Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels

Gulsah Sevimli; Matthew J. Smith; Tuba Akgul Caglar; Şükriye Bilir; Melike Secilmis; Hamza Y. Altun; Esra N. Yigit; Fan Yang; Thomas P. Keeley; Roland Malli; Gürkan Öztürk; Giovanni E. Mann; Emrah Eroglu

doi:10.1016/j.redox.2022.102319

Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels

Gulsah Sevimli, Matthew J. Smith, Tuba Akgul Caglar, Şükriye Bilir, Melike Secilmis, Hamza Y. Altun, Esra N. Yigit, Fan Yang, Thomas P. Keeley, Roland Malli, Gürkan Öztürk, Giovanni E. Mann, Emrah Eroglu^*

^*Corresponding author for this work

Vascular Biology & Inflammation

Research output: Contribution to journal › Article › peer-review

3 Citations (Scopus)

Abstract

Iron is an essential metal for cellular metabolism and signaling, but it has adverse effects in excess. The physiological consequences of iron deficiency are well established, yet the relationship between iron supplementation and pericellular oxygen levels in cultured cells and their downstream effects on metalloproteins has been less explored. This study exploits the metalloprotein geNOps in cultured HEK293T epithelial and EA.hy926 endothelial cells to test the iron-dependency in cells adapted to standard room air (18 kPa O₂) or physiological normoxia (5 kPa O₂). We show that cells in culture require iron supplementation to activate the metalloprotein geNOps and demonstrate for the first time that cells adapted to physiological normoxia require significantly lower iron compared to cells adapted to hyperoxia. This study establishes an essential role for recapitulating oxygen levels in vivo and uncovers a previously unrecognized requirement for ferrous iron supplementation under standard cell culture conditions to achieve geNOps functionality.

Original language	English
Article number	102319
Journal	Redox Biology
Volume	53
DOIs	https://doi.org/10.1016/j.redox.2022.102319
Publication status	Published - Jul 2022

Keywords

Cell culture
Culture media
Ferric iron
Ferrous iron
geNOps
Hydrogen peroxide
Hyperoxia
NO bioavailability
Normoxia
Pericellular oxygen

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10.1016/j.redox.2022.102319

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Sevimli, G., Smith, M. J., Caglar, T. A., Bilir, Ş., Secilmis, M., Altun, H. Y., Yigit, E. N., Yang, F., Keeley, T. P., Malli, R., Öztürk, G., Mann, G. E., & Eroglu, E. (2022). Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels. Redox Biology, 53, [102319]. https://doi.org/10.1016/j.redox.2022.102319

@article{51a64673ae304809a81d5154146fdf93,

title = "Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels",

abstract = "Iron is an essential metal for cellular metabolism and signaling, but it has adverse effects in excess. The physiological consequences of iron deficiency are well established, yet the relationship between iron supplementation and pericellular oxygen levels in cultured cells and their downstream effects on metalloproteins has been less explored. This study exploits the metalloprotein geNOps in cultured HEK293T epithelial and EA.hy926 endothelial cells to test the iron-dependency in cells adapted to standard room air (18 kPa O2) or physiological normoxia (5 kPa O2). We show that cells in culture require iron supplementation to activate the metalloprotein geNOps and demonstrate for the first time that cells adapted to physiological normoxia require significantly lower iron compared to cells adapted to hyperoxia. This study establishes an essential role for recapitulating oxygen levels in vivo and uncovers a previously unrecognized requirement for ferrous iron supplementation under standard cell culture conditions to achieve geNOps functionality.",

keywords = "Cell culture, Culture media, Ferric iron, Ferrous iron, geNOps, Hydrogen peroxide, Hyperoxia, NO bioavailability, Normoxia, Pericellular oxygen",

author = "Gulsah Sevimli and Smith, {Matthew J.} and Caglar, {Tuba Akgul} and {\c S}{\"u}kriye Bilir and Melike Secilmis and Altun, {Hamza Y.} and Yigit, {Esra N.} and Fan Yang and Keeley, {Thomas P.} and Roland Malli and G{\"u}rkan {\"O}zt{\"u}rk and Mann, {Giovanni E.} and Emrah Eroglu",

