Estradiol, acting through estrogen receptor alpha, restores dimethylarginine dimethylaminohydrolase activity and nitric oxide production in oxLDL-treated human arterial endothelial cells

Abstract

a b s t r a c tAsymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) synthase. ADMA accumulation, mainly due to a decreased dimethylarginine dimethylaminohydrolase (DDAH) activity, has been related to the development of cardiovascular diseases. We investigate whether estradiol prevents the changes induced by oxidized low density lipoprotein (oxLDL) on the DDAH/ADMA/NO pathway in human umbilical artery endothelial cells (HUAEC). HUAEC were exposed to estradiol, native LDL (nLDL), oxLDL and their combinations for 24 h. In some experiments, cells were also exposed to the unspecific estrogen receptor (ER) antagonist ICI 182780, the specific ERa antagonist MPP or specific agonists for ERa, ERb and GPER. ADMA concentration was measured by HPLC and concentration of NO by amperometry. Protein expression and DDAH activity were measured by immunoblotting and an enzymatic method, respectively. oxLDL, but not nLDL, increased ADMA concentration with a concomitant decrease on DDAH activity. oxLDL reduced eNOS protein and NO production. Estradiol alone had no effects on DDAH/ADMA/NO pathway, but increased the attenuated endothelial NO production induced by oxLDL by reduction in ADMA and preventing loss of eNOS protein levels. ICI 182780 and MPP completely abolished these effects of estradiol on oxLDL-exposed cells. ERa agonist, but not ERb and GPER agonists, mirrored estradiol effects on NO production. In conclusion, estradiol restores (1) DDAH activity, and therefore ADMA levels, and (2) NO production impaired by oxLDL in HUAEC acting through ERa.

Endothelial transcriptomic changes induced by oxidized low density lipoprotein disclose an up-regulation of Jak–Stat pathway

“…Our group has previously described that the oxidation state of LDL has a differential regulatory effect in several mechanisms that are of importance for the development of atherosclerosis [12]. To test whether Fig.…”

“…Human umbilical artery endothelial cells (HUAECs) were isolated from arteries of human umbilical cords, as previously described [12]. Briefly, umbilical arteries were isolated, cannulated, washed with PBS and incubated for 15 min with 0.2% collagenase type II.…”

“…The nLDL and oxLDL concentrations used in the experiments were selected based on the literature and on previous experience [12][13][14]. To discard any potential toxic effect of the compounds on HUAECs, cell viability was assessed as previously reported [15].…”

Phosphorylation of Nonmuscle Myosin Light Chain Promotes Endothelial Injury in Hyperlipidemic Rats Through a Mechanism Involving Downregulation of Dimethylarginine Dimethylaminohydrolase 2

“…Reports from other laboratories and ours have repeatedly demonstrated suggests that DDAH/ADMA pathway plays a key role in regulation of vascular endothelial functions under physiological and pathological conditions. 8,9,22,23 As an endogenous inhibitor of NOS, ADMA is emerging as a biomarker for endothelial dysfunction. 24 In this study, we have found that the plasma levels of ADMA were significantly elevated in the hyperlipidemic rats concomitant with endothelial injury.…”

“…7 Accumulating evidence has demonstrated that the elevated plasma levels of ADMA in many cardiovascular diseases including hyperlipidemia are due to, at least partially, the decrease in DDAH activity. [8][9][10][11] Although the activities of DDAH have been reported to be reduced in hyperlipidemic animals, it is not known whether the decrease in DDAH activities involves the suppression of DDAH gene expression.…”