Abstract-Epidemiological evidence suggests a role for sex-dependent mechanisms in the pathophysiology of hypertension. It has been shown that 5␣-dihydrotestosterone (DHT) administration (56 mg/kg of body weight per day IP for 14 days) increases blood pressure, cytochrome P450 4A expression, and 20-hydroxyeicosatetraenoic acid synthesis in rats.We examined whether increased vascular 20-hydroxyeicosatetraenoic acid synthesis underlies endothelial dysfunction and hypertension in DHT-treated male Sprague-Dawley rats by using HET0016, a selective cytochrome P450 4A inhibitor. Coadministration of HET0016 (10 mg/kg per day IP for 14 days) to DHT-treated rats markedly reduced DHT-induced interlobar arterial production of 20-hydroxyeicosatetraenoic acid (14.3Ϯ1.5 versus 1.5Ϯ0.5 ng/mg of protein per hour; PϽ0.05), superoxide anion (246Ϯ47 versus 31Ϯ8 cpm/g of protein), and the levels of gp91-phox, p47-phox, and 3-nitrosylated proteins. Moreover, the maximal relaxing response to acetylcholine in phenylephrinepreconstricted renal interlobar arteries from DHT-treated rats (42.8Ϯ4.8%) significantly (PϽ0.05) increased in the presence of HET0016 (81.5Ϯ10.8%). Importantly, the administration of HET0016 negated DHT-induced hypertension; systolic blood pressure was reduced from 146Ϯ2 mm Hg in DHT-treated rats to 130Ϯ1 mm Hg ( The synthesis of 20-HETE is catalyzed primarily by enzymes of the cytochrome P450 (CYP) 4A family. 6,7 CYP4A proteins are present in vascular tissues and show distinct distribution along the vascular tree. 8 Suppression and overexpression of CYP4A proteins in small arteries and arterioles decreases and increases, respectively, vascular reactivity and myogenic tone 7,9,10 ; these effects can be reversed by the addition of 20-HETE or inhibition of its synthesis.CYP4A and 20-HETE synthesis have been linked to hypertension in numerous experimental models. In the spontaneously hypertensive rat, depletion or inhibition of CYP4A activity lowers blood pressure (BP). 11,12 Inhibition of vascular 20-HETE synthesis by intravenous administration of CYP4A1 or CYP4A2 antisense oligonucleotides decreases BP in normotensive and hypertensive rats, 6,7 whereas transduction with adenoviruses expressing the CYP4A2 protein increases vascular CYP4A expression and 20-HETE levels and augments BP. 13 A role for androgens in promoting elevation of BP is well recognized 14 and, according to recent studies, such a role may rely on increased synthesis of vascular 20-HETE. Hence, mice deficient in cyp4a14 (the mouse homologue of CYP4A2) displayed androgen-sensitive hypertension, which was reversed by castration. 15 In these mice, cyp4a12 expression (the mouse homologue of CYP4A8) is elevated and so is renal microsomal 20-HETE synthesis. Similarly, androgeninduced hypertension in rats treated with 5␣-dihydrotestosterone (DHT) has been associated with increased CYP4A8 expression and renal vascular 20-HETE synthesis. 16 The mechanisms by which 20-HETE promotes hypertension are primarily linked to its ability to sensitize constrictor responsi...

Endothelial dysfunction is associated with endothelial cell activation, i.e., up-regulation of surface cell adhesion molecules and the release of proinflammatory cytokines. 20-Hydroxyeicosatetraenoic acid (HETE), a major vasoactive eicosanoid in the microcirculation, has been implicated in the regulation of endothelial cell function through its angiogenic and pro-oxidative properties. We examined the effects of 20-HETE on endothelial cell activation in vitro. Cells transduced with adenovirus containing either CYP4A1 or CYP4A2 produced higher levels of 20-HETE, and they demonstrated increased expression levels of the adhesion molecule intercellular adhesion molecule (ICAM) (4 -7-fold) and the oxidative stress marker 3-nitrotyrosine (2-3-fold) compared with cells transduced with control adenovirus. Treatment of cells with 20-HETE markedly increased levels of prostaglandin (PG) E 2 and 8-epi-isoprostane PGF 2␣ , commonly used markers of activation and oxidative stress, and most prominently, interleukin-8, a potent neutrophil chemotactic factor whose overproduction by the endothelium is a key feature of vascular injury. 20-HETE at nanomolar concentrations increased inhibitor of nuclear factor-B phosphorylation by 2 to 5-fold within 5 min, which was followed with increased nuclear translocation of nuclear factor-B (NF-B). Likewise, 20-HETE activated the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway by stimulating phosphorylation of ERK1/2. Inhibition of NF-B activation and inhibition of ERK1/2 phosphorylation inhibited 20-HETE-induced ICAM expression. It seems that 20-HETE triggers NF-B and MAPK/ERK activation and that both signaling pathways participate in the cellular mechanisms by which 20-HETE activates vascular endothelial cells.

