Glioma-derived endothelial cells promote glioma cells migration via extracellular vesicles-mediated transfer of MYO1C

https://doi.org/10.1016/j.bbrc.2020.02.017Get rights and content

Highlights

  • GhEC-EV significantly increases GC migration than NhEC-EV.

  • MYO1C is specifically expressed in GhEC-EV.

  • MYO1C plays an important role in GhEC-EV-induced GC migration.

  • MYO1C was detected in glioma cerebrospinal fluid.

Abstract

Extracellular vesicles (EV), as the intercellular information transfer molecules which can regulate the tumor microenvironment, promote migration and tumor progression. Previous studies reported that EV from endothelial cells was used to guide the fate and survival of gliomas, but many researches focus on normal human endothelial cells (NhEC) rather than tumor-derived endothelial cells. Our laboratory isolated human endothelial cells from glioma issue (GhEC). We have previously demonstrated that EV from GhEC and NhEC, which both can promote glioma stem cells (GSC) proliferation and tumorsphere formation in vitro and tumourigenicity in vivo by the transfer of CD9. However, NhEC-EV or GhEC-EV could suppress glioma cells (GC) proliferation in vitro. It demonstrates the undifferentiated impact of EV. Here, we first compared GhEC-EV proteins with NhEC-EV (Screening criteria: GhEC-EV/NhEC-EV, FC > 1.5), and obtained 70 differential expression proteins, most of which were associated with invasion and migration. We found that GhEC or GhEC-EV preferred promoting GC migration than treating with NhEC or NhEC-EV. In terms of mechanism, we further revealed that EV-mediated transfer of MYO1C induced glioma cell LN229 migration. Knockdown of MYO1C in GhEC or GhEC-EV suppressed this effect. Overexpression of MYO1C promoted migration on the contrary. MYO1C was also detected in glioma cerebrospinal fluid (CSF), which is more suitable as a liquid biopsy biomarker and contributes to early diagnosis and monitoring in glioma. Our findings provide a new protein—MYO1C in EV to target tumor blood vessels, and bring a new point-cut to the treatment of gliomablastoma (GBM).

Introduction

GBM is a highly vascularized tumor in which the tumor vascular system is abnormal in almost every aspect of its structure and function [1]. Affected by tumor microenvironment, tumor blood vessels and their endothelial cells are significantly different from normal blood vessels and normal endothelial cells in morphology, function, protein expression, and gene level [[2], [3], [4]]. The interaction between tumor cells and stromal cells (such as vascular endothelial cells) plays a vital role in the growth, migration, and metastasis of cancer [[5], [6], [7]]. Previous studies indicated that endothelial cells, peripheral cells and astrocytes could form neurovascular units to support the progression of gliomas [8].

Extracellular vesicles (EV), which diameter is between 50 nm and 1000 nm [9]. It is widely distributed in body fluids and mediates many biological and cellular functions, such as cell-to-cell communication. It can carry and transmit important signal molecules to neighboring cells, such as RNA, proteins, nucleic acids, etc [9]. EV is related to the occurrence and progress of various diseases [10]. It has also been reported that EV is involved in the transmission of information between cells [11], and possess diagnosis and therapeutic potential [12]. There are many articles on RNA components of EV [13], few studies pay attention on proteomics of EV. And the mechanism of EV is very complicated.

Our laboratory previously illustrated that both NhEC-EV and GhEC-EV could promote GSC self-renewal, proliferation and tumor globule formation in vitro, and promote tumor growth in vivo by passing CD9. Nevertheless, NhEC-EV or GhEC-EV could suppress glioma cells (GC) proliferation in vitro [14]. It demonstrates the undifferentiated impact of EV. The purpose of this article is to compare the functionally different effects of GC treated with NhEC-EV and GhEC-EV in order to find a new method to target the elimination of tumor vascular endothelium. We found that MYO1C was specifically expressed in GhEC-EV and promoted GC migration.

Section snippets

Human samples and cell culture

Cerebrospinal fluid (CSF) samples (Glioma, n = 8; Meningioma, n = 1; Schwannoma, n = 1) were provided by Yanwei Liu from Beijing Tiantan Hospital. Primary Human Brain Microvascular Endothelial Cells (ACBRI 376; NhEC) were purchased from Cell systems. Glioma vascular endothelial cells (GhEC) were separated by our lab [14]. EC were maintained in EBM-2 medium containing 2% fetal bovine serum (FBS)-EV-free, 0.1% hydrocortisone, 0.1% R3-IGF, 0.4% hFGF-b, 0.1% VEGF, 0.1% ascorbic acid, 0.1% GA-1000,

Proteomic analysis of GhEC-EV and NhEC-EV

Our lab isolated endothelial cells from glioma tissue previously [14]. Here we named them as glioma human endothelial cells (GhEC). To find differences between GhEC-EV and NhEC-EV. We compared both their appearance and size. Firstly, we detected ALIX, TSG101, CD9, CD63 and Flotillin-1 for EV typical markers, Calnexin for endoplasmic reticulum marker in EC and EC-derived EV (Fig. S1A). Furthermore, EV was identified by Transmission Electron Microscope (TEM) and Nanoparticle-tracking analysis

Discussion

There are significant differences between tumor vascular endothelial cells and normal vascular endothelial cells. Previous studies indicated that brain tumor vascular endothelial cells showed a decrease in tight junction proteins expression (such as claudin-1 and occludin), and an increase in pinocytosis and permeability [18]. Tumor endothelial cells preferred to promote proliferation and motility than normal endothelial cells [19]. These studies suggest that the emergence of vascular

Declaration of competing interest

The authors declare that they have no competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This research was supported by the National Key Research and Development Program of China (2016YFC0902502), the National Sciences Foundation of China (31671316, 21874156), Beijing Nova Program of Science and Technology (Z191100001119137), the CAMS Innovation Fund for Medical Sciences (CIFMS;2016-I2M-1-001, 2017-I2M-2-004, 2017-I2M-3-010). We also thankful to Yanwei Liu from Beijing Tiantan hospital for providing CSF samples.

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