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M2-polarization and the tumoricidal to tumor-promoting transition are commonly observed with tumor-infiltrating macrophages after interplay with cancer cells or/and other stroma cells. Our previous study indicated that macrophage M2-polarization can be induced by extracellular HSP90α (eHSP90α) secreted from endothelial-to-mesenchymal transition-derived cancer-associated fibroblasts. To extend the finding, we herein validated that eHSP90α-induced M2-polarized macrophages exhibited a tumor-promoting activity and the promoted tumor tissues had significant increases in microvascular density but decreases in CD4 + T-cell level. We further investigated the signaling pathways occurring in eHSP90α-stimulated macrophages. When macrophages were exposed to eHSP90α, CD91 and toll-like receptor 4 (TLR4) functioned as the receptor/co-receptor for eHSP90α binding to recruit interleukin (IL)-1 receptor-associated kinases (IRAKs) and myeloid differentiation factor 88 (MyD88), and next elicited a canonical CD91/MyD88-IRAK1/4-IκB kinase α/β (IKKα/β)-nuclear factor-κB (NF-κB)/interferon regulatory factor 3 (IRF3) signaling pathway. Despite TLR4-MyD88 complex-associated activations of IKKα/β, NF-κB and IRF3 being well-known as involved in macrophage M1-activation, our results demonstrated that the CD91-TLR4-MyD88 complex-associated IRAK1/4-IKKα/β-NF-κB/IRF3 pathway was not only directly involved in M2-associated CD163, CD204, and IL-10 gene expressions but also required for downregulation of M1 inflammatory cytokines. Additionally, Janus kinase 2 (JAK2) and tyrosine kinase 2 (TYK2) were recruited onto MyD88 to induce the phosphorylation and activation of the transcription factor signal transducer and activator of transcription-3 (STAT-3). The JAK2/TYK2-STAT-3 signaling is known to associate with tumor promotion. In this study, the MyD88-JAK2/TYK2-STAT-3 pathway was demonstrated to contribute to eHSP90α-induced macrophage M2-polarization by regulating the expressions of M1- and M2-related genes, proangiogenic protein vascular endothelial growth factor, and phagocytosis-interfering factor Sec22b.

Background: Endothelial-to-mesenchymal transition (EndoMT) can provide a source of cancer-associated fibroblasts which contribute to desmoplasia of many malignancies including pancreatic ductal adenocarcinoma (PDAC). We investigated the clinical relevance of EndoMT in PDAC, and explored its underlying mechanism and therapeutic implication., Methods: Expression levels of 29 long non-coding RNAs were analyzed from the cells undergoing EndoMT, and an EndoMT index was proposed to survey its clinical associations in the PDAC patients of The Cancer Genome Atlas database. The observed clinical correlation was further confirmed by a mouse model inoculated with EndoMT cells-involved PDAC cell grafts. In vitro co-culture with EndoMT cells or treatment with the conditioned medium were performed to explore the underlying mechanism. Because secreted HSP90α was involved, anti-HSP90α antibody was evaluated for its inhibitory efficacy against the EndoMT-involved PDAC tumor., Results: A combination of low expressions of LOC340340, LOC101927256, and MNX1-AS1 was used as an EndoMT index. The clinical PDAC tissues with positive EndoMT index were significantly correlated with T4-staging and showed positive for M2-macrophage index. Our mouse model and in vitro cell-culture experiments revealed that HSP90α secreted by EndoMT cells could induce macrophage M2-polarization and more HSP90α secretion to promote PDAC tumor growth. Furthermore, anti-HSP90α antibody showed a potent therapeutic efficacy against the EndoMT and M2-macrophages-involved PDAC tumor growth., Conclusions: EndoMT cells can secrete HSP90α to harness HSP90α-overproducing M2-type macrophages to promote PDAC tumor growth, and such effect can be targeted and abolished by anti-HSP90α antibody.

