Bone Marrow Mesenchymal Stem Cells Roles In Prostate Cancer Bone Metastasis

Objective:Advanced prostate cancer is often accompanied by bone metastasis,seriously affecting the quality of patients' life.In this study,we investigated the causes and mechanisms of bone metastasis in prostate cancer by in vitro cell experiments.The role of bone marrow mesenchymal stem cells in prostate cancer bone metastasis can explore new strategies for clinical prevention and treatment of bone metastasis,take intervention to inhibit the recurrence and metastasis in prostate cancer and improve the quality of life of patients.Methods:1.(1)Murine bone marrow were isolated and purified according to the different ways of cell adherent growth.And observe the morphological changes of the cells closely.(2)In vitro,mesenchymal stem cells were inducted into adipocytes and osteoblasts by inducing agents,and were identified by Re-oil red O staining and alizarin red staining.(3)The mesenchymal stem cell supernatants were collected and cultured as mesenchymal stem cell conditioned medium(MSC-CM).The prostate cancer cells,C42 cells,were cultured with MSC-CM for 1-5 days.The absorbance of each well was measured by MTT method,and drew the growth curve.(4)Wound scratch assay was performed to observe the cell scavenging of C42 cells after 48 hours of treatment with competent medium of mesenchymal stem cells.The area of scratches was calculated by ImageJ software.Transwell experiments were performed by adding a certain number of cells in the upper chamber.The lower chamber was added by the mesenchymal stem cell conditioned medium.After 24 hours of culture,the number of cells transferred to the lower cells was observed by Giemsa staining.2.(1)CD133+ prostate cancer stem cells were sorted by magnetic beads cells sorting(MACS),and the stem cells was further confirmed by RT-PCR and Western Blot experiments.(2)CD133+ C42 cells were cultured in DMEM medium for 1-5days.The absorbance of each well was measured by MTT method,and the growth curve was drawn.(3)The prostate cancer stem cells CD133 + C42 were cultured withmesenchymal stem cell conditioned medium,and the RNA and protein of the cells were extracted on day 2 and day 4 respectively.The expression of CD133 and CD44 on the surface markers of stem cells was observed by RT-PCR and Western Blot.Results:1.(1)Mesenchymal stem cells began to adhere to the well 24 hours later,showing short spindle,round or polygonal cells.Cell growth on day 8 was intensive.(2)Mesenchymal stem cells induce the formation of fat cells in adipocytes,after oil red O staining,lipid droplets were stained orange,and the nucleus were light blue.Mesenchymal stem cells induce osteoblast morphology mostly polygonal,and the volume increases.After alizarin red staining,resulting in dark red compounds.(3)The growth curve showed that the cell proliferation ability of MSC-CM group was significantly increased.(4)Wound scratch assay showed that the scratched area of MSC-CM cells was significantly larger than that of the control group.Transwell experiments showed that the number of cells in MSC-CM group was significantly larger than that in the control group.2.(1)CD133 + C42 was sorted by magnetic beads cells sorting method,and it was detected by gene and protein level.The results were consistent with the biological characteristics of tumor stem cells.(2)MTT assay showed that the proliferative capacity of CD133 + C42 in prostate cancer stem cells was significantly enhanced compared with that of untreated prostate cancer cells C42.(3)The expression of CD133 and CD44 in the control group was decreased,while the MSC-CM group was stable.Concludions:1.Bone marrow mesenchymal stem cells can promote the proliferation of prostate cancer cells,migration and invasive ability.2.Bone marrow mesenchymal stem cells can maintain stable expression of stem cells in stem cells.