M. D. Anderson Morgan Welch Inflammatory Breast Cancer Research Program and Clinic, Houston, Texas, USA; Department of Radiation Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas, USA.
Department of Experimental Therapeutics, The University of Texas M. D. Anderson Cancer Center, Houston, Texas.
Int J Radiat Oncol Biol Phys. 2018 Mar 15;100(4):1034-1043. doi: 10.1016/j.ijrobp.2017.11.043. Epub 2017 Dec 7.
To determine the role of macrophage polarization on the response of inflammatory breast cancer (IBC) cells to radiation and whether modulation of macrophage plasticity can alter radiation response.
The human THP-1 monocyte cell line and primary human monocytes isolated from peripheral blood mononuclear cells were differentiated into macrophages and polarized to either an "antitumor" (M1) or a "protumor" (M2) phenotype. These polarized macrophages were co-cultured with IBC cells (SUM149, KPL4, MDA-IBC3, or SUM190) without direct contact for 24 hours, then subjected to irradiation (0, 2, 4, or 6 Gy). Interleukin (IL)4/IL13-induced activation of STAT6 signaling was measured by Western blotting of phospho-STAT6 (Tyr641), and expression of M2 polarization gene markers (CD206, fibronectin, and CCL22) was measured by quantitative polymerase chain reaction.
Expression of M2 polarization markers was higher in M2-polarized macrophages after IL4/IL13 treatment than in control (M0) or M1-polarized macrophages. Co-culture of IBC cell lines with M1-polarized THP-1 macrophages mediated radiosensitivity of IBC cells, whereas co-culture with M2-polarized macrophages mediated radioresistance. Phosphopeptide mimetic PM37, targeting the SH2 domain of STAT6, prevented and reversed IL4/IL13-mediated STAT6 phosphorylation (Tyr641) and decreased the expression of M2 polarization markers. Pretreatment of M2-THP1 macrophages with PM37 reduced the radioresistance they induced in IBC cells after co-culture. Targeted proteomics analysis of IBC KPL4 cells using a kinase antibody array revealed induction of protein kinase C zeta (PRKCZ) in these cells only after co-culture with M2-THP1 macrophages, which was prevented by PM37 pretreatment. KPL4 cells with stable short hairpin RNA knockdown of PRKCZ exhibited lower radioresistance after M2-THP1 co-culture.
These data suggest that inhibition of M2 polarization of macrophages by PM37 can prevent radioresistance of IBC by down-regulating PRKCZ.
确定巨噬细胞极化在炎性乳腺癌（IBC）细胞对辐射反应中的作用，以及调节巨噬细胞可塑性是否可以改变辐射反应。
人 THP-1 单核细胞系和从外周血单核细胞中分离的原代人单核细胞分化为巨噬细胞，并被极化至“抗肿瘤”（M1）或“促肿瘤”（M2）表型。这些极化的巨噬细胞与 IBC 细胞（SUM149、KPL4、MDA-IBC3 或 SUM190）无直接接触共培养 24 小时，然后进行照射（0、2、4 或 6 Gy）。通过 Western 印迹法检测磷酸化 STAT6（Tyr641）测量白细胞介素（IL）4/IL13 诱导的 STAT6 信号激活，通过定量聚合酶链反应测量 M2 极化基因标志物（CD206、纤维连接蛋白和 CCL22）的表达。
IL4/IL13 处理后，M2 极化巨噬细胞中 M2 极化标志物的表达高于对照（M0）或 M1 极化巨噬细胞。IBC 细胞系与 M1 极化 THP-1 巨噬细胞共培养介导 IBC 细胞的放射敏感性，而与 M2 极化巨噬细胞共培养介导放射抵抗。针对 STAT6 的 SH2 结构域的磷酸肽模拟物 PM37 可阻止和逆转 IL4/IL13 介导的 STAT6 磷酸化（Tyr641），并降低 M2 极化标志物的表达。M2-THP1 巨噬细胞用 PM37 预处理可降低共培养后诱导的 IBC 细胞的放射抵抗性。使用激酶抗体阵列对 IBC KPL4 细胞进行靶向蛋白质组学分析显示，仅在与 M2-THP1 巨噬细胞共培养后，这些细胞中才诱导蛋白激酶 C ζ（PRKCZ），而 PM37 预处理可防止其诱导。与 M2-THP1 共培养后具有稳定短发夹 RNA 敲低 PRKCZ 的 KPL4 细胞表现出较低的放射抵抗性。
这些数据表明，PM37 通过抑制巨噬细胞的 M2 极化，可以通过下调 PRKCZ 来防止 IBC 的放射抵抗性。
