Biomedical subjects

C A Gunnett

Publications and source records attributed to C A Gunnett.

10 recordsLinked to original sources

PubMed2005

Mechanisms of inducible nitric oxide synthase-mediated vascular dysfunction.

OBJECTIVE: Inducible nitric oxide synthase (iNOS) is expressed in arteries during inflammation and may contribute to vascular dysfunction. Effects of gene transfer of iNOS to carotid arteries were examined in vitro in the absence of systemic inflammation to allow examination of mechanisms by which iNOS impairs contraction and relaxation. METHODS AND RESULTS: After gene transfer of iNOS with an adenovirus (AdiNOS), constrictor responses to phenylephrine (PE) and U46619 were impaired. After AdiNOS, inhibition of soluble guanylate cyclase (sGC) with 1H-[1,2,4]oxadiazolo-[4,3,2]quinoxalin-1-one (ODQ) reduced the EC50 for PE from 4.33+/-0.78 micromol/L to 1.15+/-0.43 micromol/L (mean+/-SEM). These results imply that iNOS impairs contraction by activation of the NO/cGMP pathway. Relaxation to acetylcholine (ACh) also was impaired after AdiNOS. Sepiapterin (300 micromol/L), the precursor for tetrahydrobiopterin (BH4), improved relaxation to Ach. Because BH4 is an essential cofactor for production of NO by both iNOS and endothelial nitric oxide synthase (eNOS), these results suggest that iNOS may reduce production of NO by eNOS by limiting availability of BH4. Next, we examined effects of expression of iNOS in endothelium and adventitia. Selective expression of iNOS in endothelium, but not adventitia, impaired contraction to phenylephrine and relaxation to acetylcholine. CONCLUSIONS: We conclude that: (1) iNOS may impair contraction in part by activation of sGC; (2) iNOS impairs relaxation, at least in part, by limiting availability of BH4; and (3) expression of iNOS in endothelium may be a more important mediator of vascular dysfunction than expression of iNOS in adventitia.

Adenoviridae↗

PubMed2002

Gene transfer of inducible nitric oxide synthase impairs relaxation in human and rabbit cerebral arteries.

BACKGROUND AND PURPOSE: These studies evaluated whether gene transfer of inducible nitric oxide synthase (iNOS) is a sufficient stimulus to produce vascular dysfunction in cerebral arteries. METHODS: Intracranial (pial) arteries were dissected from human brain tissue obtained during elective surgery. Isolated human arteries were incubated in vitro with adenovirus containing iNOS (AdiNOS) or a nonexpressive transgene (control, AdBglII) (500 micro L, 3x10(9) plaque-forming units per milliliter), and vascular function was examined 24 hours later. In anesthetized rabbits, AdiNOS or AdBglII (300 microL 1x10(10)) was injected into the cisterna magna. Three days later, the basilar artery was removed, and reactivity was examined ex vivo. RESULTS: In submaximally precontracted vessels, we observed impairment of NO-dependent relaxation in human cerebral arteries after gene transfer of iNOS. Maximum relaxation to bradykinin (1 micromol/L, an endothelium-dependent agonist) was 77+/-11% (mean+/-SE) after AdBglII and 31+/-22% (P<0.05) after AdiNOS. After AdiNOS, responses to nitroprusside (an endothelium-independent NO donor) also were impaired. Responses to both nitroprusside and bradykinin were improved by aminoguanidine (300 micromol/L), an inhibitor of iNOS. AdiNOS produced no change in vasoconstrictor responses to U46619. In basilar arteries from rabbits examined in vitro after gene transfer in vivo, responses to histamine, serotonin, and nitroprusside all were similar after AdiNOS or AdBglII. In contrast, relaxation to acetylcholine was significantly depressed after AdiNOS. Maximum relaxation to acetylcholine (10 micromol/L) was 90+/-3% after AdBglII and 68+/-5% (P<0.05) after AdiNOS. Relaxation of arteries after AdiNOS was improved by aminoguanidine. CONCLUSIONS: These studies suggest that expression of iNOS may impair NO-dependent relaxation in both human and rabbit cerebral arteries.

Animals↗

PubMed2001

The future of gene therapy for stroke.

