20-Hydroxyeicosatetraenoic Acid Contributes to the Inhibition of K+ Channel Activity and Vasoconstrictor Response to Angiotensin II in Rat Renal Microvessels

The present study examined whether 20-hydroxyeicosatetraenoic acid (HETE) contributes to the vasoconstrictor effect of angiotensin II (ANG II) in renal microvessels by preventing activation of the large conductance Ca²⁺-activated K⁺ channel (K_Ca) in vascular smooth muscle (VSM) cells. ANG II increased the production of 20-HETE in rat renal microvessels. This response was attenuated by the 20-HETE synthesis inhibitors, 17-ODYA and HET0016, a phospholipase A₂ inhibitor AACOF₃, and the AT₁ receptor blocker, Losartan, but not by the AT₂ receptor blocker, PD123319. ANG II (10^-11 to 10^-6 M) dose-dependently decreased the diameter of renal microvessels by 41 ± 5%. This effect was blocked by 17-ODYA. ANG II (10^-7 M) did not alter K_Ca channel activity recorded from cell-attached patches on renal VSM cells under control conditions. However, it did reduce the NPo of the K_Ca channel by 93.4 ± 3.1% after the channels were activated by increasing intracellular calcium levels with ionomycin. The inhibitory effect of ANG II on K_Ca channel activity in the presence of ionomycin was attenuated by 17-ODYA, AACOF₃, and the phospholipase C (PLC) inhibitor U-73122. ANG II induced a peak followed by a steady-state increase in intracellular calcium concentration in renal VSM cells. 17-ODYA (10^-5 M) had no effect on the peak response, but it blocked the steady-state increase. These results indicate that ANG II stimulates the formation of 20-HETE in rat renal microvessels via the AT₁ receptor activation and that 20-HETE contributes to the vasoconstrictor response to ANG II by blocking activation of K_Ca channel and facilitating calcium entry.     

TNFalpha-induced cyclooxygenase 2 not only increases the vasopermeability of blood-brain barrier but also enhances the neutrophil survival in Escherichia coli-induced brain inflammation.

In Escherichia coli-induced brain inflammation, cyclooxygenase-2 was induced not only on brain arterioles at 3 h, but also on infiltrating neutrophils at 9 h post-intracerebral injection. Intravenous injection of E. coli or recombinant TNFalpha also induced cyclooxygenase-2 expression on arterioles. Cyclooxygenase-2 and TNFalpha were co-localized on the arterioles as well as the infiltrating neutrophils by serial-section staining, indicating that cyclooxygenase-2 was induced by TNFalpha. NS398 (a cyclooxygenase-2 selective inhibitor) not only inhibited the increase of blood-brain barrier permeability, but also enhanced the apoptosis of the infiltrating neutrophils after E. coli stimulation. This suggests that TNFalpha-stimulated cyclooxygenase-2 induction play an important role on E. coli-induced brain inflammation. Its inhibition would help the resolution of neutrophil-mediated brain inflammation.     

Suppression of osteoprotegerin expression by prostaglandin E2 is crucially involved in lipopolysaccharide-induced osteoclast formation

LPS is a potent stimulator of bone resorption in inflammatory diseases. The mechanism by which LPS induces osteoclastogenesis was studied in cocultures of mouse osteoblasts and bone marrow cells. LPS stimulated osteoclast formation and PGE(2) production in cocultures of mouse osteoblasts and bone marrow cells, and the stimulation was completely inhibited by NS398, a cyclooxygenase-2 inhibitor. Osteoblasts, but not bone marrow cells, produced PGE(2) in response to LPS. LPS-induced osteoclast formation was also inhibited by osteoprotegerin (OPG), a decoy receptor of receptor activator of NF-kappaB ligand (RANKL), but not by anti-mouse TNFR1 Ab or IL-1 receptor antagonist. LPS induced both stimulation of RANKL mRNA expression and inhibition of OPG mRNA expression in osteoblasts. NS398 blocked LPS-induced down-regulation of OPG mRNA expression, but not LPS-induced up-regulation of RANKL mRNA expression, suggesting that down-regulation of OPG expression by PGE(2) is involved in LPS-induced osteoclast formation in the cocultures. NS398 failed to inhibit LPS-induced osteoclastogenesis in cocultures containing OPG knockout mouse-derived osteoblasts. IL-1 also stimulated PGE(2) production in osteoblasts and osteoclast formation in the cocultures, and the stimulation was inhibited by NS398. As seen with LPS, NS398 failed to inhibit IL-1-induced osteoclast formation in cocultures with OPG-deficient osteoblasts. These results suggest that IL-1 as well as LPS stimulates osteoclastogenesis through two parallel events: direct enhancement of RANKL expression and suppression of OPG expression, which is mediated by PGE(2) production.     

