The use of E-cigarettes as a risk factor for oral potentially malignant disorders and oral cancer: a rapid review of clinical evidence.

BACKGROUND: The popularity of e-cigarettes has increased rapidly in the last decade, particularly among teens and young adults, being advertised as a less harmful alternative to conventional tobacco products. However, in vitro and in vivo studies have evidenced a variable quantity of potentially harmful components and some recognized carcinogens which may cause DNA damage in oral cells. Additionally, evidence suggests that e-cigarettes may play active roles in the pathogenesis of other malignancies, such as lung and bladder cancers. Therefore, this rapid review aimed to assess the available clinical evidence about using e-cigarettes as a risk factor for oral potentially malignant disorders (OPMD) and oral cancer. MATERIAL AND METHODS: A systematic search for English language articles published was performed in PubMed (MEDLINE), Embase, Scopus, and Web of Science. After the study selection process, the authors included twelve clinical studies about OPMD and oral cancer risk in e-cigarette users. RESULTS: The main findings showed the presence of carcinogenic compounds in saliva and morphologic changes, DNA damage, and molecular pathways related to carcinogenesis in the oral cells of e-cigarette users. However, results were inconsistent compared to tobacco smokers and control groups. CONCLUSIONS: the current clinical evidence on this topic is limited and insufficient to support using e-cigarettes as a risk factor for OPMD and oral cancer. Nevertheless, dental care professionals should advise patients responsibly about the potentially harmful effects of e-cigarettes on the oral mucosa cells. Future long-term and well-designed clinical studies are needed.

Clinical manifestations of head and neck cancer in pediatric patients, an analysis of 253 cases in a single Brazilian center.

BACKGROUND: Pediatric head and neck cancer (PHNC) is rare and its nonspecific clinical manifestations may often lead to delayed diagnosis. We aimed to describe the signs, symptoms, and clinicopathological characteristics of PHNC. MATERIAL AND METHODS: Medical records were retrospectively reviewed for all PHNC cases diagnosed from 1986 to 2016 affecting patients aged 19-years and younger from a tertiary referral center in Brazil. Demographic variables, anatomical site of primary tumors, histopathological diagnoses, signs and symptoms, and patterns of misdiagnosis were collected and interpreted by statistical and descriptive analysis. RESULTS: A total of 253 PHNC cases were included. The mean age was 9.3 years and male patients were more frequently affected (60.9%). Burkitt lymphoma (23.7%), nasopharyngeal carcinoma (15.8%), and rhabdomyosarcoma (15.4%) were the most common cancer types. The nasopharynx (28.9%), cervical/lymph node region (25.3%), and craniofacial bones (8.3%) were the predominant anatomical sites. Tumor/swelling (68.4%), was the clinical finding often presented. The univariable analysis showed association between tumor histology and clinical variables such as sex (p=0.022), age (p<0.0001), anatomical location (p<0.0001) tumor/swelling (p=0.034), pain (p=0.031), systemic/general manifestations (p=0.004), nasal/breathing alterations (p=0.012), orbital/ocular alterations (p<0.0001). Misdiagnosis such as tonsillitis, otitis, and abscess were frequent. CONCLUSIONS: Although the clinical findings of PHNC are often unspecific, this study provided signs and symptoms with significant correlations between tumor histology. The suspicion of malignancy should be considered when the main signs and symptoms reported here appear and persist, in order to conduct a timely diagnosis.

Studies on coronary arteriopathy in dogs following administration of CI-1020, an endothelin A receptor antagonist.