New diagnostic and treatment strategies are being developed for stroke. Gene therapy has several potential advantages over classical pharmacologic therapy. Direct administration of DNA into the brain offers the advantage of producing high concentrations of therapeutic agents in a relatively localized environment. Gene transfer also provides longer duration of effect than traditional drug therapy. Recent studies indicate that gene transfer can produce functional proteins in brain parenchyma and cerebral blood vessels after stroke. In animal models, gene transfer may reduce effects of cerebral ischemia or subarachnoid hemorrhage. This review summarizes some current methods of gene transfer to the brain and recent progress that may lead to gene therapy for stroke.

Forecasting↗

PubMed2001

NO-dependent vasorelaxation is impaired after gene transfer of inducible NO-synthase.

Proinflammatory stimuli produce expression of inducible NO-synthase (iNOS) within blood vessels and are associated with impaired endothelium-dependent relaxation. Gene transfer of iNOS was used to test the hypothesis that expression of iNOS in blood vessels produces impairment of NO-dependent relaxation as well as contraction. An adenoviral vector containing cDNA for murine iNOS, AdCMViNOS, and a control virus, AdCMVBglII, were used for gene transfer to rabbit carotid arteries in vitro and in vivo. After gene transfer of iNOS in vitro, contractile responses to KCl, phenylephrine, and U46619 were impaired. Relaxation in response to acetylcholine, ADP, A23187, and nitroprusside was also impaired. For example, maximum relaxation of vessels to acetylcholine (10 micromol/L) was 78+/-4% (mean+/-SE) after AdBglII (10(10.5) plaque-forming units) and 34+/-5% after AdiNOS (10(10.5) plaque-forming units, P<0.05). NO-independent relaxation in response to 8-bromo-cGMP and papaverine was not impaired after AdiNOS. Contraction and relaxation were improved in carotid arteries expressing iNOS by aminoguanidine and L-N-iminoethyl lysine, inhibitors of iNOS. After intraluminal gene transfer of iNOS in vivo, contraction of vessels in vitro was normal, but responses to acetylcholine were impaired. In summary, the major finding is that NO-dependent relaxation is impaired in arteries after gene transfer of iNOS in vitro and in vivo. Thus, expression of iNOS per se impairs NO-dependent relaxation.

Adenoviridae↗

PubMed2000

IL-10 deficiency increases superoxide and endothelial dysfunction during inflammation.

Little is known about the role of interleukin-10 (IL-10), an anti-inflammatory cytokine, in blood vessels. We used IL-10-deficient mice (IL-10 -/-) to examine the hypothesis that IL-10 protects endothelial function after lipopolysaccharide (LPS) treatment. The responses of carotid arteries were studied in vitro 6 h after injection of a relatively low dose of LPS (10 microgram ip). In IL-10 -/- mice, the maximum relaxation to ACh (3 microM) was 56 +/- 6% (means +/- SE) after LPS injection and 84 +/- 4% after vehicle injection (P < 0.05). Thus endothelium-dependent relaxation was impaired in carotid arteries from IL-10 -/- mice after LPS injection. In contrast, this dose of LPS did not alter relaxation to ACh in vessels from wild-type (IL-10 +/+) mice. Relaxation to nitroprusside and papaverine was similar in arteries from both IL-10 -/- and IL-10 +/+ mice after vehicle or LPS injection. Because inflammation is associated with increased levels of reactive oxygen species, we also tested the hypothesis that superoxide contributes to the impairment of endothelial function by LPS in the absence of IL-10. Results using confocal microscopy and hydroethidine indicated that levels of superoxide are elevated in carotid arteries from IL-10 -/- mice compared with IL-10 +/+ mice after LPS injection. The impaired relaxation of arteries from IL-10 -/- mice after LPS injection was restored to normal by polyethylene glycol-suspended superoxide dismutase (50 U/ml) or allopurinol (1 mM), an inhibitor of xanthine oxidase. These data provide direct evidence that IL-10 protects endothelial function after an acute inflammatory stimulus by limiting local increases in superoxide. The source of superoxide in this model may be xanthine oxidase.

Animals↗

PubMed2000

Tumor necrosis factor-alpha impairs contraction but not relaxation in carotid arteries from iNOS-deficient mice.

