Department of Medical Oncology, Wuxi People's Hospital, Affiliated to Nanjing Medical University, Wuxi, PRC.
Lab Invest. 2013 Jul;93(7):844-54. doi: 10.1038/labinvest.2013.69. Epub 2013 Jun 10.
M2-polarized tumor-associated macrophages (TAMs) are key regulators of the link between inflammation and cancer. A negative correlation between infiltration intensity of M2-polarized TAMs and prognosis of pancreatic cancer has been reported. Epithelial-mesenchymal transition (EMT) is an important biological process in the progression of primary tumors toward metastasis. Inflammation-induced EMT has been previously shown, therefore, we hypothesized M2-polarized TAMs could induce EMT in pancreatic cancer. Toll-like receptor 4 (TLR4) signaling has an active role in tumor progression during chronic inflammation and the receptor is primarily expressed on macrophages. Activation of TLR4 on M2-polarized TAMs stimulates an increase in the cytokine interleukin-10 (IL-10); consequently, another aim was to investigate the potential role of TLR4/IL-10 signaling in the EMT of pancreatic cancer. Treatment with IL-4 (20 ng/ml) for 24 h successfully induced the polarization of macrophage cell line RAW 264.7 to M2 phenotype, IL-10(high), IL-12(low), and IL-23(low), and high expression of CD204 and CD206. A coculture system allowed investigation of the roles of M2-polarized TAMs and TLR4/IL-10 signaling in the EMT of Panc-1 and BxPC-3 pancreatic cancer cell lines. Our results showed that coculture with M2-polarized TAMs increased fibroblastic morphology, upregulated mesenchymal markers vimentin and snail at the mRNA and protein levels, and increased proliferation, migration, and metalloproteinase (MMP)2 and MMP9 proteolytic activity in pancreatic cancer cells. Simultaneously, coculture with M2-polarized TAMs decreased the expression of the epithelial marker E-cadherin. Coculture with pancreatic cancer cells increased TLR4 mRNA and protein expression in M2-polarized TAMs. Application of TLR4 siRNA and neutralizing antibodies against TLR4 and IL-10 markedly inhibited E-cadherin reduction and the upregulation of snail and vimentin. Furthermore, activation of TLR4 signaling by lipopolysaccharide profoundly increased the EMT of pancreatic cancer cells. In conclusion, M2-polarized TAMs promoted EMT in pancreatic cancer cells partially through TLR4/IL-10 signaling, suggesting novel therapeutic strategies and enhancing our understanding of M2-polarized TAMs.
M2 极化的肿瘤相关巨噬细胞（TAMs）是炎症与癌症之间联系的关键调节因子。有报道称，M2 极化 TAMs 的浸润强度与胰腺癌的预后呈负相关。上皮-间充质转化（EMT）是原发性肿瘤向转移进展的重要生物学过程。先前已经表明炎症诱导的 EMT，因此，我们假设 M2 极化的 TAMs 可以诱导胰腺癌中的 EMT。Toll 样受体 4（TLR4）信号在慢性炎症中的肿瘤进展中具有积极作用，并且该受体主要在巨噬细胞上表达。M2 极化 TAMs 上 TLR4 的激活刺激细胞因子白细胞介素-10（IL-10）的增加；因此，另一个目的是研究 TLR4/IL-10 信号在胰腺癌 EMT 中的潜在作用。用 IL-4（20ng/ml）处理 24 小时成功地将巨噬细胞系 RAW 264.7 诱导为 M2 表型，IL-10（高），IL-12（低）和 IL-23（低），并且 CD204 和 CD206 的高表达。共培养系统允许研究 M2 极化 TAMs 和 TLR4/IL-10 信号在 Panc-1 和 BxPC-3 胰腺癌细胞 EMT 中的作用。我们的结果表明，与 M2 极化的 TAMs 共培养增加了成纤维细胞形态，在 mRNA 和蛋白水平上上调了间充质标记物波形蛋白和 snail，并增加了胰腺癌细胞的增殖、迁移和金属蛋白酶（MMP）2 和 MMP9 的蛋白水解活性。同时，与 M2 极化的 TAMs 共培养降低了上皮标记物 E-钙粘蛋白的表达。与胰腺癌细胞共培养增加了 M2 极化的 TAMs 中 TLR4 mRNA 和蛋白的表达。应用 TLR4 siRNA 和针对 TLR4 和 IL-10 的中和抗体显著抑制了 E-钙粘蛋白的减少和 snail 和波形蛋白的上调。此外，脂多糖激活 TLR4 信号显著增加了胰腺癌细胞的 EMT。总之，M2 极化的 TAMs 通过 TLR4/IL-10 信号促进了胰腺癌细胞的 EMT，提示了新的治疗策略，并增强了我们对 M2 极化的 TAMs 的理解。
