Tags

Type your tag names separated by a space and hit enter

Nitrite-derived nitric oxide reduces hypoxia-inducible factor 1α-mediated extracellular vesicle production by endothelial cells.
Nitric Oxide 2017; 63:1-12NO

Abstract

INTRODUCTION

Extracellular vesicles (EVs) are small, spherical particles enclosed by a phospholipid bilayer (∼30-1000 nm) released from multiple cell types, and have been shown to have pathophysiological roles in a plethora of disease states. The transcription factor hypoxia-inducible factor-1 (HIF-1) allows for adaptation of cellular physiology in hypoxia and may permit the enhanced release of EVs under such conditions. Nitric oxide (NO) plays a pivotal role in vascular homeostasis, and can modulate the cellular response to hypoxia by preventing HIF-1 accumulation. We aimed to selectively target HIF-1 via sodium nitrite (NaNO2) addition, and examine the effect on endothelial EV, size, concentration and function, and delineate the role of HIF-1 in EV biogenesis.

METHODS

Endothelial (HECV) cells were exposed to hypoxic conditions (1% O2, 24 h) and compared to endothelial cells exposed to normoxia (21% O2) with and without the presence of sodium nitrite (NaNO2) (30 μM). Allopurinol (100 μM), an inhibitor of xanthine oxidoreductase, was added both alone and in combination with NaNO2 to cells exposed to hypoxia. EV and cell preparations were quantified by nanoparticle tracking analysis and confirmed by electron microscopy. Western blotting and siRNA were used to confirm the role of HIF-1α and HIF-2α in EV biogenesis. Flow cytometry and time-resolved fluorescence were used to assess the surface and intravesicular protein content.

RESULTS

Endothelial (HECV) cells exposed to hypoxia (1% O2) produced higher levels of EVs compared to cells exposed to normoxia. This increase was confirmed using the hypoxia-mimetic agent desferrioxamine. Treatment of cells with sodium nitrite (NaNO2) reduced the hypoxic enhancement of EV production. Treatment of cells with the xanthine oxidoreductase inhibitor allopurinol, in addition to NaNO2 attenuated the NaNO2-attributed suppression of hypoxia-mediated EV release. Transfection of cells with HIF-1α siRNA, but not HIF-2α siRNA, prior to hypoxic exposure prevented the enhancement of EV release.

CONCLUSION

These data provide evidence that hypoxia enhances the release of EVs in endothelial cells, and that this is mediated by HIF-1α, but not HIF-2α. Furthermore, the reduction of NO2- to NO via xanthine oxidoreductase during hypoxia appears to inhibit HIF-1α-mediated EV production.

Authors+Show Affiliations

School of Medicine, Cardiff University, Cardiff, CF24 4HQ, UK.Division Newborn Medicine, Boston Children's Hospital, Harvard Medical School, Harvard University, Boston, 02115, MA, USA.Institute of Cancer & Genetics, Cardiff University, Cardiff, CF14 4XN, UK.Neurosciences and Mental Health Research Institute, School of Medicine, Cardiff University, Cardiff, CF24 4HQ, UK.Cardiff School of Health Sciences, Cardiff Metropolitan University, Cardiff, CF5 2SG, UK. Electronic address: PJames@cardiffmet.ac.uk.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

