Molecular mechanisms of IL-33–mediated stromal interactions in cancer metastasis
Patrik Andersson, Yunlong Yang, Kayoko Hosaka, Yin Zhang, Carina Fischer, Harald Braun, Shuzhen Liu, Guohua Yu, Shihai Liu, Rudi Beyaert, Mayland Chang, Qi Li, Yihai Cao
Patrik Andersson, Yunlong Yang, Kayoko Hosaka, Yin Zhang, Carina Fischer, Harald Braun, Shuzhen Liu, Guohua Yu, Shihai Liu, Rudi Beyaert, Mayland Chang, Qi Li, Yihai Cao
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Research Article Cell biology Oncology

Molecular mechanisms of IL-33–mediated stromal interactions in cancer metastasis

Abstract

Molecular mechanisms underlying the cancer stroma in metastasis need further exploration. Here, we discovered that cancer-associated fibroblasts (CAFs) produced high levels of IL-33 that acted on tumor-associated macrophages (TAMs), causing them to undergo the M1 to M2 transition. Genomic profiling of metastasis-related genes in the IL-33–stimulated TAMs showed a >200-fold increase of MMP9. Signaling analysis demonstrated the IL-33-ST2-NF-κB-MMP9-laminin pathway that governed tumor stroma–mediated metastasis. In mouse and human fibroblast-rich pancreatic cancers, genetic deletion of IL-33, ST2, or MMP9 markedly blocked metastasis. Pharmacological inhibition of NF-κB and MMP9 also blocked cancer metastasis. Deletion of IL-33, ST2, or MMP9 restored laminin, a key basement membrane component associated with tumor microvessels. Together, our data provide mechanistic insights on the IL-33-NF-κB-MMP9-laminin axis that mediates the CAF-TAM–committed cancer metastasis. Thus, targeting the CAF-TAM-vessel axis provides an outstanding therapeutic opportunity for cancer treatment.

Authors

Patrik Andersson, Yunlong Yang, Kayoko Hosaka, Yin Zhang, Carina Fischer, Harald Braun, Shuzhen Liu, Guohua Yu, Shihai Liu, Rudi Beyaert, Mayland Chang, Qi Li, Yihai Cao

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Figure 3

Transcriptional regulation of the Mmp9 promoter activity by IL-33- NF-κB signaling.

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Transcriptional regulation of the Mmp9 promoter activity by IL-33- NF-κB...
(A) Western immunoblot analysis of phosphorylation of IκBα, Erk, and p38 in vehicle- and IL-33–treated macrophages in the presence or absence of their specific inhibitors. β-Actin served as a loading control (n = 3 samples per group). WA, withaferin A; SB, SB203580. (B) qPCR quantification of Mmp9 mRNA levels of IL-33–stimulated macrophages in the presence or absence of their specific inhibitors (n = 6 samples per group). (C) Western immunoblot analysis of phosphorylation of IκBα in IL-33–stimulated macrophages in the presence or absence of a soluble ST2 (sST2). β-Actin served as a loading control (n = 3 samples per group). (D) A network scheme demonstrating protein-protein interactions linking IL-33 to NF-κB. Irak, IL-1 receptor–associated kinase; MyD88, myeloid differentiation primary response 88. (E) Immunostaining of NF-κB (green) in IL-33–stimulated macrophages at 5-, 30-, or 60-minute time points. Cell nuclei were counterstained with DAPI (red; scale bar: 25 μm). Overlapping yellow signals were randomly quantified (n = 3 fields per group). (F) Immunostaining of NF-κB (green) in IL-33–stimulated macrophages in the presence or absence of different concentrations of withaferin A. Cell nuclei were counterstained with DAPI (red; scale bar: 50 μm). Overlapping yellow signals were quantified (n = 4 random fields per group). (G) Detection of MMP9 protease activity by a gelatin-based zymography assay in conditional medium derived from IL-33–stimulated macrophages in the presence or absence of different concentrations of withaferin A (n = 4 samples per group). (H) qPCR quantification of Mmp9 mRNA levels in IL-33–stimulated macrophages in the presence or absence of different concentrations of withaferin A (n = 6 samples per group). (I) Schematic indicating 5 NF-κB–binding sites located in the proximal region of the mouse Mmp9 promoter. (J) Mmp9 promoter activity in macrophages by the luciferase reporter assay. Relative luciferase activity was measured after IL-33 stimulation in the presence or absence of different concentrations of withaferin A (n = 3 samples per group). (K) Schematic showing the IL-33-ST2-NF-κB-MMP9 signaling pathway. IKK, IκB kinase. Mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001, Student’s t test.