We used mice deficient in expression of inducible NO synthase (iNOS -/-) to directly examine the role of iNOS in impaired vasoconstrictor responses following tumor necrosis factor-alpha (TNF-alpha). In iNOS +/+ mice, contraction of carotid arteries in response to prostaglandin F(2alpha) (PGF(2alpha)) was impaired following TNF-alpha (100 microg/kg ip)(n = 10, P < 0.01). In contrast to responses in wild-type mice, contraction to low concentrations of PGF(2alpha) were normal, but maximum contraction to PGF(2alpha) was impaired in arteries from iNOS -/- mice treated with TNF-alpha [0.35 +/-.0.02 g (n = 8) following vehicle and 0.25 +/- 0.02 g (n = 7) following TNF-alpha (P < 0.05)]. Aminoguanidine, a relatively selective inhibitor of iNOS, partially restored contraction to PGF(2alpha) in vessels from iNOS +/+ mice but had no effect in iNOS -/- mice injected with TNF-alpha, suggesting that a mechanism(s) other than iNOS contributes to impaired responses. In contrast to contractile responses, relaxation of the carotid artery in response to acetylcholine and nitroprusside was not altered following TNF-alpha in iNOS +/+ or iNOS -/-mice. Responses of carotid arteries from iNOS -/- mice and effects of aminoguanidine suggest that both iNOS-dependent and iNOS-independent mechanisms contribute to impaired contractile responses following TNF-alpha.

Acetylcholine↗

PubMed1999

Vascular effects of lipopolysaccharide are enhanced in interleukin-10-deficient mice.

BACKGROUND AND PURPOSE: The role in blood vessels of interleukin-10 (IL-10), a potent anti-inflammatory cytokine, is not known. Using mice with targeted deletion of the gene for IL-10 (IL-10(-/-)), we examined the hypothesis that IL-10 is a major modulator of the vascular effects of lipopolysaccharide (LPS). Methods-We examined in vitro responses of carotid arteries obtained from wild-type (129/SvEv or C57BL/6; IL-10(+/+)) and IL-10-deficient mice 6 hours after injection of a relatively low dose of LPS (10 microgram). RESULTS: Contraction of the carotid artery in response to U46619 was impaired in IL-10-deficient mice treated with LPS compared with LPS-treated controls. After LPS, U46619 (0.03 and 0.1 microgram/mL) contracted the carotid artery by 0.11+/-0.02 (mean+/-SEM) and 0.38+/-0.03 g in wild-type (n=10) and 0.03+/-0.01 and 0.19+/-0.03 g in IL-10-deficient (n=8) mice (P<0.05 versus control). Aminoguanidine, an inhibitor of inducible nitric oxide synthase (iNOS), had no significant effect on contraction of the carotid artery from LPS-treated control mice but restored contraction of the carotid artery in response to U46619 in IL-10-deficient mice to levels seen in wild-type mice. Similar findings were obtained when phenylephrine was used as a vasoconstricting agent. These findings indicate that LPS produces much greater impairment of contractile responses of the carotid artery in IL-10-deficient mice than in control mice. Impaired contractile function was eliminated by aminoguanidine, suggesting that expression of iNOS is enhanced in arteries from IL-10-deficient mice. In carotid arteries from animals injected with LPS, reverse transcription-polymerase chain reaction (RT-PCR) products for iNOS were found more frequently in IL-10-deficient mice than in wild-type mice. RT-PCR products for iNOS were not present in arteries from vehicle-treated animals (IL-10-deficient or wild-type mice). CONCLUSIONS: This is the first evidence that endogenous IL-10 is a major determinant of the effects of LPS on vascular tone. The results suggest that impaired constrictor responses of the carotid artery after LPS in IL-10-deficient mice are mediated by enhanced expression of iNOS.

Animals↗

PubMed1998

Vascular effects of LPS in mice deficient in expression of the gene for inducible nitric oxide synthase.

