PGE2/MFG-E8正反馈环路调控乳腺癌耐药细胞与巨噬细胞交互对话的机制研究

The exploration between the drug resistance and the macrophage phenotype change in tumor microenvironment is intensively studied. However, the key regulators and molecular mechanisms are still undefined. In our study, we found that there is a induction and promotion between the breast-resistant cells and macrophage. PGE2 secretion from adriamycin treated-MCF-7/ADR resistant cell promote M2 polarization. Milk fat globule- EGF factor 8 (MFG-E8) secretions is substantially elevated in PGE2-induce M2-like macrophages. In turn, MFG-E8 promotes drug resistance and PGE2 secretion of breast cancer cells. Based on the above work, we propose an innovative notion and idea of that the continuous positive feedback loop between PGE2 and MFG-E8 might be the molecular mechanisms of interaction between resistant cells and macrophages. In this project, to clarify underlying mechanisms of the positive feedback interaction of PGE2 and MFG-E8 regulate the cross talk between breast cancer resistant cells and macrophages, we explored the molecular mechanism of PGE2-induced M2-like macrophages through which the regulation of C/EBPβ binding with MFG-E8 and MFG-E8 promotes the maintain of PGE2-mediated breast cancer cell resistance by sustained activation of STAT3 / NF-κB / COX-2 pathway. This study will lay the foundation of theory and experiment from a new perspective for further research in the tumor microenvironment-mediated drug resistance of breast cancer cells.

肿瘤微环境中巨噬细胞表型转化影响药物敏感性受到关注，但巨噬细胞与耐药细胞间作用方式、关键调控因子及机制仍未阐明。课题组发现乳腺癌耐药细胞与巨噬细胞间存在相互诱导和促进作用；阿霉素通过促进MCF7/ADR耐药细胞PGE2分泌诱导巨噬细胞M2极化和MFG-E8分泌，而巨噬细胞分泌MFG-E8反馈诱导乳腺癌耐药和PGE2分泌；基于上述基础，申请者创新性提出PGE2/MFG-E8间相互促进、不断循环的正反馈作用可能是耐药细胞与巨噬细胞间互动调控的分子机制。本项目拟通过证实PGE2调控C/EBPβ促进MFG-E8表达诱导巨噬细胞M2极化的分子机制、MFG-E8通过持续激活STAT3/NF-κB/COX-2途径促进PGE2介导乳腺癌细胞耐药性维持，阐明PGE2与MFG-E8相互作用正反馈调控乳腺癌耐药细胞与巨噬细胞交互对话的分子机制，从全新角度为深入探讨肿瘤微环境介导的乳腺癌耐药性奠定理论与实验基础。

本项目拟研究乳腺癌阿霉素耐药性产生与巨噬细胞表型转化之间的潜在联系。我们在体外共培养TAM和乳腺癌细胞用于评估TAM对乳腺癌阿霉素耐药性的影响，使用流式细胞术，ELISA及WB等以揭示其潜在机制，发现乳腺癌阿霉素耐药细胞条件培养巨噬细胞后诱导巨噬细胞M2表型转化，同时乳腺癌耐药细胞上清液中前列腺素E2（PGE2）明显高表达，PGE2诱导的M2型巨噬细胞又进一步增加乳腺癌细胞对阿霉素的化学抗性。PGE2处理的巨噬细胞后C/EBPβ和MFG-E8的表达增加， MFG-E8启动子区存在C/EBPβ的结合位点，M2巨噬细胞高分泌的MFG-E8进一步促进乳腺癌细胞阿霉素耐药的同时COX-2，NF-kBp65和STAT3表达增加， COX-2启动子区存在NF-kBp65和STAT3的结合位点。以上研究结果提示乳腺癌阿霉素耐药细胞通过PGE2促进巨噬细胞M2表型极化，PGE2激活巨噬细胞中的C/EBPβ促进MFG-E8表达，MFG-E8通过NF-kB/STAT3-COX-2通路进一步诱导乳腺癌细胞对阿霉素的化学抗性和PGE2的分泌，形成正反馈循环，揭示乳腺癌微环境中巨噬细胞与肿瘤细胞之间的新的交互对话。