Endothelin-1 (ET-1) and vein graft failure and the therapeutic potential of ET-1 receptor antagonists

Despite the exploration of a large number of disparate drugs in animal models and clinical trials, no pharmacological intervention, with the exception of aggressive lipid lowering therapy has reduced late vein graft failure in man. The importance of devising more effective strategies is exemplified by the enormous economic consequences of vein graft failure. Worldwide, there are currently more than 1,000,000 coronary artery bypass graft surgery (CABG) operations a year, the same number of patients undergoing infrainguinal bypass for vascular diseases of the lower limb. The pathophysiology of vein graft failure is complex, involving disparate factors that include adhesion of platelets and leukocytes, rheological forces, metalloproteinase expression, proliferation and migration of vascular smooth muscle cells, neointima formation, oxidative stress, hypoxia and neural re-organisation. Although this diverse etiology may seem to preclude any single drug type as being effective in mediating vein graft failure: one factor that is involved in every facet of vein graft pathobiology is endothelin-1 (ET-1). As such a single drug type (ET_A antagonist) may prove to be the magic bullet in this scenario. Thus, in this review, we will consider the etiology of vein graft disease in relation to ET-1 and will then present an argument (with evidence) that specific ET_A receptor antagonists constitute a potentially effective means of preventing vein graft failure.     

ET-1 actions in the kidney: evidence for sex differences

Hypertension and chronic kidney disease are more common in men than in premenopausal women at the same age. In animal models, females are relatively protected against genetic or pharmacological procedures that produce high blood pressure and renal injury. Overactivation or dysfunction of the endothelin (ET) system modulates the progression of hypertension or kidney diseases with the ET_A receptor primarily mediating vasoconstriction, injury and anti-natriuresis, and ET_B receptors having opposite effects. The purpose of this review is to examine the role of the ET system in the kidney with a focus on the inequality between the sexes associated with the susceptibility to and progression of hypertension and kidney diseases. In most animal models, males have higher renal ET-1 mRNA expression, greater ET_A-mediated responses, including renal medullary vasoconstriction, and increased renal injury. These differences are reduced following gonadectomy suggesting a role for sex hormones, mainly testosterone. In contrast, females are relatively protected from high blood pressure and kidney damage via increased ET_B versus ET_A receptor function. Furthermore, ET_A receptors may have a favourable effect on sodium excretion and reducing renal damage in females. In human studies, the genetic polymorphisms of the ET system are more associated with hypertension and renal injury in women. However, the knowledge of sex differences in the efficacy or adverse events of ET_A antagonists in the treatment of hypertension and kidney disease is poorly described. Increased understanding how the ET system acts differently in the kidneys between sexes, especially with regard to receptor subtype function, could lead to better treatments for hypertension and renal disease.

LINKED ARTICLES

This article is part of a themed section on Endothelin. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2013.168.issue-1     

Pathophysiology of saphenous vein graft failure: a brief overview of interventions

Coronary artery bypass graft surgery (CABG) is widely used for the treatment of atheromatous stenosis of coronary arteries. However, as many as 50% of grafts fail within 10 years after CABG due to neointima (NI) formation, a process involving the proliferation of vascular smooth muscle cells (VSMCs) and superimposed atherogenesis. To date no therapeutic intervention has proved successful in treating late vein graft failure. However, several diverse approaches aimed at preventing neointimal formation have been devised which have yielded promising results. In this review, therefore, we will summarise the pathophysiology of vein graft disease and then briefly consider interventional approaches to prevent late vein graft failure which include surgical technique, conventional pharmacology, external sheaths, cytostatic drugs and gene transfer.     

Animal models for studying vein graft failure and therapeutic interventions

Vein grafts have been extensively used to bypass blockages in arteries, but are themselves subject to early closure by thrombosis or later obstruction by vein graft disease (neointimal hyperplasia and remodelling). Animal models are a crucial means of testing potential therapeutic and surgical interventions to prevent graft stenosis and occlusion. This review outlines many of the animal models of vein grafting. Recent studies include targeted gene therapy to prevent acute vein graft thrombosis and the use of folic acid to limit graft failure in diabetic pigs.     

