Nonselective Endothelin Research Articles

Vascular endothelial growth factor induced functional and morphologic signs of endothelial dysfunction in isolated arteries from normal pregnant women

Objective: The purpose of this study was to determine the effects of vascular endothelial growth factor on basal tone, endothelium-dependent dilatation, permeability, and morphologic features of endothelium in isolated arteries from normal pregnant women. We hypothesized that vascular endothelial growth factor might induce signs of endothelial dysfunction. Study Design: Arteries (approximately 200 μm) were dissected from subcutaneous fat biopsy specimens that were obtained at cesarean delivery and mounted on a pressure arteriograph. Changes in basal tone, dilatation to bradykinin (1 nmol/L to 3 μmol/L) before, during, and after 3 hours of incubation with vascular endothelial growth factor (0.5 or 1 nmol/L), vascular endothelial growth factor (0.5 nmol/L) plus bosentan (a nonselective endothelin receptor A and B antagonist, 1 μmol/L), or vehicle were compared. Scanning electron microscopy was applied for endothelial morphologic features. Permeability to Evans blue dye was evaluated in arteries after incubation with vascular endothelial growth factor, vascular endothelial growth factor plus angiopoietin-1, or vehicle, and in arteries that were obtained from women with preeclampsia. Results: Basal tone was higher after 60 minutes of incubation with vascular endothelial growth factor (0.5 nmol/L) compared with vehicle (29% ± 5% [n = 10] vs 10% ± 4% [n = 7], P = .006). Combination of vascular endothelial growth factor with bosentan failed to increase the tone (n = 4). Bradykinin-mediated dilatation was impaired in arteries that were incubated with vascular endothelial growth factor 0.5 nmol/L (max dilatation: 287% ± 16% vs 160% ± 23% [n = 10], P = .0001) or vascular endothelial growth factor 1 nmol/L (max dilatation: 207% ± 21% vs 88% ± 4% [n = 3], P = .003). Bradykinin-mediated dilatation was similar after incubation with vehicle (n = 7) or the combination of vascular endothelial growth factor plus bosentan (n = 4). Evans blue dye staining was higher after incubation with vascular endothelial growth factor but was reversed by the addition of angiopoietin-1. Scanning electron microscopy demonstrated the development of intercellular gaps. Conclusion: Vascular endothelial growth factor impaired bradykinin-mediated dilatation and enhanced basal tone and permeability. This might indicate a potential role for vascular endothelial growth factor in the development of endothelial dysfunction in pregnancy. Angiopoietin-1 inhibited the vascular endothelial growth factor–induced vascular leakage, which may have therapeutic implications in preeclampsia. (Am J Obstet Gynecol 2003;188:168-76.)

Effects of endothelin receptor antagonists on endothelin-1 and inducible nitric oxide synthase genes in a rat endotoxic shock model.

Levels of the endothelium-derived vasoconstrictor endothelin (ET)-1 and the vasodilator nitric oxide (NO) are markedly increased in endotoxic shock, although the pathophysiological role of ET-1 and its relation to NO under septic conditions remains obscure. To delineate the roles of ET-1 and the ET receptors, and the NO/inducible NO synthase (iNOS) system in endotoxic shock, we examined the gene expression of ET-1, ET receptors A and B (ETA and ETB) and iNOS in the heart and the liver of a rat endotoxic shock model, and we studied the effects of ET receptor antagonists on haemodynamics, survival rate and expression of ET-1, ET receptors and iNOS. Administration of bacterial lipopolysaccharide (LPS) into rats caused a profound hypotension with resultant death. However, these effects were blocked by a non-selective ETA/ETB receptor antagonist (TAK044), but not by an ETA-selective antagonist (BQ123). Injection of LPS caused a marked elevation in the plasma levels of both ET-1 and NO, which were not affected by treatment with either ET receptor antagonist. Administration of LPS caused increases in levels of ET-1, ETB and iNOS mRNA in the heart and the liver, whereas ETA mRNA expression was markedly downregulated in these organs. These results suggest that ET receptor subtype genes are differentially regulated in major organs from endotoxic shock rats, and that non-selective ET receptor antagonists ameliorate endotoxin-induced hypotension and death irrespective of iNOS-derived NO.

Long-term cold liver storage induces endothelin-1 release and a time-dependent increase in portal pressure at reperfusion in the rat.

