Angiotensin II receptor one (AT1) mediates dextrose induced endoplasmic reticulum stress and superoxide production in human coronary artery endothelial cells.

BACKGROUND

Renin-angiotensin-aldosterone system (RAAS) has been implicated in diabetes-related vascular complications partly through oxidative stress.

OBJECTIVE

To determine the role of angiotensin II receptor subtype one (AT1) in dextrose induced endoplasmic reticulum (ER) stress, another cellular stress implicated in vascular disease.

METHODS

Human coronary artery endothelial cells with or without AT1 receptor knock down were treated with 27.5mM dextrose for 24h in the presence of various pharmacologic blockers of RAAS and ER stress and superoxide (SO) production were measured. Transfection of cells with AT1 antisense RNA knocked down cellular AT1 by approximately 80%. The ER stress was measured using the placental alkaline phosphatase (ES-TRAP) assay and western blot analysis of glucose regulated protein 78 (GRP78), c-jun-N-terminal kinase 1 (JNK1), phospho-JNK1, eukaryotic translation initiation factor 2α (eIF2α) and phospho-eIF2α measurements. Superoxide (SO) generation was measured using the superoxide-reactive probe 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-A]pyrazin-3-one hydrochloride (MCLA) chemiluminescence.

RESULTS

In cells with AT1 knock down, dextrose induced ER stress was significantly blunted and treatment with 27.5mM dextrose resulted in significantly smaller increase in SO production compared to 27.5mM dextrose treated and sham transfected cells. Dextrose induced ER stress was reduced with pharmacologic blockers of AT1 (losartan and candesartan) and mineralocorticoid receptor blocker (spironolactone) but not with angiotensin converting enzyme inhibitors (captopril and lisinopril). The dextrose induced SO generation was inhibited by all pharmacologic blockers of RAAS tested.

CONCLUSIONS

The results indicate that dextrose induced ER stress and SO production in endothelial cells are mediated at least partly through AT1 receptor activation.