Contribution of cytochrome P-450 omega-hydroxylase to altered arteriolar reactivity with high-salt diet and hypertension

The present study evaluated the contribution of cytochrome P-450 omega-hydroxylase in modulating the reactivity of cremaster muscle arterioles in normotensive rats on high-salt (HS) and low-salt (LS) diet and in rats with reduced renal mass hypertension (RRM-HT). Changes in arteriolar diameter in response to ACh, sodium nitroprusside (SNP), ANG II, and elevated O(2) were measured via television microscopy under control conditions and following cytochrome P-450 omega-hydroxylase inhibition with 17-octadecynoic acid (17-ODYA) or N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS). In normotensive rats on either LS or HS diet, resting tone was unaffected and arteriolar reactivity to ACh or SNP was minimally affected by cytochrome P-450 omega-hydroxylase inhibition. In RRM-HT rats, cytochrome P-450 omega-hydroxylase inhibition reduced resting tone and significantly enhanced arteriolar dilation to ACh and SNP. Treatment with 17-ODYA or DDMS inhibited arteriolar constriction to ANG II and O(2) in all the groups, although the degree of inhibition was greater in RRM-HT than in normotensive animals. These results suggest that metabolites of cytochrome P-450 omega-hydroxylase contribute to the altered reactivity of skeletal muscle arterioles to vasoconstrictor and vasodilator stimuli in RRM-HT.     

Angiotensin II modulates ion transport in rat proximal tubules through CYP metabolites

To assess the effect of angiotensin II on ion transport in rat isolated proximal tubules and establish the arachidonic acid cytochrome P450 metabolites' role mediating angiotensin II effect and to analyze whether corticosteroids play a role modulating this effect, we studied the effect of low (10 and 100 pM) and high (0.1-1 microM) angiotensin II concentrations on proximal tubule ion transport, measured as (86)Rb uptake. Low angiotensin II produced a stimulation on the (86)Rb uptake (195.79 +/- 35, 377.9 +/- 81, and 300 +/- 49 pg (86)Rb/microg protein/2 min, for control and 10 and 100 pM angiotensin II, respectively). High angiotensin II concentration inhibited ion transport (0.1 microM, 57.9 +/- 5 and 1 microM, 47.3 +/- 4 pg (86)Rb/microg protein/2 min), this effect was prevented by 17-ODYA and by losartan, while indomethacin had no effect. Dexamethasone treatment increased angiotensin II-induced (86)Rb uptake inhibition and arachidonic acid metabolism (19-, 20-HETE and 12-HETE), while adrenalectomy partly prevented angiotensin II-induced inhibition and decreased cytochrome P450-dependent arachidonic acid metabolism. In conclusion, high doses of angiotensin II produce inhibition of ion transport in rat isolated proximal tubules; this effect is mediated by AT(1) receptors, involves cytochrome P450-dependent arachidonic acid metabolites, and is upregulated by corticosteroids.     