Nitric oxide (NO), generated from L-arginine by endothelial nitric oxide synthase (eNOS), is a key endothelial-derived factor whose bioavailability is essential to the normal function of the endothelium. Endothelium dysfunction is characterized by loss of NO bioavailability because of either reduced formation or accelerated degradation of NO. We have recently reported that overexpression of vascular cytochrome P-450 (CYP) 4A in rats caused hypertension and endothelial dysfunction driven by increased production of 20-hydroxyeicosatetraenoic acid (20-HETE), a major vasoconstrictor eicosanoid in the microcirculation. To further explore cellular mechanisms underlying CYP4A-20-HETE-driven endothelial dysfunction, the interactions between 20-HETE and the eNOS-NO system were examined in vitro. Addition of 20-HETE to endothelial cells at concentrations as low as 1 nM reduced calcium ionophore-stimulated NO release by 50%. This reduction was associated with a significant increase in superoxide production. The increase in superoxide in response to 20-HETE was prevented by N(G)-nitro-L-arginine methyl ester, suggesting that uncoupled eNOS is a source of this superoxide. The response to 20-HETE was specific in that 19-HETE did not affect NO or superoxide production, and, in fact, the response to 20-HETE could be competitively antagonized by 19(R)-HETE. 20-HETE had no effect on phosphorylation of eNOS protein at serine-1179 or threonine-497 following addition of calcium ionophore; however, 20-HETE inhibited association of eNOS with 90-kDa heat shock protein (HSP90). In vivo, impaired acetylcholine-induced relaxation in arteries overexpressing CYP4A was associated with a marked reduction in the levels of phosphorylated vasodilator-stimulated phosphoprotein, an indicator of bioactive NO, that was reversed by inhibition of 20-HETE synthesis or action. Because association of HSP90 with eNOS is critical for eNOS activation and coupled enzyme activity, inhibition of this association by 20-HETE may underlie the mechanism, at least in part, by which increased CYP4A expression and activity cause endothelial dysfunction.

Toxin production during cyanobacterial blooms poses a significant public health threat in water bodies globally and requires the development of effective bloom management strategies. Previously, synthesis of the hepatotoxin microcystin has been proposed to be regulated by iron availability, but the contribution of the toxin to the adaptation of cyanobacteria to environmental stresses, such as changing light intensity and nutrient limitation, remains unclear. The aim of this study was to compare the iron stress response in toxic and non-toxic strains of Microcystis aeruginosa subjected to moderate and severe iron limitation. The transcription of a number of genes involved in iron uptake, oxidative stress response, toxin synthesis and transcriptional control of these processes was accessed by quantitative real-time PCR (qRT-PCR). The process of adaptation of M. aeruginosa to iron stress was found to be highly dynamic and strain-specific. Toxin production in PCC 7806 increased in an iron-dependent manner and appeared to be regulated by FurA. The inability to produce microcystin, either due to natural mutations in the mcy gene cluster or due to insertional inactivation of mcyH, affected the remodelling of the photosynthetic machinery in iron-stressed cells, the transport of Fe(II) and transcription of the Fur family of transcriptional regulators. The presence of the toxin appears to give an advantage to microcystin-producing cyanobacteria in the early stages of exposure to severe iron stress and may protect the cell from reactive oxygen species-induced damage.

Although the mechanism underlying the effect of androgen on BP and cardiovascular disease is not well understood, recent studies suggest that 8,11,, a primary cytochrome P450 4 (Cyp4)-derived eicosanoid, may mediate androgen-induced hypertension. Here, treatment of normotensive mice with 5a-dihydrotestosterone increased BP and induced both Cyp4a12 expression and 20-HETE levels in preglomerular microvessels. Administration of a 20-HETE antagonist prevented and reversed the effects of dihydrotestosterone on BP. Cyp4a14(2/2) mice, which exhibit androgen-sensitive hypertension in the male mice, produced increased levels of vascular 20-HETE; furthermore, administration of a 20-HETE antagonist normalized BP. To examine whether androgen-independent increases in 20-HETE are sufficient to cause hypertension, we studied Cyp4a12-transgenic mice, which express the CYP4A12-20-HETE synthase under the control of a doxycycline-sensitive promoter. Administration of doxycycline increased BP by 40%, and administration of a 20-HETE antagonist prevented this increase. Levels of CYP4A12 and 20-HETE in preglomerular microvessels of doxycycline-treated transgenic mice approximately doubled, correlating with increased 20-HETE-dependent sensitivity to phenylephrine-mediated vasoconstriction and with decreased acetylcholine-mediated vasodilation in the renal microvasculature. We observed a similar contribution of 20-HETE to myogenic tone in the mesenteric microvasculature. Taken together, these results suggest that 20-HETE both mediates androgeninduced hypertension and can cause hypertension independent of androgen. 24: 128824: -129624: , 201324: . doi: 10.1681 The v-hydroxylation of arachidonic acid (AA) to 20-hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE) is catalyzed by members of the cytochrome P450 4 (CYP4) gene family and regulated by factors such as age, sex hormones, and dietary lipids. 1,2 CYP4 expression and 20-HETE synthesis have been implicated in the regulation of vascular and tubular function and the development of hypertension in experimental models. [3][4][5] Studies demonstrating that 20-HETE is a vasoconstrictor [6][7][8] suggest that increased 20-HETE synthesis and/or effects in the renal vasculature underlies its prohypertensive property. [9][10][11][12] This notion has been substantiated by several reports showing the following: (1) the synthesis of and vascular reactivity to 20-HETE are significantly higher in spontaneously hypertensive rats (SHRs), 13,14 (2) inhibition of vascular 20-HETE synthesis by CYP4A2 antisense oligonucleotides decreases BP in SHRs, 15,16 J Am Soc Nephrol