Statins and vein graft failure in coronary bypass surgery

Saphenous vein grafts used in coronary artery bypass graft surgery suffer from lower patency rates compared to left internal mammary artery. A number of clinical trials and observational studies have demonstrated a significant benefit of statin treatment on vein graft patency. Aside from their well-known lipid-lowering capacities, statins exert pleiotropic effects by direct inhibition of the mevalonate pathway in the wall of these grafts. This leads to reduced geranylgeranylation of small GTPases such as Rho and Rac. Through this LDL-independent mechanism, statins improve endothelial function and reduce vascular inflammation and oxidative stress, inhibiting also smooth muscle cell proliferation and migration. Although the existing evidence supports a beneficial effect of statins on vein grafts biology, more clinical trials focused on the effect of aggressive statin treatment on vein graft patency are required, in order to safely translate this strategy into clinical practice.     

Perivascular administration of drugs and genes as a means of reducing vein graft failure

Coronary artery bypass surgery is a highly effective and durable therapy of coronary artery disease. Together with internal mammary arteries the saphenous vein grafts are the most important conduits for coronary surgery. We reviewed the topic of local pharmacologic and gene therapeutic treatment approaches to prevent neointimal hyperplasia in vein grafts. Perivascular therapy of veins before arterialization would be a simple approach that avoids systemic side effects of medications. The current data available show that there are promising experimental approaches (in vitro models, animal in vivo models) for pharmacological and gene therapeutic treatment of vein graft failure.     

Erythropoietin production in healthy volunteers subjected to controlled hypobaric hypoxia: further evidence against a role for adenosine

Aims Objective of this study was to investigate whether adenosine modulates renal erythropoietin production.
Methods In the present study erythropoietin production was stimulated by hypobaric hypoxia by subjecting healthy volunteers to a simulated altitude of 4000  m in a low pressure chamber for 5.5  h. During exposure to hypoxia the subjects received i.v. in a randomized, single-blind, cross-over fashion the non-specific adenosine antagonist theophylline, the adenosine reuptake inhibitor dipyridamole and placebo.
Results Contrary to the working hypothesis, theophylline did not decrease and dipyridamole did not further boost erythropoietin concentrations.
Conclusions The results are in agreement with our earlier study using haemorrhage as a controlled physiological stimulus of erythropoietin production, and would question a major role for adenosine as a mediator of renal erythropoietin production.     

Oxidative stress and vein graft failure: a focus on NADH oxidase, nitric oxide and eicosanoids

Recent interest has focused on superoxide and the upregulation of NADPH oxidase expression in the aetiology of vein graft failure. Implantation of saphenous vein grafts promotes upregulation of NADPH oxidase through a number of distinct interrelated mechanisms: (a) endothelial denudation, (b) factors released by adherent platelets, monocytes and neutrophils, (c) hypoxia and (d) altered prostacyclin (PGI(2)) and enhanced isoprostane formation. These, in turn, impact on neointima (NI) formation (vascular smooth muscle cell [VSMC] replication and migration) and metalloproteinase (MMP) expression, key events in vein graft thickening. NADPH oxidase in the aetiology of vein graft failure will be discussed in this review with particular reference to nitric oxide and eicosanoids and related drugs that inhibit its activity and expression.     

The role of established and emerging risk factors in peripheral vascular graft occlusion

Several studies have evaluated the association between established as well as emerging vascular risk factors with peripheral graft occlusion. There is evidence for a link between the risk for graft occlusion and total serum cholesterol, low-density lipoprotein cholesterol and triglyceride levels. The overall effect of smoking shows a 2.35-fold increase in risk of graft failure. Studies involving diabetic patients undergoing peripheral bypass may have failed to detect higher occlusion rates, possibly due to increased morbidity and mortality as well as higher amputation rates even if the graft is patent. Both antiplatelet agents and anticoagulation seem to be effective in the prevention of graft occlusion. Unconvincing data have been published with regards to the effect of hypertension on graft patency. Emerging factors such as fibrinogen, lipoprotein (a), C-reactive protein and homocysteine levels should also be considered when assessing the risk of graft occlusion. More research is needed to prevent graft occlusion due to the obvious clinical relevance, quality of life and cost issues.     

