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M2 bone marrow-derived macrophage-derived exosomes shuffle microRNA-21 to accelerate immune escape of glioma by modulating PEG3.

ABSTRACT: Background:Growing studies have focused on the role of microRNA-21 (miR-21) in glioma, thus our objective was to discuss the effect of M2 bone marrow-derived macrophage (BMDM)-derived exosomes (BMDM-Exos) shuffle miR-21 on biological functions of glioma cells by regulating paternally expressed gene 3 (PEG3). Methods:Seventy-one cases of human glioma tissues and 30 cases of non-tumor normal brain tissues were collected and stored in liquid nitrogen. PEG3 and miR-21 expression in glioma tissues was tested. The fasting venous blood of glioma patients and healthy control was collected and centrifuged, and then the supernatant was stored at -?80 °C refrigerator. The contents of interferon (IFN)-? and transforming growth factor-?1 (TGF-?1) in serum were tested by ELISA. Glioma cells and normal glial cells were cultured to screen the target cells for further in vitro experiments. BMDM-Exos was obtained by ultra-high speed centrifugation and then was identified. BMDM-Exos was co-cultured with U87 cells to detect the biological functions. The fasting venous blood of glioma patients was extracted and treated with ethylene diamine tetraacetic acid-K2 anti-freezing, and then CD8+T cells were isolated. CD8+T cells were co-cultured with U87 cells to detect the CD8+T proliferation, cell cytotoxic activity, U87 cell activity, as well as IFN-? and TGF-?1 levels. Moreover, BALB/c-nu/nu mice was taken, and the human-nude mouse glioma orthotopic transplantation model was established with U87 cells, and then mice were grouped to test the trends in tumor growth. The brain of mice (fixed by 10% formaldehyde) was sliced to detect the expression of Ki67 and proliferating cell nuclear antigen (PCNA). The spleen of mice was taken to prepare single-cell suspension, and the percentage of T lymphocytes in spleen to CD8+T cells was detected. Results:PEG3 expression was decreased and miR-21 expression was increased in glioma cells and tissues. Depleting miR-21 or restoring PEG3 suppressed growth, migration and invasion as well as accelerated apoptosis of glioma cells, also raised CD8+T proliferation, cell cytotoxic activity, and IFN-? level as well as decreased U87 cell activity and TGF-?1 level. BMDM-Exos shuttle miR-21 promoted migration, proliferation and invasion as well as suppressed apoptosis of glioma cells by reducing PEG3. Exosomes enhanced the volume of tumor, Ki67 and PCNA expression, reduced the percentage of CD8+T cells in glioma mice. Conclusion:BMDM-Exos shuffle miR-21 to facilitate invasion, proliferation and migration as well as inhibit apoptosis of glioma cells via inhibiting PEG3, furthermore, promoting immune escape of glioma cells.

SUBMITTER: Yang F

PROVIDER: S-EPMC7099792 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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M2 bone marrow-derived macrophage-derived exosomes shuffle microRNA-21 to accelerate immune escape of glioma by modulating PEG3.

Yang Fan F Wang Tiecheng T Du Peng P Fan Haitao H Dong Xushuai X Guo Hua H

Cancer cell international 20200327

<h4>Background</h4>Growing studies have focused on the role of microRNA-21 (miR-21) in glioma, thus our objective was to discuss the effect of M2 bone marrow-derived macrophage (BMDM)-derived exosomes (BMDM-Exos) shuffle miR-21 on biological functions of glioma cells by regulating paternally expressed gene 3 (PEG3).<h4>Methods</h4>Seventy-one cases of human glioma tissues and 30 cases of non-tumor normal brain tissues were collected and stored in liquid nitrogen. PEG3 and miR-21 expression in gl ...[more]

PMID: 32231463

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Project description:Rationale: During the development of atherosclerosis, macrophages secrete exosomes that regulate vascular smooth muscle cells (VSMCs); however, whether nicotine, a major constituent of cigarettes, can modulate this communication in the context of atherogenesis remains to be further studied. In this study, we hypothesized that nicotine induces macrophages to secrete atherogenic exosomes containing microRNAs (miRNAs) to mediate cell-to-cell crosstalk and encourage proatherogenic phenotypes of VSMCs. Methods: In an in vivo study, nicotine was administered subcutaneously to 8-week-old male ApoE-/- mice fed a high-fat diet (HFD) for 12 weeks. Oil red O and hematoxylin and eosin (HE) were used to stain atherosclerotic lesions. Lesion macrophages, VSMCs and exosomes were stained for CD68, α-smooth muscle actin (α-SMA) and CD9, and plaque exosomes were observed by transmission electron microscopy (TEM). Exosomes derived from control macrophages (M-Exos) and from nicotine-treated macrophages (NM-Exos) were isolated by ultracentrifugation, purified by sucrose density gradient centrifugation and characterized based on specific morphology and surface markers. The IVIS® Spectrum in vivo imaging system showed the biodistribution of NM-Exos and M-Exos in circulation. Chitosan hydrogel-incorporated exosomes were applied to simulate exosome secretion in situ. Scratch wound assay, transwell assay and EdU staining were conducted to assess the effects of NM-Exos on the migration and proliferation of mouse VSMCs. RNA-seq was performed to determine the miRNA profiles of M-Exos and NM-Exos. Quantitative real-time PCR (qRT-PCR) analysis was conducted to detect the expression levels of miRNAs and mRNAs. The roles of the candidate miRNA and its target gene were assessed using specific RNA inhibitors, siRNAs and miRNA mimics. Western blotting was used to detect candidate protein expression levels. A dual-luciferase reporting system was utilized to confirm the binding of a specific miRNA to its target gene. Results: Nicotine induced atherosclerotic lesion progression and resulted in plaque exosome retention in vivo. The biodistribution of NM-Exos showed that plaque-resident exosomes might be secreted in situ. VSMCs cocultured in vitro with nicotine-stimulated macrophages presented an increased capacity for migration and proliferation, which was exosome-dependent. In addition, isolated NM-Exos helped promote VSMC migration and proliferation. miRNA profiling showed that miR-21-3p was enriched in NM-Exos, and this miRNA was shown to play a key role in regulating NM-Exos-induced effects by directly targeting phosphatase and tension homologue (PTEN). Conclusion: Exosomal miR-21-3p from nicotine-treated macrophages may accelerate the development of atherosclerosis by increasing VSMC migration and proliferation through its target PTEN.

Dataset Information

M2 bone marrow-derived macrophage-derived exosomes shuffle microRNA-21 to accelerate immune escape of glioma by modulating PEG3.

Publications

M2 bone marrow-derived macrophage-derived exosomes shuffle microRNA-21 to accelerate immune escape of glioma by modulating PEG3.

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