A selective nonpeptide endothelin A (ETA) receptor antagonist, CI-1020, was administered to beagle dogs intravenously (i.v.) for 4 hours to 4 weeks. One animal/sex received CI-1020 at 1 mg/kg/hr intravenously for 4, 8, or 24 hours to investigate onset of arteriopathy. Control animals (1/sex) received the vehicle only. To determine reversibility of arteriopathy, 8 dogs/sex were given CI-1020 at 1 mg/kg/hr for 4 days. Two dogs/sex were sacrificed 1, 3, 8, and 29 days following cessation of infusion. Lesion development with prolonged exposure was investigated in 1 male dog. It was given CI-1020 by i.v. bolus at 120 mg/kg/day for 4 weeks and Monastral blue dye was administered i.v. to facilitate localization of vascular lesions. Coronary blood flow was determined in 4 dogs infused with CI-1020 at 0.3, 3, and 30 mg/kg for one hour at each dose. Macroscopically, hemorrhage or blue discoloration of Monastral blue was noted in the extramural coronary arteries along the coronary groove and atrium. Histologically, the earliest coronary changes were noted in animals sacrificed after 24 hours of treatment and characterized by medial hemorrhage and necrosis with a few infiltrating neutrophils. In the reversibility study, incidence and severity of arteriopathy was dependent on time of sacrifice following cessation of infusion. Acute necrotizing inflammation of arteries was present in all animals (n = 4) on day 1 postinfusion, whereas on day 8 postinfusion, lesions characterized by medial small pockets of trapped red cells, cell debris, and adventitial thickening were seen in 1 dog/sex. By day 29 postinfusion, coronary arteries were similar to controls. In the dog given daily i.v. bolus injections of CI-1020 for 4 weeks, arterial inflammatory lesions varied from acute to chronic, although most lesions were considered chronic active. Monastral blue pigments were noted in the wall of most arteries with chronic or chronic active lesions. Acute lesions were similar to those noted in day 1 postinfusion of the reversibility study. Medial smooth muscle necrosis and/or fibrosis with mixed inflammatory cell infiltrates characterized chronic or chronic active lesions. Smooth muscle proliferation and migration into the intima were also noted. There were no significant changes in coronary blood flow, coronary vascular resistance, or mean arterial blood pressure following CI-1020 infusion for 3 hours. In the 24-hour infusion study, plasma endothelin 1 (ET-1) levels were mildly elevated (1.5-4 fold) during CI-1020 infusion when compared to either pretest or control values. These results indicate that administration of endothelin antagonist (CI-1020) to dogs was associated with development of coronary arteriopathy, which was completely resolved within 29 days following cessation of treatment. With prolonged (4-week) CI-1020 treatment, arterial lesions at varying stages of development (acute, chronic active, chronic) were seen, suggesting that tolerance to treatment (up to 4 weeks) does not occur.

Antithrombotic effect of LB-30057 (CI-1028), a new synthetic thrombin inhibitor, in a rabbit model of thrombosis: comparison with inogatran.

LB-30057 (CI-1028) is a novel, orally bioavailable, direct thrombin inhibitor with a Ki of 0.38 nM against human thrombin. The effects of LB-30057 on thrombus formation and hemostasis were evaluated in a veno-venous shunt model of thrombosis in rabbits, and compared with inogatran, another direct inhibitor of thrombin. Each compound was studied at 5 or 6 different doses with 5 or 6 rabbits in each group. After administration as a bolus i.v. injection followed by continuous infusion, both LB-30057 and inogatran dose-dependently inhibited thrombus formation, which was measured as an increase in time to occlusion (TTO) and a decrease in thrombus weight. Both compounds also improved vena caval blood flow and reduced the overall incidence of thrombotic occlusion. LB-30057 significantly prolonged TTO from 23 +/- 4 min (before dose) to 110 +/- 10 min at the highest dose (0.7 mg/kg + 47 microg/kg/min) (p < 0.001), and reduced thrombus weight from 57 +/- 2 mg to 15 +/- 5 mg (p < 0.001). Occlusive thrombus formed in only one of six rabbits that received the highest dose of LB-30057 (vs. 13/13 in the control group, p < 0.01). At the dose that produced the maximum antithrombotic effect (0.7 mg/kg + 47 microg/kg/min), LB-30057 increased aPTT and bleeding time approximately 2-and 2.5-fold above baseline, respectively. On a gravimetric basis, LB-30057 and inogatran displayed comparable in vivo antithrombotic efficacy. When compared to equally effective anti thrombotic doses of inogatran, LB-30057 caused less prolongation in aPTT, had no effect on PT, and tended to have less of effect on bleeding time. These results indicate that LB-30057 is an effective antithrombotic compound and it appears to have a better benefit/risk profile than inogatran in this experimental model.

The antithrombotic effects of CI-1028, an orally bioavailable direct thrombin inhibitor, in a canine model of venous and arterial thrombosis.

Direct thrombin inhibitors represent a new class of drug that may offer a therapeutic alternative for the treatment and prevention of thrombembolic conditions, especially on the venous side of the systemic circulation. CI-1028 (PD 172524/LB30057) is a potent, highly selective inhibitor of thrombin that is orally bioavailable. The efficacy of this compound has been demonstrated in animal models in which intra-venous administration was used. The objective of this study was to evaluate the efficacy of CI-1028 after oral administration in a canine electrolytic injury model of venous and arterial thrombosis. CI-1028 was administered via oral gavage, and animals received either saline or 10, 15, 20, or 30 mg/kg of drug. Fifteen minutes later, the dogs were anesthetized and a femoral artery and vein were exposed and instrumented to induce electrolytic injury and thrombosis while continuously monitoring blood flow in the vessels. Maximum blood CI-1028 concentrations of 0.88+/-0.27, 1.8+/-0.3, 2.2+/-0.5, and 3.2+/-0.5 microg/mL were generally achieved 15 to 30 minutes after administering the compound in the 10-, 15-, 20-, and 30-mg/kg groups, respectively. Administration of CI-1028 increased the time to occlusion (TTO), the principal efficacy end point, in a dose-dependent manner in both arteries and veins. The TTO in the control group (n=8) averaged 66+/-11 minutes in the arteries and 69+/-6 minutes in the veins. In dogs treated with 10 mg/kg (n=8), the TTO was not significantly different from that of the control group. In the 15-mg/kg group (n=9) TTO averaged 140+/-27 minutes in the arteries (p=not significant) and 125+/-15 minutes (p<0.05) in the veins. In the 20-mg/kg group (n=8), TTO was significantly longer than controls in both types of vessels, averaging 168+/-30 minutes in the arteries (p=0.05) and 155+/-21 minutes (p<0.05) in the veins. Likewise, at 30 mg/kg (n=8) both the arterial (179+/-17 minutes) and venous (188+/-15 minutes) TTO was significantly prolonged compared with controls. Surgical blood loss and template bleeding times tended to increase in a dose-dependent manner but a statistically significant elevation was detected for template bleeding time only at the highest dose. Dramatic changes in thrombin time were detected, consistent with the CI-1028 mechanism of action. Virtually no changes were detected in prothrombin time. Maximum activated partial thromboplastin time (aPTT) and activated clotting time changes were detected approximately 30 minutes after dosing, and they were approximately twofold and fivefold baseline values, respectively, at the highest dose. In conclusion, these results demonstrate that CI-1028 provides dose-dependent antithrombotic efficacy after oral administration in a canine model of venous and arterial thrombosis.

