Transcript and protein analysis reveals better survival skills of monocyte-derived dendritic cells compared to monocytes during oxidative stress.
BACKGROUND
Dendritic cells (DCs), professional antigen-presenting cells with the unique ability to initiate primary T-cell responses, are present in atherosclerotic lesions where they are exposed to oxidative stress that generates cytotoxic reactive oxygen species (ROS). A large body of evidence indicates that cell death is a major modulating factor of atherogenesis. We examined antioxidant defence systems of human monocyte-derived (mo)DCs and monocytes in response to oxidative stress.


METHODS
Oxidative stress was induced by addition of tertiary-butylhydroperoxide (tert-BHP, 30 min). Cellular responses were evaluated using flow cytometry and confocal live cell imaging (both using 5-(and-6)-chloromethyl-2,7-dichlorodihydrofluorescein diacetate, CM-H(2)DCFDA). Viability was assessed by the neutral red assay. Total RNA was extracted for a PCR profiler array. Five genes were selected for confirmation by Taqman gene expression assays, and by immunoblotting or immunohistochemistry for protein levels.


RESULTS
Tert-BHP increased CM-H(2)DCFDA fluorescence and caused cell death. Interestingly, all processes occurred more slowly in moDCs than in monocytes. The mRNA profiler array showed more than 2-fold differential expression of 32 oxidative stress-related genes in unstimulated moDCs, including peroxiredoxin-2 (PRDX2), an enzyme reducing hydrogen peroxide and lipid peroxides. PRDX2 upregulation was confirmed by Taqman assays, immunoblotting and immunohistochemistry. Silencing PRDX2 in moDCs by means of siRNA significantly increased CM-DCF fluorescence and cell death upon tert-BHP-stimulation.


CONCLUSIONS
Our results indicate that moDCs exhibit higher intracellular antioxidant capacities, making them better equipped to resist oxidative stress than monocytes. Upregulation of PRDX2 is involved in the neutralization of ROS in moDCs. Taken together, this points to better survival skills of DCs in oxidative stress environments, such as atherosclerotic plaques.
Laboratory of Cellular and Molecular Cardiology, University of Antwerp, Wilrijk, Antwerp, Belgium. ilse.vanbrussel@ua.ac.be
Ilse
I
Van Brussel
Dorien M
DM
Schrijvers
Wim
W
Martinet
Isabel
I
Pintelon
Maartje
M
Deschacht
Kathy
K
Schnorbusch
Louis
L
Maes
Johan M
JM
Bosmans
Christiaan J
CJ
Vrints
Dirk
D
Adriaensen
Paul
P
Cos
Hidde
H
Bult
D002478:Cells, Cultured; D003713:Dendritic Cells / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D009000:Monocytes / Q000166:cytology*; D018384:Oxidative Stress / Q000235:genetics / Q000502:physiology*; D017382:Reactive Oxygen Species / Q000378:metabolism
D017382:Reactive Oxygen Species