Rabbit Basilar Artery Research Articles

What is the restorative effect of VEGF inhibitor bevacuzimab against subarachnoid hemorrhage in an experimental model?

This study investigated the effect of vascular endothelial growth factor (VEGF) inhibitor bevacuzimab (BVZ) on the rabbit basilar artery using an experimental subarachnoid hemorrhage (SAH) model. Eighteen adult male New-Zealand white rabbits were randomly divided into three groups: a control group (n = 6), SAH group (n = 6), and SAH+BVZ group (n = 6). Experimental SAH was created by injecting autologous arterial blood into the cisterna magna. In the treatment group, the subjects were administered a daily dose of 10 mg/kg, intravenous BVZ for 2 days after the SAH. Basilar artery diameters were measured with magnetic resonance angiography (MRA) 72 h after the SAH in all groups. After 72 h, whole brains, including the upper cervical region, were obtained from all the animals after perfusion and fixation of the animal. The wall thickness, luminal area, and the apoptosis at the basilar arteries were evaluated in all groups. BVZ significantly prevented SAH-induced vasospasm confirmed in vivo with MRA imaging with additional suppression of apoptosis on basilar artery wall. VEGF inhibition with BVZ has shown to have a vasospasm and apoptosis attenuating effect on basilar artery in a SAH model.

Expression of c-Jun N-terminal kinase and B cell lymphoma/leukemia-2 associated X protein in rabbit basilar artery aneurysm

Objective To observe the expression of c-Jun N-terminal kinase(JNK) and B cell lymphoma/leukemia-2 associated X protein (bax) during the formation of basilar aneurysm in the rabbit. Methods Forty New Zealand white rabbits were divided into five group, Normal group (n=5), Sham group (n=5) , Operation group(2 weeks, 8 weeks, 16 weeks)(n=10). The bilateral common carotid artery was exposed in the sham operation group by bilateral common carotid artery ligation. MRA (Magnetic Resonance Angiography) was performed at the prescribed time, All experimental animals were killed in order to observe the pathological staining of the basilar artery tip morphology, vascular intima, the middle elastic layer and the immunohistochemical staining. Results The results of immunohistochemical staining showed that the positive rate of JNK in each group was (8.600±2.074)% in the control group, (18.900±4.458)% in the 2W group (t=13.405, P=0.000), and the postoperative 8W group (25.800±3.615)% (t=22.570, P=0.000), postoperative 16W group (38.400±8.343)% (t=14.555, P=0.000); the difference between the experimental group and the control group was statistically significant. The positive expression rates of bax were: (9.400±3.050)% in postoperative 2W group, (22.100±4.748)% in postoperative 2W group (t=14.719, P=0.000) postoperative 8W group (29.200±4.894)% (t=18.866, P=0.000), postoperative 16W group (36.700±4.762)% (t=24.371, P=0.000); each experimental group was statistically significant difference from the control group. Imaging results: aneurysm The volume was: control group (7.600±3.847), (24.746±27.813) mm3,t=2.814, P=0.020), postoperative 8W group (37.083±35.695) mm3 (t=3.285, P=0.009), postoperative 16W group (105.751±97.094) mm3 (t=3.444, P=0.007); the difference between the experimental group and the control group was statistically significant. Conclusion C-Jun N-terminal kinase and bax are involved in the formation of basilar artery aneurysm in rabbits. Key words: Intracranial aneurysm; c-Jun N-terminal kinase; B cell lymphoma/leukemia-2 associated X protein

Mechanisms involved in the increased sensitivity of the rabbit basilar artery to atrial natriuretic peptide in diabetes