Spontaneous thrombosis in mice carrying the factor V Leiden mutation.

A polymorphism in coagulation factor V, factor V Leiden (FVL), is the major known genetic risk factor for thrombosis in humans. Approximately 10% of mutation carriers experience clinically significant thrombosis in their lifetime. In a small subset of patients, thrombosis is associated with coinheritance of other prothrombotic gene mutations. However, the potential contribution of additional genetic risk factors in the majority of patients remains unknown. To gain insight into the molecular basis for the variable expressivity of FVL, mice were generated carrying the homologous mutation (R504Q [single-letter amino acid codes]) inserted into the endogenous murine Fv gene. Adult heterozygous (FvQ/+) and homozygous (FvQ/Q) mice are viable and fertile and exhibit normal survival. Compared with wild-type mice, adult FvQ/Q mice demonstrate a marked increase in spontaneous tissue fibrin deposition. No differences in fetal development or survival are observed among FvQ/Q, FvQ/+ or control littermates on the C57BL/6J genetic background. In contrast, on a mixed 129Sv-C57BL/6J genetic background, FvQ/Q mice develop disseminated intravascular thrombosis in the perinatal period, resulting in significant mortality shortly after birth. These results may explain the high degree of conservation of the R504/R506 activated protein C cleavage site within FV among mammalian species and suggest an important contribution of other genetic factors to the thrombosis associated with FVL in humans. (Blood. 2000;96:4222-4226)

Chronic endothelin blockade in dogs with pacing-induced heart failure: possible modulation of sympathoexcitation.

BACKGROUND: Endothelin-1 (ET-1) is a potent vasoconstrictor peptide elaborated by many cell types. Plasma ET-1 levels are significantly augmented in patients and experimental animals with heart failure. Enhanced levels of ET-1 may contribute to myocardial depression and alterations in sympathetic nerve activity in the setting of chronic heart failure. The effects of chronic blockade of endothelin A (ET(A)) receptors on the development and severity of experimental heart failure and sympathoexcitation were evaluated in these experiments using the specific ET(A) antagonist, PD156707. METHODS AND RESULTS: Four groups of conscious, chronically instrumented mongrel dogs were administered either PD156707 (750 mg orally thrice daily) or a placebo starting 1 day before ventricular pacing or a sham (nonpaced) period. Before pacing or the sham period, baseline hemodynamic and plasma norepinephrine (NE) measurements were made. Hemodynamic and NE measurements were made every 3 to 4 days for the next 28 days. All parameters were relatively stable in nonpaced dogs administered placebo. Paced placebo dogs showed classic hemodynamic and sympathoexcitatory changes indicative of heart failure. Nonpaced dogs administered PD156707 showed a significant decrease in mean arterial pressure and total peripheral resistance beginning 3 days after drug administration. Myocardial function was not affected by PD156707 in nonpaced dogs. In paced dogs, PD156707 also reduced arterial pressure and peripheral resistance. Changes in myocardial function were small and insignificant. Paced dogs administered PD156707 showed an approximately 50% lower increase in plasma NE level from days 10 to 24 compared with paced dogs administered placebo (941.8 +/- 122.8 vs 501.1 +/- 92.6 pg/mL at 17 days; P < .01). CONCLUSIONS: These data suggest that ET-1 contributes to the maintenance of arterial pressure in both sham dogs and dogs paced into heart failure. ET-1 does not appear to have a potent effect on inotropic state, but the data strongly suggest that ET-1 may contribute to the progressive deterioration of circulatory function in heart failure by mediating sympathoexcitation and enhancing plasma NE concentration.

