Effects Of 15-HETE Research Articles

Activation of ERK pathway is required for 15-HETE-induced angiogenesis in human umbilical vascular endothelial cells

Angiogenesis plays a critical role in the progression of cardiovascular disease, retinal ischemia, or tumorigenesis. The imbalance of endothelial cell proliferation and apoptosis disturbs the establishment of the vasculogenesis, which is affected by several arachidonic acid metabolites. 15-Hydroxyeicosatetraenoic acid (15-HETE) is one of the metabolites. However, the underlying mechanisms of angiogenesis induced by 15-HETE in human umbilical vascular endothelial cells (HUVECs) are still poorly understood. Since extracellular signal-regulated kinase (ERK) is a critical regulator of cell proliferation, there may be a crosstalk between 15-HETE-regulating angiogenic process and ERK-proliferative effect in HUVECs. To test this hypothesis, we study the effect of 15-HETE on cell proliferation, angiogenesis, and apoptosis using cell viability measurement, cell cycle analysis, western blot, scratch–wound, tube formation assay, and nuclear morphology determination. We found that 15-HETE promoted HUVEC angiogenesis, which were mediated by ERK. Moreover, 15-HETE-induced proliferation and cell cycle transition from the G0/G1 phase to the G2/M + S phase. All these effects were reversed after blocking ERK with PD98059 (an ERK inhibitor). In addition, HUVEC apoptosis was relieved by 15-HETE through the ERK pathway. Thus, ERK is necessary for the effects of 15-HETE in the regulation of HUVEC angiogenesis, which may be a novel potential target for the treatment of angiogenesis-related diseases.

Cellular mechanisms and intracellular signaling pathways for the modulation of eNOS in pulmonary arteries by 15-HETE

The 15-hydroxyeicosatetraenoic acid (15-HETE), a lipid metabolite and vasoconstrictor, plays an important role in hypoxic contraction of pulmonary arteries (PAs) through working on smooth muscle cells (SMCs). Previous studies have shown that vascular endothelium is also involved in PAs tone regulation. However, little is known as to how the pulmonary artery endothelial cells (PAECs) are related to the 15-HETE-induced vasoconstriction and that which intracellular signaling systems are critical. To test this hypothesis, we examined PAs constriction in isolated rat PAs rings, the expression and activity of endothelial nitric oxide synthase (eNOS) with western blot, and nitric oxide (NO) production using the DAF-FM DA fluorescent indicator. The results showed that the 15-HETE-induced PAs constriction was diminished in endothelium-intact rings. In the presence of the eNOS inhibitor L-NAME, vasoconstrictor responses to KCl were greater than the control. The activation of eNOS was activated by Ca2+ released from intracellular stores and the PI3K/Akt pathway. Phosphorylations of the eNOS at Ser-1177 and Akt at Ser-473 were necessary for their activity. A prolonged 15-HETE treatment (30 min) led to a decrease in NO production by phosphorylation of eNOS at Thr-495, leading to augmentation of PAs constriction. Therefore, 15-HETE initially inhibited the PAs constriction through the endothelial NO system, and both Ca2+ and the PI3K/Akt signaling systems are required for the effects of 15-HETE on PAs tone regulation.

The role of ERK1/2 in 15-HETE-inhibited apoptosis in pulmonary arterial smooth muscle cells

15-Hydroxyeicosatetrenoic acid (15-HETE) is an important product of arachidonic acid catalyzed by 15-lipoxygenase (15-LO) in the wall of pulmonary vessels, which plays a key role in pulmonary arterial hypertension. The previous studies showed that 15-HETE inhibits apoptosis. It is still unknown, however, whether 15-HETE acts on the apoptotic responses through the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway. The aim of the study is to test the hypothesis that ERK1/2 pathway participates in the protective effects of 15-HETE on the cell survival. This hypothesis was validated by cell viability measurement, nuclear morphology determination, terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay, mitochondrial potentials assay and Western blot. We found that 15-HETE enhanced cell survival, suppressed the expression of phosphatase and tensin homologue deleted on chromosome ten, upregulated X-linked inhibitor of apoptosis protein and Bcl-2 and attenuated mitochondrial depolarization in pulmonary artery muscle smooth cells (PASMCs) under serum-deprived conditions. These effects were reversed by ERK1/2 inhibitor PD98059. Taken together, our data indicated that the ERK1/2 kinase is a regulator of PASMC apoptosis, and potential therapeutical strategy for pulmonary hypertension may be developed by targeting at intracellular signaling systems centered by the kinase.

