Distribution of class 1 integrons among enteropathogenic Escherichia coli.

The aim of this study was to investigate the incidence of and resistance gene content of class 1 integrons among enteropathogenic Escherichia coli (EPEC) and non-EPEC and to investigate intraspecies genetic diversity of EPEC strains isolated from children with diarrhea in Iran. Twenty-eight EPEC and 16 non-EPEC strains isolated from children with diarrhea were tested for the presence of a class 1 integron associated integrase gene (int1). Sequence analysis was performed to identify the resistance gene content of integrons. Genetic diversity and cluster analysis of EPEC isolates were also investigated using enterobacterial repetitive intergenic concensus-polymerase chain reaction (ERIC-PCR) fingerprinting. Twenty-three (82%) EPEC isolates and 11 (68.7%) non-EPEC isolates harbored the int1 gene specific to the conserved integrase region of class 1 integrons. Sequence analysis revealed the dominance of dfrA and aadA gene cassettes among the isolates of both groups. ERIC-PCR fingerprinting of EPEC isolates revealed a high diversity among these isolates. The widespread distribution of 2 resistance gene families (dfrA and aadA) among both groups of EPEC and non-EPEC isolates indicates the significance of integrons in antibiotic resistance transfer among these bacteria. Furthermore, clonal diversity of EPEC isolates harbouring a class 1 integron also suggests the circulation of these mobile elements among a diverse population of EPEC in this country.

Material properties of common suture materials in orthopaedic surgery.

Suture materials in orthopaedic surgery are used for closure of wounds, repair of fascia, muscles, tendons, ligaments, joint capsules, and cerclage or tension band of certain fractures. The purpose of this study was to compare the biomechanical properties of eleven commonly used sutures in orthopaedic surgery. Three types of braided non-absorbable and one type of braided absorbable suture material with different calibers (n=77) underwent biomechanical testing for maximum load to failure, strain, and stiffness. All samples were tied by one surgeon with a single SMC (Seoul Medical Center) knot and three square knots. The maximum load to failure and strain were highest for #5 FiberWire and lowest for #0 Ethibond Excel (p<0.001). The stiffness was highest for #5 FiberWire and lowest for #2-0 Vicryl (p<0.001). In all samples, the failure of the suture material occurred at the knot There was no slippage of the knot in any of the samples tested. This data will assist the orthopaedic surgeon in selection and application of appropriate suture materials and calibers to specific tasks.

Early experience with the bifurcated Excluder endoprosthesis for treatment of the abdominal aortic aneurysm.

PURPOSE: This report describes our initial experience with the modular, bifurcated Excluder endoprosthesis and its safety and efficacy in the primary endovascular repair of infrarenal abdominal aortic aneurysms (AAAs). METHODS: AAAs (mean diameter, 58.2 +/- 14.3 mm) were repaired in 19 patients with this device between March 1999 and January 2000. The mean age of patients was 71.8 +/- 8.4 years (range, 57-86 years). This modular device was inserted through an 18F introducer sheath placed in one femoral artery, and the contralateral artery was cannulated with a 12F introducer sheath. All procedures were performed in a standard operating room with angiographic capabilities. RESULTS: Endograft deployment was successful in all patients. The average surgical time was 135 +/- 37 minutes, with a mean blood loss of 229 +/- 138 mL. In eight patients, the use of either aortic or iliac extenders was required for enhanced sealing or additional length. An external iliac artery dissection occurring at the time of endograft insertion was successfully treated with a Wallstent. Type II leaks initially found to be present by means of intraoperative completion angiography had spontaneously thrombosed by the 1-month follow-up computed tomography scan. There was one perioperative death (5.3%). Complications included superficial wound infections (n = 3) and a nonfatal myocardial infarction (n = 1). The mean length of hospital stay was 2.9 +/- 1.2 days, and only six patients required intensive care. Endoleaks were seen in four patients (21%) by means of the 30-day computed tomography scan; three of these endoleaks had spontaneously sealed at the time of the 6-month follow-up examination (5.5% 6-month endoleak rate). Aneurysm size did not increase in the patients with leaks. CONCLUSION: The Excluder endoprosthesis was an effective means of excluding an infrarenal AAA from the systemic circulation in this selected group of patients. The smaller sheath sizes may increase the pool of potential candidates. Further study of this device is warranted, and continued assessment of the long-term durability of the device will be necessary.