Cytochrome P-450 metabolites in endothelin-stimulated cardiac hormone secretion

We examined the role of cytochrome P-450-arachidonate (CYP450-AA) metabolites in endothelin-1 (ET-1)-stimulated atrial natriuretic peptide (ANP) and pro-ANP-(1-30) secretion from the heart. 17-Octadecynoic acid (17-ODYA, 10(-5) M) significantly inhibited ANP secretion stimulated by ET-1 (10(-8) M) in the isolated perfused rat atria and inhibited pro-ANP-(1-30) secretion stimulated by ET-1 (10(-8) M) or 20-hydroxyeicosatetraenoic acid in cultured neonatal rat ventricular myocytes (NRVM). In NRVM, 17-ODYA significantly (P < 0.05) increased secretion of cAMP but had no significant effect on the secretion of cGMP from NRVM. Staurosporine, an inhibitor of protein kinase C, completely blocked the inhibitory action of 17-ODYA, whereas a protein kinase A inhibitor, H-89 (5 x 10(-5) M), did not significantly attenuate the effects of 17-ODYA. The results show that the inhibitory action of 17-ODYA on ET-1-augmented ANP secretion is mediated through cAMP and suggest that CYP450-AA may play an important role in ET-1-induced cardiac hormone secretion.     

High dietary salt reduces the contribution of 20-HETE to arteriolar oxygen responsiveness in skeletal muscle

The coupling of tissue blood flow to cellular metabolic demand involves oxygen-dependent adjustments in arteriolar tone, and arteriolar responses to oxygen can be mediated, in part, by changes in local production of 20-HETE. In this study, we examined the long-term effect of dietary salt on arteriolar oxygen responsiveness in the exteriorized, superfused rat spinotrapezius muscle and the role of 20-HETE in this responsiveness. Rats were fed either a normal-salt (NS, 0.45%) or high-salt (HS, 4%) diet for 4-5 wk. There was no difference in steady-state tissue Po(2) between NS and HS rats, and elevation of superfusate oxygen content from 0% to 10% caused tissue Po(2) to increase by the same amount in both groups. However, the resulting reductions in arteriolar diameter and blood flow were less in HS rats than NS rats. Inhibition of 20-HETE formation with N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS) or 17-octadecynoic acid (17-ODYA) attenuated oxygen-induced constriction in NS rats but not HS rats. Exogenous 20-HETE elicited arteriolar constriction that was greatly reduced by the large-conductance Ca(2+)-activated potassium (K(Ca)) channel inhibitors tetraethylammonium chloride (TEA) and iberiotoxin (IbTx) in NS rats and a smaller constriction that was less sensitive to TEA or IbTx in HS rats. Arteriolar responses to exogenous angiotensin II were similar in both groups but more sensitive to inhibition with DDMS in NS rats. Norepinephrine-induced arteriolar constriction was similar and insensitive to DDMS in both groups. We conclude that 20-HETE contributes to oxygen-induced constriction of skeletal muscle arterioles via inhibition of K(Ca) channels and that a high-salt diet impairs arteriolar responses to increased oxygen availability due to a reduction in vascular smooth muscle responsiveness to 20-HETE.     

Regulation of leptin release and lipolysis by PGE2 in rat adipose tissue

The role of eicosanoids formed by adipose tissue from rats was examined in the presence of the specific cyclooxygenase-2 inhibitor NS-398. This agent totally blocked the release of prostaglandin E2 (PGE2) by rat adipose tissue over a 24-h incubation in primary culture. The final concentration of PGE2 after 24 h was 12 nM, and half-maximal inhibition of PGE2 formation required 35 nM NS-398. While inhibition of PGE2 formation by NS-398 had no effect on basal leptin release or lipolysis, it enhanced the lipolytic action of 10 nM isoproterenol by 36%. The in vivo administration of PGE2 doubled serum leptin. PGE2 also directly stimulated leptin release by rat adipose tissue incubated in the presence of 25 nM dexamethasone, which inhibited endogenous PGE2 formation by 94%. The inhibition of lipolysis as well as the stimulation of leptin release by PGE2 were mimicked by N6-cyclopentyladenosine (CPA). These data indicate that exogenous PGE2 can stimulate leptin release by adipose tissue when the basal formation of PGE2 is blocked by dexamethasone. However, while the endogenous formation of PGE2 does not appear to regulate basal lipolysis or leptin release, it may play a role in the activation of lipolysis by catecholamines.     

