Residency training in robotic surgery: The role of simulation.

Simulation has become increasingly important in surgical teaching in recent years and the French National Authority for Health (HAS) recently underlined the goal and ethical priority: "never the first time on the patient". Simulation programs have been tested and validated for laparotomy and for laparoscopy, but there is not yet a validated program specific for robotic surgery. Due to substantial advances in this new technology, we have developed a program in Nancy dedicated to outside-the-operating room (OR) teaching of robotic surgery using the Da Vinci robot. This teaching is based on a combined program of theoretical teaching (e-learning) and learning of practical skills using virtual simulators (DV-Trainer®, Robotic Mentor®, DVSS®), mechanical simulators (Dome, Applied® abdominal simulators), microsurgery and wet lab using ex vivo animal organs, anesthetized animals, and cadavers. This program also emphasizes team training. The course is intended for residents in surgical training and is integrated into the specialized study diploma (DES) program for Visceral and Digestive Surgery; it also can be used by qualified surgeons who can integrate it with the Inter-University Diploma (DIU) in General Robotic Surgery for basic techniques and also for DIUs in other surgical specialties (digestive and gynecologic surgery) for robotic uses within their specialty. These courses are based on the concept of step-by-step skills acquisition and verification allowing a transition to safe clinical activity.

The role of oxidative damage in poor scalp health: ramifications to causality and associated hair growth.

The oxidative stress element of unhealthy scalp leads to compromised pre-emergent hair formation and poorly formed hair as it grows. Only cosmetic solutions can minimize the impact of unhealthy hair and to achieve healthy looking and feeling hair, the scalp health must be normalized first. The objectives of this research were to both investigate whether oxidative stress was a relevant aetiological element in scalp dandruff and seborrhoeic dermatitis and whether scalp condition affects the quality of hair that grows from it. Further, this research was designed to determine whether an effective anti-dandruff shampoo would repair and protect the scalp and pre-emergent hair from oxidative stress. This study demonstrated that oxidative stress is an aetiological element relevant to the dandruff condition and that a potentiated ZPT shampoo effectively improves scalp condition, including a reduction in oxidative stress. The compromised hair condition associated with dandruff is concomitantly improved when the scalp condition is improved. It appears that there is a direct link between hair quality and scalp health.

An angiosperm-wide analysis of the gynodioecy-dioecy pathway.

BACKGROUND AND AIMS: About 6 % of an estimated total of 240 000 species of angiosperms are dioecious. The main precursors of this sexual system are thought to be monoecy and gynodioecy. A previous angiosperm-wide study revealed that many dioecious species have evolved through the monoecy pathway; some case studies and a large body of theoretical research also provide evidence in support of the gynodioecy pathway. If plants have evolved through the gynodioecy pathway, gynodioecious and dioecious species should co-occur in the same genera. However, to date, no large-scale analysis has been conducted to determine the prevalence of the gynodioecy pathway in angiosperms. In this study, this gap in knowledge was addressed by performing an angiosperm-wide survey in order to test for co-occurrence as evidence of the gynodioecy pathway. METHODS: Data from different sources were compiled to obtain (to our knowledge) the largest dataset on gynodioecy available, with 275 genera that include at least one gynodioecious species. This dataset was combined with a dioecy dataset from the literature, and a study was made of how often dioecious and gynodioecious species could be found in the same genera using a contingency table framework. KEY RESULTS: It was found that, overall, angiosperm genera with both gynodioecious and dioecious species occur more frequently than expected, in agreement with the gynodioecy pathway. Importantly, this trend holds when studying different classes separately (or sub-classes, orders and families), suggesting that the gynodioecy pathway is not restricted to a few taxa but may instead be widespread in angiosperms. CONCLUSIONS: This work complements that previously carried out on the monoecy pathway and suggests that gynodioecy is also a common pathway in angiosperms. The results also identify angiosperm families where some (or all) dioecious species may have evolved from gynodioecious precursors. These families could be the targets of future small-scale studies on transitions to dioecy taking phylogeny explicitly into account.