Reduced sarco/endoplasmic reticulum Ca(2+) uptake activity can account for the reduced response to NO, but not sodium nitroprusside, in hypercholesterolemic rabbit aorta.

BACKGROUND: Hypercholesterolemia (HC) impairs acetylcholine-induced relaxation but has little effect on that caused by the NO donor sodium nitroprusside (SNP), suggesting that acetylcholine releases less NO from the endothelium in HC. The relaxation to authentic NO gas, however, is also impaired in HC aortic smooth muscle, indicating an abnormal smooth muscle response. NO relaxes arteries by both cGMP-dependent and -independent mechanisms, and the response involves calcium (Ca(2+)) store refilling via the sarco/endoplasmic reticulum calcium ATPase (SERCA). We studied the involvement of cGMP and SERCA in the smooth muscle response to NO and SNP in HC rabbit aorta. METHODS AND RESULTS: A selective guanylyl cyclase inhibitor, 1H-[1,2,4]-oxadiazole-[4,3-a]quinoxalin-1-one, eliminated SNP-induced relaxation but only partially blocked NO-induced relaxation in both normal and HC aorta. The residual relaxation to NO was still less in HC and, in both normal and HC aorta, was abolished by concomitant administration of the SERCA inhibitor cyclopiazonic acid (CPA). In contrast, CPA did not affect SNP-induced relaxation in either normal or HC aorta. SERCA activity measured by (45)Ca(2+) uptake was markedly decreased in HC, although SERCA2 protein expression did not change significantly. CONCLUSIONS: These data suggest that NO-induced relaxation but not that to SNP is partially mediated by cGMP-independent Ca(2+) uptake into sarco/endoplasmic reticulum and that reduced sarco/endoplasmic reticulum Ca(2+) pump function can account for the impaired response to NO in HC.

Detection of isolated hook fractures 36 months after implantation of the Ancure endograft: a cautionary note.

Two cases of delayed (36-month) Ancure hook fracture are reported in patients who experienced a decrease in aneurysm size and no evidence of endoleak. Both devices used redesigned hooks and are otherwise identical to those devices currently used in clinical practice. Notably, hook fractures were not visualized on all abdominal radiographic views, nor were they noted on the final "institutional" report by the reviewing radiologist. Careful clinical follow-up with multiple-view abdominal radiographs remains essential for all patients treated with an endovascular graft, with particular attention directed to the integrity of the metal components. The broader clinical significance of this observation with respect to the Ancure endograft remains to be defined.

Endovascular revascularization of renal artery stenosis: technical and clinical results.