Evaluation of angiotensins II and III as vasoconstrictors in the hepatic and hindlimb vasculatures of dogs

Previous work has demonstrated that intravenously administered angiotensin II is more potent than angiotensin III as a systemic vasopressor agent. We tested the hypothesis that this difference in potency is caused at least partially by angiotensin II being more potent than angiotensin III as a vasoconstrictor in the hindlimb and hepatic vasculatures. The effects of angiotensins II and III on hindlimb and hepatic blood flow were evaluated in 14 dogs anesthetized with pentobarbital. Blood flows were measured electromagnetically. Graded doses of angiotensins II and III were administered as bolus injections directly into the arterial supply of the hindlimb and liver. On the basis of duration and graphic integration of the flow responses, but not on the basis of absolute changes in amplitude, angiotensin II was significantly more potent than angiotensin III as a vasoconstrictor in the hindlimb vasculature. In the hepatic circulation the flow changes produced by angiotensin II and angiotensin III were not significantly different on the basis of duration, graphic integration, or amplitude. We conclude that (i) differential vasoconstrictor responses of the hindlimb, but not the hepatic circulation, to angiotensins II and III contribute to the difference in systemic vasopressor potency between these two peptides, and (ii) because flow responses are an integral event with duration and constantly varying amplitude, evaluation of vasoconstrictor potency based only upon amplitude of the flow changes can be misleading.     

The nonsteroidal anti-inflammatory drug NS398 reactivates SPARC expression via promoter demethylation to attenuate invasiveness of lung cancer cells

Nonsteroidal anti-inflammatory drugs (NSAIDs) have been shown to exhibit potent anticancer effects in vitro and in vivo. One of the mechanisms by which NSAIDs suppress tumorigenesis is inhibition of angiogenesis and metastasis. In this study, we used a microarray system to study the change of expression profile of metastasis-related genes regulated by NS398, a NSAID and a cyclooxygenase-2 (COX-2) inhibitor. We found that several negative regulators of cell invasion, including secreted protein acidic and rich in cysteine (SPARC), thrombospondin 1 (TSP-1), thrombospondin 3 (TSP-3), and tissue inhibitors of matrix metalloproteinase-2 (TIMP-2) are upregulated by NS398. In addition, we demonstrated that upregulation of SPARC expression by NS398 in human lung cancer cells is mediated by promoter demethylation and associated with a decrease in DNA methyltransferase (DNMT) expression. This is the first report to show that NS398 can inhibit the expression of DNMT1 and 3b. Functional assay indicated that SPARC is a critical mediator for NS398 to inhibit cell invasion. Our results provide new insights for the understanding of the anticancer actions of NSAIDs.     

Age-related loss of neuronal nicotinic receptor expression in the aging mouse hippocampus corresponds with cyclooxygenase-2 and PPAR gamma expression and is altered by long-term NS398 administration

Age-related changes in the mammalian dorsal hippocampus are associated with diminished expression of neuronal nicotinic acetylcholine receptors (nAChR), which is particularly severe in pathologies such as those associated with dementias, including Alzheimer's disease. Because the mouse is a useful model for age-related decline in nAChR expression in the basal forebrain and limbic system, we used immunohistochemistry to examine the influence of long-term (12-month) oral administration of nicotine and/or the cyclooxygenase-2 (COX-2) preferring non-steroidal anti-inflammatory drug (NSAID) NS398 on nAChR alpha4, alpha5, alpha7, and beta4 expression in the C57BL/6 mouse. Inhibitory neurons of the dorsal hippocampus that express nAChRs also constitutively express COX-2 and the peroxisome proliferator-antagonist receptor subtype gamma-2 (PPAR gamma2) which is also a target of NS398. Administration of NS398 correlated with retention of nAChR alpha4 and to a lesser extent nAChR beta4, but not nAChR alpha5 or alpha7, but nicotine exhibited no similar effect. Nicotine and NS398 co-administration abolished the NS398-related effect on nAChR alpha4 retention. These results provide evidence that the interaction during aging between oral administration of nicotine and NSAIDs are not straightforward and could even be antagonistic when combined.     

