Youko Nakada

Effect of Bacillus subtilis spore administration on activation of macrophages and natural killer cells in mice

The effect of Bacillus subtilis (strain A102) spores on the activation of murine macrophages and natural killer cells (NK) was examined. The macrophage activity and NK activity were enhanced by oral administration of A102 spores, and slightly enhanced by oral administration of culture supernatant. There was no difference in the results of macrophage activity and NK activity using other live or dead spores. The NK activity and macrophage activity were increased with increments of concentration up to 0.1 g per mouse, and both activities were decreased at concentration of more than 0.15 g per mouse. The NK activity was increased 1 and 2 days after oral administration of A102 spores, and the activity level 2 days after administration was about 3-fold higher than the level prior to treatment. Macrophage activity was also increased from 1 to 3 days after oral administration of A102 spores, and the activity level 3 days after administration was about 3-fold higher than the level prior to treatment. The induction of interferons at 1 day after oral administration in mouse serum was 5-fold higher than that in controls. These findings indicate that oral administration of A102 gave rise to the induction of interferons, and it is likely that macrophages and NK cells were activated by interferons.

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.

Relationship between radical production and natural killer cytotoxic factor (NKCF) in canine natural killer (NK) cell-mediated cytotoxicity

The relationship between radical production and natural killer cytotoxic factor (NKCF) release via canine natural killer (NK)-mediated cytotoxic mechanism was examined. Radical production and NKCF release was induced in NK cells stimulated with either dead target cells, or their cytoplasmic membranes, as well as live target cells. Canine NKCF evoked target cell lysis but did not induce radical production. Radical production was inhibited by the addition of Tiron or n-propyl gallate, whereas NK-mediated cytotoxicity and NKCF release were only inhibited by the addition of n-propyl gallate. These results suggested that radical production and NKCF release may be induced by the contact and binding of NK cells to the target cell cytoplasmic membrane. Therefore, the release of NKCF from NK cells attached to the target cell cytoplasmic membrane may be associated with the production of radicals, especially hydroxyl radicals.

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。