Atrial natriuretic peptide (ANP) is a vasodilator with significant regional differences and controversial effects in the cerebral circulation, a vascular bed particularly prone to diabetes-induced complications. The present study has investigated how alloxan-induced diabetes modifies the mechanisms involved in the response of the rabbit basilar artery to ANP. ANP (10–12–10−7M) relaxed precontracted basilar arteries, with higher potency in diabetic than in control rabbits. In arteries from both groups of animals, endothelium removal reduced ANP-induced relaxations. Inhibition of NO-synthesis attenuated ANP-induced relaxation but this attenuation was lower in diabetic than in control rabbits. In control rabbits, indomethacin displaced to the left the concentration-response curve to ANP, without significantly modifying the Emax value. In diabetic rabbits, indomethacin significantly enhanced arterial relaxations to ANP. In KCl-depolarised arteries, relaxation to ANP was almost abolished both in control and in diabetic rabbits. Iberiotoxin inhibited relaxations to ANP in both groups of rabbits. Glibenclamide and 4-aminopyridine inhibited the ANP-induced relaxations more in diabetic than in control rabbits. Basilar arteries from diabetic rabbits showed decreased natriuretic peptide receptor C expression and no changes in natriuretic peptide receptor A, large conductance calcium-activated K+ channels (BKCa), ATP-sensitive K+ channels (KATP) and voltage-sensitive K+ channels (KV) expression. These results suggest that diabetes enhances the sensitivity of the rabbit basilar artery to ANP by mechanisms that at least include reduced expression of natriuretic peptide receptor C, and enhanced activity of KATP and KV channels. Furthermore, diabetes reduces endothelial NO and prostacyclin which mediate arterial relaxation to ANP.

Relaxant Effects of the Selective Estrogen Receptor Modulator, Bazedoxifene, and Estrogen Receptor Agonists in Isolated Rabbit Basilar Artery.

We have previously shown that the selective estrogen receptor modulator, bazedoxifene, improves the consequences of ischemic stroke. Now we aimed to characterize the effects and mechanisms of action of bazedoxifene in cerebral arteries. Male rabbit isolated basilar arteries were used for isometric tension recording and quantitative polymerase chain reaction. Bazedoxifene relaxed cerebral arteries, as 17-β-estradiol, 4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol [estrogen receptor (ER) α agonist], and G1 [G protein-coupled ER (GPER) agonist] did it (4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol > bazedoxifene = G1 > 17-β-estradiol). 2,3-Bis(4-hydroxyphenyl)-propionitrile (ERβ agonist) had no effect. Expression profile of genes encoding for ERα (ESR1), ERβ (ESR2), and GPER was GPER > ESR1 > ESR2. As to the endothelial mechanisms, endothelium removal, N-nitro-L-arginine methyl ester, and indomethacin, did not modify the relaxant responses to bazedoxifene. As to the K channels, both a high-K medium and the Kv blocker, 4-aminopyridine, inhibited the bazedoxifene-induced relaxations, whereas tetraethylammonium (nonselective K channel blocker), glibenclamide (selective KATP blocker) or iberiotoxin (selective KCa blocker) were without effect. Bazedoxifene also inhibited both Ca- and Bay K8644-elicited contractions. Therefore, bazedoxifene induces endothelium-independent relaxations of cerebral arteries through (1) activation of GPER and ERα receptors; (2) increase of K conductance through Kv channels; and (3) inhibition of Ca entry through L-type Ca channels. Such a profile is compatible with the beneficial effects of estrogenic compounds (eg, SERMs) on vascular function and, specifically, that concerning the brain. Therefore, bazedoxifene could be useful in the treatment of cerebral disorders in which the cerebrovascular function is compromised (eg, stroke).

Nuclear factor-κB is involved in oxyhemoglobin-induced endothelin-1 expression in cerebrovascular muscle cells of the rabbit basilar artery

The present research was designed to investigate whether endothelin-1 (ET-1) secretion can be induced by oxyhemoglobin and whether nuclear factor κB (NF-κB) is involved in the regulation of ET-1 transcription in cerebrovascular muscle cells. Cerebrovascular muscle cells isolated from a rabbit basilar artery were stimulated by oxyhemoglobin (OxyHb) and ET-1 production was increased significantly in the supernatant. Inhibition of NF-κB with pyrrolidine dithiocarbamate and small interfering RNA decreased the expression of ET-1. Nuclear translocation of NF-κB and the degradation of IkB-α was observed with the stimulation of OxyHb. The supernatant obtained from cerebrovascular muscle cells stimulated by OxyHb produced contractions in arterial rings and was blocked by the ET-1 receptor antagonist (BQ-123). The time course of the OxyHb-induced contractions of the basilar artery rings correlated with the time course of the OxyHb-induced ET-1 secretion. The contraction of the basilar artery rings induced by OxyHb was attenuated when the artery rings were preincubated with pyrrolidine dithiocarbamate and SN50 (20 and 10 µM, respectively). These results indicate that cerebrovascular muscle cells may be an important source of ET-1 production after subarachnoid hemorrhage. NF-κB was involved in the expression of ET-1 and the inhibition of the NF-κB pathway may be beneficial for the treatment of cerebral vasospasm.