Long-term cold storage and reperfusion lead to great liver injury involving major microcirculatory disturbance. This study investigated the effect of storage duration on endothelin-1 production, portal pressure, and microcirculation. Rat livers were perfused before and after 15 min to 48 h of cold storage in University of Wisconsin (UW) solution. Portal pressure was unchanged for up to 12 h of storage, and increased by 40% and 70% after 24 and 48 h, respectively. Vascular space was increased by a factor of 1.7 following 48 h of storage. Endothelin-1 was released by livers stored in UW solution for 48 h (3.99 +/- 1.85 pg/100 microl). The nonselective endothelin-1 receptor antagonist TAK-044 partially prevented the increase in portal pressure and prevented the increase in vascular space. Cold storage induces a time-dependent elevated portal pressure at reperfusion. The involvement of endothelin-1 in this process offers opportunities to improve liver graft quality by using endothelin-1 inhibitors.

Endothelin in human inflammatory bowel disease: comparison to rat trinitrobenzenesulphonic acid-induced colitis

There have been suggestions that endothelins (ET-1, ET-2, ET-3) are involved in the pathogenesis of human inflammatory bowel disease (IBD). Furthermore, the non-selective endothelin receptor antagonist, bosentan, ameliorates colonic inflammation in TNBS colitis in rats. However, no studies have measured the tissue expression and release of endothelins in human IBD in direct comparison to experimental TNBS colitis. Mucosal biopsies were obtained from 114 patients (42 Crohn's colitis, 35 ulcerative colitis and 37 normal) and compared to whole colonic segments from rats with TNBS colitis. ET-1/2 levels were reduced in human IBD but greatly increased in experimental TNBS colitis. RT–PCR indicated ET-2 was the predominant endothelin isoform in human IBD whereas ET-1 prevailed in the TNBS model. No associations were found between human IBD and tissue expression, content or release of ET-1/2. Our study shows, therefore, that unlike TNBS colitis in rats, in which ET-1/2 levels are greatly elevated and ET receptor antagonists are efficacious, there is no significant link between endothelins and human IBD.

Long-term effects of selective and nonselective endothelin receptor antagonists in mice with heart failure

Background: The ETA and ETB receptors mediate vasoconstriction, aldosterone release, and fibrosis. However, the role of ETB receptors is still controversial because those expressed on endothelial cells also stimulate vasodilatation and may oppose the actions of the ETA receptor. Plasma levels of endothelin-1 (ET-1) are increased in heart failure (HF) and are associated with myocardial dysfunction. The relative efficacy of selective and nonselective ET antagonists in the treatment of HF is unclear. We hypothesized that blockade of ETA receptors may improve cardiac function and prevent left ventricular remodeling in mice with HF, and these effects may be mediated in part by activation of ETB. Methods and Results: A mouse model of chronic HF induced by myocardial infarction (MI) was used. Seven days after MI, mice were divided into vehicle, ETA-ant, or ETA/B-ant groups and treated for 23 weeks. Cardiac function, LV dimensions, and hemodynamics were evaluated in conscious mice before MI and during treatment. Histologic analysis of the heart and liver was performed at the end of the study. HF significantly decreased EF and increased LV dimensions, interstitial collagen fraction (ICF) and myocyte cross-sectional area (MCSA). Both ETA-ant and ETA/B-ant slightly increased EF but had no significant effect on LV dimensions, hypertrophy, or ICF. Both treatments decreased MCSA; however, this was only significant in the ETA/B-ant group. Conclusions: Both selective and nonselective ET-ant have similar slight effects on cardiac function and remodeling. This suggests that (1) ETB receptors do not mediate the beneficial cardiac effects of ETA-ant and (2) blockade of the ET system alone may not provide significant cardioprotection, at least in mice with HF induced by MI.