Antithrombotic effects of BCH 2763, a new direct thrombin inhibitor, in a canine model of venous thrombosis.

Deep venous thrombosis (DVT) is a common cardiovascular disease, resulting in significant mortality each year in the United States. Direct thrombin inhibitors represent a new class of drugs that could potentially be better than conventional antithrombotic therapy based on indirect inhibition of coagulation factors with heparin and warfarin. BCH 2763 is a potent, selective bifunctional thrombin inhibitor that blocks both the active catalytic site and the anion binding exosite. The objective of this study is to test the antithrombotic efficacy of BCH 2763 in a canine model of DVT induced through electrolytic injury to the femoral vein. BCH 2763 was administered at three dose levels: 0.125 mg/kg bolus followed by 10 microg/kg/min IV infusion (low-dose; n = 5), 0.25 mg/kg bolus followed by 20 microg/kg/min infusion (mid-dose; n = 5), and 0.75 mg/kg bolus followed by 60 microg/kg/min (high-dose; n = 5). The control group (n = 5) received a 5-ml intravenous bolus of saline followed by a 1 mL/kg/h infusion. The parameters evaluated were changes in activated partial thromboplastin time (aPTT), thrombin time (TT), prothrombin time (PT), time to formation of an occlusive thrombus in the femoral vein, and the amount of venous blood flow delivered over the course of the experiment. There were significant dose-dependent increases in aPTT, TT, and PT in the BCH 2763-treated animals compared with the control group. The time to formation of an occlusive thrombus in the control group averaged 69.6 +/- 9 minutes. Treatment with BCH 2763 prolonged the time to occlusion to 126.4 +/- 13 minutes in the low-dose group, 155.4 +/- 17 minutes in the mid-dose group, and 229 +/- 7 minutes in the high-dose group (80% remained patent for the duration of the study), which were all significantly greater than the controls. Femoral venous blood flow was significantly greater in the mid-dose (51 +/- 8%) and the high-dose (70 +/- 6%) groups compared with the control vessels (22 +/- 3%). In conclusion, the results of this study indicate that BCH 2763 is an effective intravenous antithrombotic agent in the canine electrolytic injury model of venous thrombosis.

Effects of a factor Xa inhibitor, DX-9065a, in a novel rabbit model of venous thrombosis.

The objective of this study was to evaluate the effects of DX-9065a, a nonpeptide, direct inhibitor of factor Xa (FXa), in a novel experimental model of venous thrombosis. The experiments were conducted on anesthetized rabbits in which a veno-venous shunt with cotton threads was inserted into the vena cava. DX-9065a was administered intravenously to the rabbits as an initial bolus followed by a maintenance infusion using the following dosing schedules: DX-I: 0.25 mg/kg + 3 micrograms/kg/min.; DX-II: 0.75 mg/kg + 9 micrograms/kg/min.; DX-III: 1.5 mg/kg + 18 micrograms/kg/min.; DX-IV: 3.0 mg/kg + 36 micrograms/kg/min.; DX-V: 6.0 mg/kg + 72 micrograms/kg/min. DX-9065a induced a dose-dependent increase in the time to occlusion and a dose-dependent decrease in thrombus weight. Because of the unique character of the model, we were also able to show a dose-dependent increase in blood flow through the shunt. In addition, there were dose-dependent increases in prothrombin time (PT) and activated coagulation time (ACT) with more variable responses in the activated partial thromboplastin time (APTT). DX-9065a had little effect on thrombin time (TT) or bleeding time at all doses tested. In conclusion, dose-dependent antithrombotic efficacy was documented with DX-9065a in this new model of venous thrombosis. Although the in vivo potency of the compound was not striking, the results support the utility of FXa inhibition in venous thrombosis and demonstrate the utility of this experimental model for evaluating the efficacy of novel anticoagulants.

A novel model of venous thrombosis in the vena cava of rabbits.

The objective of this study was to develop and validate a new experimental model of venous thrombosis in the rabbit. A 3-cm length of siliconized PE tubing was used as a veno-venous shunt inserted into the abdominal vena cava of anesthetized rabbits. The PE tubing contained six cotton threads which helped to restrict blood flow through the tubing and served as a foreign, thrombogenic surface upon which a thrombus could develop. By continuously measuring blood flow through the vena cava, the rate of thrombus development can be monitored until zero flow is achieved indicating that a completely occlusive thrombus is present. The shunt can be removed making it possible to weigh the thrombus and/or determine its composition. A second shunt can be placed in the vena cava to make a second determination of time to occlusion and thrombus weight, using the data from the first shunt as an internal control standard for comparison. Reproducibility of the technique was demonstrated in a control group (n = 7) in which two successive shunts were used without an antithrombotic intervention. In studies with the first and second shunts, time to occlusion averaged 20.6+/-5.2 min and 20.2+/-5.7 min (pNS), respectively. The net thrombus weights (less the wet weight of the cotton threads) were 49.0+/-3.5 mg and 47.0+/-3.3 mg (pNS). Histologic examination of the thrombi indicated that they were largely composed of fibrin and red blood cells, consistent with the characteristics of venous thrombi. The low molecular weight heparin (LMWH) enoxaparin was used as an antithrombotic intervention to validate the model. Dose-dependent changes in time to occlusion and thrombus weight were achieved which paralleled alterations in coagulation parameters (thrombin time and activated partial thromboplastin time) and bleeding time determined with an ear bleeding technique. The veno-venous shunt model is easy to use, reproducible, and responds appropriately to an antithrombotic intervention, indicating that it should be useful for experimental evaluation of antithrombotic agents designed for venous thromboembolic disorders.