Punica granatum L. Juice Attenuates Experimental Cerebral Vasospasm in the Rabbit Subarachnoid Hemorrhage Model: A Basilar Artery Morphometric Study and Apoptosis.

Background This study investigated the effect of Punica granatum L. (pomegranate) juice on the rabbit basilar artery in an experimental subarachnoid hemorrhage (SAH) model. Methods Eighteen adult male New Zealand white rabbits were randomly divided into three groups: a control group (n = 6), SAH group (n = 6), and SAH + treatment group (n = 6). Basilar artery diameter was measured with magnetic resonance angiography (MRA) in all groups at the beginning of the study. Experimental SAH was created by injecting autologous arterial blood into the cisterna magna. In the treatment group, the subjects were administered a daily dose of 30 ml/kg pomegranate juice via gastric gavage for 4 days after the SAH. The SAH group and SAH + treatment group underwent cerebral MRA after 72 hours. After a neurologic score assessment, all the animals were killed. The wall thickness and lumen area of the basilar artery were measured histometrically in all groups, and the apoptotic cell percentage in the artery was identified. The mean diameter of the basilar artery during MRA was measured. Results Pomegranate improved neurologic functions compared with the SAH group (p < 0.01). The mean basilar artery diameter on MRA in the SAH + treatment group was larger than in the SAH group and smaller than in the control group (p < 0.01 and p < 0.05, respectively). The mean vessel wall thickness value in the SAH + treatment group was lower than in the SAH group (p < 0.01), whereas there was no difference between the control and the SAH + treatment group (p > 0.05). The apoptotic cell rate in the SAH + treatment group was significantly lower than in the SAH group (p < 0.001). Evaluation of the basilar artery luminal area showed no difference between the three groups (p > 0.05). Discussion Pomegranate was shown to have a vasospasm- attenuating effect on the basilar artery in the rabbit SAH model for the first time in our study.

Upregulation of tissue inhibitor of metalloproteinase-1 contributes to restoration of the extracellular matrix in the rabbit basilar artery during cerebral vasospasm after subarachnoid hemorrhage

Vascular remodeling caused by extracellular matrix (ECM) metabolism contributes to the development of cerebral vasospasm after subarachnoid hemorrhage (SAH). The balance between tissue inhibitor of metalloproteinases (TIMPs) and matrix metalloproteinases (MMPs) plays an important role in ECM remodeling. We investigated the mechanism of vascular remodeling following cerebral vasospasm in a rabbit double hemorrhage model. Rabbit basilar arteries were harvested on days 3, 5, and 7 after initial hemorrhage. TIMP-1, TIMP-2, MMP-2, and MMP-9 mRNA and protein expression were investigated with microarray analysis, quantitative real-time PCR, immunoblot analysis, and enzyme-linked immunosorbent assay (ELISA). The expression and localization of TIMP-1, TIMP-2, MMP-2, MMP-9, elastin, fibronectin, laminin, and collagens I, III, and IV were investigated with immuohistochemical staining. After SAH, TIMP-1 mRNA and protein expression were significantly increased on day 3 and then decreased to the control level on days 5 and 7. MMP-9 protein expression was significantly increased on day 7. TIMP-2 and MMP-2 mRNA and protein expression were significantly increased on day 7. Elastin, fibronectin, laminin, and collagens I, III, and IV protein expression was decreased on day 3 and then restored to control levels on day 7. Upregulation of TIMP-1 during the early phase of cerebral vasospasm may contribute to the recovery of the ECM during the late phase of cerebral vasospasm, resulting in a protective role of TIMP-1 from cerebral vasospasm. Moreover, the increase in arterial compliance by the decrease in ECM during the early phase of cerebral vasospasm may facilitate vasoconstriction of the cerebral artery.

