Effects Of Osteomimicry And Proliferation Of Breast Cancer:Induced By Osteoblastic Cell In Vitro

Background and Objectives:Breast cancer is currently the first good hair tumor of women.Although the early diagnosis of the treatment effect is good,but in the event of transfer after its effect is poorer prognosis.Bone is the most popular part that the breast cancer cells prone to transfer.A lot of complications would appear after breast cancer cells metastasize to bone,such as hypercalcemia,pathological fracture,the nerve and spinal cord compression which leading a poor quality of life.Studies have found that breast cancer cells that metastasize to bone will lead a mimicry change,namely breast cancer cells can simulate the osteoblast phenotype in the bone metastases And acquire the ability to secrete specific proteins and factors secreted by osteoblasts,This feature allows tumor cells metastasize to bone to evade the immune system to identify,and at last leads to a rapid proliferation in the bone microenvironment.This feature may be become a new breakthrough in our research of tumor bone metastases.Tumor cells in the bone metastases could disguise themselves into osteoblasts,so osteoblasts play a what kind of role in mimicry,it is not clear.Osteoblasts could specific secrete osteocalcin(OCN),osteopontin(OPN),bone protection(OPG),which play an important role in the process of bone metastases.Now it have been confirmed that the expression of OCN is significantly increased in the serum of patients with breast cancer,and OPG’s expression is closely related to the tumor cells’ clone and homing,the increased expression of OPN is conducive to the proliferation and invasion of breast cancer cells.It is improved that prostate cancer cells would have a mimicry change when co-cultured with osteoblasts.The first part of this study hypothesis if the bone mimicry of tumor cells was caused by the osteoblasts,It also could up-regulates the expression of OPN,OCN and OPG which were closely related to tumor metastasis and proliferation.Second,it is well known that the growth of tumor cells in the bone microenvironment is closely related with the tumor growth factors which is released after bone destruction,But studies have shown that although bisphosphonates can inhibit osteoclast’s differentiation,maturation to prevent bone destruction and indirect reduces tumor growth factor’s release,but it can not restrain the growth of the tumor,which showed that in addition to the bony destruction after the release of growth factors to stimulate tumor cell proliferation,there must be other way in tumor cell’s proliferation.This research is aimed to investigate the effects of osteomimicry and proliferation of osteolytic breast cancer cells by co-culturing with osteoblasts.Methods:1.By transwell Chambers establish osteoblast MC3T3-E1 and breast cancer MDA –MB-231 cells co-culture system,and simulate the bone microenvironment.Using real-time quantitative PCR and Western blot to observe the expression of gene and protein of osteocalcin(OCN),osteopontin(OPN),osteoprotegerin(OPG)between the experimental group and control group,in which breast cancer cells are co-cultured with osteoblasts in the experimental group,while the breast cancer cells are cultured alone in the control group.2.To observe the difference in the colony formation velocity and size of breast cancer cells between the experimental group in which breast cancer cells are co-cultured with osteoblasts and the control group that the breast cancer cells are cultured alone by crystal violet staining observation.3.Cell Counting Kit-8(CCK-8)is used to detect the difference of the ability of proliferation between the experimental group,in which breast cancer cells are co-cultured with osteoblasts,and the control group that the breast cancer cells are cultured alone.4.To detect the expression levels of Wnt-1 and Wnt-2 after co-cultured with osteoblasts in breast cancer cells by PCR.Results:1.Successfully established osteogenesis cells MC3T3-E1 and MDA-MB-231 breast cancer cells co-culture system,to simulate the bone microenvironment.The expression of OPG,OPN and OCN are all up-regulated both in the gene and protein levels after co-cultured with osteoblasts(p<0.05,n=3).2.By crystal violet staining,MDA-MB-231 breast cancer cells which is co-cultured with MC3T3-E1 osteoblasts of experimental group compared to the control group,experimental group on the number of colony formation and size are better than the control group.3.CCK-8 experiments showed that at the time of 0 and 24 h,the difference of cell proliferation between experimental group and control group is not obvious.But at the time 48 and 72 h,the experimental group’s cell proliferation ability was significantly improved(p<0.05,n=3).4.PCR results suggest that after co-cultured with MC3T3-E1 osteoblasts,the expression of Wnt-1 and Wnt-2 are all up-regulated in MDA-MB-231 breast cancer cells(p<0.05,n=3).Conclusion:1.MDA-MB-231 breast cancer cells will get a osteoblast-like phenotype in the condition of co-cultured with MC3T3-E1 osteoblast cells2.MC3T3-E1 Osteoblasts can enhance breast cancer MDA-MB-231 cells of colony forming ability,and can promote the proliferation,it also can induce the expression of Wnt-1 and Wnt-2 in MDA-MB-231 breast cancer cells...