Primary involvement of NADPH oxidase 4 in hypoxia-induced generation of reactive oxygen species in adipose-derived stem cells

Ji Hye Kim, Seung Yong Song, Sang Gyu Park, Sun U. Song, Ying Xia, Jong Hyuk Sung

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

We have previously demonstrated that hypoxia stimulates adipose-derived stem cells (ASCs) through the generation of reactive oxygen species (ROS). However, the precise mechanism involved in the ROS generation by ASCs is not well understood. We sought to investigate in this work: (1) which subtype of NADPH oxidase (Nox) is primarily expressed in ASCs; (2) where Nox4 is localized in ASCs; and (3) whether silencing of Nox4 attenuates hypoxia-enhanced function of ASC. We used 2′,7′-dichlorofluorescin diacetate (DCF-DA) as an indicator of ROS generation and found that the fluorescence intensity of DCF-DA was significantly increased after hypoxia exposure (2% oxygen). In addition, hypoxia enhanced the proliferation and migration of ASCs and upregulated the mRNA expression of Oct4 and Rex1. Quantitative analysis of mRNA expression of Nox family in ASCs demonstrated that Nox4 is primarily expressed in ASCs, while immunofluorescence assay showed that Nox4 is mainly localized in the perinuclear region and overlaps with Mitotracker, a mitochondria marker. Silencing of Nox4 by siRNA treatment downregulated the RNA and protein expression of Nox4, which significantly reduced the ROS generation under hypoxia. In addition, Nox4 silencing significantly reduced the proliferation and migration of ASCs and downregulated the mRNA expression of Oct4 and Rex1. Phosphorylation of platelet-derived growth factor receptor-β, AKT, and ERK1/2 also diminished following Nox4 silencing. In a nutshell, these results suggest that Nox4 is primarily expressed in ASCs and plays a pivotal role in the hypoxia-enhanced stimulation of ASCs.

Original languageEnglish
Pages (from-to)2212-2221
Number of pages10
JournalStem Cells and Development
Volume21
Issue number12
DOIs
Publication statusPublished - 2012 Aug 10

Fingerprint

NADPH Oxidase
Reactive Oxygen Species
Stem Cells
Messenger RNA
Hypoxia
Down-Regulation
Platelet-Derived Growth Factor Receptors
Small Interfering RNA
Fluorescent Antibody Technique
Mitochondria
Fluorescence
Phosphorylation
RNA
Oxygen

All Science Journal Classification (ASJC) codes

  • Hematology
  • Developmental Biology
  • Cell Biology

Cite this

Kim, Ji Hye ; Song, Seung Yong ; Park, Sang Gyu ; Song, Sun U. ; Xia, Ying ; Sung, Jong Hyuk. / Primary involvement of NADPH oxidase 4 in hypoxia-induced generation of reactive oxygen species in adipose-derived stem cells. In: Stem Cells and Development. 2012 ; Vol. 21, No. 12. pp. 2212-2221.
@article{5dde0a40f9f643efa5ddd7d374ed9d68,
title = "Primary involvement of NADPH oxidase 4 in hypoxia-induced generation of reactive oxygen species in adipose-derived stem cells",
abstract = "We have previously demonstrated that hypoxia stimulates adipose-derived stem cells (ASCs) through the generation of reactive oxygen species (ROS). However, the precise mechanism involved in the ROS generation by ASCs is not well understood. We sought to investigate in this work: (1) which subtype of NADPH oxidase (Nox) is primarily expressed in ASCs; (2) where Nox4 is localized in ASCs; and (3) whether silencing of Nox4 attenuates hypoxia-enhanced function of ASC. We used 2′,7′-dichlorofluorescin diacetate (DCF-DA) as an indicator of ROS generation and found that the fluorescence intensity of DCF-DA was significantly increased after hypoxia exposure (2{\%} oxygen). In addition, hypoxia enhanced the proliferation and migration of ASCs and upregulated the mRNA expression of Oct4 and Rex1. Quantitative analysis of mRNA expression of Nox family in ASCs demonstrated that Nox4 is primarily expressed in ASCs, while immunofluorescence assay showed that Nox4 is mainly localized in the perinuclear region and overlaps with Mitotracker, a mitochondria marker. Silencing of Nox4 by siRNA treatment downregulated the RNA and protein expression of Nox4, which significantly reduced the ROS generation under hypoxia. In addition, Nox4 silencing significantly reduced the proliferation and migration of ASCs and downregulated the mRNA expression of Oct4 and Rex1. Phosphorylation of platelet-derived growth factor receptor-β, AKT, and ERK1/2 also diminished following Nox4 silencing. In a nutshell, these results suggest that Nox4 is primarily expressed in ASCs and plays a pivotal role in the hypoxia-enhanced stimulation of ASCs.",
author = "Kim, {Ji Hye} and Song, {Seung Yong} and Park, {Sang Gyu} and Song, {Sun U.} and Ying Xia and Sung, {Jong Hyuk}",
year = "2012",
month = "8",
day = "10",
doi = "10.1089/scd.2011.0561",
language = "English",
volume = "21",
pages = "2212--2221",
journal = "Stem Cells and Development",
issn = "1547-3287",
publisher = "Mary Ann Liebert Inc.",
number = "12",

}

Primary involvement of NADPH oxidase 4 in hypoxia-induced generation of reactive oxygen species in adipose-derived stem cells. / Kim, Ji Hye; Song, Seung Yong; Park, Sang Gyu; Song, Sun U.; Xia, Ying; Sung, Jong Hyuk.