Abstract T P249: Upregulation of Relaxin After Experimental Subarachnoid Hemorrhage

Background and Purpose: Relaxin is a pregnancy hormone which exerts vasodilatory effects by activating nitric oxide synthase and matrix metalloproteinases through receptor (RXFP1). Recently, this vasodilatory effect has been considered useful as a potential therapeutic target in acute heart failure. However, the role of relaxin in the cerebral artery has not been fully explored. In this study, we investigated the role of relaxin in rabbit basilar artery (BA) after subarachnoid hemorrhage (SAH). Methods: Basilar arteries were collected from a rabbit SAH model on days 3, 5, and 7 after SAH. Total RNA was extracted from the BA and a cDNA microarray containing 43,623 genes was performed. The expression of relaxin and RXFP1 mRNA was investigated by quantitative RT-PCR. Serum and cerebrospinal fluid (CSF) relaxin concentrations were measured by ELISA. Perfusion fixation was performed on days 0, 3, 5 and 7, and the localization of RXFP-1 in the BA was examined using immunohistochemistry. Results: The expression of relaxin mRNA was significantly up-regulated on days 5 and 7 compared to the control sample. The expression of RXFP1 mRNA was significantly down-regulated on day 3 and sustained on day 7. The serum relaxin concentration in the rabbit SAH model was not significantly changed after SAH, whereas the CSF relaxin concentration was significantly elevated on days 5 and 7. Immunohistochemistry revealed strong RXFP1 staining in media of the BA on day 0, and showed weak staining on days 3, 5 and 7. Conclusions: Relaxin expression in the BA and relaxin concentration in the CSF increase after SAH, and considerably, relaxin expression is negatively correlated with the RXFP1 expression in the rabbit BA after SAH.

Mechanisms underlying increased vascular smooth muscle contractility in the rabbit basilar artery following subarachnoid hemorrhage.

Increased vascular contractility plays an important role in the development of cerebral vasospasm following subarachnoid hemorrhage (SAH). Here, we summarize our current knowledge regarding molecular mechanisms that contribute to increased smooth muscle contractility of rabbit basilar artery following SAH. Our studies demonstrated that upregulation of receptor expression, impairment of feedback regulation of receptor activity, and enhancement of myofilament Ca²⁺ sensitization might lead to increased smooth muscle contractility following SAH.

Gene Expression Profiling and Bioinformatic Analysis of Rabbit Basilar Artery after Experimental Subarachnoid Hemorrhage

Objective: The molecular mechanisms which contribute to the development of vascular events including cerebral vasospasm after subarachnoid hemorrhage (SAH) in cerebral artery remain to be elucidated. In this study, we investigated the time course of changes in the gene expression of cerebral artery using rabbit SAH model and performed bioinformatic analysis of differentially expressed genes. Methods: Rabbit basilar arteries were harvested on days 3, 5, and 7 after initial hemorrhage. Changes in gene expression of the rabbit basilar artery were investigated by using Agilent rabbit oligo microarrays and analyzed the data by Ingenuity Pathway Analysis (IPA). Results: Among investigated 43,623 genes, 1,121 genes were differentially expressed at least 1 time point. We found that the number, magnitude of fold change, and gene expression pattern were most dynamically changed on day 3, whereas narrowing of the basilar artery became most severe on day 5. In microarray datasets analyzed by IPA revealed that 25 biological functions identified from differentially expressed genes were significantly upregulated. Conclusion: Our findings that were based on gene expression analysis followed by bioinformatic analysis may provide a simple basis to interlink the various presumed pathogenesis of vascular events including cerebral vasospasm after SAH.

Upregulation of relaxin after experimental subarachnoid hemorrhage in rabbits.