Angiotensin II Induces Vascular Endocannabinoid Release, Which Attenuates Its Vasoconstrictor Effect via CB1 Cannabinoid Receptors

In the vascular system angiotensin II (Ang II) causes vasoconstriction via the activation of type 1 angiotensin receptors. Earlier reports have shown that in cellular expression systems diacylglycerol produced during type 1 angiotensin receptor signaling can be converted to 2-arachidonoylglycerol, an important endocannabinoid. Because activation of CB(1) cannabinoid receptors (CB(1)R) induces vasodilation and reduces blood pressure, we have tested the hypothesis that Ang II-induced 2-arachidonoylglycerol release can modulate its vasoconstrictor action in vascular tissue. Rat and mouse skeletal muscle arterioles and mouse saphenous arteries were isolated, pressurized, and subjected to microangiometry. Vascular expression of CB(1)R was demonstrated using Western blot and RT-PCR. In accordance with the functional relevance of these receptors WIN55212, a CB(1)R agonist, caused vasodilation, which was absent in CB(1)R knock-out mice. Inhibition of CB(1)Rs using O2050, a neutral antagonist, enhanced the vasoconstrictor effect of Ang II in wild type but not in CB(1)R knock-out mice. Inverse agonists of CB(1)R (SR141716 and AM251) and inhibition of diacylglycerol lipase using tetrahydrolipstatin also augmented the Ang II-induced vasoconstriction, suggesting that endocannabinoid release modulates this process via CB(1)R activation. This effect was independent of nitric-oxide synthase activity and endothelial function. These data demonstrate that Ang II stimulates vascular endocannabinoid formation, which attenuates its vasoconstrictor effect, suggesting that endocannabinoid release from the vascular wall and CB(1)R activation reduces the vasoconstrictor and hypertensive effects of Ang II.     

Differential effects of losartan and candesartan on vasoconstrictor responses in the rat

Losartan has been reported to have inhibitory effects on thromboxane (TP) receptor-mediated responses. In the present study, the effects of 2 nonpeptide angiotensin II (AT1) receptor antagonists, losartan and candesartan, on responses to angiotensin II, the thromboxane A2 mimic, U46619, and norepinephrine were investigated and compared in the pulmonary and systemic vascular beds of the intact-chest rat. In this study, intravenous injections of angiotensin II, U46619, and norepinephrine produced dose-related increases in pulmonary and systemic arterial pressure. Losartan and candesartan, in the doses studied, decreased or abolished responses to angiotensin II. Losartan, but not candesartan, and only in a higher dose, produced small, but statistically significant, reductions in pressor responses to U46619 and to norepinephrine in the pulmonary and systemic vascular beds. Furthermore, losartan significantly reduced arachidonic acid-induced platelet aggregation, whereas candesartan had no effect. Pressor responses to angiotensin II were not changed by thromboxane and alpha-adrenergic receptor antagonists, or by cyclooxygenase and NO synthase inhibitors. These results show that losartan and candesartan are potent selective AT1 receptor antagonists in the pulmonary and systemic vascular beds and that losartan can attenuate thromboxane and alpha-adrenergic responses when administered at a high dose, whereas candesartan in the highest dose studied had no effect on responses to U46619 or to norepinephrine. The present data show that the effects of losartan and candesartan on vasoconstrictor responses are different and that pulmonary and systemic pressor responses to angiotensin II are not modulated or mediated by the release of cyclooxygenase products, activation of TP receptors, or the release of NO in the anesthetized rat.     

