Glioma exosomes mediate the expansion and function of myeloid-derived suppressor cells through microRNA-29a/Hbp1 and microRNA-92a/Prkar1a pathways
Xiaofan Guo
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
First author's name: Xiaofan Guo and Wei Qiu.Search for more papers by this authorWei Qiu
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
First author's name: Xiaofan Guo and Wei Qiu.Search for more papers by this authorJian Wang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Biomedicine, University of Bergen, Bergen, Norway
Search for more papers by this authorQinglin Liu
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorMingyu Qian
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorShaobo Wang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorZongpu Zhang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorXiao Gao
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorZihang Chen
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorQindong Guo
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorJianye Xu
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorCorresponding Author
Hao Xue
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Correspondence to: Gang Li, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, Tel.: 086-0531-82166615, Fax: 086-0531-82166615, E-mail: [email protected]; or Hao Xue, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, E-mail: [email protected]Search for more papers by this authorCorresponding Author
Gang Li
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Correspondence to: Gang Li, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, Tel.: 086-0531-82166615, Fax: 086-0531-82166615, E-mail: [email protected]; or Hao Xue, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, E-mail: [email protected]Search for more papers by this authorXiaofan Guo
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
First author's name: Xiaofan Guo and Wei Qiu.Search for more papers by this authorWei Qiu
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
First author's name: Xiaofan Guo and Wei Qiu.Search for more papers by this authorJian Wang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Biomedicine, University of Bergen, Bergen, Norway
Search for more papers by this authorQinglin Liu
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorMingyu Qian
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorShaobo Wang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorZongpu Zhang
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorXiao Gao
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorZihang Chen
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorQindong Guo
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorJianye Xu
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Search for more papers by this authorCorresponding Author
Hao Xue
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Correspondence to: Gang Li, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, Tel.: 086-0531-82166615, Fax: 086-0531-82166615, E-mail: [email protected]; or Hao Xue, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, E-mail: [email protected]Search for more papers by this authorCorresponding Author
Gang Li
Institute of Brain and Brain-Inspired Science, Shandong University, Jinan, Shandong Province, People's Republic of China
Shandong Provincial Key Laboratory of Brain Function Remodeling, Shandong University, Jinan, Shandong Province, People's Republic of China
Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, Shandong Province, People's Republic of China
Correspondence to: Gang Li, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, Tel.: 086-0531-82166615, Fax: 086-0531-82166615, E-mail: [email protected]; or Hao Xue, Department of Neurosurgery, Qilu Hospital of Shandong University, 107 Wenhua Western Road; Jinan, Shandong 250012, China, E-mail: [email protected]Search for more papers by this authorAbstract
Myeloid-derived suppressor cells (MDSCs) play a pivotal role in mediating the formation of an immunosuppressive environment and assisting tumors in evading the host immune response. However, the mechanism through which tumors manipulate the differentiation and function of MDSCs remains unclear. Here, we report that hypoxia-induced glioma cells can stimulate the differentiation of functional MDSCs by transferring exosomal miR-29a and miR-92a to MDSCs. Our results showed that glioma-derived exosomes (GEXs) can enhance the differentiation of functional MDSCs both in vitro and in vivo, and hypoxia-induced GEXs (H-GEXs) demonstrated a stronger MDSCs induction ability than did normoxia-induced GEXs (N-GEXs). A subsequent miRNA sequencing analysis of N-GEXs and H-GEXs revealed that hypoxia-induced exosomal miR-29a and miR-92a expression induced the propagation of MDSCs. miR-29a and miR-92a activated the proliferation and function of MDSCs by targeting high-mobility group box transcription factor 1 (Hbp1) and protein kinase cAMP-dependent type I regulatory subunit alpha (Prkar1a), respectively. Altogether, the results of our study provide new insights into the role of glioma exosomal miRNAs in mediating the formation of immunosuppressive microenvironments in tumors and elucidate the underlying exosomal miR-29a/miR-92a-based regulatory mechanism responsible for the modulation of functional MDSC induction.
Abstract
What's new?
Myeloid derived suppressor cells (MDSC) suppress the immune system, reducing the effectiveness of immunotherapy in various cancers. These authors investigated how hypoxia promotes the immunosuppressive activity of MDSCs. Glioma cells, they found, use exosomes to transport miRNAs to MDSC progenitor cells, stimulating their differentiation. Hypoxia-induced exosomes exerted a more powerful effect on MDSC proliferation than normoxia-induced exosomes. The miRNAs induced MDSCs by targeting two genes, Hbp1 and Prkar1a. miRNA silencing of Hbp1 induced the cell cycle progression of MDSCs. This is the first study to demonstrate a mechanism by which glioma cells stimulate MDSC expansion.