Long-term effects of selective and nonselective endothelin receptor antagonists in mice with heart failure

Background: The ETA and ETB receptors mediate vasoconstriction, aldosterone release, and fibrosis. However, the role of ETB receptors is still controversial because those expressed on endothelial cells also stimulate vasodilatation and may oppose the actions of the ETA receptor. Plasma levels of endothelin-1 (ET-1) are increased in heart failure (HF) and are associated with myocardial dysfunction. The relative efficacy of selective and nonselective ET antagonists in the treatment of HF is unclear. We hypothesized that blockade of ETA receptors may improve cardiac function and prevent left ventricular remodeling in mice with HF, and these effects may be mediated in part by activation of ETB. Methods and Results: A mouse model of chronic HF induced by myocardial infarction (MI) was used. Seven days after MI, mice were divided into vehicle, ETA-ant, or ETA/B-ant groups and treated for 23 weeks. Cardiac function, LV dimensions, and hemodynamics were evaluated in conscious mice before MI and during treatment. Histologic analysis of the heart and liver was performed at the end of the study. HF significantly decreased EF and increased LV dimensions, interstitial collagen fraction (ICF) and myocyte cross-sectional area (MCSA). Both ETA-ant and ETA/B-ant slightly increased EF but had no significant effect on LV dimensions, hypertrophy, or ICF. Both treatments decreased MCSA; however, this was only significant in the ETA/B-ant group. Conclusions: Both selective and nonselective ET-ant have similar slight effects on cardiac function and remodeling. This suggests that (1) ETB receptors do not mediate the beneficial cardiac effects of ETA-ant and (2) blockade of the ET system alone may not provide significant cardioprotection, at least in mice with HF induced by MI.

Pulmonary hypertension secondary to left ventricular dysfunction: the role of nitric oxide and endothelin-1 in the control of pulmonary vascular tone.

1. Using an in vivo model of pulmonary hypertension (PHT) secondary to left ventricular dysfunction (LVD), the pulmonary arterial response to the nitric oxide synthase (NOS) blocker L-NAME (30 micromol.min(-1) i.v.) and the subsequent responses to cumulatively administered endothelin-1 (ET-1) (0.001 -- 4 nmol.kg(-1) i.v.) or big ET-1 (0.1 -- 2.0 nmol.kg(-1) i.v.) were studied. Additionally, the effect of the non-selective ET-1 receptor antagonist, SB209670, was investigated. 2. Eight weeks after coronary artery ligation or sham operation, rabbits demonstrated increased mean pulmonary arterial pressure (PAP) accompanied by right ventricular hypertrophy. 3. Blockade of NOS caused a greater increase in basal PAP (increased by 7.7 +/- 1.1 mmHg c.f. 3.8 +/- 1.0 mmHg in controls, P<0.05) and uncovered a greater pulmonary pressor response to exogenous ET-1 in rabbits with PHT (increased by 10.2 +/- 2.3 mmHg c.f. 4.9 +/- 1.0 mmHg in controls, P<0.05). 4. Big ET-1 evoked a pulmonary pressor effect, in both groups of rabbits, that was increased following blockade of NOS and was more potent in rabbits with PHT. 5. The non-selective ET-1 receptor antagonist, SB209670, reduced basal PAP (from 16.9 mmHg to 15.9 mmHg, P < 0.05) in rabbits with PHT and blocked the response to ET-1 in the presence of L-NAME. 6. In conclusion, the results demonstrate that basal NO activity masks a pulmonary pressor response to exogenously administered ET-1. An increased responsiveness to ET-1 was shown in the pulmonary arterial bed of rabbits with PHT secondary to LVD, implicating a pathophysiological role for ET-1 in this model.

Endothelin-1 and endothelin receptor antagonists as potential cardiovascular therapeutic agents.

Endothelin (ET)-1 is an endothelium-derived peptide with potent vasoconstrictor and proliferative properties. The ET system is activated in several cardiovascular disease states associated with functional and structural vascular changes, including hypertension and heart failure. The two ET receptor subtypes are known as ET(A)R and ET(B)R. The former is located mainly on vascular smooth muscle cells and is responsible for mediating vasoconstriction and proliferation. The latter is present predominantly on endothelial cells and mediates vasorelaxation as well as ET-1 clearance. Activation of smooth muscle ET(B)R causes vasoconstriction. Selective ET(A)R antagonists as well as nonselective ET(A)R-ET(B)R antagonists have been developed. Studies with animal models and early-phase clinical trials provided strong evidence that these agents are effective in the treatment of heart failure, essential hypertension, pulmonary hypertension, and atherosclerosis. However, the complexity of biologic effects mediated by two different receptor subtypes complicates therapy with selective versus nonselective ET receptor antagonists. In addition to subtype selectivity and potency, changes in receptor subtype distribution under different pathologic conditions and different patient populations will play a crucial role in the evaluation of these potentially therapeutic drugs.