Hypoxic pulmonary vasoconstriction is modified by P-450 metabolites

20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P-450 4A (CYP4A) metabolite of arachidonic acid (AA) in human and rabbit lung microsomes and is a dilator of isolated human pulmonary arteries (PA). However, little is known regarding the contribution of P-450 metabolites to pulmonary vascular tone. We examined 1) the effect of two mechanistically distinct omega- and omega1-hydroxylase inhibitors on perfusion pressures in isolated rabbit lungs ventilated with normoxic or hypoxic gases, 2) changes in rabbit PA ring tone elicited by 20-HETE or omega- and omega1-hydroxylase inhibitors, and 3) expression of CYP4A protein in lung tissue. A modest increase in perfusion pressure (55 +/- 11% above normoxic conditions) was observed in isolated perfused lungs during ventilation with hypoxic gas (FI(O(2)) = 0.05). Inhibitors of 20-HETE synthesis, 17-oxydecanoic acid (17-ODYA) or N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS), increased baseline perfusion pressure above that of vehicle and amplified hypoxia-induced increases in perfusion pressures by 92 +/- 11% and 105 +/- 11% over baseline pressures, respectively. 20-HETE relaxed phenylephrine (PE)-constricted PA rings. Treatment with 17-ODYA enhanced PE-induced contraction of PA rings, consistent with inhibition of a product that promotes arterial relaxation, whereas 6-(20-propargyloxyphenyl)hexanoic acid (PPOH), an epoxygenase inhibitor, blunted contraction to PE. Conversion of AA into 20-HETE was blocked by 17-ODYA, DDMS, and hypoxia. CYP4A immunospecific protein confirms expression of CYP4A in male rabbit lung tissue. Our data suggest that endogenously produced 20-HETE could modify rabbit pulmonary vascular tone, particularly under hypoxic conditions.     

Enzymatic formation of angiotensins II and III in the hindlimb circulation of dogs

Experiments were performed in 14 anesthetized dogs to (1) to determine if the reductions in hindlimb blood flow produced by [des-Asp1] angiotensin I were due to its local enzymatic (kininase II) conversion to angiotensin III and (2) to quantitate the extent of conversion of angiotensin I to angiotensin II and of [des-Asp1] angiotensin I to angiotensin III in the hindlimb circulation. Graded doses of these peptides were administered as bolus injections directly into the left external iliac artery while measuring flow in this artery electromagnetically. Dose-response relationships were determined before and during the inhibition of kininase II activity with captopril or antagonism of angiotensin receptor sites with [Ile7] angiotensin III. Captopril inhibited the vasoconstrictor responses to angiotensin I and [des-Asp1] angiotensin I, but did not affect the responses to angiotensins II or III, or norepinephrine. [Ile7] angiotensin III inhibited the vasoconstrictor responses to all four angiotensin peptides but did not alter the responses to norepinephrine. These findings indicate that the hindlimb vasoconstrictor responses to [des-Asp1] angiotensin I were due to the local formation of angiotensin III. The extent of conversion of [des-Asp1] angiotensin I to angiotensin III that occurred in one transit through the hindlimb arterial circulation was estimated to be 36.7%, which was not different from the estimated 36.4% conversion of angiotensin I to angiotensin II. We conclude that angiotensin I and [des-Asp1] angiotensin I are converted to their respective vasoactive forms (angiotensins II and III) to a similar extent in the hindlimb circulation via the action of kininase II.     

Effect of angiotensin II on the apical K+ channel in the thick ascending limb of the rat kidney