Supporting Information
| Filename | Description |
|---|---|
| ijc32052-sup-0001-appendixS1.docWord document, 35 KB | Appendix S1: Supplemental Methods. |
| ijc32052-sup-0002-FigureS1.tifTIFF image, 5.2 MB |
Supplementary Figure S1 Exosomes secreted by glioma cells are internalized by MDSCs A. Electron microscopy images of exosomes secreted by different glioma cell lines stimulated with normoxia or hypoxia (N-GEXs or H-GEXs). Scale bar: 100 nm. B. Particle diameter and concentration analysis of GEXs measured by qNano. C. Western blotting analysis of GEXs and cells for the negative exosome marker Calnexin and the positive exosome markers TSG101 and CD9. D. Confocal microscopy images of MDSCs incubated with PKH67 (green)-labeled GEXs derived from the P3 and GL261 glioma cell lines for 0, 6, 24 and 48 h. Phalloidin-TRITC (red) and DAPI (blue) were used to stain MDSCs. Scale bar: 5 μm. E. Bioluminescence imaging confirmed glioma formation, and representative confocal microscopy images validated the internalization of GEXs by MDSCs in vivo at 2 or 4 weeks after the implantation of recombinase-expressing GL261 cells. MDSCs in non-injected mice or mice implanted with normal GL261 cells were used as controls. Scale bar: 100 μm. The percentage of positive cells (ratio of the number of cells expressing eGFP to the total number of cells observed) was calculated. NS, not significant. F. The bone marrow cells were treated with PKH67 labeled H-GEXs derived from GL261 and prepared for flow cytometry 7 days later. The bone marrow cells that have taken the PKH67 labeled exosomes were shown (F). The percentage of MDSCs in bone marrow cells have taken PKH67 labeled exosomes (P2) or not (P1) were shown (G). The percentage of MDSCs in all bone marrow cells was also shown (P1 and P2). *P < 0.05, **P < 0.01, ***P < 0.005, NS, not significant. (n = 3; mean ± s.e.m) |
| ijc32052-sup-0003-FigureS2.tifTIFF image, 869 KB |
Supplementary Figure S2 miRNA expression in GEXs A. The supernatants of normoxic GL261/G422 cells transfected with control, miR-29a-overexpressing or miR-92a-overexpressing lentiviruses and hypoxic GL261/G422 cells transfected with control, miR-29a-knockout or miR-92a-knockout lentiviruses were collected to isolate exosomes (N-GEXs, N + 29 GEXs, N + 92 GEXs, H-GEXs, H-29KO GEXs, and H-92KO GEXs). Supernatants of hypoxic GL261/G422 cells transfected miR-29a and miR-92a knockout lentiviruses together were also collected to isolate exosomes (H-29-92 double KO GEXs). The miR-29a and miR-92a miRNA expression in these GEXs was detected by qRT-PCR. B. Normal or Cre recombinase-expressing GL261 cells were cultured for 48 h, and the culture medium was used to isolate exosomes. The Cre mRNA expression in these GEXs was detected by qRT-PCR. *P < 0.05, **P < 0.01, ***P < 0.005, NS, not significant. (n = 3; mean ± s.e.m) |
| ijc32052-sup-0004-FigureS3.tifTIFF image, 496.9 KB |
Supplementary Figure S3 The expression of miR-29a and miR-92a in splenic MDSCs The expression of miR-29a and miR-92a in splenic MDSCs isolated from glioma bearing mouse (the mouse was intracranially injected with GL261 glioma cells one months ago) and control mouse (the mouse was intracranially injected with PBS one months ago) was detected by qRT-PCR. *P < 0.05, **P < 0.01, ***P < 0.005, NS, not significant. (n = 3; mean ± s.e.m) |
| ijc32052-sup-0005-FigureS4.tifTIFF image, 639.6 KB |
Supplementary Figure S4 Optiprep density gradient. Hypoxia induced exosomes derived from U87 cells were isolated using an iodixanol density gradient centrifugation. Fractions were subjected to qPCR analysis with primers specific to miR-29a and miR-92a as well as to western blot analysis with the exosome marker protein CD9. |
| ijc32052-sup-0006-FigureS5.tifTIFF image, 1.7 MB |
Supplementary Figure S5 siRNA, mimic, antagomirs and PCR primer sequences A. Sequences of Hbp1, Prkar1a and control siRNAs. B. Sequences of miR-29a-3p mimics, miR-92a-3p mimics, miR-29a-3p antagomir, miR-92a-3p antagomir and control. C. Sequences of PCR primers. |
| ijc32052-sup-0007-FigureS6.tifTIFF image, 3.9 MB | Supplementary Figure S6 The uncropped scans of western blots from Figure 6 |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
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