Pretreatment of Donors with Endothelin Receptor Antagonist TAK-044 Improves Cardiac Functional Recovery Following Preservation with University of Wisconsin Solution

Objective : Previous studies suggest that endothelin-1 (ET-1) plays a role in myocardial ischemia/reperfusion injury. Although administration of an endothelin receptor antagonist to the recipient has been shown to improve myocardial function after ischemia/reperfusion in a rat heart transplantation model, the effect of administering an endothelin receptor antagonist to the donor has not yet been examined. This study was designed to investigate the effects of pretreating donors with an ET A /ET B endothelin receptor antagonist (TAK-044) on myocardial function after cold preservation of a rat heart. Design : Male rats were pretreated with normal saline (control group, n = 8), TAK-044 (TAK group, n = 8, 1 mg/kg). Following cardiac arrest using cardioplegia, we washed out the coronary vascular beds with cold University of Wisconsin solution followed by 6-h preservation. After preservation, the hearts were mounted on a Langendorff apparatus to estimate aortic flow (AF), coronary flow (CF), cardiac output (CO), systolic pressure (SP), heart rate (HR), and rate-pressure product (RPP: HR 2 SP). The concentration of lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) within the coronary perfusate during reperfusion was measured. Results : AF, SP, and CO were significantly greater in the TAK group than in the control group ( p = 0.0045, 0.004, and 0.0295, respectively). Conclusion : Pretreatment of donors with a nonselective endothelin receptor antagonist (TAK-044) improved cardiac functional recovery following preservation and may be beneficial for prolonged myocardial preservation.

Role of endogenous endothelin on coronary reflow after cardioplegic arrest

Objective: Endothelin plays a role in the regulation of basal coronary tone. We hypothesized that low coronary reflow and reduced cardiac function after prolonged ischemia may be due to increased release of endogenous endothelin. Methods: Using an isolated perfused rat heart, we examined the effect of the addition of various endothelin antagonists during reperfusion after 4 hours of cardioplegic arrest at 4°C. Hearts were freeze-clamped at the end of reperfusion for analysis of high-energy phosphate levels. Results are expressed as the percentages of preischemic values. Results: The addition of bosentan or Ro61-0612 (nonselective endothelin antagonists) resulted in a significant increase in the recovery of coronary flow after 30 minutes of reperfusion (100.9% vs 85.3% [P =.03] and 122.4% vs 83.7% [P <.001], respectively, versus controls). The addition of PD155080 (endothelin A antagonist) had a similar effect (129.5% vs 91.4%, P =.008). BQ788 (endothelin B antagonist) and phosphoramidon (endothelin-converting enzyme inhibitor) had no effect. Myocardial adenosine triphosphate levels were significantly (12.1%) higher after reperfusion with Ro61-0612 (18.1 ± 0.4 μmol/g vs 16.2 ± 0.5 μmol/g, P =.01). There was no difference in the recovery of cardiac mechanical function with any of the antagonists studied. Conclusion: These results suggest that endogenous endothelin plays a role in low coronary reflow after prolonged cardioplegic arrest but does not impair recovery of myocardial function.

Role of endothelin ETB receptor activation in angiotensin II-induced hypertension: effects of salt intake.

We showed recently that endothelin (ET)A receptors are involved in the salt sensitivity of ANG II-induced hypertension. The objective of this current study was to characterize the role of endothelin ETB receptor activation in the same model. Male rats on fixed normal (2 meq/day) or high (6 meq/day) salt intake received a continuous intravenous infusion of ANG II or salt only for 15 days. During the middle 5 days of the infusion period, rats were given either the selective ETB receptor antagonist A-192621 or the nonselective endothelin receptor antagonist A-182086 (both at 24 mg x kg(-1) x day(-1) intra-arterially). Infusion of ANG II caused a greater rise in arterial pressure in rats on high-salt intake. The administration of A-192621 increased arterial pressure further in all rats. The chronic hypertensive effect of A-192621 was not significantly affected by salt intake or ANG II. The administration of A-182086 lowered arterial pressure chronically only in rats on normal salt intake receiving ANG II. Thus the salt sensitivity of ANG II-induced hypertension is not caused by changes in ETB receptor function.

