Participation of Oxidative Stress on Rat Middle Cerebral Artery Changes Induced by Focal Cerebral Ischemia: Beneficial Effects of 3,4-Dihydro-6-hydroxy-7-methoxy-2,2-dimethyl-1(2H)-benzopyran (CR-6)

Abstract

Cerebral ischemia followed by reperfusion alters vessel properties of brain arteries in rats, inducing an inflammatory response and excessive generation of reactive oxygen species. This study investigated the participation of oxidative stress on vessel properties after ischemia/reperfusion and the beneficial effects of 3,4-dihydro-6-hydroxy-7-methoxy-2,2-dimethyl-1(2H)-benzopyran (CR-6). The right middle cerebral artery was occluded (90 min) and reperfused (24 h). Sham-operated animals were used as controls. Ischemic rats were treated either with CR-6 (100 mg/kg in 1 ml olive oil) or vehicle (1 ml olive oil) administered orally at 2 and 8 h after the onset of ischemia. The structural, mechanical, and myogenic properties of the middle cerebral artery (MCA) were assessed by pressure myography. Superoxide anion (O 2 . ) production was evaluated by ethidium fluorescence, and protein tyrosine nitrosylation was determined by immunofluorescence. Infarct volume was smaller in rats treated with CR-6. In MCA, ischemia/reperfusion increased wall thickness, cross-sectional area, wall/lumen, and decreased wall stress. CR-6 treatment prevented all of these changes induced by ischemia/reperfusion. However, impaired myogenic response and larger lumen diameter in active conditions observed after ischemia/reperfusion were not modified by CR-6. Treatment with CR-6 prevented the increase in O 2 . production and partially prevented the enhanced protein tyrosine nitrosylation that occurred in response to ischemia/reperfusion. Our findings suggest that oxidative stress is involved in the alterations of MCA properties observed after ischemia/reperfusion and that CR-6 induces protection.Stroke is a complex neurodegenerative disorder characterized by interruption of blood flow supply to the brain. There are several mechanisms involved in ischemic brain injury. Increasing evidence indicates that cerebral ischemia and particularly reperfusion are responsible for oxidative stress caused by free radical production. After reperfusion, there is a surge in production of superoxide anion (O 2 . ), nitric oxide (NO), and peroxynitrite (ONOO Ϫ ). Formation of these species in the vicinity of blood vessels plays an important role in reperfusion-induced injury (Doyle et al., 2008). In healthy individuals, antioxidant activity counterbalances free radical production, but in the case of ischemia, the balance between reactive oxygen species (ROS) and antioxidant activity is shifted toward free radicals, causing oxidative stress.All vascular cells can produce ROS through different sources, including NADPH oxidase, mitochondria, cyclooxygenases, and ABBREVIATIONS: ROS, reactive oxygen species; MCA, middle cerebral artery; I/R, ischemia/reperfusion; CR-6, 3,4-dihydro-6-hydroxy-7-methoxy-2,2-dimethyl-1(2H)-benzopyran; KHS, Krebs-Henseleit solution; DHE, dihydroethidium; SOD, superoxide dismutase; ANOVA, analysis of variance; CSA, cross-sectional area; PEG, polyethylene glycol; WT, wall thickness; D iCa 2ϩ, internal diameter measured in active c...

Middle cerebral artery remodeling following transient brain ischemia is linked to early postischemic hyperemia: A target of uric acid treatment

“…The structural, mechanical, and myogenic properties of the MCA were studied with a pressure myograph (Danish Myo Tech Model P100; J.P. Trading I/S, Aarhus, Denmark), as described previously (14,15). In brief, the vessel was placed on two glass microcannulae and secured with surgical nylon suture.…”

“…The proximal segment of the ipsilateral MCA was immediately used for pressure myography and processed for nuclei distribution microscopic fluorescence studies (15). Distal ipsilateral MCA segments were used for evaluation of O 2 .-production, immunofluorescence and mRNA levels (14). Coronal brain sections for immunofluorescence studies were processed as described (23).…”

“…A simple neurological test on a nine-point scale (0=no deficit to 9=highest handicap) was performed before killing the animals, as reported (23). At 24 h, rats were euthanized under deep isoflurane anesthesia, and the brain was removed and placed in cold Krebs-Henseleit solution (KHS) (14,15). Evaluation of infarct volume was performed with the 2,3,5-triphenyltetrazolium chloride (Sigma-Aldrich, St. Louis, MO) technique, as described (23).…”

“…For instance, hypertension, which is a leading cause of stroke, induces remodeling of cerebral arteries causing larger infarcts (15,24). Several lines of evidence suggest that transient MCA occlusion in rats induces MCA wall thickening (7,14,15). Increased oxidative stress seems to be involved in such arterial remodeling since antioxidant treatment at reperfusion rescues MCA from remodeling and reduces infarct volume (14).…”

“…Several lines of evidence suggest that transient MCA occlusion in rats induces MCA wall thickening (7,14,15). Increased oxidative stress seems to be involved in such arterial remodeling since antioxidant treatment at reperfusion rescues MCA from remodeling and reduces infarct volume (14). After cerebral I/R, the inflammatory response is accompanied by an increase in macrophage infiltration (32).…”