| Summary: | 博士 === 中原大學 === 生物醫學工程研究所 === 104 === Endothelial cells (EC) do not only serve the barrier function between the blood and the vascular smooth cells, but also regulate hemostasis, vascular tone and vessel morphology/proliferation. EC release an array of vasorelaxants --- EC-derived relaxing factors (EDRF); the most prominent known relaxing factors include nitric oxide (NO). Release of nitric oxide (NO) is triggered by a rise in endothelial cell (EC) cytosolic Ca2+ concentration ([Ca2+ ]i) and is of prime importance in vascular tone regulation as NO relaxes vascular smooth muscle. Agonists could stimulate EC [Ca2+ ]i elevation by triggering Ca2+ influx via plasma membrane ion channels, one of which is the store-operated Ca2+ channel; the latter opens as a result of agonist-triggered internal Ca2+ release. EC also release vasocontracting factors called endothelium-derived contracting factors (EDCF). The latter include endothelin-1, isoprostanes and reactive oxygen species. A loss of balance between the secretion of EDRF and EDCF may lead to vascular diseases. Endotoxin (lipopolysaccharide, LPS) could cause sepsis, which is often the fatal cause in critically ill patients. One of the LPS-induced damages is EC dysfunction, eventually leading to perturbations in hemodynamics. I obtained data showing that LPS-challenged mouse cerebral cortex endothelial bEND.3 cells did not suffer from apoptotic death, and in fact had intact agonist-triggered intracellular Ca2+ release; however, they had reduced store-operated Ca2+ entry (SOCE) after LPS treatment for 3 h or more. Using real-time PCR, I did not find a decrease in gene expression of stromal interaction molecule 1 (STIM1) and Orai1 (two SOCE protein components) in bEND.3 cells treated with LPS for 15 h. LPS inhibitory effects could be largely prevented by sodium salicylate (an inhibitor of nuclear factor-κB; NF-κB) or SB203580 (an inhibitor of p38 mitogen-activated protein kinases; p38 MAPK), suggesting that the p38 MAPK–NF-κB pathway is involved in SOCE inhibition. During the investigation of the above project, I used parthenonide to inhibit NF-κB; however, substantial perturbations in Ca2+ signaling was observed. Therefore, another project was developed to investigate the actions of this drug. Parthenolide is a sesquiterpene lactone compound isolated from the leaves and flowerheads of the plant feverfew (Tanacetum parthenium). The anticancer effects of parthenolide have been well studied and this lactone compound is currently under clinical trials. Parthenolide is also a protective agent in cardiac reperfusion injury via its inhibition of nuclear factor-κB (NF-κB). Not much is known if this compound affects signal transduction in non-tumor cells. I investigated whether parthenolide affected Ca2+ signaling in endothelial cells, key components in regulating the vascular tone. In this work using mouse cortical microvascular bEND.3 endothelial cells, I found that a 15-h treatment with parthenolide resulted in amplified ATP-triggered Ca2+ signal; the latter had a very slow decay rate suggesting suppression of Ca2+ clearance. Evidence suggests parthenolide suppressed Ca2+ clearance by inhibiting the plasmalemmal Ca2+ pump; such suppression did not result from decreased expression of the plasmalemmal Ca2+ pump protein. Rather, such suppression was possibly a consequence of endoplasmic reticulum (ER) stress, since salubrinal (an ER stress protector) was able to alleviate parthenolide-induced Ca2+ clearance suppression. Given the current deployment of parthenolide as an anti-cancer drug in clinical trials and the potential usage of this lactone as a cardioprotectant, it is important to examine in details the perturbing effects of parthenolide on Ca2+ homeostasis in endothelial cells and neighboring vascular smooth muscle cells, activities of which exert profound effects on hemodynamics.
|
|---|
