Yao Zhang

Background and Aims The development and progression of hepatocellular carcinoma (HCC) is dependent on its local microenvironment. Tumor‐associated macrophages (TAMs) are deemed a key factor for the tumor microenvironment and attribute to contribute to tumor aggressiveness. However, the detailed mechanism underlying the pro‐metastatic effect of TAMs on HCC remains undefined. Approach and Results The present study proved that TAMs were enriched in HCC. TAMs were characterized by an M2‐polarized phenotype and accelerated the migratory potential of HCC cells in vitro and in vivo . Furthermore, we found that M2‐derived exosomes induced TAM‐mediated pro‐migratory activity. With the use of mass spectrometry, we identified that integrin, α M β 2 (CD11b/CD18), was notably specific and efficient in M2 macrophage–derived exosomes (M2 exos). Blocking either CD11b and/or CD18 elicited a significant decrease in M2 exos–mediated HCC cell metastasis. Mechanistically, M2 exos mediated an intercellular transfer of the CD11b/CD18, activating the matrix metalloproteinase‐9 signaling pathway in recipient HCC cells to support tumor migration. Conclusions Collectively, the exosome‐mediated transfer of functional CD11b/CD18 protein from TAMs to tumor cells may have the potency to boost the migratory potential of HCC cells, thus providing insights into the mechanism of tumor metastasis.

BACKGROUND: Metastasis is a key feature of malignant tumors and significantly contributes to their high mortality, particularly in hepatocellular carcinoma (HCC). Therefore, it is imperative to explore the mechanism of tumor metastasis. Recently, tumor-associated macrophages (TAMs) have been demonstrated to promote tumor progression, while TAM-derived molecules involved in HCC metastasis warrant further investigation. METHODS: THP-1 was treated with IL-4 (Interleukin-4) and IL-13 (Interleukin-13) for M2 polarized macrophages. Exosomes derived from M2 macrophages were characterized. Then, HCC cells or human umbilical vein endothelial cells (HUVECs) were co-cultured with M2 macrophages or treated with M2 macrophage-secreted exosomes. Next, Transwell®, Scratch assay, tube formation, and endothelial permeability assays were performed. Moreover, RT-PCR, western blotting, immunofluorescence, and ELISA were used to assess mRNA and protein expression levels. Finally, the miRNA expression profiles of exosomes derived from M2 and M0 macrophages were analyzed. RESULTS: M2 macrophage infiltration was correlated with metastasis and a poor prognosis in HCC patients. M2-derived exosomes were absorbed by HCC and HUVEC cells and promoted the epithelial-mesenchymal transition (EMT), vascular permeability, and angiogenesis. Notably, MiR-23a-3p levels were significantly higher in M2-derived exosomes and hnRNPA1 mediated miR-23a-3p packaging into exosomes. Phosphatase and tensin homolog (PTEN) and tight junction protein 1 (TJP1) were the targets of miR-23a-3p, as confirmed by luciferase reporter assays. Lastly, HCC cells co-cultured with M2-derived exosomes secreted more GM-CSF, VEGF, G-CSF, MCP-1, and IL-4, which in turn further recruited M2 macrophages. CONCLUSIONS: Our findings suggest that M2 macrophage-derived miR-23a-3p enhances HCC metastasis by promoting EMT and angiogenesis, as well as increasing vascular permeability. Video Abstract.

Exosome-based immunotherapy for cancer holds promise, but needs improvements, especially for tumor-derived exosomes. We investigated, whether exosomes derived from IL-12-anchored human renal cancer cells could enhance their immunogenicity and increase induction of specific antitumor response. A mammalian co-expression plasmid of glycolipid-anchored-IL-12 (GPI-IL-12) was constructed by subcloning IL-12A chain gene (P35 subunit) and a fusion gene containing GPI-anchor signal sequence of human placental alkaline phosphatase-1 (hPLAP-1) and IL-12B chain gene (P40 subunit) in pBudCE4.1. Then exosomes were prepared from renal cancer cells modified to express GPI-IL-12. The results showed that exosomes derived from IL-12-anchored renal cancer cells expressed renal cell carcinoma-associated antigen G250 and GPI-IL-12. The incorporation of GPI-IL-12 onto exosomes (exosomes-GPI-IL-12, EXO/IL-12) significantly promotes proliferation of T cells, and subsequently increased the release of IFN-gamma. Notably, stimulation with EXOs/IL-12 could efficiently induce antigen-specific cytotoxic T lymphocytes (CTLs), resulting in more significant cytotoxic effects in vitro. These results suggested that exosomes derived from IL-12-anchored renal cancer cells bore GPI-IL-12 and G250, which have tumor rejection antigen with enhanced immunogenicity and antitumor effects, representing a novel strategy of exosomes-based vaccine for renal cell carcinoma.