PURPOSE: The natural history of renal artery stenosis is progression with subsequent deterioration of kidney function and development of renovascular hypertension. Percutaneous transluminal renal angioplasty is effective in the treatment of nonostial lesions but less effective for ostial stenoses. Because of the poor technical success experienced with percutaneous transluminal renal angioplasty, stenting of ostial stenoses is becoming the standard of endovascular care. In this retrospective study we analyzed the technical and clinical outcomes after renal artery stenting in 73 consecutive patients. PATIENTS AND METHODS: From July 1992 to January 1999, 88 Palmaz stents were deployed in 85 renal artery stenoses in 73 patients, with a mean age of 67.9 +/- 9.4 years. Twelve patients (16%) underwent bilateral stent placement. Atheromatous lesions were the most prevalent (99%: 82% ostial, 16% nonostial). Most stents were implanted for suboptimal balloon dilation (52%) or dissection (24%). Mean percent stenosis was 86% +/- 12%. Renal insufficiency (creatinine level > or = 1.5 mg/dL) was present in 50 (68%) patients, and uncontrolled hypertension (systolic > or = 160 mm Hg or diastolic > or = 90 mm Hg with more than two medications) was present in 57 (78%). RESULTS: Primary technical success was achieved in 89%. At the initial procedure, three additional stents were placed for residual stenoses, and urokinase was used to treat one intraprocedural stent thrombosis, resulting in an assisted primary technical success rate of 94%. Major complications occurred in 9.1% of stents placed: access artery thrombosis (n = 4), renal artery extravasation (n = 1), renal artery thrombosis (n = 1), and hematoma requiring operation (n = 2). Long-term clinical data were available on 69 (95%) patients at 20 +/- 17 months. Overall, a significant decrease in systolic and diastolic pressures (P <.001) and reduction of medication (P <.01) were noted without a change in renal function (P = NS). Angiography was performed on 22 patients at 11.3 +/- 10.3 months for persistent or worsening renal function or hypertension or for other reasons; 10 patients had significant restenoses in 14 renal arteries. CONCLUSION: Our retrospective analysis demonstrates that endovascular stenting of renal artery stenosis in patients with poorly controlled hypertension or deteriorating renal function is a safe and effective alternative treatment to surgical management.

Resistance of endothelium-dependent relaxation to elevation of O(-)(2) levels in rabbit carotid artery.

Endogenous superoxide anion (O(-)(2)) interferes with the bioactivity of nitric oxide (NO) in endothelium-dependent arterial relaxation (EDR). Using the lucigenin chemiluminescence assay, we measured O(-)(2) in the thoracic and abdominal aortas and the carotid artery of rabbits to determine whether ambient O(-)(2) varies among the three arteries and differentially diminishes the effect of NO. Basal levels of O(-)(2) were significantly higher in carotid arteries than in the thoracic aorta [23 +/- 6.1 vs. 3.9 +/- 1.4 chemiluminescence units (CU); P < 0.05], whereas EDR in response to ACh (10(-8)-10(-5) M) was not significantly different on ANOVA. After treatment with the superoxide dismutase (SOD) inhibitor diethyldithiocarbamate (DDC; 10 mM), O(-)(2) levels were significantly elevated, becoming greater in the carotid artery and abdominal aorta than in the thoracic aorta (185 +/- 31.2 and 202 +/- 40.3 vs. 89 +/- 18 CU; P < 0.05). DDC significantly reversed EDR in the thoracic aorta but not in the carotid artery; at 10(-6) M ACh, the decrease seen with DDC was 48 +/- 6.2 vs. 6.8 +/- 8.0% of maximal relaxation in the thoracic aorta and carotid artery, respectively. In the thoracic aorta, exogenous SOD reversed the inhibition of EDR caused by DDC. Moreover, DDC/O(-)(2)-resistant EDR in the carotid artery was ablated by the addition of nitro-L-arginine methyl ester (300 microM; P < 0.05), an NO synthase inhibitor, consistent with peroxynitrite or an O(-)(2)-resistant NO donor being involved in carotid relaxation. Indeed, exogenous peroxynitrite caused similar relaxation of the carotid artery and thoracic aorta, which was unaffected by DDC. Our studies show a greater production of nitrite and O(-)(2) per unit area by the carotid artery, suggesting a greater amount of their product peroxynitrite. These findings support the hypothesis that peroxynitrite is the relaxing agent that resists high O(-)(2) in the carotid artery.

Nitric oxide is the mediator of both endothelium-dependent relaxation and hyperpolarization of the rabbit carotid artery.

