Fusaoki Koide

Homotypic and heterotypic protection against influenza virus infection in mice by recombinant vaccinia virus expressing the haemagglutinin or nucleoprotein gene of influenza virus

Recombinant vaccinia virus expressing the influenza virus haemagglutinin (HA) or nucleoprotein (NP) genes from A/SW/Hong Kong/1/74 (H1N1) under the control of a hybrid promoter containing the P7.5 early promoter element and promoter of the gene encoding the major protein of cowpox virus A type inclusion body was constructed to investigate protective immunity against homologous and heterologous viruses in mice. These recombinant vaccinia viruses produced authentic influenza virus HA and NP in infected cells. The recombinant vaccinia virus-influenza virus HA conferred efficient subtype-specific protection although mice challenged with heterologous influenza viruses underwent initial infection. By contrast, immunization with the recombinant vaccinia-influenza virus NP limited virus multiplication in the lungs against challenge infection with all H1N1 and H3N2 influenza viruses examined, although less efficiently. These results will prompt the re-examination of the possibility of using the recombinant vaccinia virus-influenza virus NP as a cross-protective vaccine.

Cross-protection against influenza virus infection in mice vaccinated by combined nasal/subcutaneous administration.

The effects of a combination of intranasal (i.n.) and subcutaneous (s.c.) administration of inactivated influenza vaccine for priming and boosting on the cross-protection against antigenically drifted virus challenge were examined in Balb/c mice. Mice were primed through the i.n. or s.c. route with a CTB*-A/Kumamoto/37/79 (H1N1) combined vaccine (CTB*: cholera toxin B subunit supplemented with 0.2% of the holotoxin) and boosted through the i.n. or s.c. route with another drift virus vaccine, A/Bangkok/10/83 (H1N1), 4 weeks later. Two weeks after boosting, the mice were challenged with a third drift virus, A/Yamagata/120/86 (H1N1). The combination of i.n. priming and i.n. boosting afforded the highest cross-protection, while combinations of s.c. priming and i.n. or s.c. boosting afforded little cross-protection. In parallel with the protective activity, anti-A/Yamagata haemagglutinin-reactive IgA and IgG antibodies were detected in nasal and bronchoalveolar wash specimens. These results suggest that cross-protection against a variant virus challenge is most favourably provided by i.n. priming with the CTB* combined vaccine and i.n. boosting with the vaccine, which optimally induces cross-protective IgA and IgG antibodies.

Correlation between canine NK cell mediated cytotoxicity and radical production

We investigated the relationship between natural killer (NK) mediated cytotoxicity and radical production in canine peripheral blood lymphocytes (PBLs). Canine NK cell mediated cytotoxicity, measured by 51Cr release assay, was found to be the highest in the T-cell (Thymus cell) subpopulation which partitioned into the 35-40% percoll fraction by centrifugation through a discontinuous gradient. Radical production showed a similar tendency in luminol-dependent chemiluminescence (CL) assay. Furthermore, enrichment of large granular lymphocytes (LGL) was observed in the cell fraction from the same percoll density. These results suggest that NK cell mediated cytotoxicity is related to radical production in canines.

Characterization of natural killer cytotoxic factor (NKCF) from canine NK cells.

We investigated the presence of canine natural killer cytotoxic factor (NKCF). Canine natural killer (NK) cell-mediated cytotoxicity measured by 51chromium (51Cr) release assay was found to be highest in the T-cell population, which was fractionated into the 35-40% Percoll fraction by discontinuous gradient centrifugation. The cytotoxicity of NKCF in the culture supernatant showed a similar tendency to NK activity. Release of NKCF was rapid after contact with target cells, and reached a plateau in 60 min. The cytotoxicity of NKCF could be detected within at least 15 min in coculture with CL-1 target cells, reaching a plateau in 60 min. We also characterized canine NKCF and found it to be a protein, which was stable against both heat and cold treatment. These findings suggest that canine NK cells release NKCF immediately after recognition and binding to the target cell, and that NKCF plays an important role in canine NK-mediated cytotoxicity.

Changes in Cell-mediated Immunological Function after Operation in Dogs.

外科的侵襲に対する犬の細胞性免疫機能を評価するために実験的に麻酔や手術負荷を加えた後にルミノール依存性化学発光 (CL: chemilumines cence) 測定法により免疫担当細胞である好中球, マクロファージ, 末梢血リンパ球 (PBL) の動態について検討した。その結果は麻酔群ではCL活性の変動はほとんど見られなかった。手術群では, 術後1日目から全血 (好中球) および一細胞当たりの活性増強と好中球の増多が観察された。その後3日目にかけて全血 (好中球) 活性が低下すると同時に, マクロファージ, PBLの活性増強が認められた。以上のことから本実験で用いたCL測定法は術前術後の免疫機能診断法の一つとして有用性が高いものと思われる。