irl-1620 and Disease-Models--Animal

Stimulation of endothelin B receptors by its agonist IRL-1620 (INN, sovateltide) provides neuroprotection and neurological and motor function improvement following cerebral ischemia. We investigated the effect of sovateltide on stem and progenitor cells mediated neural regeneration and its effect on the cerebral tissue repair and restoration of neurological and motor function. Sovateltide (5 μg/kg) was injected intravenously in permanent middle cerebral artery occluded (MCAO) rats at 4, 6, and 8 h at days 0, 3, and 6. Neurological and motor function tests were carried out pre-MCAO and at day 7 post-MCAO. At day 7, significantly reduced expression of neuronal differentiation markers HuC/HuD and NeuroD1 was seen in MCAO + vehicle than sham rats. Sovateltide treatment upregulated HuC/HuD and NeuroD1 compared to MCAO + vehicle and their expression was similar to sham. Expression of stem cell markers Oct 4 and Sox 2 was similar in rats of all of the groups. Significantly reduced infarct volume and DNA damage with recovery of neurological and motor function was observed in sovateltide-treated MCAO rats. These results indicate that sovateltide initiates a regenerative response by promoting differentiation of neuronal progenitors and maintaining stem cells in an equilibrium following cerebral ischemic stroke.

Topics: Animals; Brain; Cell Differentiation; Disease Models, Animal; DNA Damage; Endothelins; Humans; Infarction, Middle Cerebral Artery; Injections, Intravenous; Ischemic Stroke; Male; Nerve Regeneration; Neurons; Peptide Fragments; Rats; Receptor, Endothelin B; Stem Cells

The development of effective drugs for stroke is urgently required as it is the 2nd largest killer in the world and its incidence is likely to increase in the future. We have demonstrated cerebral endothelin B receptors (ETBR) as a potential target to treat acute cerebral ischemic stroke. However, the mechanism of ETBR mediated neural regeneration and repair remains elusive. In this study, a permanent middle cerebral artery occluded (MCAO) rat model was used. Sovateltide (an ETBR agonist) injected intravenously showed better survival and neurological and motor function improvement than control. Higher neuronal progenitor cells (NPCs) differentiation along with better mitochondrial morphology and biogenesis in the brain of sovateltide rats were noted. Exposure of cultured NPCs to hypoxia and sovateltide also showed higher NPC differentiation and maturation. This study shows a novel role of ETBR in NPCs and mitochondrial fate determination in cerebral ischemia, and in improving neurological deficit after stroke.

Topics: Administration, Intravenous; Animals; Antigens, Nuclear; Basic Helix-Loop-Helix Transcription Factors; Cell Differentiation; Cell Hypoxia; Disease Models, Animal; Dynamins; Endothelins; GTP Phosphohydrolases; Infarction, Middle Cerebral Artery; Male; Mitochondria; Mitochondrial Dynamics; Mitochondrial Proteins; Nerve Tissue Proteins; Neural Stem Cells; Peptide Fragments; Rats; Stroke

Enhancing blood flow to tumors is a prominent strategy for improving the tumor accumulation of macromolecular drugs through the enhanced permeability and retention (EPR) effect. IRL-1620 is an agonist of the endothelin B receptor, and is a promising molecule to enhance tumor blood flow by activating endothelial nitric oxide synthase. However, contradictory effects on tumor blood flow modulation have been reported because the effects of IRL-1620 may differ in different animal models. Here, we examined for the first time the effect of IRL-1620 on the EPR effect for PEGylated liposomes in a CT-26 murine colon cancer model. Co-injection of IRL-1620 at an optimum dose (3 nmol/kg) nearly doubled the tumor accumulation of liposomes compared with controls, indicating that IRL-1620 enhanced the EPR effect in the present colon cancer model. Co-injection of IRL-1620 is a promising strategy to improve the therapeutic effects of macromolecular drugs while reducing their side effects.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Colon; Colonic Neoplasms; Disease Models, Animal; Endothelin B Receptor Antagonists; Endothelins; Humans; Intestinal Mucosa; Liposomes; Male; Mice; Peptide Fragments; Permeability; Receptor, Endothelin B

ETB receptor agonist, IRL-1620 (or SPI-1620) presently in US Phase 1 clinical trial, has been demonstrated to selectively and transiently increase tumor blood flow. The present study was conducted to determine the effect of IRL-1620 on radiation therapy in tumor bearing mice inoculated with Dalton's Lymphoma Ascites cells. Tumors were allowed to grow for 30 days to a size of 1.10-1.29 cm3 before starting the treatment. The animals with or without IRL-1620 treatment were exposed to radiation (4 Gy/dose) on every alternate day for a total of 5 doses. Tumor volume was determined twice every week till the end of study. Radiation alone did not affect the tumor volume; however, animals treated with IRL-1620 followed by radiation produced a significant (64%) reduction in tumor volume. Survival of mice improved from 0/10 at 56 days after tumor inoculation in vehicle plus radiation group to 6/10 at 70 days in IRL-1620 (9 nmol/kg) plus radiation group. It is concluded that IRL-1620 improves the efficacy of radiation treatment in tumor bearing mice. (These findings have been earlier presented as an abstract ).