The influence of an inhibitor of lipoxygenases on the modulation of the plasticity of cholinoreceptors by 15-HETE.

The role of acyclic eicosanoids in the modulation of the plasticity of somatic cholinoreceptors by 15(S)-hydroxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid (15-HETE) was investigated in identified RPa3 and LPa3 neurons of Helix lucorum using the two-electrode voltage clamping technique. It was demonstrated that the inhibitor of lipoxygenases, nordihydroguaiaretic acid (2-20 microM), completely blocks the short- and long-latency effects of 15-HETE (2-20 microM) on the depth of damping of the inward current induced by rhythmic applications of acetylcholine (ACh) to the soma. It is concluded that the short-latency effect of 15-HETE on the plasticity of cholinoreceptors is determined by its inhibition mainly of 5-lipoxygenase, which leads to a reduction in the level of acyclic eicosanoids that are formed under the influence of this enzyme. The potentiation of the effects of the acyclic eicosanoids, whose synthesis is resumed in the second phase, probably underlies the direct long-latency modulatory effect of 15-HETE.

Cutaneous Inflammation: Effects of Hydroxy Acids and Eicosanoid Pathway Inhibitors on Vascular Permeability

Four metabolic products of arachidonic acid lipoxygenation, 5-hydroxyeicosatetraenoate (5-HETE), 12-HETE, 15-HETE, 5(S),12(S)-DiHETE, were injected intradermally into depilated dorsae of albino guinea pigs. The presence of intravenously injected 125I-bovine serum albumin (10uCi/kg) in 13-mm punch biopsy specimens served as a marker for altered vascular response; histologic changes were evaluated at 6 and 24 h after the injection in 1-micron-thick sections. Thirty minutes after the injections of 15 nanomoles and 60 nanomoles of 5-HETE, the ratios of radioactivity in HETE-injected to that in buffer-injected sites were 1.35 +/- 0.06 (mean +/- SE) and 2.80 +/- 0.27, respectively. Corresponding effects of 15-HETE were 1.39 +/- 0.17 and 1.63 +/- 0.21, respectively. Values for 60 nanomoles of 12-HETE and 5,12-DiHETE were intermediate in comparison with the above eicosanoids. The most notable histologic changes were a neutrophilic infiltrate induced by 12-HETE at 6 and 24 h, and neutrophilic and eosinophilic infiltrates in response to 5,12-DiHETE injection at 6 and 24 h. Effects of topically applied eicosanoid pathway inhibitors were also evaluated, using intradermally injected sodium arachidonate (AA) as agonist. Three mixed cyclooxygenase/lipoxygenase inhibitors, BW755C, phenidone, and nordihydroguaiaretic acid, suppressed vascular response by 9%, 9%, and 6% for 150 nmol of AA, and by 9%, 13%, and 12% for 300 nmol of AA, respectively. The cyclooxygenase inhibitor, indomethacin, induced suppressions of 39% for 150 nmol AA and 22% for 300 nmol AA, respectively. These data demonstrate that metabolites of both cyclooxygenase and lipoxygenase eicosanoid pathways are involved in alteration in vascular response accompanying cutaneous inflammation.

Vascular Effects of 15-Hydroperoxyeicosatetraenoic Acid and 15-Hydroxyeicosatetraenoic Acid on Canine Arteries

The vascular effects of 15-hydroperoxyeicosatetraenoic acid (15-HPETE) and 15-hydroxyeicosatetraenoic acid (15-HETE) were investigated on isolated helical strips of canine cerebral and coronary arteries. 15-HPETE caused strong concentration-related contraction of cerebral arteries under resting tension. After contraction with prostaglandin F2α (PGF2α), 15-HPETE caused marked relaxation of coronary arteries. The effects of 15-HETE on isolated canine arteries were similar to those of 15-HPETE. The relaxation of coronary arteries caused by both 15-HPETE and 15-HETE was completely inhibited in the presence of aspirin, but not in the presence of tranylcypromine. Preincubation of coronary and cerebral arterial strips with 15-HPETE or 15-HETE resulted in suppression of the production of 6-keto-PGF1α from exogenously added arachidonic acid; and 15-HPETE, but not 15-HETE, enhanced the production of HETE(s) significantly. Aspirin blocked 15-HPETE induced HETE(s) production in coronary arteries. On cerebral angiography, strong contraction of intracranial arteries was observed after intracisternal injection of 15-HPETE. On the other hand, 15-HETE.had little effect on intracranial arteries in vivo. The mechanism of the vascular effects of 15-HPETE and 15-HETE will be discussed.