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Hydrogen peroxide induces mRNA for tumour necrosis factor alpha in human endothelial cells

Valen, G; Erl, W; Eriksson, P; Wuttge, Dirk LU ; Paulsson, G and Hansson, G K (1999) In Free Radical Research 31(6). p.503-512
Abstract
Reactive oxygen intermediates are important mediators of inflammation. We investigated if hydrogen peroxide (H2O2) induces tumour necrosis factor alpha (TNFalpha) expression in cultured human cells from umbilical vein endothelium (HUVEC), aortic smooth muscle cells (SMC), peripheral blood mononuclear cells (PBMC), or the cell line Mono Mac 6. Cultures were stimulated with 200 micromol/L H2O2 for 15 min. After 4 h cells were harvested, mRNA extracted, and amplified by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) with histone (H3) as reference gene. In HUVECs, mRNA for TNFalpha increased with a factor of 4 after stimulation (p < 0.001), in PBMC with a factor of 2 (p < 0.05), while mRNA from SMC and Mono Mac... (More)
Reactive oxygen intermediates are important mediators of inflammation. We investigated if hydrogen peroxide (H2O2) induces tumour necrosis factor alpha (TNFalpha) expression in cultured human cells from umbilical vein endothelium (HUVEC), aortic smooth muscle cells (SMC), peripheral blood mononuclear cells (PBMC), or the cell line Mono Mac 6. Cultures were stimulated with 200 micromol/L H2O2 for 15 min. After 4 h cells were harvested, mRNA extracted, and amplified by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) with histone (H3) as reference gene. In HUVECs, mRNA for TNFalpha increased with a factor of 4 after stimulation (p < 0.001), in PBMC with a factor of 2 (p < 0.05), while mRNA from SMC and Mono Mac 6 did not increase significantly. Cellular TNFalpha protein in HUVECs was measured with flow cytometry (FACS) before and 6, 12, and 24 h after stimulation. TNFalpha protein was detectable in small, but reproducible amounts 12 h after stimulation, and increased further after 24 h. However, no secretion of TNFalpha was detected by ELISA. FACS analysis of the passaged HUVEC cultures did not reveal any contamination with non-endothelial cells. In conclusion, H2O2 induces TNFalpha mRNA in HUVECs and PBMC. In HUVECs an increase of intracellular TNFalpha protein was also detected, indicating that endothelial cells can produce small amounts of TNFalpha. (Less)
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author
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Endothelial cells, hydrogen peroxide, tumour necrosis factor agr, mononuclear blood cells, Mono Mac 6, reactive oxygen intermediates, smooth muscle cells
in
Free Radical Research
volume
31
issue
6
pages
503 - 512
publisher
Harwood Academic
external identifiers
  • pmid:10630674
  • scopus:0032740681
ISSN
1029-2470
DOI
10.1080/10715769900301071
language
English
LU publication?
no
id
3fcb6611-e23c-4465-aba8-af6b10536e53 (old id 1115125)
date added to LUP
2008-07-07 14:30:58
date last changed
2017-08-06 04:42:25
@article{3fcb6611-e23c-4465-aba8-af6b10536e53,
  abstract     = {Reactive oxygen intermediates are important mediators of inflammation. We investigated if hydrogen peroxide (H2O2) induces tumour necrosis factor alpha (TNFalpha) expression in cultured human cells from umbilical vein endothelium (HUVEC), aortic smooth muscle cells (SMC), peripheral blood mononuclear cells (PBMC), or the cell line Mono Mac 6. Cultures were stimulated with 200 micromol/L H2O2 for 15 min. After 4 h cells were harvested, mRNA extracted, and amplified by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) with histone (H3) as reference gene. In HUVECs, mRNA for TNFalpha increased with a factor of 4 after stimulation (p &lt; 0.001), in PBMC with a factor of 2 (p &lt; 0.05), while mRNA from SMC and Mono Mac 6 did not increase significantly. Cellular TNFalpha protein in HUVECs was measured with flow cytometry (FACS) before and 6, 12, and 24 h after stimulation. TNFalpha protein was detectable in small, but reproducible amounts 12 h after stimulation, and increased further after 24 h. However, no secretion of TNFalpha was detected by ELISA. FACS analysis of the passaged HUVEC cultures did not reveal any contamination with non-endothelial cells. In conclusion, H2O2 induces TNFalpha mRNA in HUVECs and PBMC. In HUVECs an increase of intracellular TNFalpha protein was also detected, indicating that endothelial cells can produce small amounts of TNFalpha.},
  author       = {Valen, G and Erl, W and Eriksson, P and Wuttge, Dirk and Paulsson, G and Hansson, G K},
  issn         = {1029-2470},
  keyword      = {Endothelial cells,hydrogen peroxide,tumour necrosis factor agr,mononuclear blood cells,Mono Mac 6,reactive oxygen intermediates,smooth muscle cells},
  language     = {eng},
  number       = {6},
  pages        = {503--512},
  publisher    = {Harwood Academic},
  series       = {Free Radical Research},
  title        = {Hydrogen peroxide induces mRNA for tumour necrosis factor alpha in human endothelial cells},
  url          = {http://dx.doi.org/10.1080/10715769900301071},
  volume       = {31},
  year         = {1999},
}