The Isolation And Preliminary Identification Of Uterine Leiomyosarcoma Cancer Stem Cell Like Cells

Objective:To isolate and culture cancer stem cells of uterine leiomyosarcoma from human uterine leiomyosarcoma cell line SK-UT-1,and study their biological characteristics to reveal the important role of CSCs in the occurrence and development of uterine leiomyosarcoma.Methods:SK-UT-1cells were cultured in serum free medium(SFM) to derive tumor spheres,defining as Uterine leiomyosarcoma stem cell like cells(ULSSCLCs); Flow cytometry, Limited dilution method, being cultured in serum-supplemented medium (SSM) and SFM several times, Trypan blue staining, Transwell invasive method and Flow cytometry were applied to examine the expression of cancer stem cell surface marker-CD133, the forming tumor spheres ability of a single-cell, the ability of differentiation of ULSSCLCs, the difference of cell proliferation between SK-UT-1cells and ULSSCLCs, the cell invasive and migration ability of both SK-UT-1cells and ULSSCLCs and the apoptosis rate and cell cycle of SK-UT-1cells and ULSSCLCs after being induced by DDP for48hours. Results:1. ULSSCLCs were isolated and expanded in SFM, and have the possibility of continuous passage.2. CD133antigens of ULSSCLCs were significantly higher than those of SK-UT-1cells (P <0.05), and With the increase of ULSSCLCs generation, the expression quantity of CD133is increasing.3. Through limited dilution method, we observed that a single-cell proliferated into ULSSCLCs under the microscope.4. When ULSSCLCs were put back into SSM, they could attach to culture plates and differentiate into SK-UT-1cells.5. The cell proliferation ability of ULSSCLCs is much higher than SK-UT-1cells (P<0.05).6. The migration(215±14) and invasive(161±15) ability of ULSSCLCs is much stronger than SK-UT-1cells(106±11)、(61±7),(P <0.05).7. When the DDP concentration were40nmol/L、80μmol/L150μmol/L, the apoptosis rate of SK-UT-1cells is higher than that of ULSSCLCs (P<0.05); The distribution of cell cycle of SK-UT-1cells and ULSSCLCs had no difference before DDP was added,48hours after DDP was added, SK-UT-1cells in S phase cell proportion were reduced compared with the pre treatment cells (P<0.05), ULSSCLCs in G2phase cell proportion were increased compared to the pre treatment cells (P<0.05)Conclusion:1. We screen out ULSSCLCs from SK-UT-1cells successfully. 2. ULSSCLCs highly express cancer stem cells antigen CD133an have stronger ability of self-renewal, differentiation, proliferation, migration, invasion and drug resistance.