ShearWave™ Elastography BE1 multinational breast study: additional SWE™ features support potential to downgrade BI-RADS®-3 lesions.

PURPOSE: To determine the benefit of ShearWave™ Elastography (SWE™) in the ultrasound characterization of BI-RADS® 3 breast lesions in a diagnostic population. MATERIALS AND METHODS: 303 BI-RADS® 3 lesions (mean size: 13.2 mm, SD: 7.5 mm) from the multicenter BE1 prospective study population were analyzed: 201 (66%) had cytology or core biopsy, and the remaining 102 had a minimum follow-up of one year; 8 (2.6%) were malignant. 7 SWE features were evaluated with regard to their ability to downgrade benign BI-RADS® 3 masses. The performance of each SWE feature was assessed by evaluating the number of lesions correctly reclassified and the impact on cancer rates within the new BI-RADS® 3' lesion group. RESULTS: No malignancies were found with an E-color "black to dark blue", which allowed the downgrading of 110/303 benign masses (p < 0.0001), with a non-significant increase in BI-RADS® 3' malignancy rate from 2.6% to 4.1%. E-max ≤ 20 kPa (2.6 m/s) was able to downgrade 48/303 (p < 0.0001) lesions with a lower increase in BI-RADS® 3' malignancy rate (3.1%). No other SWE features were useful for reclassifying benign BI-RADS® 3 lesions. CONCLUSION: Applying simple reclassification rules, SWE assessment of the maximum stiffness of lesions allowed the downgrading of a sub-group of benign BI-RADS® 3 lesions. This was accompanied by a non-significant increase in the malignancy rate in the new BI-RADS® 3 class.

Serum cardiac troponin I concentrations in cattle with cardiac and noncardiac disorders.

BACKGROUND: Making a clinical diagnosis of pericarditis in cattle is difficult and additional diagnostic tests are needed to evaluate cattle with suspected pericarditis. Serum cardiac troponin I (cTnI) concentrations are increased in cattle with pericarditis, but the utility of measuring serum cTnI concentrations in cattle with suspected pericarditis in cattle remains unclear. OBJECTIVES: To determine if serum cTnI concentrations in cattle can be used to differentiate pericarditis from other cardiac disorders and noncardiac thoracic diseases. ANIMALS: Seventy-seven clinically diseased cattle and 19 healthy control cattle. METHODS: Serum cTnI concentrations were measured using an Immunlite Troponin I immunometric chemiluminescent assay in consecutive cases of postmortem-confirmed pericarditis (n=18), endocarditis (n=15), chronic suppurative pneumonia (n=13), congenital heart disease (n=10), reticulitis (n=3), mediastinal abscess (n=7), thymic lymphoma (n=6), and caudal vena cava thrombosis (n=5). Serum cTnI concentrations were measured in 19 healthy cattle. RESULTS: Although serum cTnI concentrations were significantly higher in cattle with pericarditis compared with healthy cattle, they were not significantly different from concentrations in cattle with endocarditis, congenital cardiac disease, mediastinal abscess, reticulitis, caudal vena cava thrombosis, or chronic suppurative pneumonia. CONCLUSIONS: Serum cTnI cannot be used to distinguish cattle with pericarditis from cattle with other primary cardiac diseases. In addition, serum cTnI concentrations cannot distinguish between cattle with primary cardiac diseases and those with other noncardiac, intrathoracic disorders.

Chronic decrease in flow contributes to heart failure-induced endothelial dysfunction in rats.