It is controversial whether the endothelial cell release of nitric oxide (NO) or a different factor(s) accounts for endothelium-dependent hyperpolarization, because in many arteries endothelium-dependent relaxation and hyperpolarization resists inhibitors of NO synthase. The contribution of NO to acetylcholine-induced endothelium-dependent hyperpolarization and relaxation of the rabbit carotid artery was determined by measuring NO with electrochemical and chemiluminescence techniques. In the presence of phenylephrine to depolarize and contract the smooth muscle cells, acetylcholine caused concentration-dependent hyperpolarization and relaxation which were closely correlated to the release of NO. N(omega)-nitro-L-arginine methyl ester (30 microM) partially reduced the release of NO and caused a similar reduction in smooth muscle cell relaxation and hyperpolarization. To determine if the residual responses were mediated by another endothelium-derived mediator or NO released despite treatment with N(omega)-nitro-L-arginine methyl ester, N(omega)-nitro-L-arginine (300 microM) was added. The combined inhibitors further reduced, but did not eliminate, NO release, smooth muscle relaxation, and hyperpolarization. Hyperpolarization and relaxation to acetylcholine remained closely correlated with the release of NO in the presence of the inhibitors. In addition, the NO donor, SIN-1, caused hyperpolarization and relaxation which correlated with the concentrations of NO that it released. These studies indicate that (i) the release of NO by acetylcholine is only partially inhibited by these inhibitors of NO synthase when used even at high concentrations, and (ii) NO rather than another factor accounts fully for endothelium-dependent responses of the rabbit carotid artery.

Enhanced role of K+ channels in relaxations of hypercholesterolemic rabbit carotid artery to NO.

Endothelium-dependent relaxations to acetylcholine remain normal in the carotid artery of hypercholesterolemic rabbits, but unlike endothelium-dependent relaxations of normal rabbits, they are inhibited by charybdotoxin, a specific blocker of Ca(2+)-dependent K+ channels. Because nitric oxide (NO) is the mediator of endothelium-dependent relaxation and can activate Ca(2+)-dependent K+ channels directly or via guanosine 3',5'-cyclic monophosphate, the present study investigated the role of Ca(2+)-dependent K+ channels in relaxations caused by NO, sodium nitroprusside, and 8-bromoguanosine 3',5'-cyclic monophosphate (8-Brc-GMP) in hypercholesterolemic rabbit carotid artery. Isometric tension was measured in rabbit carotid artery denuded of endothelium from normal and hypercholesterolemic rabbits which were fed 0.5% cholesterol for 12 wk. Under control conditions, relaxations to all agents were similar in normal and hypercholesterolemic rabbit arteries. Charybdotoxin had no significant effect on relaxations of normal arteries to NO, sodium nitroprusside, or 8-BrcGMP, but the Ca(2+)-dependent K+ channel blocker significantly inhibited the relaxations caused by each of these agents in the arteries from hypercholesterolemic rabbits. By contrast, relaxations to the calcium channel blocker nifedipine were potentiated to a similar extent by charybdotoxin in both groups. In addition, arteries from hypercholesterolemic rabbits relaxed less than normal to sodium nitroprusside when contracted with depolarizing potassium solution. These results indicate that although nitrovasodilator relaxations are normal in the hypercholesterolemic rabbit carotid artery, they are mediated differently, and to a greater extent, by Ca(2+)-dependent K+ channels. These data also suggest that K+ channel-independent mechanism(s) are impaired in hypercholesterolemia.

Induction of cytochrome P4501A1 by aryl hydrocarbon receptor agonists in porcine aorta endothelial cells in culture and cytochrome P4501A1 activity in intact cells.