In: Stem Cells and Development, Vol. 21, No. 12, 10.08.2012, p. 2212-2221.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Primary involvement of NADPH oxidase 4 in hypoxia-induced generation of reactive oxygen species in adipose-derived stem cells

AU - Kim, Ji Hye

AU - Song, Seung Yong

AU - Park, Sang Gyu

AU - Song, Sun U.

AU - Xia, Ying

AU - Sung, Jong Hyuk

PY - 2012/8/10

Y1 - 2012/8/10

N2 - We have previously demonstrated that hypoxia stimulates adipose-derived stem cells (ASCs) through the generation of reactive oxygen species (ROS). However, the precise mechanism involved in the ROS generation by ASCs is not well understood. We sought to investigate in this work: (1) which subtype of NADPH oxidase (Nox) is primarily expressed in ASCs; (2) where Nox4 is localized in ASCs; and (3) whether silencing of Nox4 attenuates hypoxia-enhanced function of ASC. We used 2′,7′-dichlorofluorescin diacetate (DCF-DA) as an indicator of ROS generation and found that the fluorescence intensity of DCF-DA was significantly increased after hypoxia exposure (2% oxygen). In addition, hypoxia enhanced the proliferation and migration of ASCs and upregulated the mRNA expression of Oct4 and Rex1. Quantitative analysis of mRNA expression of Nox family in ASCs demonstrated that Nox4 is primarily expressed in ASCs, while immunofluorescence assay showed that Nox4 is mainly localized in the perinuclear region and overlaps with Mitotracker, a mitochondria marker. Silencing of Nox4 by siRNA treatment downregulated the RNA and protein expression of Nox4, which significantly reduced the ROS generation under hypoxia. In addition, Nox4 silencing significantly reduced the proliferation and migration of ASCs and downregulated the mRNA expression of Oct4 and Rex1. Phosphorylation of platelet-derived growth factor receptor-β, AKT, and ERK1/2 also diminished following Nox4 silencing. In a nutshell, these results suggest that Nox4 is primarily expressed in ASCs and plays a pivotal role in the hypoxia-enhanced stimulation of ASCs.

AB - We have previously demonstrated that hypoxia stimulates adipose-derived stem cells (ASCs) through the generation of reactive oxygen species (ROS). However, the precise mechanism involved in the ROS generation by ASCs is not well understood. We sought to investigate in this work: (1) which subtype of NADPH oxidase (Nox) is primarily expressed in ASCs; (2) where Nox4 is localized in ASCs; and (3) whether silencing of Nox4 attenuates hypoxia-enhanced function of ASC. We used 2′,7′-dichlorofluorescin diacetate (DCF-DA) as an indicator of ROS generation and found that the fluorescence intensity of DCF-DA was significantly increased after hypoxia exposure (2% oxygen). In addition, hypoxia enhanced the proliferation and migration of ASCs and upregulated the mRNA expression of Oct4 and Rex1. Quantitative analysis of mRNA expression of Nox family in ASCs demonstrated that Nox4 is primarily expressed in ASCs, while immunofluorescence assay showed that Nox4 is mainly localized in the perinuclear region and overlaps with Mitotracker, a mitochondria marker. Silencing of Nox4 by siRNA treatment downregulated the RNA and protein expression of Nox4, which significantly reduced the ROS generation under hypoxia. In addition, Nox4 silencing significantly reduced the proliferation and migration of ASCs and downregulated the mRNA expression of Oct4 and Rex1. Phosphorylation of platelet-derived growth factor receptor-β, AKT, and ERK1/2 also diminished following Nox4 silencing. In a nutshell, these results suggest that Nox4 is primarily expressed in ASCs and plays a pivotal role in the hypoxia-enhanced stimulation of ASCs.

UR - http://www.scopus.com/inward/record.url?scp=84864674993&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84864674993&partnerID=8YFLogxK

U2 - 10.1089/scd.2011.0561

DO - 10.1089/scd.2011.0561

M3 - Article

C2 - 22181007

AN - SCOPUS:84864674993

VL - 21

SP - 2212

EP - 2221

JO - Stem Cells and Development

JF - Stem Cells and Development

SN - 1547-3287

IS - 12

ER -