Blockade of Endothelin Receptors Reduces Diet-Induced Hypercholesterolemia and Atherosclerosis in Apolipoprotein E-Deficient Mice

Objective: Endothelin (ET)-1 has proatherogenic properties, since ET receptor antagonists reduce atherosclerotic lesions in animals. However, we recently demonstrated that ET-1 and ET_B receptors are increased in atherosclerotic lesions. To further examine the effects of ET_B receptor antagonism on atherogenesis, we investigated the chronic effects of the nonselective ET_A/ET_B receptor antagonist SB209670 on the development of atherosclerosis in apolipoprotein E (ApoE)-deficient mice. Methods: Ninety-four male mice (10 weeks of age) were randomly divided into four groups: mice fed a Western-type diet or a chow diet with SB209670 treatment (10 mg/kg/day) or placebo for 12 weeks. Results: In mice fed the Western-type diet, but not in mice fed the chow diet, treatment with SB209670 significantly attenuated the increase in plasma total cholesterol, predominantly in the very-low-density lipoprotein and intermediate-density lipoprotein fractions, without altering the plasma triglyceride level. Furthermore, treatment with SB209670 significantly reduced the extent of aortic atherosclerosis, by 53% in mice fed the Western-type diet and by 38% in mice fed the chow diet. Histological analysis revealed that SB209670 prevented the formation of atheromatous plaque lesions by inhibiting the fibroproliferative process. Conclusion: We found that chronic administration of SB209670 reduced diet-induced hypercholesterolemia and atherosclerosis in ApoE-deficient mice. Thus, nonselective ET receptor antagonists may have a therapeutic potential in the treatment of human atherosclerotic disease.

Nonselective endothelin receptor antagonist initiated soon after the onset of myocardial infarction may deteriorate 24-hour survival.

To investigate the effects of endothelin blockade initiated immediately after the onset of myocardial infarction on survival and left ventricular remodeling, treatment with the nonselective receptor antagonist TAK-044 (n = 22) or saline (n = 19) for 3 weeks was initiated immediately after coronary ligation in rats. The 24-h survival rate was significantly lower in the TAK-044 group than in the saline group. The systolic blood pressure 24 h after the onset of myocardial infarction was similar in the saline and TAK-044 groups, although it was significantly lower in the TAK-044 group during the 3-week protocol. Heart weight/tibial length was significantly increased in the TAK-044 group compared with the saline group. As all deaths in the TAK-044 group occurred within 24 h after myocardial infarction, we performed additional experiments using a separate group of rats 12-16 h after myocardial infarction. Plasma and myocardial endothelin-1 levels were significantly increased, and a bolus injection of TAK-044 significantly reduced left ventricular dP/dtmax in these rats that had had a myocardial infarction compared with sham-operated rats. Endothelin receptor blockade initiated immediately after the onset of myocardial infarction may deteriorate acute-phase survival and left ventricular remodeling. Inhibition of the positive inotropic action of endothlin-1 may partially explain the increased 24-h mortality.

Insulin-induced biphasic responses in rat mesenteric vascular bed: role of endothelin.

The vasodilatory capacity of insulin has been widely reported, yet some investigators have not noted this effect. Because insulin has been shown to enhance endothelin release, we speculated that endothelin could be attenuating insulin-evoked vasodilation. We examined the effect of ex vivo insulin perfusion on vascular resistance by using the Sprague-Dawley rat mesenteric vascular bed. In methoxamine-preconstricted preparations, insulin (3.0 pmol/L to 10 nmol/L) evoked a concentration-dependent decrease in perfusion pressure (PP) with a maximal response of 42.0+/-9.2%, whereas continuous exposure to 10 nmol/L insulin induced a 51.8+/-3.5% relaxation. Further exposure to 10 nmol/L insulin resulted in the generation of endothelin and a subsequent loss of the vasodilatory response. Indomethacin had no effect on vascular responses. The vasodilatory response was significantly inhibited by nitric oxide synthase inhibition (20.5+/-4.2%; P<0.01) and calcium-activated potassium channel blockade (28.5+/-3.7%; P<0.05). Endothelial denudation attenuated the vasodilatory component (20.3+/-7.1%; P<0.01) and altered the biphasic pattern of the response. The decline in insulin-evoked vasodilation was significantly prevented by an endothelin-A antagonist (BQ123), an endothelin-B antagonist (BQ788), and nonselective endothelin blockade with both BQ123 and BQ788. These results demonstrate that the endothelium is intimately involved in regulating the vascular response to insulin. Insulin promotes the release of nitric oxide and endothelium-derived hyperpolarizing factor. During sustained exposure to higher concentrations, this vasodilatory effect is countered by the pathological generation of endothelin. Endothelin receptor blockade facilitates the maintenance of vasodilation despite high insulin concentrations.