28017872

Citation

Burnley-Hall, Nicholas, et al. "Nitrite-derived Nitric Oxide Reduces Hypoxia-inducible Factor 1α-mediated Extracellular Vesicle Production By Endothelial Cells." Nitric Oxide : Biology and Chemistry, vol. 63, 2017, pp. 1-12.
Burnley-Hall N, Willis G, Davis J, et al. Nitrite-derived nitric oxide reduces hypoxia-inducible factor 1α-mediated extracellular vesicle production by endothelial cells. Nitric Oxide. 2017;63:1-12.
Burnley-Hall, N., Willis, G., Davis, J., Rees, D. A., & James, P. E. (2017). Nitrite-derived nitric oxide reduces hypoxia-inducible factor 1α-mediated extracellular vesicle production by endothelial cells. Nitric Oxide : Biology and Chemistry, 63, pp. 1-12. doi:10.1016/j.niox.2016.12.005.
Burnley-Hall N, et al. Nitrite-derived Nitric Oxide Reduces Hypoxia-inducible Factor 1α-mediated Extracellular Vesicle Production By Endothelial Cells. Nitric Oxide. 2017 Feb 28;63:1-12. PubMed PMID: 28017872.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Nitrite-derived nitric oxide reduces hypoxia-inducible factor 1α-mediated extracellular vesicle production by endothelial cells. AU - Burnley-Hall,Nicholas, AU - Willis,Gareth, AU - Davis,Jessica, AU - Rees,D Aled, AU - James,Philip E, Y1 - 2016/12/23/ PY - 2016/10/04/received PY - 2016/12/07/revised PY - 2016/12/15/accepted PY - 2016/12/27/pubmed PY - 2018/7/6/medline PY - 2016/12/27/entrez KW - Extracellular vesicles KW - Hypoxia KW - Hypoxia-inducible factor KW - Nitric oxide KW - Nitrite SP - 1 EP - 12 JF - Nitric oxide : biology and chemistry JO - Nitric Oxide VL - 63 N2 - INTRODUCTION: Extracellular vesicles (EVs) are small, spherical particles enclosed by a phospholipid bilayer (∼30-1000 nm) released from multiple cell types, and have been shown to have pathophysiological roles in a plethora of disease states. The transcription factor hypoxia-inducible factor-1 (HIF-1) allows for adaptation of cellular physiology in hypoxia and may permit the enhanced release of EVs under such conditions. Nitric oxide (NO) plays a pivotal role in vascular homeostasis, and can modulate the cellular response to hypoxia by preventing HIF-1 accumulation. We aimed to selectively target HIF-1 via sodium nitrite (NaNO2) addition, and examine the effect on endothelial EV, size, concentration and function, and delineate the role of HIF-1 in EV biogenesis. METHODS: Endothelial (HECV) cells were exposed to hypoxic conditions (1% O2, 24 h) and compared to endothelial cells exposed to normoxia (21% O2) with and without the presence of sodium nitrite (NaNO2) (30 μM). Allopurinol (100 μM), an inhibitor of xanthine oxidoreductase, was added both alone and in combination with NaNO2 to cells exposed to hypoxia. EV and cell preparations were quantified by nanoparticle tracking analysis and confirmed by electron microscopy. Western blotting and siRNA were used to confirm the role of HIF-1α and HIF-2α in EV biogenesis. Flow cytometry and time-resolved fluorescence were used to assess the surface and intravesicular protein content. RESULTS: Endothelial (HECV) cells exposed to hypoxia (1% O2) produced higher levels of EVs compared to cells exposed to normoxia. This increase was confirmed using the hypoxia-mimetic agent desferrioxamine. Treatment of cells with sodium nitrite (NaNO2) reduced the hypoxic enhancement of EV production. Treatment of cells with the xanthine oxidoreductase inhibitor allopurinol, in addition to NaNO2 attenuated the NaNO2-attributed suppression of hypoxia-mediated EV release. Transfection of cells with HIF-1α siRNA, but not HIF-2α siRNA, prior to hypoxic exposure prevented the enhancement of EV release. CONCLUSION: These data provide evidence that hypoxia enhances the release of EVs in endothelial cells, and that this is mediated by HIF-1α, but not HIF-2α. Furthermore, the reduction of NO2- to NO via xanthine oxidoreductase during hypoxia appears to inhibit HIF-1α-mediated EV production. SN - 1089-8611 UR - https://www.unboundmedicine.com/medline/citation/28017872/Nitrite_derived_nitric_oxide_reduces_hypoxia_inducible_factor_1α_mediated_extracellular_vesicle_production_by_endothelial_cells_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1089-8603(16)30182-3 DB - PRIME DP - Unbound Medicine ER -