Background. Although relaxin causes vasodilatation in systemic arteries, little is known about its role in cerebral arteries. We investigated the expression and role of relaxin in basilar arteries after subarachnoid hemorrhage (SAH) in rabbits. Methods. Microarray analysis with rabbit basilar artery RNA was performed. Messenger RNA expression of relaxin-1 and relaxin/insulin-like family peptide receptor 1 (RXFP1) was investigated with quantitative RT-PCR. RXFP1 expression in the basilar artery was investigated with immunohistochemistry. Relaxin concentrations in cerebrospinal fluid (CSF) and serum were investigated with an enzyme-linked immunosorbent assay. Using human brain vascular smooth muscle cells (HBVSMC) preincubated with relaxin, myosin light chain phosphorylation (MLC) was investigated with immunoblotting after endothelin-1 stimulation. Results. After SAH, RXFP1 mRNA and protein were significantly downregulated on day 3, whereas relaxin-1 mRNA was significantly upregulated on day 7. The relaxin concentration in CSF was significantly elevated on days 5 and 7. Pretreatment with relaxin reduced sustained MLC phosphorylation induced by endothelin-1 in HBVSMC. Conclusion. Upregulation of relaxin and downregulation of RXFP1 after SAH may participate in development of cerebral vasospasm. Downregulation of RXFP1 may induce a functional decrease in relaxin activity during vasospasm. Understanding the role of relaxin may provide further insight into the mechanisms of cerebral vasospasm.

Nitric oxide suppresses L-type calcium currents in basilar artery smooth muscle cells in rabbits

Objectives: Nitric oxide (NO) is well known to be a vasodilator, and NO donor compounds are currently used for treating vasospasm following subarachnoid hemorrhage. However, the action mechanism of cerebral vascular relaxation is not yet clear. L-type calcium channels have been determined to play an essential role in smooth muscle contraction. To investigate the role of L-type calcium channels in NO-induced relaxation of basilar smooth muscle cells, we examined the effect of the NO donor, sodium nitroprusside (SNP) on calcium (Ca2+) currents using smooth muscle cells isolated from a rabbit basilar artery.Method: The smooth muscle cells were isolated from rabbit basilar artery by enzyme treatment. To identify L-type Ca2+ currents, we used cesium chloride, a potassium channel blocker and Bay K8644, an activator of L-type Ca2+ channel.Results: The L-type calcium currents (91±13·0 pA; n = 11) were significantly reduced by SNP (32±5 pA; n = 11; P<0·05). 1H-[1,2,4] Oxadiazolo [4,3-a] quinoxalin-1-one, a 3',5'-cyclic guanosine monophosphate inhibitor, blocked the effect of SNP on L-type Ca2+ currents, and similar results were obtained after the application of 7-nitroindazole, a specific NO synthase inhibitor. Furthermore, inward currents were enhanced by Bay K8644 (170±22 pA; n = 5) and were suppressed by SNP (54±13 pA; n = 5; P<0·05).Discussion: These results demonstrate that NO suppresses the L-type Ca2+ currents in rabbit basilar smooth muscle cells, and suggest that L-type Ca2+ channels may play a pivotal role in NO-induced vascular relaxation.

Study the effects of zonisamide on fine structure of rabbit basilar artery and hippocampus in rabbit subarachnoid hemorrhage model

In this study, we investigated the effect of a novel antiepileptic drug, zonisamide (ZNS), on the basilar artery and hippocampus in a rabbit subarachnoid hemorrhage (SAH) model. Three groups of New Zealand white rabbits were used: a sham (non-SAH) group, an SAH + saline group, and SAH + drug treatment group that received ZNS. In the treatment group, the subjects were given ZNS for 3 days after the SAH. Hippocampal sections were evaluated for neural tissue degeneration. Basilar artery lumen areas and arterial wall thickness were also measured in all groups. The mean luminal area of the SAH + ZNS was significantly greater than the SAH + saline group. In addition, the arterial wall thickness of SAH + ZNS group was significantly thinner than the SAH + saline group. The neuronal degeneration scores of the hippocampal CA1 regions in the SAH + ZNS group were significantly lower than the SAH + saline treatment animals. These results indicate that ZNS has a vasodilatatory effect on the basilar artery and a neuronal protective effect in the CA1 region of the hippocampus in a rabbit SAH model.