Nitric oxide and endothelin relationship in intestinal ischemia/reperfusion injury (II)

Endothelins ( ETs ) are potent vasoconstrictors derived from vascular endothelium. They have primary roles in many pathophysiologic states including ischemia/reperfusion (I/R) injury. The relationships between nitric oxide (NO) and ETs are still under investigation. In this study on rats we want to focus on the interaction of NO and ET especially in I/R injury. For this purpose ET-1 and PD-156252, a nonselective ET receptor blocker, were given in a mesenteric I/R model and reactive oxygen species were detected directly using chemiluminescence of the ileal tissue. ET administrations to sham and I/R groups caused significant increases in NO concentrations whereas, in terms of peroxynitrite, which is a highly reactive group of free radicals, its increasing effects were seen only in I/R groups. This suggests that in I/R where superoxide levels increase together with NO, the conversion to peroxynitrite is likely and this effect is augmented with ET administration. On the other hand PD administration decreases superoxide and thereby peroxynitrite levels and this study shows that the effect of PD-156252 is established through this mode of action. These data suggest therapeutic approaches that may be beneficial in the treatment of I/R injury.

Important Role for Endothelins in Acute Hepatic Ischemia/Reperfusion Injury

The aim of this study was to explore the complex role of endothelins (ETs) in hepatic ischemia-reperfusion injury and to minimize this type of injury by nonselective ET receptor blockade. In an in vivo rat model, hepatic ischemia was induced for 30 min. The rats were divided into three groups: (1) sham operated, (2) untreated ischemic, and (3) group treated with the nonselective ET receptor antagonist bosentan (1 mg/kg body weight iv). Blockage of the ET system during ischemia-reperfusion was assessed by: (1) in vivo microscopic analysis, (2) measurement of local tissue pO2

Endothelin Modulation of Choroidal Blood Flow in the Rabbit

Based on the previous finding that locally produced nitric oxide (NO) and endothelin (ET) exert competing effects on choroidal resistance vessels, the present study sought to further characterize the pharmacology of ET in the choroid. The specific goal was to quantify the choroidal blood flow responses to acute changes in perfusion pressure before and after administering endothelin 1 (ET1), a non-selective ET antagonist, and selective antagonists for the endothelin A (ETA) and endothelin B (ETB) receptor subtypes. Anesthetized rabbits were instrumented with an ear artery cannula to measure mean arterial pressure (MAP), occluders on the aorta and vena cava to control MAP, and a vitreous cannula to measure intraocular pressure (IOP). Choroidal blood flow was measured by laser Doppler flowmetry with a vitreous fiber optic probe. The protocol entailed changing the ocular perfusion pressure by varying MAP before and after ET1 (0.9μ gkg−1, i.v.,n=14), non-selective ET blockade (A-182086, 3mgkg−1, i.v., n=10), selective ETA blockade (FR-139317, 3mgkg−1, i.v., n=12), and selective ETB blockade (A-192621, 3mgkg−1, i.v., n=14). ET1 and ETB blockade shifted the choroidal pressure-flow relation downward, while the non-selective antagonist and the selective ETA antagonist had no effect. The choroid had a biphasic response to exogenous ET1 as seen in other tissues (i.e. initial brief dilation followed by prolonged constriction) that was blocked by the non-selective antagonist whereas the ETA antagonist enhanced the dilation and blocked the constriction, and the ETB antagonist blocked the dilation and enhanced the constriction. These results indicate that ETA and ETB receptors are present and mediate opposing effects on choroidal vascular resistance. The results also suggest that endogenous ET preferentially elicits ETB vasodilation, most likely by stimulating endothelial nitric oxide release.