We have used the patch-clamp technique to study the effect of angiotensin II (AII) on the activity of the apical 70 pS K+ channel and used Na(+)-sensitive fluorescent dye (SBFI) to investigate the effect of AII on intracellular Na+ concentration (Na+i) in the thick ascending limb (TAL) of the rat kidney. Addition of 50 pM AII reversibly reduced NPo, a product of channel open probability (Po) and channel number (N), to 40% of the control value and reduced the Na+i by 26%. The AII (50 pM)-induced decrease in channel activity defined by NPo was partially reversed by addition of 5 microM 17-octadecynoic acid (17-ODYA), an agent which blocks the cytochrome P450 monooxygenase. The notion that P450 metabolites of arachidonic acid (AA) may mediate the inhibitory effect of AII was further suggested by experiments in which addition of 10 nM of 20-hydroxyeicosatetraenoic acid (20-HETE) blocked the channel activity in cell-attached patches in the presence of 17-ODYA. We have used gas chromatography mass spectrometry (GC/MS) to measure the production of 20-HETE, a major AA metabolite of the P450-dependent pathway in the TAL of the rat. Addition of 50 pM AII increased the production of 20-HETE to 260% of the control value, indicating that 20- HETE may be involved in mediating the effect of AII (50 pM). In contrast to the inhibitory effect of 50 pM AII, addition of 50-100 nM AII increased the channel activity to 270% of the control value and elevated the Na+i by 45%. The effect of AII on the activity of the 70 pS K+ channel was also observed in the presence of 5 microM 17-ODYA and 5 microM calphostin C, an inhibitor of protein kinase C. However, addition of 100 microM NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase, abolished completely the AII (50- 100 nM)-induced increase in channel activity and addition of an exogenous nitric oxide (NO) donor, S-nitroso-N-acetyl-penicillamine (SNAP), increased channel activity in the presence of L-NAME. These data suggest that the stimulatory effect of AII is mediated by NO. We conclude that AII has dual effects on the activity of the apical 70 pS K+ channel. The inhibitory effect of AII is mediated by P450-dependent metabolites whereas the stimulatory effect may be mediated via NO.     

The reaction of vascular endothelial cells in old animals to an increased blood level of angiotensin II

The comparative morphological investigation of endothelium (E) of femoral arteries in old and young rabbits has revealed quantitative differences in the content of cells differing in their structure and function, and a four-fold increase in the number of cells having some signs of malfunction or injury in the E of old vs. young animals. These peculiarities, as well as different initial functional state of groups of cells, their location in the intima and degree of their compression during a 30 min vasoconstriction induced by angiotensin II (0.5 microgram/kg-1.min-1) predetermine a different degree of injury of endothelial cells. A phasic response of endothelial cells to angiotensin II administration, as well as slow restoration of permeability and autolysis of part of the injured endothelial cells in old animals are revealed. A peculiar pattern of the endothelial injury in old animals at a sharp increase of the blood pressure may account for the causes of the accelerated formation of fibrous-muscular thickenings and lipid strips in arteries of old rabbit at chronic hypertension.     

Vasomotor Reaction to Cyclooxygenase-1-Mediated Prostacyclin Synthesis in Carotid Arteries from Two-Kidney-One-Clip Hypertensive Mice

This study tested the hypothesis that in hypertensive arteries cyclooxygenase-1 (COX-1) remains as a major form, mediating prostacyclin (prostaglandin I₂; PGI₂) synthesis that may evoke a vasoconstrictor response in the presence of functional vasodilator PGI₂ (IP) receptors. Two-kidney-one-clip (2K1C) hypertension was induced in wild-type (WT) mice and/or those with COX-1 deficiency (COX-1^-/-). Carotid arteries were isolated for analyses 4 weeks after. Results showed that as in normotensive mice, the muscarinic receptor agonist ACh evoked a production of the PGI₂ metabolite 6-keto-PGF_1α and an endothelium-dependent vasoconstrictor response; both of them were abolished by COX-1 inhibition. At the same time, PGI₂, which evokes contraction of hypertensive vessels, caused relaxation after thromboxane-prostanoid (TP) receptor antagonism that abolished the contraction evoked by ACh. Antagonizing IP receptors enhanced the contraction to the COX substrate arachidonic acid (AA). Also, COX-1^-/- mice was noted to develop hypertension; however, their increase of blood pressure and/or heart mass was not to a level achieved with WT mice. In addition, we found that either the contraction in response to ACh or that evoked by AA was abolished in COX-1^-/- hypertensive mice. These results demonstrate that as in normotensive conditions, COX-1 is a major contributor of PGI₂ synthesis in 2K1C hypertensive carotid arteries, which leads to a vasoconstrictor response resulting from opposing dilator and vasoconstrictor activities of IP and TP receptors, respectively. Also, our data suggest that COX-1^-/- attenuates the development of 2K1C hypertension in mice, reflecting a net adverse role yielded from all COX-1-mediated activities under the pathological condition.