note = "Funding Information: We gratefully acknowledge support from Scientific and Technological Research Council of Turkey (Grant 118C242, E.E., T.A.C., H.Y.A., M.S., and G.S.), Integration Projects of Sabancı University ( EPD-2019-1 , EE), Heart Research U.K. (RG2673, G.E.M.), British Heart Foundation ( FS/16/67/32548 , G.E.M.), COST Action CA20121 (G.E.M.), and King's Together Strategic Award (G.E.M.). We thank Professors Helmut Sies and Richard J. Naftalin for their helpful discussions. Publisher Copyright: {\textcopyright} 2022",

year = "2022",

month = jul,

doi = "10.1016/j.redox.2022.102319",

language = "English",

volume = "53",

journal = "Redox Biology",

issn = "2213-2317",

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TY - JOUR

T1 - Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels

AU - Sevimli, Gulsah

AU - Smith, Matthew J.

AU - Caglar, Tuba Akgul

AU - Bilir, Şükriye

AU - Secilmis, Melike

AU - Altun, Hamza Y.

AU - Yigit, Esra N.

AU - Yang, Fan

AU - Keeley, Thomas P.

AU - Malli, Roland

AU - Öztürk, Gürkan

AU - Mann, Giovanni E.

AU - Eroglu, Emrah

N1 - Funding Information: We gratefully acknowledge support from Scientific and Technological Research Council of Turkey (Grant 118C242, E.E., T.A.C., H.Y.A., M.S., and G.S.), Integration Projects of Sabancı University ( EPD-2019-1 , EE), Heart Research U.K. (RG2673, G.E.M.), British Heart Foundation ( FS/16/67/32548 , G.E.M.), COST Action CA20121 (G.E.M.), and King's Together Strategic Award (G.E.M.). We thank Professors Helmut Sies and Richard J. Naftalin for their helpful discussions. Publisher Copyright: © 2022

PY - 2022/7

Y1 - 2022/7

N2 - Iron is an essential metal for cellular metabolism and signaling, but it has adverse effects in excess. The physiological consequences of iron deficiency are well established, yet the relationship between iron supplementation and pericellular oxygen levels in cultured cells and their downstream effects on metalloproteins has been less explored. This study exploits the metalloprotein geNOps in cultured HEK293T epithelial and EA.hy926 endothelial cells to test the iron-dependency in cells adapted to standard room air (18 kPa O2) or physiological normoxia (5 kPa O2). We show that cells in culture require iron supplementation to activate the metalloprotein geNOps and demonstrate for the first time that cells adapted to physiological normoxia require significantly lower iron compared to cells adapted to hyperoxia. This study establishes an essential role for recapitulating oxygen levels in vivo and uncovers a previously unrecognized requirement for ferrous iron supplementation under standard cell culture conditions to achieve geNOps functionality.

AB - Iron is an essential metal for cellular metabolism and signaling, but it has adverse effects in excess. The physiological consequences of iron deficiency are well established, yet the relationship between iron supplementation and pericellular oxygen levels in cultured cells and their downstream effects on metalloproteins has been less explored. This study exploits the metalloprotein geNOps in cultured HEK293T epithelial and EA.hy926 endothelial cells to test the iron-dependency in cells adapted to standard room air (18 kPa O2) or physiological normoxia (5 kPa O2). We show that cells in culture require iron supplementation to activate the metalloprotein geNOps and demonstrate for the first time that cells adapted to physiological normoxia require significantly lower iron compared to cells adapted to hyperoxia. This study establishes an essential role for recapitulating oxygen levels in vivo and uncovers a previously unrecognized requirement for ferrous iron supplementation under standard cell culture conditions to achieve geNOps functionality.

KW - Cell culture

KW - Culture media

KW - Ferric iron

KW - Ferrous iron

KW - geNOps

KW - Hydrogen peroxide

KW - Hyperoxia

KW - NO bioavailability

KW - Normoxia

KW - Pericellular oxygen

UR - http://www.scopus.com/inward/record.url?scp=85129621442&partnerID=8YFLogxK

U2 - 10.1016/j.redox.2022.102319

DO - 10.1016/j.redox.2022.102319

M3 - Article

C2 - 35525027

AN - SCOPUS:85129621442

SN - 2213-2317

VL - 53

JO - Redox Biology

JF - Redox Biology

M1 - 102319

ER -

Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels

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Keywords

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