irl-1620 and Infarction--Middle-Cerebral-Artery

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

Manipulation of the central endothelin (ET) system via either ETA antagonists or ETB agonists following experimental cerebral ischemia has been shown to be beneficial in previous experimental studies. In order to further explore the involvement of these receptors in cerebral ischemia, we determined changes in binding affinity (Kd) and density (Bmax) of ETA and ETB receptors in the rat brain at 24 h and 1 week following middle cerebral artery occlusion (MCAO). Rats subject to MCAO exhibited significant neurological and motor function deficit as well as large infarct volumes (126.41±40.12 and 152.82±21.67 mm(3) on days 1 and 7, respectively). Bmax increased (P<0.01) and Kd decreased (P<0.01) for ETA receptors in the infarcted right cerebral hemisphere compared to sham 24 h post MCAO. However, after 7 days of MCAO, Bmax of ETA receptors was similar, while Kd in the infarcted hemisphere increased (P<0.05) compared to sham. Binding characteristics for brain ETB receptors were not altered 24 h post MCAO. However, 7days following MCAO, there was a significant decrease (P<0.001) in Kd values and an increase (P<0.001) in Bmax values for ETB receptors in the ischemic cerebral hemisphere. The initial increase in ETA receptors is damaging and may be aggravating cerebral ischemia due to its vasoconstrictive actions. On the other hand, since ETB receptors have been shown to enhance brain angiogenesis, it is possible that an increase in binding characteristics of these receptors is part of a natural defense mechanism to repair the ischemic brain.

Topics: Animals; Brain Ischemia; Endothelins; Infarction, Middle Cerebral Artery; Iodine Radioisotopes; Male; Peptide Fragments; Radioligand Assay; Rats; Receptor, Endothelin A; Receptor, Endothelin B; Time Factors

Endothelin B receptor agonist, IRL-1620, has been shown in previous studies, conducted in our lab, to provide significant neuroprotection at both 24h and 1 week following permanent cerebral ischemia. It is possible that IRL-1620 may be neuroprotective due to angiogenesis and neurogenesis. However, the effect of IRL-1620 on neurovascular remodeling following cerebral ischemia has not been established. The present study was conducted to determine the effect of IRL-1620 [Suc-[Glu9,Ala11,15]-Endothelin-1(8-12)] on astrocytes, neurons, and vascular endothelial cells after induction of cerebral ischemia. Male Sprague-Dawley rats undergoing permanent middle cerebral artery occlusion (MCAO) received three intravenous injections of either vehicle or IRL-1620 at 2, 4, and 6h post occlusion. At 24h post occlusion, IRL-1620 treatment preserved neuronal numbers in the cortex, striatum and subventricular zone (SVZ) of the ischemic rat brain, while simultaneously enhancing the number of blood vessels labeled with vascular endothelial growth factor (VEGF) compared to vehicle treatment. By 1 week following MCAO, VEGF-positive vessels/30 µm brain slice in the IRL-1620 group numbered 11.33±2.13 versus 4.19±0.79 in the vehicle group (P<0.01). Additionally, animals receiving IRL-1620 displayed increased number of proliferating cells (P<0.0001) and cells positively staining for nerve growth factor (NGF; P<0.0001) in the infarcted brain. VEGF and NGF protein expression significantly increased at 1 week post MCAO in the infarcted hemisphere of IRL-1620 treated rats as compared to sham (P<0.01). Pretreatment with BQ788 blocked the effects of IRL-1620, thus confirming the role of ETB receptors in the neurovascular remodeling actions of IRL-1620. Results of the present study indicate that IRL-1620, administered on the day of infarct, is neuroprotective and enhances angiogenic and neurogenic remodeling following cerebral ischemia.

Topics: Angiogenesis Inducing Agents; Animals; Astrocytes; Endothelins; Infarction, Middle Cerebral Artery; Male; Neurogenesis; Neurons; Neuroprotective Agents; Peptide Fragments; Rats; Rats, Sprague-Dawley; Receptor, Endothelin B

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