RESUMEN
Fragments encoding amino acids 76-130 in the linear conserved region (LCR) of A/Hubei/1/2010 (H5N1) HA2 was fused to hepatitis B core antigen (HBc) to generate a LCR-HBe virus-like particle (VLP). Results showed that the fusion protein of LCR-HBc was highly expressed in this prokaryotic expression system. The purified LCR-HBc particle stimulated high levels of IgG production in mice with a titer of > 1:12 800, and provided 50% cross-protection against lethal challenge by H1N1 viruses.
Asunto(s)
Animales , Femenino , Ratones , Secuencia de Aminoácidos , Glicoproteínas Hemaglutininas del Virus de la Influenza , Alergia e Inmunología , Subtipo H5N1 del Virus de la Influenza A , Alergia e Inmunología , Vacunas contra la Influenza , Alergia e Inmunología , Interferón gamma , Pulmón , Patología , Ratones Endogámicos BALB C , Datos de Secuencia MolecularRESUMEN
Nucleoprotein (NP) of influenza virus is highly conserved and type-specific. NP can trigger strong cell-mediated immune responses in host and is involved in the protection against the challenges with different subtype influenza viruses. Here, NP of an avian H5N1 (A/Hubei/1/2010, HB) was expressed by baculovirus surface-display technology and its immunogenicity as well as protective mechanism was investigated in mice infection model. Western blot and immunolabeled electron microscopy assay showed NP was displayed on baculovirus surface. ELISA results showed NP could induce high level of anti-NP IgG in the sera from NP-Bac-inoculated mice. Two cellular immune peptides (NP57-74 IQNSITIERMVLSAFDER and NP441-458 RTEIIKMMESARPEDLSF) were identified by IFN-gamma ELISPOT assay. NP57-66 and NP441-450 and NP protein could be able to trigger the activation of CD4+ and CD8+ T cells, and the response of CD8+ T was more predominant. The challenge study of mice-adapted virus A/PR/8/34 (H1N1) showed that NP-Bac could reduce viral load and attenuate the damage to lung tissue. 50% protection ratio against the virus could be detected.
Asunto(s)
Animales , Femenino , Humanos , Ratones , Anticuerpos Antivirales , Alergia e Inmunología , Baculoviridae , Genética , Metabolismo , Protección Cruzada , Ensayo de Immunospot Ligado a Enzimas , Inmunidad Celular , Subtipo H1N1 del Virus de la Influenza A , Genética , Alergia e Inmunología , Subtipo H5N1 del Virus de la Influenza A , Genética , Alergia e Inmunología , Gripe Humana , Alergia e Inmunología , Virología , Ratones Endogámicos BALB C , Proteínas de Unión al ARN , Genética , Alergia e Inmunología , Linfocitos T , Alergia e Inmunología , Proteínas del Núcleo Viral , Genética , Alergia e InmunologíaRESUMEN
To establish the mouse-lethal model for pandemic H1N1 influenza virus, provide an animal model for studying the pathogenicity and host adaptation of 2009 pandemic H1N1 influenza virus, and find out the key amino acid mutations which may affect viral virulence and replication. A pandemic H1N1 influenza virus strain, A/Sichuan/SWL1/2009 (H1N1, SC/1) was passaged in mouse lung by 15 cycles with intranasal infection. The passaged viruses were all propagated in MDCK cells and sequenced. Based on the sequencing results, four mice in each group were inoculated with 6 selected viruses and their weight and survival rate were monitored during the following 14 days after infection. Additionally, SC/1-MA P14 and P15 viruses were sequenced after purification by Plague Assay. Viral virulence was increased after serial passages and the mortality of 100% was detected after 7 passages. Several amino acid residue mutations of passaged viruses which may contribute to the enhanced virulence were observed. The increased virulence of passaged viruses and mammalian host adaptation maybe associated with amino acid mutations in viral functional proteins. Finally, we established a mouse-lethal model.