Topics: Animals; Disease Models, Animal; Endothelins; Lymphoma; Male; Mice; Peptide Fragments; Receptor, Endothelin B; Tumor Burden

Endothelin and its receptors have long been considered therapeutic targets in the treatment of ischemic stroke. Recent studies indicate that ET(B) receptors may provide both vasodilatation and neuroprotection. The purpose of this study was to determine the effect of selectively activating the ET(B) receptors following permanent middle cerebral artery occlusion in rats. IRL-1620 [Suc-[Glu9,Ala11,15]-Endothelin-1(8-12)], a highly selective ET(B) agonist, was used alone and in conjunction with BQ788, an ET(B) antagonist, to determine the role of ET(B) receptors in cerebral ischemia. Rats were assessed for neurological deficit and motor function, and their brains were evaluated to determine infarct area, oxidative stress parameters, and ET receptor protein levels. Animals treated with IRL-1620 showed significant improvement in all neurological and motor function tests when compared with both vehicle-treated and BQ788-treated middle cerebral artery occluded groups. In addition, there was a significant decrease in infarct volume 24h after occlusion in animals treated with IRL-1620 (24.47±4.37mm(3)) versus the vehicle-treated group (153.23±32.18mm(3)). Blockade of ET(B) receptors by BQ788 followed by either vehicle or IRL-1620 treatment resulted in infarct volumes similar to those of rats treated with vehicle alone (163.51±25.41 and 139.21±15.20mm(3), respectively). Lipid peroxidation, as measured by malondialdehyde, increased and antioxidants (superoxide dismutase and reduced glutathione) decreased following infarct. Treatment with IRL-1620 reversed these effects, indicating that ET(B) receptor activation reduces oxidative stress injury following ischemic stroke. Animals pretreated with BQ788 showed similar oxidative stress damage as those in the vehicle-treated group. No significant difference was observed in ET(B) receptor levels in any of the groups. The present study demonstrates that ET(B) receptor activation may be a novel neuroprotective therapy in the treatment of focal ischemic stroke.

Topics: Analysis of Variance; Animals; Brain Infarction; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelin B Receptor Antagonists; Endothelins; Gene Expression Regulation; Glutathione; Infarction, Middle Cerebral Artery; Male; Malondialdehyde; Motor Activity; Muscle Strength; Nervous System Diseases; Neurologic Examination; Oligopeptides; Peptide Fragments; Piperidines; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Receptor, Endothelin B; Rotarod Performance Test; Superoxide Dismutase

Chronic post-ischemic pain (CPIP) is an animal model of CRPS-I developed using a 3-h ischemia-reperfusion injury of the rodent hind paw. The contribution of local endothelin to nociception has been evaluated in CPIP mice by measuring sustained nociceptive behaviors (SNBs) following intraplantar injection of endothelin-1 or -2 (ET-1, ET-2). The effects of local BQ-123 (ETA-R antagonist), BQ-788 (ETB-R antagonist), IRL-1620 (ETB-R agonist) and naloxone (opioid antagonist) were assessed on ET-induced SNBs and/or mechanical and cold allodynia in CPIP mice. ETA-R and ETB-R expression was assessed using immunohistochemistry and Western blot analysis. Compared to shams, CPIP mice exhibited hypersensitivity to local ET-1 and ET-2. BQ-123 reduced ET-1- and ET-2-induced SNBs in both sham and CPIP animals, but not mechanical or cold allodynia. BQ-788 enhanced ET-1- and ET-2-induced SNBs in both sham and CPIP mice, and cold allodynia in CPIP mice. IRL-1620 displayed a non-opioid anti-nociceptive effect on ET-1- and ET-2-induced SNBs and mechanical allodynia in CPIP mice. The distribution of ETA-R and ETB-R was similar in plantar skin of sham and CPIP mice, but both receptors were over-expressed in plantar muscles of CPIP mice. This study shows that ETA-R and ETB-R have differing roles in nociception for sham and CPIP mice. CPIP mice exhibit more local endothelin-induced SNBs, develop a novel local ETB-R agonist-induced (non-opioid) analgesia, and exhibit over-expression of both receptors in plantar muscles, but not skin. The effectiveness of local ETB-R agonists as anti-allodynic treatments in CPIP mice holds promise for novel therapies in CRPS-I patients.

