The Effect Of Rh G-CSF On Mammary Gland Development In MMTV-erbB2 Transgenic Mouse

Objective: 1. Study of the proliferation effect of recombinant human granulocyte colony-stimulating factor(Rh G-CSF) on normal mammary cell line MCF-10 A and breast cancer cell line SK-BR-3. 2. Explore the Rh G-CSF effect on mammary gland development in MMTV-erbB2 transgenic mouse.Method: 1. Culture MCF-10 A and SK-BR-3 cells with different concentrations of RhG-CSF in vitro at different times, then using the MTT colorimetric to analyze the cell proliferation rate. 2. 60 healthy female aged 12 weeks MMTV-er Bb2 transgenic mice were randomly divided into three groups, the low-dose group, namely, RhG-CSF drug was administered by subcutaneous injection at dose of 0.125μg every day, control group that injected with normal saline 0.25μg every day and high dose group that RhG-CSF was administered by subcutaneous injection at dose of 0.25μg every day. After eight weeks treatment with RhG-CSF drug by subcutaneous injection, six mice were dissected from each group, taking breast tissue in the bottom left side of mice, observing the changes of mice mammary gland in morphological structure by whole mount staining, and the ductal structures of mice mammary gland were observed by HE staining. Immunohistochemistry was using to analysis the PCNA protein expression in breast tissue of each group. Real-time quantitative PCR was used to measure the cyclinD1, c-myc,VEGF, G-CSF3 R mRNA expression in mouse mammary tissue.Results: 1. After the treatment of different concentrations of RhG-CSF in normal breast epithelial cells and breast cancer epithelial cells,cells proliferation was significantly higher at low doses group compare to the control group. However, high dose group was not obvious. Whole mount results show that compared to the control group, the low-dose group(P=0.000)had more mammary small leaf exuberant than the high-dose mouse after eight weeks treatment with Rh G-CSF drug. However, ductal proliferation was not obvious in high dose group(P=0.027). 3. Compare to control group, PCNA expression was higher in the low-dose group(P=0.000). RT-PCR showed cyclinD1, c-myc, VEGF, G-CSF3 R were highly expressed at low dose(P<0.05), high dose group was not obvious.Conclusion: 1. RhG-CSF can significantly promote the proliferation of breast cancer cells in low dose group.While, the proliferation effect on human normal mammary gland cell MCF-10 A is not obvious. 2. RhG-CSF effect on mammary gland development in MMTV-erbB2 transgenic mouse, low dose Rh G-CSF can obviously promote mammary tissue proliferation, while the effect was not obvious at high doses.