Extraction of Exosomes from Glioblastoma Multiforme Patients’ Blood Plasma

Introduction. Glioblastoma multiforme (GBM) is the most common and aggressive form of primary malignant brain tumour in adults associated with a poor prognosis. Exosomes have been shown to be useful non-invasive biomarkers for the diagnosis and prognosis of tumours, GBM included. Exosomes play a role of biological carriers which can perform various tasks through various signalling pathways of carcinogenesis, such as PI3K/AKT, SOX2, PTEN, ERK and STAT3.

Materials and methods. Exosomes were isolated from blood plasma taken from patients diagnosed with GBM prior to surgical resection.

Results and discussion. Plasma exosomes from patients with GBM had spherical shape and varied in size from 40 to 100 nm matching the exosomes’ morphological characteristics. The combination of ultrafiltration and double ultracentrifugation makes it possible to extract exosome examples from plasma without the presence of contaminating particles over 100 nm in size; the shape and size of these vesicles match the characteristics of exosomes isolated from other biological fluids.

Conclusion. The experimental protocol for the extraction of exosomes from GBM patients’ plasma described here proves effective as a method used to ensure the purity of exosomes. Applying this method offers further opportunities for research into the role of exosomes in GBM pathogenesis. Equally this method can be used in research involving other human pathologies.

1. Tian W., Liu S., Li B. Potential role of exosomes in cancer metastasis. Biomed Res Int. 2019;2019:4649705. DOI: 10.1155/2019/4649705

2. Osaki M., Okada F. Exosomes and their role in cancer progression. Yonago Acta Med. 2019;62(2):182–90. DOI: 10.33160/yam.2019.06.002

3. Zhang H., Wang L., Li C., Yu Y., Yi Y., Wang J., et al. Exosomeinduced regulation in inflammatory bowel disease. Front Immunol. 2019;10:1464. DOI: 10.3389/fimmu.2019.01464

4. Lema D.A., Burlingham W.J. Role of exosomes in tumour and transplant immune regulation. Scand J Immunol. 2019 Jul 8:e12807. DOI: 10.1111/sji.12807

5. Kelemen E., Danis J., Göblös A., Bata-Csörgő Z., Széll M. Exosomal long non-coding RNAs as biomarkers in human diseases. EJIFCC. 2019;30(2):224–36. PMID: 31263395

6. Wang X., Zhong W., Bu J., Li Y., Li R., Nie R., et al. Exosomal protein CD82 as a diagnostic biomarker for precision medicine for breast cancer. Mol Carcinog. 2019;58(5):674–85. DOI: 10.1002/mc.22960

7. Laidlaw K.M.E., MacDonald C. Endosomal trafficking of yeast membrane proteins. Biochem Soc Trans. 2018;46(6):1551–58. DOI: 10.1042/ BST20180258

8. Chutipongtanate S., Greis K.D. Multiplex biomarker screening assay for urinary extracellular vesicles study: a targeted label-free proteomic approach. Sci Rep. 2018;8(1):15039. DOI: 10.1038/s41598-018-33280-7

9. Bowers K. RNA interference-mediated inhibition of ESCRT in mammalian cells. Methods Mol Biol. 2019;1998:305–18. DOI: 10.1007/978- 1-4939-9492-2_22

10. Sharma S., Scholz-Romero K., Rice G.E., Salomon C. Methods to enrich exosomes from conditioned media and biological fluids. Methods Mol Biol. 2018;1710:103–15. DOI: 10.1007/978-1-4939-7498-6_8

11. Koritzinsky E.H., Street J.M., Star R.A., Yuen P.S. Quantification of exosomes. J Cell Physiol. 2017;232(7):1587–90. DOI: 10.1002/jcp.25387

12. Luarte A., Bátiz L.F., Wyneken U., Lafourcade C. Potential therapies by stem cell-derived exosomes in CNS diseases: focusing on the neurogenic niche. Stem Cells Int. 2016;2016:5736059. DOI: 10.1155/2016/5736059

13. Moon J.M., Xu L., Giffard R.G. Inhibition of microRNA-181 reduces forebrain ischemia-induced neuronal loss. J Cereb Blood Flow Metab. 2013;33(12):1976–82. DOI: 10.1038/jcbfm.2013.157

14. Elsharkawi F., Elsabah M., Shabayek M., Khaled H. Urine and serum exosomes as novel biomarkers in detection of bladder cancer. Asian Pac J Cancer Prev. 2019;20(7):2219–24. DOI: 10.31557/APJCP.2019.20.7.2219

15. Zhou N., Li C., Zhou Y., Liu J., Su X., Qin H., et al. Potential markers from serum-purified exosomes for detecting oral squamous cell carcinoma metastasis. Cancer Epidemiol Biomarkers Prev. 2019 Jul 26;pii: cebp.1122.2018. DOI: 10.1158/1055-9965.EPI-18-1122

16. Doyle L.M., Wang M.Z. Overview of extracellular vesicles, their origin, composition, purpose, and methods for exosome isolation and analysis. Cells. 2019;8(7):E727. DOI: 10.3390/cells8070727

17. Malla B., Aebersold D.M., Dal Pra A. Protocol for serum exosomal miRNAs analysis in prostate cancer patients treated with radiotherapy. J Transl Med. 2018;16(1):223. DOI: 10.1186/s12967-018-1592-6