The role of established and emerging risk factors in peripheral vascular graft occlusion

Several studies have evaluated the association between established as well as emerging vascular risk factors with peripheral graft occlusion. There is evidence for a link between the risk for graft occlusion and total serum cholesterol, low-density lipoprotein cholesterol and triglyceride levels. The overall effect of smoking shows a 2.35-fold increase in risk of graft failure. Studies involving diabetic patients undergoing peripheral bypass may have failed to detect higher occlusion rates, possibly due to increased morbidity and mortality as well as higher amputation rates even if the graft is patent. Both antiplatelet agents and anticoagulation seem to be effective in the prevention of graft occlusion. Unconvincing data have been published with regards to the effect of hypertension on graft patency. Emerging factors such as fibrinogen, lipoprotein (a), C-reactive protein and homocysteine levels should also be considered when assessing the risk of graft occlusion. More research is needed to prevent graft occlusion due to the obvious clinical relevance, quality of life and cost issues.     

DSP4 and Herrnstein's equation: further evidence for a role of noradrenaline in the maintenance of operant behaviour by positive reinforcement

The effect of the selective noradrenaline neurotoxin DSP4 on steady-state operant behaviour was examined using a quantitative behavioural paradigm based on Herrnstein's (1970) equation, which defines a hyperbolic relationship between steady-state response rate and reinforcement frequency in variable-interval schedules. Eleven rats received injections of DSP4 (two doses of 50 mg/kg, intraperitoneally), and 12 rats received injections of the vehicle alone. The rats were trained to steady state in a series of six variable-interval schedules of sucrose reinforcement, affording scheduled reinforcement frequencies of 4–360 reinforcers per hour. Herrnstein's equation was fitted to the data obtained from each rat and to the averaged data obtained from the two groups. The value ofK _H (the parameter expressing the reinforcement frequency needed to maintain the half-maximal response rate) was higher in the DSP4-treated rats than in the control rats; the value ofR _max (the parameter expressing the maximum response rate) did not differ significantly between the two groups. At the end of the behavioural experiment the rats were sacrificed for determination of the concentrations of catecholamines in the brain by high-performance liquid chromatography. The levels of noradrenaline in the parietal cortex, hippocampus and cerebellum of the DSP4-treated rats were less than 20% of those of the control rats. The results provide further evidence that central noradrenergic neurones are involved in the maintenance of operant behaviour by positive reinforcement.     

Lack of evidence for a role of serotonin in interleukin-1-induced hypophagia

Interleukin-1 (IL-1) administration depresses food intake in rodents. IL-1 is known to increase the metabolism of serotonin, which is known to affect feeding behavior. Thus, serotonin is an obvious candidate for a mediator of the hypophagic response to IL-1. Therefore, we tested the ability of serotonergic agonists and antagonists to alter the hypophagic responses to IL-1 and bacterial lipopolysaccharide (LPS). Hypophagia was assessed in ad lib-fed mice by recording the intake of sweetened milk in a 30-min period. Acute intraperitoneal administration of mouse IL-1beta reliably decreased milk intake. This hypophagic response was not affected by any of the serotonin antagonists tested, including 5-HT(1A) (WAY100135 and propranolol), 5-HT(1B) (GR127935), 5-HT(2) (ritanserin, ketanserin, SB206553, and RS102221), mixed 5-HT(1/2) (methysergide and metergoline), and 5-HT(3) (tropisetron) receptor antagonists. The 5-HT(1A) agonists (8-OH-DPAT and ipsapirone) and a 5-HT(1B) agonist (CGS12066B) known to decrease the activity of serotonergic neurons, also had no effect. Mice pretreated with 5,7-dihydroxytryptamine to deplete brain serotonin ate less, but, nevertheless, displayed similar hypophagic responses to mIL-1beta or LPS. The results suggest that serotonin is not involved in the decrease in short-term milk intake induced by mIL-1beta or LPS in mice that have been fed ad lib.     

Characterization of bovine phenol sulfotransferases: evidence of a major role for SULT1B1 in the liver

Abstract

1.?Cattle are an important component of the human food chain. Drugs used either legally or illegally in cattle may therefore enter the food chain and it is thus important to understand pathways of drug metabolism in this species, including sulfation catalyzed by the sulfotransferases (SULTs).

2.?In this study, we have analyzed the sulfation of 4-nitrophenol and other compounds in male and female bovine liver and characterized recombinant bovine SULT isoforms 1A1 and 1B1 expressed in Escherichia coli.

3.?We found that, in contrast to most other mammalian species, the major phenol sulfotransferase SULT1A1 is not expressed in bovine liver. Rather SULT1B1 seems to be a major form in both male and female bovine liver.