In vivo subcellular elemental dynamics in liver graft: With special reference to effect of non-selective endothelin receptor antagonist, TAK-044, on the graft injury

Background: No data are available concerning the in vivo subcellular dynamics of elements in liver grafts and the effect of endothelin receptor antagonist, TAK-044, against graft injury. Methods: Liver transplantation was performed in porcine under active veno-venous bypass. The grafts stored in chilled preservation solution were recirculated following reflush with lactated Ringer's solution with or without TAK-044 (10 mg/kg). Cold and warm ischemic times of the grafts were comparable between the two groups. Elements (Na, K, Cl, Ca, P and S) were measured in three fractions of cytoplasm, mitochondria and nucleus by electron probe X-ray microanalysis for the graft biopsy specimens obtained at various time from donor laparotomy to 1 week after liver grafting. Liver functions also were compared between the two groups. Results: In both groups, concentration of each element changed in parallel among the three subcellular fractions and their changes were less marked in the nucleus. In the control group, there were significant increases in cytoplasmic Na and Cl after portal reperfusion and in cytoplasmic and mitochondrial Ca after hepatic artery reperfusion. These were accompanied by K and mitochondrial S decreases without a statistical significance. In the TAK group, such postreperfusion elemental alterations were significantly suppressed and early deterioration of the liver functions was alleviated, as compared with the control group. Conclusion: A supplemental use of TAK-044 in a rinse solution before reflush contributed to stability of subcellular elements after reperfusion and better preservation of early graft function.

Blockade of endothelin receptors markedly reduces atherosclerosis in LDL receptor deficient mice: role of endothelin in macrophage foam cell formation.

We evaluated the direct effects of long-term blockade of ET(A) and ET(B) receptors using a mixed endothelin (ET) receptor antagonist, LU224332, in the low density lipoprotein receptor (LDL-R) knockout mouse model of atherosclerosis. Four groups of LDL-R deficient mice were studied: control mice fed normal chow (group I); mice fed a high cholesterol (HC, 1.25%) diet alone (group II), HC fed animals treated with LU224332 (group III); and mice fed normal chow treated with the LU compound (group IV). All treatments were continued for 8 weeks at which time the animals were sacrificed and the aortae were removed and stained with oil red O. Atherosclerotic area (AA) was determined by quantitative morphometry and normalized relative to total aortic area (TA). Cholesterol feeding resulted in a marked increased in total plasma cholesterol ( approximately 15 fold) and widespread aortic atherosclerosis (AA/TA: group I: 0.013+/-0.007; group II: 0.33+/-0. 11; P<0.001). Atherosclerotic lesions were characterized by immunohistochemistry as consisting mainly of macrophages which also showed high levels of ET-1 expression. Treatment with ET antagonist significantly reduced the development of atherosclerosis (AA/TA: group III: 0.19+/-0.07, P<0.01 vs. group II), without altering plasma cholesterol levels and blood pressure. The direct effect of LU224332 on macrophage activation and foam-cell formation was determined in vitro using a human macrophage cell line, THP-1. Treatment of the THP-1 cells with LU224332 significantly reduced cholesterol ester and triacylglycerol accumulation and foam-cell formation on exposure to oxidized LDL (P<0.01 and P<0.05, respectively). We conclude that a nonselective ET receptor antagonist substantially inhibited the development of atherosclerosis in a genetic model of hyperlipidemia, possibly by inhibiting macrophage foam-cell formation, suggesting a role for these agents in the treatment and prevention of atherosclerotic vascular disease.

Endothelin receptor blockade decreases lung water in young rats exposed to viral infection and hypoxia.

Viral respiratory infections may increase the susceptibility of young animals to hypoxia-induced pulmonary edema. Because hypoxia stimulates endothelin production, we hypothesized that an increase in lung endothelin contributes to these alterations in lung water. Weanling rats were infected with Sendai virus, causing a mild respiratory infection. At day 7 after infection, animals were exposed to hypoxia (inspired O(2) fraction = 0.1) for 24 h. Exposure to virus plus hypoxia led to increases in lung water compared with control groups (P < 0.001). Lung endothelin levels were significantly higher in the virus plus hypoxia group than in control groups (P < 0.001). A second group of infected animals received bosentan, a nonselective endothelin receptor antagonist, during exposure to hypoxia. Bosentan-treated animals showed less lung water accumulation, less lung lavage fluid protein, and less perivascular fluid cuffing than untreated animals (P < 0.01). We conclude that the combination of a recent viral respiratory infection and exposure to moderate hypoxia led to increases in endothelin in the lungs of young rats and that endothelin receptor blockade ameliorates the hypoxia-induced increases in lung water found in these animals.