In the synchronized switching damping (SSD) techniques, the voltage on the piezoelectric element is switched synchronously with the vibration to be controlled using an inductive shunt circuit (SSDI). The inherent capacitance and the inductance in the shunt circuit comprise an electrically resonant circuit. In this study, a negative capacitance is used in the shunt circuit instead of an inductance in the traditional SSD technique. The voltage on the piezoelectric element can be effectively inverted although the equivalent circuit is capacitive and no resonance occurs. In order to investigate the principle of the new SSD method based on a negative capacitance (SSDNC), the variation of the voltage on the piezoelectric element and the current in the circuit are analyzed. Furthermore, the damping effect using the SSDNC is deduced, and the energy balance and stability of the new system are investigated analytically. The method is applied to the single-mode control and two-mode control of a composite beam, and its control performance was confirmed by the experimental results. For the first mode in single-mode control, the SSDNC is much more effective than SSDI. In other cases, the SSDNC is also more effective than the SSDI, although not significantly.

Abstract-We have shown previously that increased vascular endothelial expression of CYP4A2 leads to 20-hydroxyeicosatetraenoic (20-HETE)-dependent hypertension. The renin-angiotensin system is a key regulator of blood pressure.In this study, we examined possible interactions between 20-HETE and the renin-angiotensin system. In normotensive (110Ϯ3 mm Hg) Sprague-Dawley rats transduced with a lentivirus expressing the CYP4A2 cDNA under the control of an endothelial-specific promoter (VECAD-4A2), systolic blood pressure increased rapidly, reaching 139Ϯ1, 145Ϯ3, and 150Ϯ2 mm Hg at 3, 5, and 10 days after transduction; blood pressure remained elevated, thereafter, with maximum levels of 163Ϯ3 mm Hg. Treatment with lisinopril, losartan, or the 20-HETE antagonist 20-hydroxyeicosa-6(Z), 15(Z)-dienoic acid decreased blood pressure to control values, but blood pressure returned to its high levels after cessation of treatment. Endothelial-specific overexpression of CYP4A2 resulted in increased expression of vascular angiotensin-converting enzyme (ACE) and angiotensin II type 1 receptor and increased levels of plasma and tissue angiotensin II; all were attenuated by treatment with HET0016, an inhibitor of 20-HETE synthesis, or with 20-hydroxyeicosa-6(Z), 15(Z)-dienoic acid. In cultured endothelial cells, 20-HETE specifically and potently induced ACE expression without altering the expression of ACE2, angiotensinogen, or angiotensin II receptors. This is the first study to demonstrate that 20-HETE, a key constrictor eicosanoid in the microcirculation, induces ACE and angiotensin II type 1 receptor expression and increases angiotensin II levels, suggesting that the mechanisms by which 20-HETE promotes hypertension include activation of the renin-angiotensin system that is likely initiated at the level of ACE induction. It is endowed with unique biological activities relevant to the regulation of vascular tone, renal function, and blood pressure. The involvement of 20-HETE in hypertension has been significantly documented in animal models and humans. 20-HETE is a vasoconstrictor and natriuretic eicosanoid; hence, its contribution to the regulation of blood pressure depends on its sites of synthesis and action. In the vasculature, 20-HETE enhances the responsiveness to constrictor stimuli, including pressure, oxygen, phenylephrine, and endothelin 1, as well as inhibition of NO synthesis by mechanisms that include production of constrictor eicosanoids, inhibition of the smooth muscle cell large conductance Ca 2ϩ -activated K ϩ channel, and the sensitization of the contractile apparatus to [Ca 2ϩ ] through phosphorylation of MLC20. 1 We and others 2-4 demonstrated that, in circulatory districts other than the pulmonary, 20-HETE reduces endothelial-dependent vasorelaxation by uncoupling endothelial NO synthase. 5,6 In the kidney, 20-HETE biosynthesis is prominent in the proximal tubule and thick ascending limb. It promotes natriuresis by inhibiting Na ϩ /K ϩ -ATPase activity in the proximal tubule and inhibiting the Na ϩ -K ϩ -...

Carbon fiber nanoelectrodes (tip diameter = ca. 100 nm) have been first used to monitor real-time dopamine release from single living vesicles of single rat pheochromocytoma (PC12) cells. The experiments show that active and inactive release sites exist on the surface of cells, and the spatial distributions have been differentiated even in the same active release zone. It is first demonstrated that multiple vesicles can sequentially release dopamine at the same site of the cell surface, which possibly plays the main role in the dopamine release from PC12 cells.