Impact of resting and ischemic blood flow on infarct probability in ischemic preconditioning--a new approach to infarct size-blood flow data by logistic regression.

The linear regression analysis of infarct size (IS) v ischemic myocardial blood flow (MBF) does not account for the heterogeneity of MBF and infarcted tissue; moreover, it cannot assess a blood flow threshold for infarction (MBFT) accurately, as with ischemic preconditioning (IP) the close relationship between ischemic MBF and IS otherwise observed is lost. Finally, the impact of resting blood flow on myocardial infarction cannot be considered in such analysis. Therefore, in a retrospective data analysis of 32 enflurane-anaesthetized swine undergoing 90 min severe ischemia and 120 min reperfusion without (CON, n = 12) or with IP induced by either 3 (IP3, n = 8) or 10 min ischemia (IP10, n = 12) and 15 min reperfusion, a MBFT was assessed by logistic regression (LR) in individual tissue pieces. MBFT was arbitrarily defined as that ischemic MBF (microspheres) at which infarct probability was 0.2, derived from the ratio of infarcted (n = 141, TTC) to all tissue samples (n = 684). The duration of the preconditioning ischemia and MBF both at rest and during the sustained ischemia were significant predictors of infarct probability. Ischemic MBFT at an infarct probability of 0.2, was 0.089 +/- 0.023 ml/min/g in CON. MBFT was decreased to 0.051 +/- 0.03 ml/min/g with IP3 (P < 0.05 v CON) and further to 0.004 +/- 0.037 ml/min/g with IP10 (P < 0.05 v CON, IP3). Corresponding to the leftward shift of MBFT, the relationships between infarct probability and MBF were shifted in parallel by IP with no change in their slopes.

Effects of ischemia, preconditioning, and adenosine deaminase inhibition on interstitial adenosine levels and infarct size.

In order to examine the relationship between local adenosine concentrations before, during, and after ischemia and the extent of ischemic myocardial damage, measurements of interstitial fluid (ISF) nucleosides were made using microdialysis probes implanted in the ischemic region of isoflurane anesthetized Micropigs undergoing 60' coronary artery occlusion (CAO) and 3 h of reperfusion (REP). Nucleoside concentrations in the dialysate collected from the microdialysis probes were used as an index of ISF levels. Dialysate nucleoside concentrations (ADO, inosine and hypoxanthine), myocardial infarct size, and myocardial blood flow (MBF) were determined in control animals (n = 6), animals preconditioned with a single 10' cycle of CAO and REP (PC, n = 6), and those treated with the adenosine deaminase inhibitor pentostatin (n = 6, 0.2 mg/Kg i.v. 30' prior to CAO). The brief PC occlusion resulted in a transient but significant increase in dialysate ADO (6.7 +/- 1.8 microM vs. 0.67 +/- 0.1 microM at baseline). Pentostatin administration had no significant effect on either dialysate nucleosides or MBF at baseline. During the 60' CAO, dialysate ADO increased in control animals. In PC animals, however, dialysate ADO during CAO was lower than control. Pretreatment with pentostatin resulted in a six-fold augmentation in dialysate ADO during the 60 min CAO when compared to the control values (110.62 +/- 30.2 microM vs. 16.31 +/- 2.1 microM at 60 min of ischemia). Pentostatin also resulted in a significant reduction in the accumulation of inosine and hypoxanthine, indicating inhibition of adenosine deaminase activity. There were no significant differences in MBF between groups at any time point. Following 3 h REP, infarct size was 35.4 +/- 5.5%, 8.1 +/- 1.5% and 8.3 +/- 1.8% of the region at risk in control, PC, and pentostatin groups, respectively. These data suggest that marked increase in ISF ADO during CAO, may be as effective in reducing INF as a modest increase in ISF ADO prior to prolonged CAO.

Concomitant endothelin receptor subtype-A blockade during the progression of pacing-induced congestive heart failure in rabbits. Beneficial effects on left ventricular and myocyte function.