The inducible isoform of nitric oxide synthase (iNOS) is expressed after systemic administration of lipopolysaccharide (LPS). The importance of expression of iNOS in blood vessels is poorly defined. Because nitric oxide from iNOS may alter vasomotor function, we examined effects of LPS on vasomotor function in carotid arteries from iNOS-deficient mice. We studied contraction of the carotid artery from wild-type and iNOS-deficient mice in vitro 12 h after injection of LPS (20 mg/kg ip). Contractile responses to PGF2alpha (3-30 microM) and thromboxane A2 analog (U-46619; 3-100 nM) were evaluated using vascular rings from mice treated with vehicle or LPS. Maximum force of contraction generated by rings in response to PGF2alpha was 0.39 +/- 0.02 and 0.25 +/- 0.01 (SE) g (n = 14) in vehicle and LPS-treated wild-type mice, respectively (P < 0.001 vs. vehicle). Thus LPS reduced constrictor responses in wild-type mice. Thiocitrulline and aminoguanidine (inhibitors of iNOS) improved contractile responses from LPS-treated wild-type vessels. Indomethacin also improved constrictor responses in arteries from wild-type mice injected with LPS. In contrast, contraction of the carotid arteries in response to PGF2alpha and U-46619 was not impaired in LPS-treated iNOS-deficient mice, and contraction was not altered by inhibitors of iNOS. Expression of iNOS mRNA was confirmed using RT-PCR in carotid arteries from wild-type mice after injection of LPS but not vehicle. PCR products for iNOS were not observed in iNOS-deficient mice. These findings provide the first direct evidence that iNOS mediates impairment of vascular contraction after treatment with LPS.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗

PubMed1994

High fish oil diet increases oxidative stress potential in mammary gland of spontaneously hypertensive rats.

1. The purpose of this study was to determine whether high omega-3 (19% menhaden oil, 1% corn oil) or high omega-6 (20% corn oil) fatty acid diets would decrease expression of hypertension in the female spontaneously hypertensive rat (SHR), promote tumourigenesis in the rat 7,12-dimethylbenz[a]anthracene (DMBA) model of mammary cancer or increase the susceptibility of the mammary gland to lipid peroxidation. A group of rats on a 5% corn oil diet served as the low fat control group. 2. We found that the high omega-3 and high omega-6 fatty acid diets did not significantly decrease mean arterial pressure. Marked differences occurred between the effects of omega-3 and omega-6 high fatty acid diets on baseline oxidation, auto-oxidation and iron-ascorbate catalyzed oxidation. The omega-3 diet showed 675% increase in basal oxidation, a 2624% increase in auto-oxidation and a 4244% increase in iron-ascorbate catalyzed oxidation compared to the omega-6 diet in mammary tissue homogenates. Although all rats were given 5 mg DMBA (i.g.), no mammary tumours were observed in any of the dietary groups. 3. We conclude that: (i) high polyunsaturated fatty acid diets do not decrease blood pressure in the female SHR; (ii) high fish oil diet markedly increases oxidative potential in the mammary gland; and (iii) the female SHR is resistant to DMBA-induced tumourigenesis.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1993

The effects of patterned calorie-restricted diets on mammary tumor incidence and plasma endothelin levels in DMBA-treated rats.

Chronic caloric restriction has been shown to inhibit mammary tumor promotion in the 7,12-dimethyl-benz[a]anthracene (DMBA) rat mammary tumor model. The objectives of this study were to determine (i) the effects of chronic caloric cycling (yo-yo dieting) on mammary tumor promotion by high fat diets and (ii) the effect of three dietary regimens +/- superimposed mammary tumor burden on plasma endothelin-1,2 (ET) levels. Female Sprague-Dawley rats were treated with DMBA (5 mg/rat) and divided into three dietary groups: ad libitum (AL) (containing 15% corn oil); 40% calorie restricted (CR) (containing 20% corn oil so consumption of fat was equivalent between AL and CR); a calorie cycled (CC) group fed alternatively AL and CR diets each 48 h period. After 10 weeks, tumor incidences were: AL, 63%; CR, 27%; CC, 57% (AL versus CR, P < 0.05; CC versus CR, P < 0.05; AL versus CC, NSD). ET levels (pg/ml plasma) were: AL, 16.0 +/- 6.54; CR, 32.31 +/- 0.34; CC, 23.44 +/- 5.04 (AL versus CR, P < 0.01; CC versus CR, P < 0.01; AL versus CC, P < 0.05). Plasma ET levels were independent of tumor incidence and tumor burden, but plasma ET levels were significantly increased in rats with a prior history of calorie restriction. As expected, maintained caloric restriction reduced mammary tumor incidence but intermittent caloric restriction (caloric cycling or yo-yo dieting) was without similar benefit.

9,10-Dimethyl-1,2-benzanthracene↗