Studies on a novel immunostimulation factor in fermented milk.

发酵乳中新型免疫刺激因子的研究。

基本信息

批准号：
16580102
负责人：
SUGAHARA Takuya
金额：
$ 2.05万
依托单位：
Ehime University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

Fat-free bovine milk fermented by 12 kinds of lactic acid bacteria and yeast enhanced monoclonal antibody production of human hybridoma HB4C5 cells 2.8-fold in serum-free medium. Immunoglobulin production of human peripheral blood lymphocytes (PBL) was also stimulated in vitro. IgM and IgG production of human PBL was accelerated up to 2.8-fold and 5.4-fold, respectively. Interferon-y production of human PBL was also accelerated 6.0-fold by 50 μg/ml of the fermented milk. However, interleukin-4 production of PBL was not affected, and tumor necrosis factor-a production was suppressed. The activity was enhanced 2.5-fold by the thermal treatment for 30 min at 65 ℃, and was completely lost by trypsin digestion. The findings suggested that the active substance in the fermented milk was heat stable protein. Gel-filtration and the SDS-PAGE analysis revealed that the molecular weight of the active substance was estimated as 19.0 kDa, which was not detected in fat-free bovine milk before fermentation. N-terminal amino acid sequence of the 19.0 kDa protein was highly homologous to proteose-peptone component 3 (PP3). Since molecular weight of PP3 is 28 kDa, it is suggested that the 19.0 kDa protein is derived from degradation of PP3 during fermentation of fat-free milk. Moreover, PP3 purified from fat-free milk also enhanced IgM production of HB4C5 cells.PP3 did not enhanced IgM-mRNA transcription level in HB4C5 cells. However, PP3 stimulated IgM production of HB4C5 cells treated with Actinomycin D, transcription inhibitor.Moreover, PP from fermented milk suppressed IgE production by human myeloma U266 cells. As mentioned above, PP3 enhance IFN-y production. These facts suggest that PP may suppress allergic reaction.

由12种乳酸细菌和酵母发酵的无脂肪牛奶增强了人类杂交瘤HB4C5细胞的单克隆抗体产生2.8倍无血清培养基。在体外还刺激了人外周血淋巴细胞（PBL）的免疫球蛋白产生。人类PBL的IgM和IgG产生分别加速了2.8倍和5.4倍。通过50μg/ml的发酵牛奶，人类PBL的干扰素-y产生也加速了6.0倍。然而，白介素-4的PBL产生不受影响，肿瘤坏死因子A产生被抑制。热处理在65℃时进行热处理30分钟增强了活性，并因胰蛋白酶消化而完全失去。研究结果表明，发酵牛奶中的活性物质是热稳定蛋白。凝胶过滤和SDS-PAGE分析表明，活性物质的分子量估计为19.0 kDa，在发酵之前未在无脂肪的牛奶中检测到。 19.0 kDa蛋白的N末端氨基酸序列与蛋白质蛋白酶成分3（PP3）高度同源。由于PP3的分子量为28 kDa，因此建议19.0 kDa蛋白在发酵的无脂肪牛奶发酵过程中源自PP3的降解。此外，从无脂肪牛奶中纯化的PP3还增强了HB4C5细胞的IgM产生。PP3并未提高HB4C5细胞中的IgM-MRNA转录水平。然而，PP3刺激了用放线菌素D，转录抑制剂处理的HB4C5细胞的IgM产生。此外，发酵牛奶中的PP抑制了人骨髓瘤U266细胞的IgE产生。如上所述，PP3增强了IFN-Y产生。这些事实表明，PP可能抑制过敏反应。