Estradiol augments while progesterone inhibits arginine transport in human endothelial cells through modulation of cationic amino acid transporter-1

Abstract

Decreased generation of nitric oxide (NO) by endothelial NO synthase (eNOS) characterizes endothelial dysfunction (ECD). Delivery of arginine to eNOS by cationic amino acid transporter-1 (CAT-1) was shown to modulate eNOS activity. We found in female rats, but not in males, that CAT-1 activity is preserved with age and in chronic renal failure, two experimental models of ECD. In contrast, during pregnancy CAT-1 is inhibited. We hypothesize that female sex hormones regulate arginine transport. Arginine uptake in human umbilical vein endothelial cells (HUVEC) was determined following incubation with either 17β-estradiol (E2) or progesterone. Exposure to E2 (50 and 100 nM) for 30 min resulted in a significant increase in arginine transport and reduction in phosphorylated CAT-1 (the inactive form) protein content. This was coupled with a decrease in phosphorylated MAPK/extracellular signal-regulated kinase (ERK) 1/2. Progesterone (1 and 100 pM for 30 min) attenuated arginine uptake and increased phosphorylated CAT-1, phosphorylated protein kinase Cα (PKCα), and phosphorylated ERK1/2 protein content. GO-6976 (PKCα inhibitor) prevented the progesterone-induced decrease in arginine transport. Coincubation with both progesterone and estrogen for 30 min resulted in attenuated arginine transport. While estradiol increases arginine transport and CAT-1 activity through modulation of constitutive signaling transduction pathways involving ERK, progesterone inhibits arginine transport and CAT-1 via both PKCα and ERK1/2 phosphorylation, an effect that predominates over estradiol.

Does l‐citrulline supplementation improve exercise blood flow in older adults?

“…Moreover, oestrogen is found to increase CAT‐1 activity by decreasing phosphorylated ERK (Bentur et al . ), and oestrogen decreases arginase levels (enzyme that metabolizes arginine) in vascular cells (Hayashi et al . ), thus older women with low oestrogen levels may have impaired Arg transport and/or intracellular activity compared with older men.…”

“…Older female rats are shown to have higher phosphorylated PKCα protein levels than older male rats (Schwartz et al 2009). Moreover, oestrogen is found to increase CAT-1 activity by decreasing phosphorylated ERK (Bentur et al 2015), and oestrogen decreases arginase levels (enzyme that metabolizes arginine) in vascular cells (Hayashi et al 2006), thus older women with low oestrogen levels may have impaired Arg transport and/or intracellular activity compared with older men. Although this study cannot address whether this was a contributing factor in our results, we cannot discount the possibility The change in blood flow and FVC was different between L-citrulline (Cit) and placebo only for exercise at the higher workload in men.…”

Vascular Endothelial Growth Factor Augments Arginine Transport and Nitric Oxide Generation via a KDR Receptor Signaling Pathway

“…We chose to examine PKC-α, JNK, and ERK1/2, which have been previously shown to modulate eNOS or CAT-1 signaling pathways [16] Utilizing Western blotting we found that the expression of PKC-α, JNK, and ERK1/2 and their phosphorylated forms were unchanged following incubation of HUVEC with VEGF 100 ng/ml for 60 (data not shown).…”

“…We have previously reported, in four different experimental models characterized by diminished arginine transport, namely hypercholesterolemia, chronic renal failure, pregnancy, and aging in the male rat, a post-translational modulation of CAT-1 which was associated with upregulation of PKC-α [8, 11, 12]. In addition, both MAPK's JNK and ERK1/2 were found to modulate CAT-1 [16]. Therefore, we pursued with experiments looking at these proteins as a possible link between VEGF and CAT-1.…”

“…Endothelial CAT-1, PKC-α, phosphorylated PKC-α (p-PKC-α), MAPK extracellular-signal regulated kinase (ERK), phosphorylated ERK (p-ERK), MAPK Jun N-terminal kinase (JNK) and phosphorylated JNK (p-JNK) protein expression were determined by immunoblotting as previously described [16]. In brief, cells were separately placed in ice-cold PBS lysis buffer (pH 7.4), containing protease inhibitors, then mechanically homogenized and left on ice for 45 min.…”