Magnesium induced vascular relaxation and role of Calcium-dependent K+ Channels

Background and objectives: Magnesium is established as a neuro-protective agent and now also known as a vasodilator. It has been known for treating vasospasm following subarachnoid hemorrhage. However, its action mechanism in cerebral vascular relaxation is not clear. Potassium channels play a pivotal role in the relaxation of smooth muscle cells. To investigate their role in magnesium-induced relaxation of basilar smooth muscle cells, we examined the effect of magnesium on potassium channels using the patch clamp technique on cells from rabbit basilar artery. Material and Methods: Fresh smooth muscle cells were isolated from the basilar artery by enzyme treatment. Whole cell current recording was done using patch-clamp technique. Appropriate bath solution was used to have potassium current. The effect of Magnesium was observed and to identify the potassium (K+) channel involved in the magnesium-induced currents, different potassium channel blockers were used. Results: Magnesium increased the step pulse-induced outward K+ currents by more than fortyfive percent over control level (p&lt;0.01). The outward K+ current was decreased significantly by application of tetraethylammonium, a non-specific K+ channel blocker, and by iberiotoxin, a largeconductance Ca2+-activated K+ (BKCa) channel blocker, but was not inhibited by glibenclamide an ATP-sensitive K+ (KATP) channel blocker. Magnesium failed to increase the outward K+ currents in the presence of IBX. Conclusion: These results demonstrate that calcium dependent pottassium (BKCa) channels has role in magnesium induced vascular relaxation in rabbit basilar smooth muscle cells and needs to be worked out for human. DOI: http://dx.doi.org/10.3126/jmcjms.v1i1.7880 Janaki Medical College Journal of Medical Sciences (2013) Vol. 1 (1):9-13

Magnesium sulfate suppresses L-type calcium currents on the basilar artery smooth muscle cells in rabbits

Objectives: Although magnesium is a well-known treatment for vasospasm following subarachnoid hemorrhage, its mechanism of action for cerebral vascular relaxation is not clear. In addition, it is known that L-type calcium (Ca2+) channels play a pivotal role in smooth muscle contraction. To investigate the role of L-type Ca2+ channels in the magnesium-induced relaxation of basilar smooth muscle cells, we examined the effect of magnesium sulfate on L-type Ca2+ currents using freshly isolated smooth muscle cells from rabbit basilar arteries.Method: Rabbits were anesthetized with ketamine (50 mg/kg) with xylazine (25 mg/kg) and exsanguinated. Smooth muscle cells were isolated from rabbit basilar arteries by enzyme treatment. Cells were stored at 4°C before use. Whole-cell patch clamp technique was used to identify L-type Ca2+ currents, using the potassium channel blocker, cesium chloride, and nimodipine and Bay K8644 as a blocker and activator of L-type Ca2+ channels, respectively.Results: Inward currents induced by step pulses were significantly reduced by nimodipine (n = 5, P<0·05) and increased by Bay K8644 (n = 5, P<0·05). The L-type Ca2+ currents (122±14·0 pA, n = 12) were significantly reduced by the application of 5 mM magnesium sulfate (28±4 pA, n = 12, P<0·05). The inward currents enhanced by Bay K8644 were further suppressed by the application of magnesium sulfate.Discussion: These results demonstrate that L-type Ca2+ channels are functionally expressed in rabbit basilar smooth muscle cells and suggest that L-type Ca2+ channels may play a pivotal role for magnesium-induced relaxation.

Altered expression of connexin43 and its possible role in endothelin - 1 - induced contraction in rabbit basilar artery

