| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2006: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2005: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
We found further up-regulation of MFG-E8 in involuting mammary glands, where the glands undergo a substantial increase in the rate of epithelial cell apoptosis. The MFG-E8 signal was detected around the epithelium of such involuting mammary glands. Total MFG-E8 in milk was also increased in the post-weaning mammary glands and, furthermore, the free MFG-E8 content in the post-weaning milk, as measured by in vitro phosphatidylserine (PS)-binding and apoptotic HC11 cell-binding activities, was much higher than that of lactation. Sucrose density-gradient ultracentrifugation analyses revealed that such enhanced PS-binding activity of MFG-E8 was present in membrane vesicle fractions. The weaning-induced MFG-E8 might play an important role in the recognition and engulfment of apoptotic epithelial cells by the neighboring phagocytotic epithelial cells in involuting mammary glands.
Proteomic analysis revealed that in addition to MFG-E8, butyrophilin and caveolin-1 were components of membrane vesicle fractions of milks from involuting mammary glands. It also was suggested that MFG-E8 was immobilized on MFGs during lactation and become mobilized in the accumulated breast milk at early involution, resulting in the rapid increase in soluble MFG-E8 with PS-binding activity in the alveolar lumen. Such soluble MFG-E8 might play an important role in subsequent labeling of apoptotic epithelial cells during the next stage of mammary gland involution. Unexpectedly, up-regulation of MFG-E8 in apoptotic mammary epithelial cells in culture was not obvious. To further elucidate a precise mechanisms of MFG-E8 up-regulation, the promoter region (approx. 2 kbp) was cloned into a plasmid vector bearing a reporter luciferase gene, and the promoter assay is in progress.
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