The recent demonstration that
bone sialoprotein (BSP) can be detected in human
breast cancer tissue by immunoperoxidase suggests that this
phosphoprotein is ectopically expressed by malignant mammary epithelial cells. Its detection in human
breast cancer cells raises questions about its potential role(s) during
breast cancer progression. Because BSP is secreted and is present in the serum, the positivity of
breast cancer cells for BSP could have been the result of an uptake of the circulating
phosphoprotein by the cells rather than of an intrinsic expression. We examined the expression of BSP at both the
protein and
mRNA levels in nine human
breast cancer samples as well as in three human
breast cancer cell lines (MCF-7, T47-D, and MDA-MB-231) using immunohistochemistry, flow cytometric analysis, immunoblot, and
reverse-transcriptase PCR. BSP was detected at both
protein and
mRNA levels in human
breast cancer tissue and in the three human
breast cancer cell lines. Using a specific polyclonal anti-BSP antibody, we showed by both fluorescence-activated cell sorter analysis and immunohistochemistry experiments that all of the human
breast cancer cell lines studied express BSP. This was localized at the cell surface and in the cytosol of the
estrogen receptor-positive MCF-7 and T47-D cell lines, whereas it was detected only in the cytosol of the
estrogen receptor-negative MDA-MB-231 cells. Using the same polyclonal anti-BSP antibody, we were able to identify an approximately 97-kd band on total
protein extracts from the three cell lines by immunoblotting.
Reverse-transcriptase PCR reactions using specific
oligonucleotides performed on total
RNA of nine human
breast cancer biopsy samples and the three cell lines demonstrated the presence of BSP
mRNA in all of the samples examined. This study is the first demonstration that human malignant breast epithelial cell lines express BSP at the
protein and
mRNA levels. Our study identified MCF-7, T47-D, and MDA-MB-231 cells as useful models for the examination of the molecular mechanisms involved in the ectopic expression of BSP in breast malignant lesions.