Objective To investigate the role of connexin43 in ET- 1 -induced contraction in rabbit basilar artery.Methods The ET - 1 - induced contraction without or with carbenoxolone was studied with an isometric tension system.The expression of connexin43 protein in ET - t stimulated basilar arteries was studied with Western blot.Scrape/scratch method was used to analyze the function of gap junction in cultured rabbit cerebrovascular smooth muscle ceils.Results ET - 1 produced a concentration - dependent contraction.Carbenoxolone inhibited ET- 1 induced contraction.The connexin43 protein level was increased in ET- 1stimulated basilar arteries.Carbenoxolone decreased the connexin43 protein level increased by ET - 1.Cells treated with ET - 1 appeared positive communication and the dye transfer was increased in a time - dependent fashion.Carbenoxolone suppressed the ET - 1 - induced increasement of dye transfer.Conclusions The enhancement of gap junction intercellular communication is activated by ET - 1 via modulating the expression of connexin43,and plays an important role in the pathogenesis of cerebral vasospasm.Inhibition of vascular GJIC as a means of reducing cerebral vasospasm after SAH may have therapeutic advantage. Key words: Connexin43 ; Gap junction ; ET - 1 ; Cerebral vasospasm

Endothelial cells express a unique transcriptional profile under very high wall shear stress known to induce expansive arterial remodeling

Chronic high flow can induce arterial remodeling, and this effect is mediated by endothelial cells (ECs) responding to wall shear stress (WSS). To assess how WSS above physiological normal levels affects ECs, we used DNA microarrays to profile EC gene expression under various flow conditions. Cultured bovine aortic ECs were exposed to no-flow (0 Pa), normal WSS (2 Pa), and very high WSS (10 Pa) for 24 h. Very high WSS induced a distinct expression profile compared with both no-flow and normal WSS. Gene ontology and biological pathway analysis revealed that high WSS modulated gene expression in ways that promote an anti-coagulant, anti-inflammatory, proliferative, and promatrix remodeling phenotype. A subset of characteristic genes was validated using quantitative polymerase chain reaction: very high WSS upregulated ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motif-1), PLAU (urokinase plasminogen activator), PLAT (tissue plasminogen activator), and TIMP3, all of which are involved in extracellular matrix processing, with PLAT and PLAU also contributing to fibrinolysis. Downregulated genes included CXCL5 and IL-8 and the adhesive glycoprotein THBS1 (thrombospondin-1). Expressions of ADAMTS1 and uPA proteins were assessed by immunhistochemistry in rabbit basilar arteries experiencing increased flow after bilateral carotid artery ligation. Both proteins were significantly increased when WSS was elevated compared with sham control animals. Our results indicate that very high WSS elicits a unique transcriptional profile in ECs that favors particular cell functions and pathways that are important in vessel homeostasis under increased flow. In addition, we identify specific molecular targets that are likely to contribute to adaptive remodeling under elevated flow conditions.

Analysis of rabbit vascular responses to DBI, an ingol derivative isolated from Euphorbia canariensis.

We have analysed the effects of 7,12-O-diacetyl-8-O-benzoil-2,3-diepiingol (DBI), an ingol derivative isolated from E. canariensis, on isometric tension developed by isolated rabbit basilar and carotid arteries. Concentration-response curves to DBI (10(-8) - 3 x 10(-5) M) were obtained cumulatively in both arteries at resting tension and active tone (KCI, 50 mM). At resting tension, DBI induced a concentration-dependent contraction, which was not inhibited in Ca(2+)-free medium. H7 (1-(5-isoquinoline sulphonyl)-2-methylpiperazine dichloride) (10(-4) M) inhibited the DBI-induced contraction both in basilar and in carotid arteries. Calmidazolium (10(-4) M) inhibited the maximum contraction of the carotid artery to DBI, and completely abolished the response in the basilar artery. In pre-contracted basilar arteries DBI induced a concentration-dependent relaxation that was not modified by incubation with NG-nitro-L-arginine (L-NOARG; 10(-5) M) or indomethacin (10(-5) M). In the carotid artery with active tone DBI induced further contractions, which were not significantly modified by L-NOARG (10(-5) M) and were potentiated by indomethacin (10(-5) M). These results suggest that DBI contracts rabbit basilar and carotid arteries by a mechanism that is independent of extracellular Ca2+ and involves the participation both of protein kinase C and of calmodulin. DBI relaxes basilar but not carotid arteries by a mechanism independent of the liberation of nitric oxide and prostacyclin. In the carotid artery prostacyclin but not nitric oxide partially counteracts the contractile action of DBI.

