摘要
OBJECTIVE To study the potential relationship between the expression of PPT-I, NK-1, NK2 and the development of breast cancer cells in bone marrow stroma and to provide evidence of potential molecular mechanisms of breast cancer patients. of bone metastasis in early stage METHODS The cocultures of breast cancer cell line T-47D and marrow-derived mesenchymal stem cells （MSC） were established with equal numbers. T-47D cells were separated from the coculture system at 48 h and 96 h after coculture by MACS magnetic cell sorting （MicroBeads）. The expression of PPT-I, NK-1, NK-2 in T-47D was then examined before and after coculture by real-time PCR and by Western blot. Alterations in cellular ultrastructure of T-47D cells were detected before and after coculture under electron microscope. Finally, changes in cell cycle distribution were examined by flow cytometry, and growth curves from before and after coculture were drawn and analyzed. RESULTS Following coculture of T-47D and MSC, the expression of PPT-I mRNA and protein was significantly upregulated, while the expression of NK-1 and NK-2 mRNA and protein was greatly downregulated. The analysis of cell cycle distribution by flow cytometry showed that the proportion of T-47D during S phase was increased, and the duration of the G2/M phase was sharply decreased. Under electron microscope, we observed that the synthesis of hereditary material was increased, but the hepatin granules were shown prominent stacking in T-47D cells. These results suggest that although the synthesis of DNA was increased, the proliferation of cells was inhibited after coculture. The cell growth curve confirmed the findings from the observation under the electron microscope and flow cytometry. CONCLUSION Tumor cells could survive through the upregulation in expression of preprotachykinin-I gene during early bone metastasis in breast cancer. The phenomenon of growth suppression in breast cancer cells after coculture in the current study could be induced by downregulation in expression of NK-1 and NK-2.
OBJECTIVE To study the potential relationship between theexpression of PPT-I, NK-1, NK2 and the development of breastcancer cells in bone marrow stroma and to provide evidence ofpotential molecular mechanisms of bone metastasis in early stageof breast cancer patients.METHODS The cocultures of breast cancer cell line T-47D andmarrow-derived mesenchymal stem cells (MSC) were establishedwith equal numbers. T-47D cells were separated from the coculturesystem at 48 h and 96 h after coculture by MACS magnetic cellsorting (MicroBeads). The expression of PPT-I, NK-1, NK-2 inT-47D was then examined before and after coculture by real-timePCR and by Western blot. Alterations in cellular ultrastructureof T-47D cells were detected before and after coculture underelectron microscope. Finally, changes in cell cycle distributionwere examined by flow cytometry, and growth curves from beforeand after coculture were drawn and analyzed.RESULTS Following coculture of T-47D and MSC, the expressionof PPT-I mRNA and protein was significantly upregulated, whilethe expression of NK-1 and NK-2 mRNA and protein was greatlydownregulated. The analysis of cell cycle distribution by flowcytometry showed that the proportion of T-47D during S phasewas increased, and the duration of the G_2/M phase was sharplydecreased. Under electron microscope, we observed that thesynthesis of hereditary material was increased, but the hepatingranules were shown prominent stacking in T-47D cells. Theseresults suggest that although the synthesis of DNA was increased,the proliferation of cells was inhibited after coculture. The cellgrowth curve confirmed the findings from the observation underthe electron microscope and flow cytometry.CONCLUSION Tumor cells could survive through theupregulation in expression of preprotachykinin-I gene duringearly bone metastasis in breast cancer. The phenomenon of growthsuppression in breast cancer cells after coculture in the currentstudy could be induced by downregulation in expression of NK-1and NK-2.
基金
supported by a grant from the National Natural Science Foundation of China(No.30670802).