4.?We also identified kinetic differences between bovine and human SULT1A1 and, using the human SULT1A1 crystal structure, identified two amino acid positions in the active site of bovine SULT1A1 (Ile₈₉Val and Phe₂₄₇Val) that may be responsible for these differences.     

Agonist activity of sumatriptan and metergoline at the human 5-HT1D beta receptor: further evidence for a role of the 5-HT1D receptor in the action of sumatriptan.

We have recently cloned a novel human 5-HT1D receptor subtype termed 5-HT1D beta. CHO K1 cells expressing the human serotonin 5-HT1D beta receptor were assayed to determine the second messenger system of this receptor. Cyclic AMP radioimmunoassays revealed that the 5-HT1D beta receptor is negatively coupled to adenylate cyclase in this cell system. A maximum of 50% inhibition of forskolin stimulated cAMP production was obtained with 5-HT1 receptor agonists which was blocked by the non-selective 5-HT receptor antagonist methiothepin (pKB = 100 nM). The novel anti-migraine drug sumatriptan, a putative 5-HT1D selective compound, acted as an agonist at the 5-HT1D beta receptor. Most notably metergoline, a putative 5-HT1 receptor antagonist, did not block the effects of 5-HT and was found to be acting as a full agonist at the 5-HT1D beta receptor. The ability of metergoline to act as an agonist at the 5-HT1D beta receptor may explain why it does not inhibit 5-HT and sumatriptan induced contraction of dog saphenous vein and other large conducting arteries. These results suggest that the 5-HT1D beta receptor may be the site of action of sumatriptan in preventing migraine, and that metergoline's actions on the dog saphenous vein are not contradictory to that hypothesis, as previously reported.     

Yohimbine interacts with the dopaminergic system to reverse sexual satiation: further evidence for a role of sexual motivation in sexual exhaustion.

The possible interaction of yohimbine with the dopaminergic system in the mediation of sexual behaviour expression in sexually exhausted male rats was investigated. The behavioural effects of the simultaneous injection of yohimbine (500 microg/kg) plus apomorphine (50 microg/kg) and those of the combined treatment of haloperidol (125 microg), a nonspecific dopamine receptor antagonist, with an effective dose of yohimbine (2000 microg/kg) on sexually satiated rats were evaluated. Data show that yohimbine and apomorphine, per se, dose-dependently reverse sexual exhaustion by increasing the percentage of sexually satiated rats copulating and resuming copulation after ejaculation. Injection of haloperidol simultaneous to an effective dose of yohimbine, blocked the ability of the latter to reverse sexual satiation. The combined treatment with subthreshold doses of apomorphine and yohimbine synergised to reverse the sexual inhibition characteristic of sexual exhaustion. Data suggest that the dopaminergic system might be the final pathway for the yohimbine-induced sexual behaviour expression in satiated rats. The possible role of sexual motivation in the sexual exhaustion phenomenon is discussed.     

Sulfation of resveratrol in human liver: evidence of a major role for the sulfotransferases SULT1A1 and SULT1E1

Sulfation of resveratrol, a polyphenolic compound present in grapes and wine with anticancer and cardioprotective activities, was studied in human liver cytosol. In the presence of 3'-phosphoadenosine-5'-phosphosulfate, three metabolites (M1-3) whose structures were identified by mass spectrometry and NMR as trans-resveratrol-3-O-sulfate, trans-resveratrol-4'-O-sulfate, and trans-resveratrol-3-O-4'-O-disulfate, respectively. The kinetics of M1 formation in human liver cytosol exhibited an pattern of substrate inhibition with a K(i) of 21.3 +/- 8.73 microM and a V(max)/K(m) of 1.63 +/- 0.41 microLmin(-1)mg(-1) protein. Formation of M2 and M3 showed sigmoidal kinetics with about 56-fold higher V(max)/K(m) values for M3 than for M2 (2.23 +/- 0.14 and 0.04 +/- 0.01 microLmin(-1)mg(-1)). Incubation in the presence of human recombinant sulfotransferases (SULTs) demonstrated that M1 is almost exclusively catalysed by SULT1A1 and only to a minor extent by SULT 1A2, 1A3 and 1E1, whereas M2 is selectively formed by SULT1A2. M3 is mainly catalysed by SULT1A2 and 1A3. In conclusion, the results elucidate the enzymatic pathways of resveratrol in human liver, which must be considered in humans following oral uptake of dietary resveratrol.     