Hypercholesterolemia increases vasospasm resulting from basilar artery subarachnoid hemorrhage in rabbits which is attenuated by Vitamin E

Background:Aneurysm rupture results in subarachnoid hemorrhage (SAH) with subsequent vasospasm in the cerebral and cerebellar major arteries. In recent years, there has been increasing evidence that hypercholesterolemia plays a role in the pathology of SAH. It is known that hypercholesterolemia is one of the major risk factors for the development of atherosclerosis. Among the factors that have been found to retard the development of atherosclerosis is the intake of a sufficient amount of Vitamin E. An inverse association between serum Vitamin E and coronary heart disease mortality has been demonstrated in epidemiologic studies. Therefore, we tested, in an established model of enhanced cholesterol feed in rabbits, the effects of hypercholesterolemia on vasospasm after SAH by using computed tomography (CT) angiograms of the rabbit basilar artery; in addition, we tested the effects of Vitamin E on these conditions, which have not been studied up to now.Methods:In this study rabbits were divided into 3 major groups: control, cholesterol fed, and cholesterol + Vitamin E fed. Hypercholesterolemia was induced by a 2% cholesterol-containing diet. Three rabbit groups were fed rabbit diet; one group was fed a diet that also contained 2% cholesterol and another group was fed a diet containing 2% cholesterol and they received i.m. injections of 50 mg/kg of Vitamin E. After 8 weeks, SAH was induced by the double-hemorrhage method and distilled water was injected into cisterna magna. Blood was taken to measure serum cholesterol and Vitamin E levels. Basilar artery samples were taken for microscopic examination. CT angiography and measurement of basilar artery diameter were performed at days 0 and 3 after SAH.Results:Two percent cholesterol diet supplementation for 8 weeks resulted in a significant increase in serum cholesterol levels. Light microscopic analysis of basilar artery of hypercholesterolemic rabbits showed disturbances in the subendothelial and medial layers, degeneration of elastic fibers in the medial layer from endothelial cell desquamation, and a reduction of waves in the endothelial layer. However, the cholesterol + Vitamin E group did not exhibit these changes. The mean diameter of the basilar artery after SAH induction in the cholesterol-treated group was decreased 47% compared with the mean diameter of the control group. This value was less affected in cholesterol + Vitamin E-treated rabbits, which decreased 18% compared with the mean diameter of the control group.Conclusions:Hypercholesterolemia-related changes in the basilar artery aggravate vasospasm after SAH. Adding Vitamin E to cholesterol-treated rabbits decreased the degree of vasospasm following SAH in the rabbit basilar artery SAH model. We suggest that Vitamin E supplements and a low cholesterol diet may potentially diminish SAH complicated by vasospasm in high-risk patients.

Aminoguanidine inhibition of iNOS activity ameliorates cerebral vasospasm after subarachnoid hemorrhage in rabbits via restoration of dysfunctional endothelial cells

Background This study was to delineate the therapeutic efficacy and potential cellular and molecular mechanisms of aminoguanidine (AG), a relatively selective inhibitor of iNOS activity, in cerebral vasospasm after subarachnoid hemorrhage (SAH) in rabbits. Methods SAH was induced by a single injection of autologous arterial blood into the cisterna magna of adult male rabbits. An intravenous bolus injection of AG (150 mg/kg) was administrated 1 h after SAH, and this dosage was repeated on the following day. Vasospasm was verified by computed tomography angiography (CTA) day 2 after SAH. Rabbit basilar arteries were harvested for transmission electron microscopy (TEM), immunohistochemical examination, RT-PCR, and western blot analysis. Results CTA data revealed that cerebral vasospasm of SAH rabbits was significantly prevented via AG treatment. TEM results demonstrated the ultrastructural morphological changes of endothelial cells of SAH rabbits were ameliorated by AG treatment. In parallel, AG treatment increased eNOS mRNA and protein levels along with the reduced immunoreactivity of nitrotyrosine in rabbit basilar arteries. Conclusions Our discovery suggested AG inhibition of iNOS activity could significantly reverse cerebral vasospasm after SAH via restoration of dysfunctional endothelial cells by the upregulation of eNOS, indicating a regulatory cross-talk between eNOS and iNOS in the pathogenesis of SAH.