NAGAOKA Satoshi Gifu University, Faculty of Agriculture, Associate Professor, 農学部, 助教授 (50202221)
EBINA Takusaburo Research Institute Miyagi Cancer Center, Department of Immunology, Director, 免疫部, 部長(研究職)
|Budget Amount *help
¥17,300,000 (Direct Cost: ¥17,300,000)
Fiscal Year 2003: ¥4,400,000 (Direct Cost: ¥4,400,000)
Fiscal Year 2002: ¥5,200,000 (Direct Cost: ¥5,200,000)
Fiscal Year 2001: ¥7,700,000 (Direct Cost: ¥7,700,000)
Potent inhibition of cell proliferation was found for the commercial preparations of a major bovine whey protein α-lactalbumin (α-LA) on cultured intestinal cell line IEC-6 albeit lot-dependent. The inhibition was irreversible and only a single exposure to the culture medium containing α-LA of an active lot for a period as short as 30 min was enough to provoke cells death, possibly through apoptosis. The oligomer fraction from size exclusion chromatography was significantly robust, while the monomer fraction remained inert, in inducing cell death.
The commercial preparations of another major bovine whey protein β-lactoglobulin (β-LG) were found to be either promotive or inhibitory on the growth of IEC-6 cells depending on its molecular forms. Thus, native dimer of β-LG promote the cell proliferation significantly even in the medium supplemented with 10% fetal calf serum, whereas the oligomeric form of this protein totally inhibited the cell growth. However, the inhibition was due to ind
ucing growth arrest, not through inducing cell death.
Incubation at 37℃ for 5 days with 30% trifluoroethanol (TFE) in acetate, pH 5.5, in a slowly rotating test tube (TFE treatment) rendered the native monomer of α-LA cytotoxic. The occurrence of SDS-stable 30-kDa or higher Mr bands on SDS-PAGE was a common feature for α-LA with the activity inducing cell death. Similarly, TFE treatment in Tris-HCl, pH 8.0 for 2 days rendered the native dimer of β-LG inhibitory. Again, the occurrence of SDS-stable 37kDa or higher Mr bands on SDS-PAGE was a common feature for β-LG with the inhibitory activity.
Finally, the therapeutic effect of α-LA was studied on murine model of gastroenteritis. Thus, human rotavirus MO was orally inoculated in 5-day-old BALB/c suckling mice, and diarrhea was observed in mice as soon as 24 hr after inoculation. When given orally 24, 48, 72 hr after inoculation, native monomer α-LA prepared from normal bovine milk either before or after the TFE treatment could reduce the duration of diarrhea.
In conclusion, the regulative effect on the intestinal cell growth in vitro of the two major bovine milk whey proteins, either with promotive or with inhibitory action, could have an efficacy on maintaining gastrointestinal health either through rapid elimination of cells in adverse state or quick repair of the tissue by the stimulation of supplying normal mature intestinal cells. Less