Topics: Analysis of Variance; Animals; Behavior, Animal; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Endothelin Receptor Antagonists; Endothelin-1; Endothelin-2; Endothelins; Hyperalgesia; Keratinocytes; Male; Mice; Muscle, Skeletal; Naloxone; Narcotic Antagonists; Oligopeptides; Pain Measurement; Pain Threshold; Peptide Fragments; Peptides, Cyclic; Peripheral Nervous System; Physical Stimulation; Piperidines; Receptors, Endothelin; Reflex Sympathetic Dystrophy; Skin

Endothelin (ET)-1 is a likely candidate for a key role in diabetic vascular complications. However, no abnormalities in the vascular responsiveness to ET-1 have been identified in the chronic stage of type 2 diabetes. Our goal was to look for abnormalities in the roles played by ET receptors (ET(A) and ET(B)) in the mesenteric artery of the type 2 diabetic Goto-Kakizaki (GK) rat and to identify the molecular mechanisms involved. Using mesenteric arteries from later-stage (32-38 wk old) individuals, we compared the ET-1-induced contraction and the relaxation induced by the selective ET(B) receptor agonist IRL1620 between GK rats and control Wistar rats. Mesenteric artery ERK activity and the protein expressions for ET receptors and MEK were also measured. In GK rats (vs. age-matched Wistar rats), we found as follows. 1) The ET-1-induced contraction was greater and was attenuated by BQ-123 (ET(A) antagonist) but not by BQ-788 (ET(B) antagonist). In the controls, BQ-788 augmented this contraction. 2) Both the relaxation and nitric oxide (NO) production induced by IRL1620 were reduced. 3) ET-1-induced contraction was enhanced by N(G)-nitro-l-arginine (l-NNA; NO synthase inhibitor) but suppressed by sodium nitroprusside (NO donor). 4) The enhanced ET-1-induced contraction was reduced by MEK/ERK pathway inhibitors (PD-98059 or U0126). 5) ET-1-stimulated ERK activation was increased, as were the ET(A) and MEK1/2 protein expressions. 6) Mesenteric ET-1 content was increased. These results suggest that upregulation of ET(A), a defect in ET(B)-mediated NO signaling, and activation of the MEK/ERK pathway together represent a likely mechanism mediating the hyperreactivity to ET-1 examined in this study.

Topics: Angiotensin II; Animals; Arginine Vasopressin; Butadienes; Diabetes Mellitus, Type 2; Diabetic Angiopathies; Disease Models, Animal; Disease Progression; Dose-Response Relationship, Drug; Endothelin-1; Endothelins; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Flavonoids; Male; MAP Kinase Kinase Kinases; Mesenteric Arteries; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitriles; Nitroarginine; Nitroprusside; Oligopeptides; Peptide Fragments; Peptides, Cyclic; Piperidines; Rats; Rats, Wistar; Receptor, Endothelin A; Receptor, Endothelin B; Signal Transduction; Vasoconstriction; Vasodilation

The susceptibility of changes in responsiveness to noxious cold stimulation of rats submitted to chronic constriction of the infraorbital nerve (CION) or carrageenan to drug inhibition was compared. Nocifensive responses were measured as total time rats engaged in bilateral facial grooming with both forepaws over the first 2 min following tetrafluoroethane spray application to the snout. Carrageenan (50 microg, s.c. into upper lip) caused short-lived ipsilateral cold hyperalgesia (peak at 3 h: vehicle 8.4+/-1.3, carrageenan 21.2+/-3.0 s) which was markedly suppressed by i.p. indomethacin (4 mg/kg), celecoxib (10mg/kg) or s.c. dexamethasone (0.5 mg/kg), endothelin ET(A) or ET(B) receptor antagonists (BQ-123 and BQ-788, respectively; 10 nmol/lip). CION caused ipsilateral cold hyperalgesia between Days 2 and 12, which peaked on Days 4 (sham 15.3+/-1.8, CION 32.4+/-5.3s) to 6. Established peak CION-induced cold hyperalgesia was unaffected by indomethacin and celecoxib, whereas dexamethasone, BQ-123, BQ-788, and i.v. injections of selective antagonists of ET(A) (atrasentan, 3-10 mg/kg) or ET(B) (A-192621, 5-20 mg/kg) receptors caused significant inhibitions lasting 1-2.5h (peaks approximately 65-90%). Bosentan (dual ET(A)/ET(B) receptor antagonist, 10 mg/kg, i.v.) abolished CION-induced cold hyperalgesia for up to 6h. Thus, once established, CION-induced orofacial hyperalgesia to cold stimuli appears to lack an inflammatory component, but is alleviated by endothelin ET(A) and/or ET(B) receptor antagonists. If this CION injury model bears predictive value to trigeminal neuralgia (i.e., paroxysmal orofacial pain triggered by various stimuli), endothelin receptors might constitute new targets for treatment of this disorder.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Atrasentan; Bosentan; Carrageenan; Celecoxib; Cold Temperature; Dexamethasone; Disease Models, Animal; Drug Evaluation, Preclinical; Endothelin-1; Endothelins; Grooming; Hyperalgesia; Indomethacin; Male; Maxillary Nerve; Nerve Compression Syndromes; Oligopeptides; Peptide Fragments; Peptides, Cyclic; Piperidines; Pyrazoles; Pyrrolidines; Rats; Rats, Wistar; Receptor, Endothelin A; Receptor, Endothelin B; Sulfonamides; Trigeminal Neuralgia