BACKGROUND: Plasma levels of endothelin-1 (ET-1) are increased in patients and animals with severe congestive heart failure (CHF). It remains unknown, however, whether ET-1 plays a direct and contributory role in the progression of CHF. Accordingly, the present project tested the hypothesis that chronic blockade of the ETA receptor would have direct and beneficial effects on left ventricular (LV) and myocyte function in a model of CHF. METHODS AND RESULTS: Global LV and isolated myocyte function were examined in rabbits in the following groups (12 per group): chronic rapid ventricular pacing (RVP; 400 bpm, 3 weeks), RVP and concomitant administration of the selective ETA receptor antagonist (PD 156707 24 mg/d), and sham controls. LV fractional shortening decreased after RVP (17 +/- 5 versus 42 +/- 3%) and end-diastolic dimension increased (2.36 +/- 0.44 versus 1.24 +/- 0.18 cm) compared with controls (P < .05). With RVP plus ETA blockade, LV fractional shortening was increased (33 +/- 6%) and end-diastolic dimension decreased (2.02 +/- 0.30 cm) compared with RVP-only values (P < .05). Plasma norepinephrine and endothelin increased twofold in the RVP group. In the RVP plus ETA blockade group, plasma endothelin increased threefold compared with RVP values. Isolated myocyte shortening velocity declined after RVP (42 +/- 13 versus 72 +/- 10 microns/s, P < .05) compared with controls but was normalized with RVP plus ETA blockade (77 +/- 16 microns/s). Myocyte inotropic response to extracellular Ca2+, beta-receptor stimulation, and ET-1 was reduced in the RVP group and returned to control levels with RVP and concomitant ETA receptor blockade. CONCLUSIONS: The results from this study suggest that chronically elevated ET-1 levels and subsequent activation of the ETA receptor play a direct and contributory role in the progression of the CHF process. Thus, specific ETA receptor blockade may provide a new and useful therapeutic modality in the setting of CHF.

Effects of endothelin ETA receptor antagonism with PD 156707 on hemodynamics and renal vascular resistance in rabbits.

The objective of this study was to determine the in vivo effectiveness of the selective endothelin ETA receptor antagonist PD 156707 (sodium 2-benzo[1,3]dioxol-5-yl-4-(4-methoxy-phenyl)-4-oxo-3-(3,4,5-trimet hoxy- benzyl)-but-2-enoate). Effectiveness was defined by the ability of the compound to block increases in renal vascular resistance and mean arterial blood pressure induced by an intravenous bolus of 0.3 nmol/kg of human endothelin-1 in pentobarbital anesthetized rabbits. Different groups of rabbits received hour long intravenous infusions of PD 156707 at doses of 0.003, 0.01, 0.03 or 0.3 mg/kg per h. During baseline conditions, mean arterial blood pressure, heart rate, renal blood flow, and renal vascular resistance were similar among the groups. The intravenous bolus of endothelin-1 significantly decreased mean arterial blood pressure (82 +/- 3 mmHg to 65 +/- 3 mmHg, P < 0.05) and increased renal vascular resistance (2.8 +/- 0.3 mmHg/ml per min to 9.2 +/- 1.1 mmHg/ml per min, P < 0.05) in untreated control animals. At doses of 0.3 and 0.03 mg/kg per h, PD 156707 virtually abolished endothelin-1 induced increases in renal vascular resistance, but did not affect the endothelin-1 induced decrease in mean arterial blood pressure. At 0.01 and 0.003 mg/kg per h, PD 156707 also inhibited endothelin-1 induced increases in renal vascular resistance but the effects were less striking, leading to the conclusion that the minimum effective intravenous dose of the compound in rabbits is in the range of 0.01-0.03 mg/kg per h. The results of this study demonstrate that PD 156707 is an extremely potent and highly selective endothelin ETA receptor antagonist. In addition, this study demonstrates the utility of renal vascular resistance as an in vivo bioassay for evaluating the selective vascular effects of endothelin receptor antagonists in this species.

Heat stress increases cardiac HSP72i but fails to reduce myocardial infarct size in rabbits 24 hours later.

Recent reports suggest that delayed myocardial protection ("second window of preconditioning") occurs 24 hours after brief ischemic or thermal stress. In order to test this hypothesis, we subjected New Zealand White rabbits to a heating regimen (42 degrees C for 15-20 minutes). Twenty four hours later, the effect of heat stress on infarct size was determined by conducting a 30 minute ischemia/3 hour reperfusion protocol. In a separate group of rabbits, Western blot analysis was used to verify that the heating regimen increased expression of HSP72i. The size of the region at risk was delineated by infusion of Unisperse blue and infarcted myocardium was identified by incubation of left ventricular slices in triphenyl tetrazolium chloride. In contrast to expectations, induction of HSP72i with thermal stress was not effective in limiting infarct size in rabbits 24 hours later, calling into question the concept that heat stress induces delayed or "second window" myocardial protection.

The therapeutic potential of novel anticoagulants.