Chronic heart failure (CHF) impairs endothelium-dependent, nitric oxide (NO)-mediated dilation. This decreased dilation may be partly secondary to the chronic decrease in blood flow, but this hypothesis has not yet been tested. Thus, we assessed whether a localized, chronic increase in blood flow in vivo reverses endothelial dysfunction of small arteries in rats with CHF. Two months after coronary artery ligation or sham surgery, second-order side branches of the superior mesenteric artery were ligated in order to obtain persistently elevated blood flow (HF) in the adjacent first-order side branch compared with normal vessels (NF). One month later, responses to acetylcholine and flow-mediated vasodilatation (FMD) were assessed in vitro in an arteriograph. Chronic heart failure induced a decrease in mesenteric blood flow (374 +/- 25 and 305 +/- 27 micro L/min for sham and CHF, respectively; P < 0.05). Neither CHF nor the chronic increase in flow affected the responses to acetylcholine. Chronic heart failure decreased FMD (maximal response in sham and control 34 +/- 6 and 13 +/- 4%, respectively; P < 0.05). Chronic increases in blood flow did not modify FMD in sham, but restored FMD in CHF rats (28 +/- 4%; P < 0.05 vs CHF NF). The restored response was abolished by an inhibitor of NO synthesis (N(G)-nitro-l-arginine). Chronic heart failure did not affect the abundance of mesenteric endothelial NO synthase (eNOS) mRNA. A chronic increase in flow significantly increased the abundance of eNOS mRNA in sham rats, but only moderately and non-significantly in CHF rats. Thus, endothelial dysfunction of small arteries in CHF appears to be largely the consequence of the chronic decrease in flow.

Interest of the association clonidine-spironolactone in cirrhotic patients with ascites and activation of sympathetic nervous system.

BACKGROUND: The aim of this study was to examine the effects of spironolactone, clonidine and the association of clonidine-spironolactone on renin-aldosterone and sympathetic systems, renal function, systemic hemodynamics and mobilization of ascites in 32 alcoholic cirrhotic patients with marked increase in sympathetic system. METHODS: Measurements were taken before and after an 8-day treatment with spironolactone (200 mg/day), after an 8-day treatment with clonidine (0.150 mg/day) and 10 days after adjunction of spironolactone (200 mg/day) to clonidine. RESULTS: Three patients abandoned the treatment or were excluded because lack of compliance. Spironolactone alone induced an increase in renin-aldosterone and sympathetic systems without any remarkable increase of natriuresis and body weight loss. Given for 8 days, clonidine alone induced a significant decrease in plasma norepinephrine associated with a significant increase in glomerular filtration rate without effect on natriuresis. In contrast, 10 days after adjunction of spironolactone to clonidine, plasma renin and aldosterone significantly decreased, natriuresis increased (from 7.4 +/- 0.7 to 41.6 +/- 3.2 mEq/24 h) and body weight decreased (from 66.03 +/- 2.3 to 63.5 +/- 2.3 kg) without adverse effects. CONCLUSION: In cirrhotic patients with ascites and marked activation of sympathetic nervous system, spironolactone (200 mg/day) is unable to mobilize ascites. In these patients, after 8 days, clonidine decreases sympathetic activity, increases glomerular filtration rate and after 18 days, decreases plasma renin and aldosterone concentrations allowing a better action of spironolactone. The association clonidine-spironolactone enhances natriuresis and body weight loss.

Lack of impairment of nitric oxide-mediated responses in a rat model of high-renin hypertension.

1. Angiotensin (Ang) II triggers the expression of a pro- oxidant phenotype in the vascular wall, suggesting that activation of the renin-angiotensin system (RAS) causes endothelial dysfunction in various pathological situations, such as hypertension. However, this hypothesis has been mostly tested in a setting of exogenous administration of AngII. 2. We tested the hypothesis of a role for endogenous activation of the RAS leading to oxidant stress and endothelial dysfunction in a high-renin model of hypertension (i.e. two-kidney, one-clip hypertension) in rats. One month after clipping or sham surgery, aorta were isolated from untreated rats or rats treated by the angiotensin AT1 receptor antagonist irbesartan (10 mg/kg per day). Mesenteric artery segments were also isolated from normotensive or hypertensive rats. 3. Hypertension reduced the relaxations to acetylcholine but did not affect the ratio of contractions to phenylephrine in the presence compared with the absence of a nitric oxide (NO) synthase inhibitor, used as an index of basal release of NO. 4. The free radical scavenger tempol reduced the contractions to phenylephrine in the absence, but not in the presence, of an inhibitor of NO synthesis. This index of free radical-mediated degradation of NO was not affected by hypertension. In parallel, hypertension did not affect the expression of p22phox, a component of the free radical generating enzyme reduced nicotinamide adenine dinucleotide phosphate oxidase. 5. Chronic treatment with the AT1 receptor antagonist decreased blood pressure, moderately improved the response to acetylcholine, but did not affect basal NO release in hypertensive rats, although it did increase basal NO release in normotensive rats. 6. Thus, this model of hypertension is characterized by an impaired stimulated NO release but not of basal NO release in isolated arteries. Furthermore, there was no functional evidence of an increased oxidative stress-mediated impairment of NO release. This is not in favour of a direct link between activation of the RAS and development of endothelial dysfunction in experimental hypertension.

