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Interplay Between V-ATPase G1 and Small EV-miRNAs Modulates ERK1/2 Activation in GBM Stem Cells and Nonneoplastic Milieu.

ABSTRACT: The ATP6V1G1 subunit (V1G1) of the vacuolar proton ATPase (V-ATPase) pump is crucial for glioma stem cells (GSC) maintenance and in vivo tumorigenicity. Moreover, V-ATPase reprograms the tumor microenvironment through acidification and release of extracellular vesicles (EV). Therefore, we investigated the role of V1G1 in GSC small EVs and their effects on primary brain cultures. To this end, small EVs were isolated from patients-derived GSCs grown as neurospheres (NS) with high (V1G1^HIGH-NS) or low (V1G1^LOW-NS) V1G1 expression and analyzed for V-ATPase subunits presence, miRNA contents, and cellular responses in recipient cultures. Our results show that NS-derived small EVs stimulate proliferation and motility of recipient cells, with small EV derived from V1G1^HIGH-NS showing the most pronounced activity. This involved activation of ERK1/2 signaling, in a response reversed by V-ATPase inhibition in NS-producing small EV. The miRNA profile of V1G1^HIGH-NS-derived small EVs differed significantly from that of V1G1^LOW-NS, which included miRNAs predicted to target MAPK/ERK signaling. Mechanistically, forced expression of a MAPK-targeting pool of miRNAs in recipient cells suppressed MAPK/ERK pathway activation and blunted the prooncogenic effects of V1G1^HIGH small EV. These findings propose that the GSC influences the brain milieu through a V1G1-coordinated EVs release of MAPK/ERK-targeting miRNAs. Interfering with V-ATPase activity could prevent ERK-dependent oncogenic reprogramming of the microenvironment, potentially hampering local GBM infiltration. IMPLICATIONS: Our data identify a novel molecular mechanism of gliomagenesis specific of the GBM stem cell niche, which coordinates a V-ATPase-dependent reprogramming of the brain microenvironment through the release of specialized EVs.

SUBMITTER: Bertolini I

PROVIDER: S-EPMC7642188 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Deletion polymorphism of glutathione S-transferase M1 (GSTM1), a phase II detoxification and antioxidant enzyme, increases susceptibility to end-stage renal disease (ESRD) as well as the development of cardiovascular diseases (CVD) among ESRD patients and leads to their shorter cardiovascular survival. The mechanisms by which GSTM1 downregulation contributes to oxidative stress and inflammation in endothelial cells in uremic conditions have not been investigated so far. Therefore, the aim of the present study was to elucidate the effects of GSTM1 knockdown on oxidative stress and expression of a panel of inflammatory markers in human umbilical vein endothelial cells (HUVECs) exposed to uremic serum. Additionally, we aimed to discern whether GSTM1-null genotype is associated with serum levels of adhesion molecules in ESRD patients. HUVECs treated with uremic serum exhibited impaired redox balance characterized by enhanced lipid peroxidation and decreased antioxidant enzyme activities, independently of the GSTM1 knockdown. In response to uremic injury, HUVECs exhibited alteration in the expression of a series of inflammatory cytokines including retinol-binding protein 4 (RBP4), regulated on activation, normal T cell expressed and secreted (RANTES), C-reactive protein (CRP), angiogenin, dickkopf-1 (Dkk-1), and platelet factor 4 (PF4). GSTM1 knockdown in HUVECs showed upregulation of monocyte chemoattractant protein-1 (MCP-1), a cytokine involved in the regulation of monocyte migration and adhesion. These cells also have shown upregulated intracellular and vascular cell adhesion molecules (ICAM-1 and VCAM-1). In accordance with these findings, the levels of serum ICAM-1 and VCAM-1 (sICAM-1 and sVCAM-1) were increased in ESRD patients lacking GSTM1, in comparison with patients with the GSTM1-active genotype. Based on these results, it may be concluded that incubation of endothelial cells in uremic serum induces redox imbalance accompanied with altered expression of a series of cytokines involved in arteriosclerosis and atherosclerosis. The association of GSTM1 downregulation with the altered expression of adhesion molecules might be at least partly responsible for the increased susceptibility of ESRD patients to CVD.

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Interplay Between V-ATPase G1 and Small EV-miRNAs Modulates ERK1/2 Activation in GBM Stem Cells and Nonneoplastic Milieu.

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