Numerous men develop postprostatectomy erectile dysfunction (PPED), due to surgery-related nervous damage. PPED is often refractory to phosphodiesterase type 5 (PDE5) inhibitors therapy.. To verify whether chronic tadalafil (CT) preserves bilateral cavernous neurotomy (BCN)-induced penile damage and hypo-oxygenation.. In a rat model of BCN we evaluated in vitro and ex vivo effect of CT treatment (2 mg/kg, daily for 3 months).. Bilateral cavernous neurotomy induced massive hypoxia and decreased muscle/fiber ratio, completely restored by CT. Hypersensitivity of hypoxic tissues to the relaxant effect of the endothelin type B receptor (ETB) agonist IRL-1620 was observed, along with ETB mRNA and protein overexpression. CT restored sensitivity to IRL-1620, and normalized ETB expression. Hypoxic penis showed increased sensitivity to the relaxant effect of the nitric oxide donor sodium nitroprusside (SNP), while acute tadalafil (100 nM) did not amplify the SNP effect. Accordingly, PDE5 mRNA and protein were reduced in BCN penile tissues. By restoring PDE5, CT decreased SNP-induced relaxation and rescued sensitivity to acute tadalafil. However, in hypoxic penis, CT normalizes neither acetylcholine hyporesponsiveness nor neuronal nitric oxide synthase-endothelial nitric oxide synthase expression.. Chronic tadalafil restores some of the investigated BCN-induced alterations, including PDE5 and tadalafil efficacy.

Topics: Animals; Carbolines; Cell Hypoxia; Disease Models, Animal; Endothelins; Erectile Dysfunction; Male; Nitric Oxide Donors; Nitroprusside; Penile Erection; Penis; Peptide Fragments; Phosphodiesterase Inhibitors; Prostatectomy; Rats; Rats, Sprague-Dawley; Tadalafil; Vasodilator Agents

This study was designed to investigate the effects of the selective endothelin-B (ET-B) receptor agonist IRL1620 and the selective ET-B receptor antagonist BQ788 on myocardial and endothelial function after reversible deep hypothermic ischemia and reperfusion.. Isogenic intraabdominal heterotopic heart transplantation was performed on Lewis rats. After one hour of cold ischemic preservation, reperfusion was started either after application of saline vehicle or IRL1620 or BQ788 or both. Left ventricular pressure-volume relations and myocardial blood flow were assessed after one and 24 hours of reperfusion. Endothelium-dependent vasodilatation to acetylcholine (ACH) and endothelium-independent vasodilatation to sodium nitroprusside (SNP) were also determined.. IRL1620 attenuated and BQ788 improved myocardial contractility significantly as indicated by the left or upward shift of the systolic pressure-volume relation, respectively, and significantly changed myocardial blood flow during early reperfusion (p<0.05). Although myocardial function and baseline myocardial blood flow were similar in both groups after 24 hours of reperfusion, endothelium-dependent vasodilatation was still significantly lower in the IRL1620 and higher in the BQ788 group (p<0.05).. These results suggest that activation of the ET-B receptors contributes to reperfusion injury after cardiac preservation in a rat heart transplant model.

