Cultivation and identification of human colon cancer stem-like cells in the serum-free culture medium in vitro
10.3969/j.issn.1673-4130.2015.15.040
- VernacularTitle:结肠癌干细胞样细胞的无血清法培养及特性鉴定
- Author:
Ting YANG
- Publication Type:Journal Article
- Keywords:
colorectal cancer;
cancer stem cells;
serum-free culture
- From:
International Journal of Laboratory Medicine
2015;(15):2221-2223
- CountryChina
- Language:Chinese
-
Abstract:
Objective To generate suspended spheres ,Human colon cancer cell line SW620 cells were acclimated gradually in the free serum culture medium ,1 week later ,a great quantity of spheres were obtained for the further research .Methods SW620 cells and spheres were incubated with the following Abs :anti‐SSEA‐4 Ab and anti‐TRA‐1‐60 Ab and detected with confocal microscopy . SW620 cells and spheres were collected and stained by PI ,Cell cycle distribution was then acquired by flow cytometry .SW620 cells and spheres were transferred into Adipocyte Differentiation Medium and cultured for 7 d and then lipid droplets were counted . SW620 cells and sphere cells were injected subcutaneously into the lateral root of one posterior limb of a nude mouse ,tumor volume was calculated .Results Spheres formed when grown under serum‐free conditions in 7 days ,and the ability of SW620 cells to form spheres was sustained .A few of SW620 adherent cells be cultured in DF+10% FBS clearly showed the expression of pluripotent stem cell markers Ssea‐4(25 .739 ± 7 .62)% ,Tra‐1‐60(27 .742 ± 4 .311)% ,but almost all spheres expressed the 2 markers and there was a significant difference between the two groups in the expression ratio (P<0 .05) .It had been shown that the spheres exhibited a relatively high proportion of cells in G0/G1 phase(84 .19 ± 2 .52)% and low proportion in S phase(7 .18 ± 1 .35)% ;when com‐pared to SW620 cells ,G0/G1 phase(63 .02 ± 6 .73)% ,S phase(20 .89 ± 3 .84)% (P<0 .05) .Adipocytes drops from spheres and ad‐herent cells were calculated under microscope after been cultured in adipocytes liquid for 7 d .The number of adipocytes drops in spheres was (583 .80 ± 77 .69) ,but(169 .20 ± 26 .43)in SW620 adherent(P<0 .05) .2 × 104 sphere cells injected subcutaneously into the lateral root of one posterior limb of a nude mouse fromed transplantation tumor after 4 weeks ,and the tumor size was (2 279 .98 ± 346 .27) mm3 ;2 × 105 adherent cells injected subcutaneously into the lateral root of one posterior limb of a nude mouse fromed transplantation tumor after 4 weeks ,and the tumor size was(889 .75 ± 78 .79) mm3 ,(P<0 .05) .Conclusion In this study , it is successfully observed that SW 620 cells being cultured in serum‐free medium resulted in spheres rapidly and the spheres are se‐rially passaged with relatively high efficiency .The spheres show the characteristics of the cancer stem cell .