Endothelium is a single-cell layer lining blood vessels and constituting capillaries and could be a primary site of chemical effects in the cardiovasculature and systemically. Cytochrome P4501A1 (CYP1A1) is strongly inducible in vertebrate endothelium in vivo by aryl hydrocarbon receptor (AhR) agonists [Mol. Pharmacol. 36:723-729 (1989); Mol. Pharmacol. 41:1039-1046 (1992)]. We investigated CYP1A expression and activity in porcine aorta endothelial cells (PAEC) exposed in culture to the AhR agonists 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3,3',4,4'-tetrachlorobiphenyl (TCB), benzo[a]pyrene (BP), or beta-naphthoflavone (BNF). Immunoblotting with monoclonal anti-CYP1A1 and polyclonal anti-CYP1A1 and anti-CYP1A2 antibodies showed that CYP1A1 was induced in cultures exposed to TCDD, TCB, BP, or BNF but was not detectable in untreated or dimethylsulfoxide-exposed cultures. CYP1A1 was strongly induced at intermediate concentrations (0.1 microM or 1.0 microM) of TCB, BP, or BNF, but induction was suppressed by higher concentrations, a response not due to general toxicity; cell viability (trypan blue exclusion) was > 97% with BNF or TCB at up to 10 microM. CYP1A1 induction by TCDD was maximal at 0.3-1.0 nM. ED50 values for induction of CYP1A1 by TCDD, TCB, and BP were 0.016 nM, 3-10 nM, and 180 nM, respectively. Immunohistochemical analysis confirmed CYP1A1 induction in PAEC but also showed that only some cells in the cultures were induced. Subcellular fractionation, marker enzyme analysis, and immunoblot analysis showed that PAEC had a typical complement of microsomal electron-transport components. NADPH-cytochrome P450 reductase showed comparable rates (approximately 40 nmol/min/mg) in induced and control cultures. Cultures maximally induced by 0.1 microM TCB had microsomal CYP1A1 [ethoxyresorufin-O-deethylase (EROD)] activity averaging 25 pmol/min/mg. Addition of purified rat reductase to PAEC microsomes increased the EROD rates 3-fold. EROD rates measured in intact cells maximally induced by BP, TCB, or TCDD ranged from 15 to 30 pmol/min/mg of whole-cell protein. Methoxyresorufin O-demethylase activity induced by TCDD was 2 pmol/min/mg, i.e., < 10% of the EROD activity. In cultures in which CYP1A1 was strongly induced, CYP1A2 was not detectably expressed. The CYP1A2 inducer acenaphthylene did not induce EROD or methoxyresorufin O-demethylase in intact cells. The results show that CYP1A1 but not CYP1A2 is strongly induced in mammalian endothelial cells in culture and that CYP1A1 is active in intact cells, although the catalytic rates are low.(ABSTRACT TRUNCATED AT 400 WORDS)

Enhanced role of potassium channels in relaxations to acetylcholine in hypercholesterolemic rabbit carotid artery.

The effect of hypercholesterolemia for 10 wk on endothelium-dependent relaxations to acetylcholine was studied in isolated rings of rabbit carotid artery and abdominal aorta contracted with phenylephrine or elevated potassium. In these arteries obtained from hypercholesterolemic rabbits, endothelium-dependent relaxations to acetylcholine were not significantly different from those of normal rabbits. In normal and hypercholesterolemic arteries, partial relaxation persisted in the presence of NG-nitro-L-arginine methyl ester (L-NAME), which blocked acetylcholine-induced increases in arterial guanosine 3',5'-cyclic monophosphate (cGMP). Combined treatment with L-NAME and the calcium-dependent potassium-channel inhibitor, charybdotoxin, blocked relaxations in both groups, suggesting that L-NAME-resistant relaxations are mediated by an endothelium-derived hyperpolarizing factor. Charybdotoxin alone or depolarizing potassium had no significant effect on normal carotid artery or normal and hypercholesterolemic abdominal aorta but significantly inhibited relaxations of the carotid artery from cholesterol-fed rabbits. The enhanced role of calcium-dependent potassium channels and the hyperpolarizing factor in relaxation of the hypercholesterolemic carotid artery suggested by these results was likely related to the fact that acetylcholine failed to stimulate cGMP only in that artery. These data suggest that endothelium-dependent relaxation in these rabbit arteries is mediated by nitric oxide-cGMP-dependent and -independent mechanisms. In hypercholesterolemia, the contribution of nitric oxide-cGMP in the carotid artery is reduced, but a hyperpolarizing factor and calcium-dependent potassium channels maintain normal acetylcholine-induced relaxation.

