细胞密度对卵巢癌干细胞悬浮球诱导的影响

发布时间：2018-03-05 17:50

本文选题：卵巢腺癌　切入点：卵巢透明细胞癌　出处：《四川大学学报(医学版)》2017年05期 　论文类型：期刊论文

【摘要】：目的研究细胞密度是否会对从人卵巢透明细胞癌系ES-2和卵巢腺癌细胞A2780中分离的癌干细胞产生影响。方法将ES-2、A2780细胞置于含生长因子、牛血清白蛋白和人胰岛素的无血清培养基中培养,采用连续传代诱导培养癌干细胞。观察不同细胞密度下ES-2、A2780细胞形态变化,流式细胞术分别检测含血清培养基(SSM)组、无血清培养基(SFM)组细胞诱导前后细胞表面标志CD133、CD44表达的改变。软琼脂克隆形成实验检测不同诱导条件下ES-2癌干细胞球生长速度和成瘤能力。结果 2×10~4 mL~(-1)密度下,ES-2细胞能在SFM中存活,但不能形成癌干细胞球;5×10~4、1×10~4 mL~(-1) ES-2细胞能在SFM中存活、增殖并形成癌干细胞球。与贴壁的细胞及诱导前相比,诱导后形成的癌干细胞球(1×10~5 mL~(-1)组、5×10~5 mL~(-1)组)高表达CD133+、CD44+(P0.05),传代后具有更高的增殖能力和克隆形成能力,且5×10~4 mL~(-1)密度下,癌干细胞球成瘤能力更强。A2780细胞分别以1×10~4、3×10~4、5×10~4 mL~(-1)的密度接种于SFM中,10d后,1×10~4、3×10~4 mL~(-1)组中形成细胞团,但3×10~4 mL~(-1)组中形成的细胞团更大,透光性更强,细胞更致密,5×10~4 mL~(-1)组未形成细胞悬浮球。流式细胞术发现A2780细胞3×10~4 mL~(-1)组中细胞与1×10~4 mL~(-1)组相比,CD133~+细胞阳性率更高,差异有统计学意义(P0.05)。3×10~4 mL~(-1)组的细胞生长速度最快。结论细胞密度5×10~4 mL~(-1)和3×10~4 mL~(-1)分别是诱导ES-2、A2780细胞株卵巢肿瘤干细胞球的合适条件。
[Abstract]:Objective to study whether cell density may affect cancer stem cells isolated from human ovarian clear cell carcinoma (ES-2) and ovarian adenocarcinoma cell line A2780. Cancer stem cells were cultured in serum-free medium of bovine serum albumin (BSA) and human insulin. The morphological changes of ES-2A2780 cells were observed under different cell densities, and the serum-containing medium (SSM) was detected by flow cytometry. Before and after induction, the expression of CD133hCD44, a cell surface marker, in serum-free medium (SFM) group was changed. Soft Agar Clone formation assay was used to detect the growth rate and tumorigenic ability of ES-2 cancer stem cell spheres under different induction conditions. Results: at 2 脳 10 ~ 4 mL / L ~ (-1) density, the expression of CD133T CD44 was detected. Cells can survive in SFM, However, ES-2 cells could survive, proliferate and form cancer stem cell spheres in SFM. Compared with adherent cells and pre-induction cells, cancer stem cells could not be formed in 5 脳 10 ~ (4) C ~ (-1) or 1 脳 10 ~ (4) mL ~ (-1) ES-2 cells. The high expression of CD133 CD44 and P0.05 was found in the cancer stem cell spheroids (1 脳 10 ~ (5) mL / L ~ (-1)) group. The cells were subcultured with higher proliferative and clone forming ability, and 5 脳 10 ~ (4) mL / L ~ (-1) density, and the expression of CD _ (44) CD _ (44) was higher than that in the control group (P _ (0.05)), and at 5 脳 10 ~ (4) mL / L ~ (-1) density. The tumor forming ability of cancer stem cells was stronger. A2780 cells were inoculated with 1 脳 10 ~ (4) ~ (3 脳 10 ~ (4)) ~ (5 脳 10 ~ (4) L ~ (-1)) density to form a cell mass in SFM group 10 days after inoculation, but in 3 脳 10 ~ (4) mLL ~ (-1) group, the cell mass was larger and the light transmittance was stronger than that in the group of 3 脳 10 ~ (4) mL ~ (-1) ~ (-1). The positive rate of CD133~ cells in the A2780 cell group was higher than that in the 1 脳 10 ~ 4 mL / L ~ (-1) group, and that in the A2780 cell group was higher than that in the 1 脳 10 ~ (4) mL / L ~ (-1) group, and the positive rate of CD133~ cells in the A2780 cell group was higher than that in the 1 脳 10 ~ (4) mL / L ~ (-1) group. Conclusion the cell density of 5 脳 10 ~ 4 mL ~ (-1) and 3 脳 10 ~ (4) mL / L ~ (-1) are the suitable conditions for inducing ovarial tumor stem cell spheres of ES-2A2780 cell line, respectively.
【作者单位】：四川大学华西第二医院妇产科;出生缺陷与相关妇儿疾病教育部重点实验室(四川大学);
【基金】：四川省学术带头人培养基金(No.2012DTPY0028,No.2013RST004) 四川省卫计委基金(No.130113) 四川省科技厅科技支撑计划项目(No.2014KJT057-2014SZ)资助
【分类号】：R737.31

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