Conventional anticoagulant therapy has been based on indirect inhibition of coagulation factors with heparin and warfarin. These agents display liabilities prompting the development of new anticoagulants over the last two decades. The first to be developed was a series of low molecular weight heparins(LMWHs). Their favourable pharmacokinetic profiles and risk/benefit ratios led to widespread use in Europe and, more recently, approval for their use in the USA. Paralleling the development of LMWHs has been the pursuit of a different strategy focused on direct rather than indirect inhibition of enzymes in the coagulation cascade. In contrast to heparin, LMWHs, or other glycosaminoglycans, direct inhibitors exert their effects independent of either antithrombin III (ATIII) or heparin cofactor II (HCII) and more effectively inhibit clot-bound thrombin or FXa. Highly potent, selective (versus other serine proteases)direct thrombin and FXa inhibitors have been identified and isolated from natural sources, such as leeches, ticks and hookworms. The recombinant forms and analogues of the senatural proteins have been produced using molecular biology techniques, i.e., rHirudin, Hirulogs, recombinant tick anticoagulant peptide (rTAP), recombinant antistasin (rATS) and recombinant nematode anticoagulant peptide-5 (rNAP-5). The design of novel structures or the modification of existing chemicals has led to the synthesis of many non-peptide, low molecular weight inhibitors of thrombin and FXa. Some of them are orally active and may be suitable for long-term clinical use. In addition, considerable progress has been made in developing specific TF/VIIa complex inhibitors. The anticoagulation properties of the new agents are being characterised in experimental studies. Some of them have been advanced to large scale clinical trials and their effectiveness, and sometimes relative ineffectiveness,in arterial and venous thromboembolic disorders has been demonstrated. They are being tested for their potential as new antithrombotic agents that act via direct enzyme inhibition. Thus,the clinician should in future be able to target different thrombotic conditions with proven, specific anticoagulant interventions.

Effects of endothelin-receptor antagonism with PD156707 on myocardial stunning in swine.

Endothelin (ET) has been proposed to play a role in pathogenesis of myocardial ischemia/reperfusion injury. The potential role of ET in myocardial stunning has not been examined. Therefore we tested the hypothesis that selective blockade of ETA receptors with PD156707 {sodium 2-benzo[1,3]dioxol-5-yl-4-(4-methoxy-phenyl) -4-oxo-3-(3,4,5-trimethoxy-benzyl)-but-2-enoate} could improve postischemic contractile dysfunction in open-chest pigs. Myocardial stunning was achieved by a sequence of three 10-min left anterior descending (LAD) occlusions interspersed with 15 min of reperfusion. All pigs received either an intravenous saline vehicle (n =6) or PD156707 (n = 6) at a loading dose infusion of 10 mg/kg/h for 1 h before the first occlusion followed by a maintenance dose of 7 mg/kg/h for 4 h. Systolic wall thickening (percentage of baseline) was measured with sonomicrometers. There was no significant difference in systolic thickening between groups at baseline, at the end of the final stunning occlusion, or at any of the time points during reperfusion. PD156707 significantly reduced arterial blood pressure before myocardial ischemia and throughout reperfusion. There was no significant difference in size of the region at risk between groups. In conclusion, selective blockade of ETA receptors with PD156707 did not significantly alter postischemic contractile function in open-chest pigs. These results suggest that activation of ETA receptors by endogenous ET does not play a significant role in the pathogenesis of myocardial stunning.

Myocardial infarct size is smaller in dogs with pacing-induced heart failure.

OBJECTIVE: Determine if ischemic tolerance is reduced in the setting of experimental heart failure (HF). METHODS: Dogs were paced for 3 weeks at 240 BPM to induce heart failure which was confirmed with hemodynamic and echocardiographic measurements. The pacemaker was turned off 30 min prior to the ischemia study. Normal (n = 9) and HF dogs (n = 12) were anesthetized with sodium pentobarbital, instrumented for cardiovascular assessment through a left lateral thoracotomy, and myocardial blood flow was measured with radioactive microspheres. The left circumflex (LCX) artery was occluded for 90 min followed by 3 h of reperfusion. Infarct size was determined with triphenyl tetrazolium chloride staining. RESULTS: Two-dimensional echocardiograms were obtained before and after 3 weeks of pacing in the HF group. Ejection fraction was reduced from 67 +/- 1 to 32 +/- 2% (P < 0.001) and left ventricular end-diastolic volume (LVEDV) increased from 29 +/- 4 ml before pacing to 47 +/- 5 ml (P < 0.001). HF dogs were characterized by a smaller peak positive dP/dt (1110 +/- 72 vs. 2546 +/- 41 mmHg/s, P < 0.01), a greater LV end-diastolic pressure (34 +/- 3 vs. 9 +/- 2 mmHg, P < 0.01), and lower LV end-systolic pressure (99 +/- 5 vs. 130 +/- 5 mmHg, P < 0.05) compared to control dogs. Heart rate was not significantly different between the two groups throughout the experiment. More HF dogs died from ventricular fibrillation (4/12) than control dogs (1/9), but this difference was not statistically significant (P > 0.2). The LCX occlusion produced a comparable decrease in blood flow in HF and normal dogs (0.08 +/- 0.01 vs. 0.09 +/- 0.01 ml/min/g), but infarct size as a percentage of the region at risk was smaller in HF dogs compared to normal dogs (21 +/- 4 vs. 45 +/- 4%, P < 0.01). Region at risk size was also smaller in HF versus normal dogs (29 +/- 3 vs. 40 +/- 2%, P < 0.05). Accordingly, a subgroup analysis of 6 HF and 5 control dogs with similar RAR sizes (35 +/- 2% vs. 37 +/- 2%) was performed and it also demonstrated that infarct size in HF dogs was smaller than in control dogs (19 +/- 5 vs. 40 +/- 4%, P < 0.01), suggesting that disparities in risk region size did not explain the differences in infarct size. CONCLUSION: Infarct size produced by a standardized ischemia-reperfusion protocol was smaller in dogs with pacing-induced HF. The reduced extent of infarction could not be attributed to differences in collateral blood flow or the size of the region at risk. Although the hearts in HF dogs were dilated, LV systolic blood pressure and the strength of contraction were lower than controls potentially reducing myocardial oxygen demand and explaining the smaller infarct size in HF dogs. Other mechanisms, however, cannot be discounted. Thus, ischemic tolerance is not reduced and may be augmented in dogs with pacing-induced heart failure.

