The mechanism underlying oxidant-induced intracellular Ca2+ ([Ca2+]i) increase was studied in cultured bovine aortic endothelial cells (BAECs) using fura-2 AM. In the presence of 2 mM extracellular Ca2+, the application of DTBNP (20 μM), a membrane-permeable oxidant, caused an increase in [Ca2+]i, and DTT (2 mM) as a reductant completely reversed the effect of DTBNP. The [Ca2+]i increase induced by DTBNP was also observed in an extracellular Ca2+-free/2 mM EGTA solution, indicating the release of Ca2+ from intracellular store(s). After endoplasmic reticulum was depleted by an IP3-generating agonist, ATP (30 μM) or an ER Ca2+ pump inhibitor, thapsigargin (1 μM), DTBNP-stressed BAECs showed an increase of [Ca2+]i in Ca2+-free/2 mM EGTA solution. Ratio-differences before and after the application of DTBNP after pretreatment with ATP or thapsigargin were 0.42±0.15 and 0.49±0.07, respectively (n=7), which are significantly reduced, compared to the control value of 0.72±0.07 in a Ca2+-free/2 mM EGTA solution. After the protonophore CCCP (10 μM) challenge to release mitochondrial Ca2+, the similar result was obtained. Ratio-difference before and after the application of DTBNP after pretreatment with CCCP was 0.46±0.09 (n=7). Simultaneous application of thapsigargin and CCCP completely abolished the DTBNP-induced [Ca2+]i increase. The above results together indicate that the increase of [Ca2+]i by DTBNP resulted from the release of Ca2+ from both endoplasmic reticulum and mitochondria.
|Number of pages||6|
|Journal||Korean Journal of Physiology and Pharmacology|
|State||Published - 1 Dec 2005|
- Bovine aortic endothelial cells
- Endoplasmic reticulum