RESUMO
Objective To observe the effects of dynamic pressure for the ability of endothelial progenitor cells (EPCs) to form blood vessels, when EPCs seeded into DBM with load. Methods Use the Ficoll density gradient centrifuge combined with difference-speed adherence screening method to separate MNCs from rat bone marrow. Identify the induced EPCs by means of immunohistochemistry and immunofluorescence. Through the organization of fixed, defatted, decalcified and other steps use of spine vertebral body,demineralized bone matrix (DBM) samples of pig were prepared in vitro. Divided scaffolds into two groups A and groups B. Induced EPCs were seeded into DBM. The cell-seeded scaffolds of groups A were dynamically loaded in compression using a sine wave at 1 Hz, 5% strain in the media-filled chamber for 4 h on days 5 of culture. and cell-seeded scaffolds of groups B were cultured directly without any load. Both of two groups were cultured two weeks. Then the ability of EPCs to form blood vessels was observed. Primer desig;Extract total RNA from cells with Trlzol; Reverse transcription reaction; PCR. Results Two groups of cells in HE staining and fluorescent staining showed the formation of vascular bundles. There were formation of blood vessels. It was obvious that the formation in group A was more than that in group B. Test the mRNA expression of vWF and Flk-1 during the EPCs differentiationby RT-PCR. Group A was significantly stronger than that of group B. Conclusion When DBM combines together with EPC, it has become organization engineering bone, then with press on it, the bone graft has been vascularized, so it has clinical application on the direction of repair bone defect.
RESUMO
DNA-PKcs, the catalytic subunit of DNA-dependent protein kinase (DNA-PK), plays an important role in the nonhomologous end-joining (NHEJ) pathway of DNA double-strand breaks (DSBs) repair. To investigate the effects of DNA-PKcs down-regulation on cell growth and sensitization to low dose radiation (LDR), we transfected DNA-PKcs siRNA into human mammary epithelia cells MCF10F, then, detected the proliferation curve of the cells and the expression of protiens in DNA repair pathways. The results showed that DNA-PKcs gene silencing, induced by the transfection of DNA-PKcs siRNA could suppress significantly the cell proliferation and inhibit the expression of the DNA repair proteins, such as Ku80, ATM and P53 after 50 cGy 137Cs gamma-irradiation.The results suggested that DNA-PKcs gene silencing could increase the sensitivity of human breast epithelial cells to the LDR, which might be relative with the decrease of the proteins.