Endothelin A receptor antagonism by PD 156707 does not reduce infarct size after coronary artery occlusion/reperfusion in pigs.

Episodes of myocardial ischemia are associated with increases in cardiac venous plasma endothelin (ET) concentrations, suggesting that ET may play a role in the development of myocardial infarction. The purpose of this study was to determine if selective blockade of ET(A) receptors by PD 156707 reduces infarct size caused by coronary artery occlusion and reperfusion in pentobarbital-anesthetized micropigs. A PD 156707 dose which selectively blocks the ET(A)-mediated vasopressor response, but not the ET(B)-mediated vasodepressor response to i.v. ET-1 challenges (0.3 nmol/kg), was established in dose ranging studies in anesthetized micropigs. In myocardial infarction studies, micropigs received either saline vehicle (n = 7) or PD 156707 (n = 8) at a loading dose of 10 mg/kg/1 hr, followed by a maintenance dose of 7 mg/kg/hr. Coinciding with the start of the maintenance dose, the left anterior descending coronary artery was occluded for 1 hr followed by 3 hr of reperfusion. PD 156707 caused a significant (29 mm Hg) decrease in arterial blood pressure before occlusion. PD 156707 had no effect on infarct size (61.1 +/- 5.6% of the region at risk in the PD 156707 treatment group vs. 70.1 +/- 3.9% in the control group). These results suggest that ET(A) receptor activation does not substantially contribute to coronary artery occlusion/reperfusion-induced myocardial infarction.

Pretreatment with endothelin-1 mimics ischemic preconditioning against infarction in isolated rabbit heart.

We have proposed that ischemic preconditioning in rabbit hearts is initiated by adenosine receptor stimulation resulting in activation of protein kinase C. If this theory is correct then any agonist which can activate PKC should also put the heart into a preconditioned state. This study sought to determine whether endothelin-1 (ET-1), which is known to activate protein kinase C can also mimic ischemic preconditioning. Isolated rabbit hearts experienced 30 min of regional ischemia followed by 120 min of reperfusion. Infarct size was measured with triphenyltetrazolium chloride. In control hearts infarction was 30.3 +/- 2.5% of the risk zone. Preconditioning with 5 min global ischemia and 10 min reperfusion reduced infarct size to 5.6 +/- 0.7% (P < 0.01). Perfusion with either 10 PM ET-1 at constant coronary artery flow for 5 min in lieu of ischemia or 50 PM ET-1 with 10 nM nicardipine to block the former's coronary constructive effect was quite protective and equipotent with preconditioning. Infarction averaged 7.2 +/- 0.8% and 5.8 +/- 1.7% of the risk zone, respectively. This protection could be blocked by PD 156 707 (10 microM), a highly specific endothelin receptor antagonist. Chelerythrine (5 microM), a PKC inhibitor, also aborted protection (22.0 +/- 1.7% infarction). However, 8-(p-sulfophenyl)theophylline (100 microM), an adenosine receptor blocker, given during ET-1 administration did not block ET-1's protective effect indicating that adenosine was not involved in the effect. PD 156707 failed to block the protection from ischemic preconditioning (12.6 +/- 2.3% infarction) revealing that endothelin is not an important physiological mediator of ischemic preconditioning. We conclude that ET-1 can mimic ischemic preconditioning in isolated rabbit hearts as would be predicted since its receptors are PKC-coupled, but that endogenous endothelin contributes little to ischemic preconditioning.