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Modulation of thioredoxin reductase-2 expression in EAhy926 cells: implications for endothelial selenoprotein
Michael S Crane1, Alexander F Howie, John R Arthur
1Department of Clinical Biochemistry, Royal Infirmary of Edinburgh, Little France, EH16 4SA Edinburgh, UK. mike.crane@luht.scot.nhs.uk
Background:
We examined the expression of the mitochondrial selenoenzyme TrxR2 in the endothelial cell line EAhy926 under conditions known to modify its cytoplasmic counterpart TrxR1.
Methods:
Cells were cultured with varying concentrations of selenite, sulforaphane or the Ca2+ ionophore A23187 for 72-h, prior to assay of TrxR concentration and activity. Further cultures underwent prolonged (7-day) Se-depletion before selenoprotein measurement.
Results:
In Se-deficient cultures, neither Se, A23187 or sulforaphane affected TrxR2 concentration, while these treatments induced TrxR1 concentration (p<0.05). When co-incubated, optimal concentrations of Se (40 nM) and sulforaphane (4 microM) only modestly increased TrxR2 protein (approximately 1.3-fold), compared with TrxR1 (approximately 4-fold). In Se-deficient cells, TrxR activity was unaffected by sulforaphane or A23187. Prolonged Se-depletion caused a comparatively small reduction in TrxR2 (66% TrxR2 retained) against TrxR1 and glutathione peroxidase-1 activity (38% and 17% retained, respectively).
Conclusions:
The relative resistance of TrxR2 to Se-deprivation and induction by sulforaphane and A23187 suggests TrxR2 lies near the top of the selenoprotein hierarchy in EAhy926 cells and exhibits near maximum expression under a range of culture conditions. In Se deficiency an inactive (possibly truncated) TrxR1 is produced in response to stimulus by sulforaphane and A23187.
General Significance:
These observations underpin a likely critical antioxidant role for TrxR2 and TrxR1 in the endothelium.
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