Elevated macrophage infiltration in tumor tissues is associated with breast cancer metastasis. Cancer cell migration/invasion toward angiogenic microvasculature is a key step in metastatic spread. We therefore studied how macrophages stimulated breast cancer cell interactions with endothelial cells. Macrophages produced cytokines, such as interleukin-8 and tumor necrosis factor-α, to stimulate endothelin (ET) and ET receptor (ETR) expression in breast cancer cells and human umbilical vascular endothelial cells (HUVECs). ET-1 was induced to a greater extent from HUVECs than from breast cancer cells, resulting in a density difference that facilitated cancer cell chemotaxis toward HUVECs. Macrophages also stimulated breast cancer cell adhesion to HUVECs and transendothelial migration, which were repressed by ET-1 antibody or ETR inhibitors. The ET axis induced integrins, such as αV and β1, and their counterligands, such as intercellular adhesion molecule-2 and P-selectin, in breast cancer cells and HUVECs, and antibodies against these integrins efficiently suppressed macrophage-stimulated breast cancer cell interactions with HUVECs. ET-1 induced Ets-like kinase-1 (Elk-1), signal transducer and activator of transcription-3 (STAT-3), and nuclear factor-κB (NF-κB) phosphorylation in breast cancer cells. The use of inhibitors to prevent their phosphorylation or ectopic overexpression of dominant-negative IκBα perturbed ET-1-induced integrin αV and integrin β1 expression. The physical associations of these three transcriptional factors with the gene promoters of the two integrins were furthermore evidenced by a chromatin immunoprecipitation assay. Finally, our mouse orthotopic tumor model revealed an ET axis-mediated lung metastasis of macrophage-stimulated breast cancer cells, suggesting that the ET axis was involved in macrophage-enhanced breast cancer cell endothelial interactions.

M2-polarization and the tumoricidal to tumor-promoting transition are commonly observed with tumor-infiltrating macrophages after interplay with cancer cells or/and other stroma cells. Our previous study indicated that macrophage M2-polarization can be induced by extracellular HSP90α (eHSP90α) secreted from endothelial-to-mesenchymal transition-derived cancer-associated fibroblasts. To extend the finding, we herein validated that eHSP90α-induced M2-polarized macrophages exhibited a tumor-promoting activity and the promoted tumor tissues had significant increases in microvascular density but decreases in CD4+ T-cell level. We further investigated the signaling pathways occurring in eHSP90α-stimulated macrophages. When macrophages were exposed to eHSP90α, CD91 and toll-like receptor 4 (TLR4) functioned as the receptor/co-receptor for eHSP90α binding to recruit interleukin (IL)-1 receptor-associated kinases (IRAKs) and myeloid differentiation factor 88 (MyD88), and next elicited a canonical CD91/MyD88–IRAK1/4–IκB kinase α/β (IKKα/β)–nuclear factor-κB (NF-κB)/interferon regulatory factor 3 (IRF3) signaling pathway. Despite TLR4-MyD88 complex-associated activations of IKKα/β, NF-κB and IRF3 being well-known as involved in macrophage M1-activation, our results demonstrated that the CD91-TLR4-MyD88 complex-associated IRAK1/4−IKKα/β−NF-κB/IRF3 pathway was not only directly involved in M2-associated CD163, CD204, and IL-10 gene expressions but also required for downregulation of M1 inflammatory cytokines. Additionally, Janus kinase 2 (JAK2) and tyrosine kinase 2 (TYK2) were recruited onto MyD88 to induce the phosphorylation and activation of the transcription factor signal transducer and activator of transcription-3 (STAT-3). The JAK2/TYK2−STAT-3 signaling is known to associate with tumor promotion. In this study, the MyD88−JAK2/TYK2−STAT-3 pathway was demonstrated to contribute to eHSP90α-induced macrophage M2-polarization by regulating the expressions of M1- and M2-related genes, proangiogenic protein vascular endothelial growth factor, and phagocytosis-interfering factor Sec22b. [ABSTRACT FROM AUTHOR]