The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200030, P.R. China.
Shanghai iCELL Biotechnology Co., Ltd., Shanghai 200333, P.R. China.
Int J Oncol. 2019 Jul;55(1):179-190. doi: 10.3892/ijo.2019.4794. Epub 2019 May 3.
Epithelial ovarian cancer (EOC) is the most lethal of all gynecologic tumors. Cancer spheroid culture is a widely used model to study cancer stem cells. Previous studies have demonstrated the effectiveness of cytokine‑induced killer (CIK) cell‑based therapies against cancer and cancer stem cells. However, it is not clear how EOC spheroid cells respond to CIK‑mediated cellular lysis, and the mechanisms involved have never been reported before. A flow cytometry‑based method was used to evaluate the anti‑cancer effects of CIK cells against adherent A2780 cells and A2780 spheroids. To demonstrate the association between hypoxia inducible factor‑1α (HIF1A) and intercellular adhesion molecule‑1 (ICAM‑1), two HIF1A short hairpin RNA (shRNA) stable transfected cell lines were established. Furthermore, the protein expression levels of hypoxia/HIF1A‑associated signaling pathways were evaluated, including transforming growth factor‑β1 (TGF‑β1)/mothers against decapentaplegic homologs (SMADs) and nuclear factor‑κB (NF‑κB) signaling pathways, comparing A2780 adherent cells and cancer spheroids. Flow cytometry revealed that A2780 spheroid cells were more resistant to CIK‑mediated cellular lysis, which was partially reversed by an anti‑ICAM‑1 antibody. HIF1A was significantly upregulated in A2780 spheroids compared with adherent cells. Using HIF1A shRNA stable transfected cell lines and cobalt chloride, it was revealed that hypoxia/HIF1A contributed to downregulation of ICAM‑1 in A2780 spheroid cells and adherent cells. Furthermore, hypoxia/HIF1A‑associated signaling pathways, TGF‑β1/SMADs and NF‑κB, were activated in A2780 spheroid cells by using western blotting. The findings indicate that EOC stem‑like cells resist the CIK‑mediated cellular lysis via HIF1A‑mediated downregulation of ICAM‑1, which may be instructive for optimizing and enhancing CIK‑based therapies.
上皮性卵巢癌（EOC）是所有妇科肿瘤中最致命的一种。癌细胞球体培养是研究癌症干细胞的常用模型。先前的研究已经证明了细胞因子诱导的杀伤（CIK）细胞疗法对癌症和癌症干细胞的有效性。然而，目前尚不清楚 EOC 球体细胞如何对 CIK 介导的细胞裂解产生反应，并且以前从未报道过涉及的机制。本研究采用基于流式细胞术的方法评估 CIK 细胞对贴壁 A2780 细胞和 A2780 球体的抗癌作用。为了证明缺氧诱导因子-1α（HIF1A）与细胞间黏附分子-1（ICAM-1）之间的关联，建立了两个 HIF1A 短发夹 RNA（shRNA）稳定转染细胞系。此外，评估了缺氧/HIF1A 相关信号通路的蛋白表达水平，包括转化生长因子-β1（TGF-β1）/母亲对抗 decapentaplegic 同源物（SMADs）和核因子-κB（NF-κB）信号通路，比较 A2780 贴壁细胞和癌症球体。流式细胞术显示，A2780 球体细胞对 CIK 介导的细胞裂解的抵抗力更强，而抗 ICAM-1 抗体部分逆转了这种作用。与贴壁细胞相比，A2780 球体细胞中 HIF1A 明显上调。使用 HIF1A shRNA 稳定转染细胞系和氯化钴揭示了缺氧/HIF1A 导致 A2780 球体细胞和贴壁细胞中 ICAM-1 的下调。此外，通过 Western blot 分析发现，缺氧/HIF1A 相关信号通路 TGF-β1/SMADs 和 NF-κB 在 A2780 球体细胞中被激活。研究结果表明，EOC 类干细胞通过 HIF1A 介导的 ICAM-1 下调抵抗 CIK 介导的细胞裂解，这可能有助于优化和增强基于 CIK 的治疗方法。