Topics: Acetylcholine; Animals; Circadian Rhythm; Disease Models, Animal; Endothelin Receptor Antagonists; Endothelins; Endothelium, Vascular; Heart Rate; Heart Transplantation; Models, Cardiovascular; Myocardial Contraction; Myocardial Reperfusion Injury; Nitroprusside; Oligopeptides; Peptide Fragments; Piperidines; Rats; Receptor, Endothelin B; Receptors, Endothelin; Recovery of Function; Regional Blood Flow; Reperfusion; Time Factors; Vasodilation; Vasodilator Agents; Ventricular Function, Left; Ventricular Pressure

1. The aim of the study was to visualize endothelin-1 (ET-1)-mediated constriction in renal vessels of cortical and juxtamedullary glomeruli in the split hydronephrotic rat kidney in vivo and to functionally characterize the ET receptor subtypes involved. 2. ET-1 (10(-9) M) constricted preglomerular vessels (by 6-18%) and efferent arterioles (by 11-13%), and decreased glomerular blood flow (GBF, by 55%) of cortical and juxtamedullary glomeruli. 3. The ETA antagonist BQ-123 (10(-6) M), as well as the ETB antagonist BQ-788 (2 x 10(-7) M) and IRL 1038 (10(-6) M), shifted the concentration-response curve of GBF for ET-1 to the right by one order of magnitude. While BQ-123 antagonized ET-1 constriction only in preglomerular vessels, BQ-788 and IRL 1038 were effective both in preglomerular vessels and efferent arterioles. 4. The ETB agonist IRL 1620 (10(-8) M) reduced GBF by 50% and constricted efferent arterioles (by 20-33%) about two times more than preglomerular vessels (by 6-14%). 5. Our results suggest that in renal cortical and juxtamedullary vessels of rats, ET-1-induced preglomerular vasoconstriction is mediated by ETA and ETB receptors, while efferent vasoconstriction is predominantly mediated by ETB receptors, which might have important consequences for the regulation of glomerular filtration pressure by ET.

Topics: Animals; Blood Pressure; Body Weight; Disease Models, Animal; Endothelin Receptor Antagonists; Endothelin-1; Endothelins; Female; Glomerular Filtration Rate; Kidney; Kidney Glomerulus; Microcirculation; Nephrosis; Peptide Fragments; Peptides, Cyclic; Rats; Rats, Wistar; Receptor, Endothelin A; Receptor, Endothelin B; Receptors, Endothelin; Vasoconstriction

The hemodynamic effects of endothelin-1 (ET-1) are mediated by at least two distinct receptors: ETa and ETb receptors. Recently, ETb receptor agonists (4 Ala ET-1 and IRL 1620) were developed. To investigate the role of ETb receptor activation on the pulmonary and systemic circulations, we studied the hemodynamic effects of intrapulmonary arterial injections of these receptor agonists in 10 intact newborn lambs. At rest, 4 Ala ET-1 (290-1,725 ng/kg) changed no hemodynamic variables. IRL 1620 (180-1,095 ng/kg) decreased mean pulmonary arterial pressure (PAP, 16.8% +/- 15.0 and 17.8% +/- 8.5, p < 0.05) and left pulmonary artery blood flow (21.6% +/- 22.1 and 33.4% +/- 27.7, p < 0.05) at the two highest doses only. During U46619-induced pulmonary hypertension, both 4 Ala ET-1 (3.2% +/- 8.0 to 15.9% +/- 6.4, p < 0.05) and IRL 1620 (8.7% +/- 6.3 to 21.9% +/- 4.1, p < 0.05) produced selective dose-dependent decreases in PAP. The decrease in mean PAP induced by 4 Ala ET-1 and IRL 1620 was attenuated by N omega-nitro-L-arginine [an inhibitor of endothelium-derived nitric oxide (EDNO) synthesis] (16.6% +/- 3.5 vs. 5.9% +/- 2.3 and 16.2% +/- 3.4 vs. 6.6% +/- 2.8, p < 0.05) and by glybenclamide (a blocker of ATP-dependent potassium channels) (18.2% +/- 7.9 vs. 7.5% +/- 8.3 and 14.7% +/- 3.6 vs. 6.3% +/- 3.2, p < 0.05). ETb receptor activation produces selective pulmonary vasodilation during pulmonary hypertension in intact newborn lambs. The vasodilating properties are mediated in part by release of ENDO and by potassium channel activation.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Animals, Newborn; Arginine; Blood Pressure; Disease Models, Animal; Endothelins; Endothelium, Vascular; Glyburide; Hypertension, Pulmonary; Injections, Intra-Arterial; Nitroarginine; Peptide Fragments; Prostaglandin Endoperoxides, Synthetic; Pulmonary Artery; Pulmonary Circulation; Regional Blood Flow; Sheep; Thromboxane A2; Vascular Resistance; Vasoconstrictor Agents; Vasodilation