein.Schematic diagram of the GcfAB recombinant protein. (A) A pET-21d plasmid containing both a.a 131 to 230 of RSV A2 G protein (GcfA) and a.a. 131 to 230 of RSV B1 G protein (GcfB) was constructed. (B) The GcfAB protein was purified by His-tag affinity chromatography (lane 1) and size-exclusion chromatography (Sephacryl S-200 HR) with (lane 2) / without (lane 3) dithiothreitol (DTT). Purification of GcfAB was confirmed by SDS-PAGE at each purification step, as indicated by the arrow.
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pone.0175384.g001: Expression and purification of recombinant GcfAB protein.Schematic diagram of the GcfAB recombinant protein. (A) A pET-21d plasmid containing both a.a 131 to 230 of RSV A2 G protein (GcfA) and a.a. 131 to 230 of RSV B1 G protein (GcfB) was constructed. (B) The GcfAB protein was purified by His-tag affinity chromatography (lane 1) and size-exclusion chromatography (Sephacryl S-200 HR) with (lane 2) / without (lane 3) dithiothreitol (DTT). Purification of GcfAB was confirmed by SDS-PAGE at each purification step, as indicated by the arrow.

Mentions: To develop the dual subunit vaccine covering both subtypes (A and B) of RSV, we employed a strategy that fuses a highly-conserved central region (a.a. 131–230) of the RSV A2 G sequence with that of the RSV B1 G sequence, resulting in a GcfAB fusion in the pET-21d vector (Fig 1A). It was previously shown that a.a. residues 183–195 of GcfA functions as a CD4 T-cell epitope that is necessary for induction of a strong antibody response, while the same region of GcfB lacks T-cell epitope functionality and induces a relatively weak antibody response [29]. Thus, we reasoned that fusion of the GcfA sequence to the GcfB sequence might simultaneously provoke T-cell activity against both GcfA and GcfB, and thus, that immunization with the GcfAB fusion antigen might induce strong humoral responses against both subtypes. To this end, the GcfAB protein was purified from E.coli by His-tag affinity chromatography and size-exclusion chromatography, and its purity was confirmed using SDS-PAGE. The approximate molecular weight of the monomeric form of GcfAB is ~33 kDa (black arrow) on SDS-PAGE under reducing conditions (Fig 1B).


Universal vaccine against respiratory syncytial virus A and B subtypes
Expression and purification of recombinant GcfAB protein.Schematic diagram of the GcfAB recombinant protein. (A) A pET-21d plasmid containing both a.a 131 to 230 of RSV A2 G protein (GcfA) and a.a. 131 to 230 of RSV B1 G protein (GcfB) was constructed. (B) The GcfAB protein was purified by His-tag affinity chromatography (lane 1) and size-exclusion chromatography (Sephacryl S-200 HR) with (lane 2) / without (lane 3) dithiothreitol (DTT). Purification of GcfAB was confirmed by SDS-PAGE at each purification step, as indicated by the arrow.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5383302&req=5

pone.0175384.g001: Expression and purification of recombinant GcfAB protein.Schematic diagram of the GcfAB recombinant protein. (A) A pET-21d plasmid containing both a.a 131 to 230 of RSV A2 G protein (GcfA) and a.a. 131 to 230 of RSV B1 G protein (GcfB) was constructed. (B) The GcfAB protein was purified by His-tag affinity chromatography (lane 1) and size-exclusion chromatography (Sephacryl S-200 HR) with (lane 2) / without (lane 3) dithiothreitol (DTT). Purification of GcfAB was confirmed by SDS-PAGE at each purification step, as indicated by the arrow.
Mentions: To develop the dual subunit vaccine covering both subtypes (A and B) of RSV, we employed a strategy that fuses a highly-conserved central region (a.a. 131–230) of the RSV A2 G sequence with that of the RSV B1 G sequence, resulting in a GcfAB fusion in the pET-21d vector (Fig 1A). It was previously shown that a.a. residues 183–195 of GcfA functions as a CD4 T-cell epitope that is necessary for induction of a strong antibody response, while the same region of GcfB lacks T-cell epitope functionality and induces a relatively weak antibody response [29]. Thus, we reasoned that fusion of the GcfA sequence to the GcfB sequence might simultaneously provoke T-cell activity against both GcfA and GcfB, and thus, that immunization with the GcfAB fusion antigen might induce strong humoral responses against both subtypes. To this end, the GcfAB protein was purified from E.coli by His-tag affinity chromatography and size-exclusion chromatography, and its purity was confirmed using SDS-PAGE. The approximate molecular weight of the monomeric form of GcfAB is ~33 kDa (black arrow) on SDS-PAGE under reducing conditions (Fig 1B).

View Article: PubMed Central - PubMed

ABSTRACT

Respiratory syncytial virus (RSV) is a major cause of acute lower respiratory tract infection in infants, young children, and the elderly. Two subtypes of RSV, A and B, circulate alternately at 1-2-year intervals during epidemics. The attachment glycoprotein (G protein) of RSV is one of the major targets for immune responses. In this study, we generated a recombinant fusion protein, GcfAB, which consists of the central regions (a.a. residues 131–230) of the G proteins of both RSV A (A2 strain) and B (B1 strain) subtypes, and investigated immunogenicity, protective efficacy, and immunopathology. We immunized mice with GcfAB plus cholera toxin as a mucosal adjuvant via intranasal (IN) or sublingual (SL) routes. The IN group showed higher levels of RSV G-specific antibody responses, including serum IgG and mucosal IgA, compared with the SL group. On the contrary, more vigorous RSV G-specific CD4+ T-cell responses were elicited in the SL group than in the IN group after RSV-A but not RSV-B viral challenge. Furthermore, the SL group showed more pulmonary eosinophil recruitment and body weight loss than did the IN group after RSV-A challenge. Both IN and SL immunization with GcfAB provided potential protection against both subtypes of infections. Together, these results suggest that vaccination with GcfAB via an IN route could be a universal vaccine regimen preventing both RSV A and B infections.

No MeSH data available.