Catecholamines and self-stimulation: evidence suggesting a reinforcing role for noradrenaline and a motivating role for dopamine.

Investigation of the role of noradrenaline (NA) and dopamine (DA) in self-stimulation showed that d-amphetamine (which releases more DA than does l-amphetamine, but not more NA) was much more effective than l-amphetamine in enhancing self-stimulation of NA sites in the locus coeruleus and near-lateral hypothalamus. In DA sites in the substantia nigra and far-lateral hypothalamus the effects of the 2 isomers were confirmed to be more nearly equal. Thymoxamine HCl (10 mg/kg IP), a specific alpha-adrenergic receptor blocker, depressed self-stimulation at all sites, but significantly more severely at DA sites. Thus the drugs most effective in influencing self-stimulation at a particular site were those acting predominantly on the unstimulated system. These findings were interpreted in terms of a hypothesis that DA and NA play complementary roles in self-stimulation and that both are essential; or, more specifically, that DA pathways, implicated in other motivational activites, contribute to a state of drive or arousal necessary for self-stimulation; while response-contingent noradrenergic activity (elicited by the electrodes directly via a transsynaptic route) mediates reinforcement. Further predictions from this hypothesis were tested as follows: (1) Direct pharmacological stimulants of adrenergic alpha-receptors should disrupt self-stimulation by acting randomly on the reinforcement system and disrupting response-reward contingencies; this was confirmed by the finding that the alpha-receptor stimulant clonidine HCl (0.05 mg/kg) depressed self-stimulation at all sites tested. (2) Drect stimulants of DA receptors should enhance self-stimulation of NA sites by augmenting dopaminergic motivational activity; but in rats with DA electrodes, noncontingent stimulation of DA receptors would also impose similar noncontingent activity on the transsynaptic noradrenergic reinforcement pathways and thus depress self-stimulation; this was confirmed by the finding that apomorphine (0.3-1.0 mg/kg) was strongly stimulant for NA electrodes but strongly depressant for DA electrodes, and that the degree and direction of these effects was highly correlated with the differential effects of d- l-amphetamine (rho = .65, p less than 0.01). Neither effect of apomorphine depended on the occurrence of motor stereotypy. These results can be interpreted in terms of 2-component models for self-stimulation, with the predominant transmitter of the drive component being identified as DA and that g the reinforcing component as NA.     

Fentanyl abuse and dependence: further evidence for second hand exposure hypothesis

We have proposed a novel hypothesis regarding the potential role of occupational or second-hand exposure in physician substance use, abuse, and addiction. While only 5.6% of licensed physicians in Florida are anesthesiologists, nearly 25% of physicians followed for substance abuse/dependence are anesthesiologists. When we sort by drug of choice, anesthesiologists have more opioid abuse and dependence than other physicians and appropriate controls. Abuse of one opioid, fentanyl, appears to be increasing and has been noted among the State of Florida's causes of opioid deaths. Fentanyl and sufentanyl are commonly administered highly potent opioid analgesics, as much as 80-800 times as potent as morphine. We have recent data from the State of Florida impaired physicians database, which has allowed us to categorize all fentanyl abusing and/ or dependent physicians. Just knowing that a physician abuses fentanyl gives you a good clue as to their specialty; 75% are anesthesiologists! While drug abuse researchers, oncologists and others who handle drugs of abuse everyday, have no greater incidence of opioid abuse or dependence, anesthesiologists are at the top of every list. Can this be due to just access and stress? We have proposed an alternative hypothesis of second hand exposure. To test this hypothesis, we developed a sensitive LC/MS/MS assay to measure the intravenous anesthetic and analgesic agents, propofol and fentanyl in air. Not only did we detect propofol and fentanyl in cardiovascular surgery operating room air, we also found the highest concentrations were close to the patient's mouth where anesthesiologists work for hours. Like tobacco, second hand opioid exposure can sensitize and change the brain making abuse, dependence and behavioral disorders more likely. Thus environmental exposure and sensitization may be an important risk factor in physician addiction. Second hand exposure may affect treatment outcome and explain anesthesiologist's inability to return to work in the operating room. We are developing an animal model for second hand exposure and additional studies of the operating room and cardiac anesthesiologists are underway.