Nitric oxide directly activates calcium-dependent potassium channels in vascular smooth muscle.

Nitric oxide is the major endothelium-derived relaxing factor (EDRF), and it is thought to relax smooth muscle cells by stimulation of guanylate cyclase, accumulation of its product cyclic GMP, and cGMP-dependent modification of several intracellular processes, including activation of potassium channels through cGMP-dependent protein kinase. Here we present evidence that both exogenous nitric oxide and native EDRF can directly activate single Ca(2+)-dependent K+ channels (K+Ca) in cell-free membrane patches without requiring cGMP. Under conditions when guanylate cyclase was inhibited by methylene blue, considerable relaxation of rabbit aorta to nitric oxide persisted which was blocked by charybdotoxin, a specific inhibitor of K+Ca channels. These studies demonstrate a novel direct action of nitric oxide on K+Ca channels.

Potassium channel-mediated relaxation to acetylcholine in rabbit arteries.

Endothelium-dependent relaxation is associated with smooth muscle hyperpolarization in many arteries which may account for relaxation that persists in the presence of nitric oxide inhibitors such as NG-nitro-L-arginine methyl ester (L-NAME). Acetylcholine (ACh)-induced relaxations of the rabbit thoracic and abdominal aorta and iliac and carotid arteries were studied for the relative contribution of nitric oxide-dependent and -independent mechanisms in rings suspended for measurement of isometric tension. Although relaxation of the thoracic aorta to ACh (10(-6) M) was almost blocked completely by L-NAME (3 x 10(-5) M), the maximal relaxation in the abdominal aorta, carotid and iliac arteries was only reduced by 28, 26 and 62%, respectively. In rings of abdominal aorta, L-NAME blocked the ACh-stimulated (10(-6) M) rise in cyclic GMP verifying that relaxation which persists in L-NAME-treated rings is not mediated by nitric oxide. The L-NAME resistant response was nearly abolished by elevated external K+ in rings of abdominal aorta and carotid artery, suggesting this relaxation may be mediated by a membrane potential sensitive mechanism. Furthermore, tetraethylammonium (10(-3) M) partially and charybdotoxin (5 x 10(-8) M) completely inhibited the remaining L-NAME-resistant relaxation in both abdominal aorta and carotid artery, suggesting a role for Ca(++)-activated K(+)-channels. Blockers of ATP-sensitive K+ channels also inhibited the L-NAME resistant relaxation in the abdominal aorta only.(ABSTRACT TRUNCATED AT 250 WORDS)

Protection of the reperfused heart by L-propionylcarnitine.

The effects of L-propionylcarnitine on mechanical function, creatine phosphate and ATP content, and lactate dehydrogenase leakage were studied in isolated perfused rat hearts exposed to global no-flow ischemia for 30 min followed by reperfusion for 20 min. Five and 10 mM L-propionylcarnitine resulted in a 100% recovery of left ventricular-developed pressure, whereas the recovery was only 40% in the hearts perfused without this agent. Ischemia-reperfusion caused a 85% loss of creatine phosphate and a 77% loss of ATP, which was prevented by 10 mM L-propionylcarnitine. Five millimolar L-propionylcarnitine protected the heart from the loss of creatine phosphate but not from the loss of ATP. Ten millimolar L-propionylcarnitine failed to improve the postischemic left ventricular-developed pressure, when it was added to the perfusate only after ischemia. L-propionylcarnitine alleviated the decrease of coronary flow in the reperfused hearts. Lactate dehydrogenase leakage was aggravated in the beginning of the reperfusion period by 10 mM L-propionylcarnitine. This adverse effect was, however, transient. L-Propionylcarnitine provides protection for the postischemic reperfused heart in a dose-dependent manner. The optimal time for administration is before the ischemic insult. High doses of this compound may perturb cell membrane integrity. Moreover, the present data point to an intracellular, metabolic, and perhaps anaplerotic mechanism of action of L-propionylcarnitine in cardiac ischemia-reperfusion injury.