[Long term surveillance of polyps and colorectal cancers]. / Surveillance à long terme des polypes et cancers colorectaux.

It is necessary to assume the colorectal cancer follow-up after curative operation to detect cancer recurrence and new polyps or cancers. A good follow-up by endoscopy is also necessary for patients with colorectal polyps and for patients with familial adenomatous polyposis.

[Biological factors influencing response to diuretics in patients with cirrhosis and ascites]. / Facteurs biologiques influençant la réponse aux diurétiques des malades atteints de cirrhose en décompensation ascitique.

PURPOSES: To examine the biological factors influencing response to diuretics in patients with cirrhosis and ascites. METHODS: Sixty-nine patients were evaluated. Patients were classified into 3 groups: group 1: "good responders" (responding to spironolactone 200 mg/day), group 2: "bad responders" (responding to spironolactone doses above 200 mg/day or requiring addition of furosemide), and group 3: "non-responders" (not responding to spironolactone 400 mg/day and furosemide 160 mg/day). RESULTS: There were 30 patients in group 1, 24 in group 2 and 15 in group 3. The degree of activation of the renin-aldosterone and sympathetic system in group 2 was significantly higher than in group 1 and lower than in group 3. Natriuresis in group 2 (11 +/- 0.7 mEq/24h) was significantly below group 1 (20 +/- 2 mEq/24h) and above group 3 (5 +/- 0.6 mEq/24h). CONCLUSIONS: In patients with cirrhosis and ascites, the degree of activation of the renin-aldosterone and sympathetic nervous system influences diuretic response of ascites and is estimated by measured baseline natriuresis.

Cholesterol regulates membrane binding and aggregation by annexin 2 at submicromolar Ca(2+) concentration.

Annexin 2 is a member of the annexin family which has been implicated in calcium-regulated exocytosis. This contention is largely based on Ca(2+)-dependent binding of the protein to anionic phospholipids. However, annexin 2 was shown to be associated with chromaffin granules in the presence of EGTA. A fraction of this bound annexin 2 was released by methyl-beta-cyclodextrin, a reagent which depletes cholesterol from membranes. Restoration of the cholesterol content of chromaffin granule membranes with cholesterol/methyl-beta-cyclodextrin complexes restored the Ca(2+)-independent binding of annexin 2. The binding of both, monomeric and tetrameric forms of annexin 2 was also tested on liposomes of different composition. In the absence of Ca(2+), annexin 2, especially in its tetrameric form, bound to liposomes containing phosphatidylserine, and the addition of cholesterol to these liposomes increased the binding. Consistent with this observation, liposomes containing phosphatidylserine and cholesterol were aggregated by the tetrameric form of annexin 2 at submicromolar Ca(2+) concentrations. These results indicate that the lipid composition of membranes, and especially their cholesterol content, is important in the control of the subcellular localization of annexin 2 in resting cells, at low Ca(2+) concentration. Annexin 2 might be associated with membrane domains enriched in phosphatidylserine and cholesterol.

[Diagnosis and treatment of gastroesophageal reflux in the adult: guidelines recommended by French and Belgian consensus]. / Diagnostic et traitement du reflux gastro-oesophagien de l'adulte: les orientations suggérées par les consensus français et belge.

Usual gastroesophageal reflux (GER) presentations are heartburn and acid regurgitation. The prevalence in occidental population ranges from 5 to 45% according to symptoms frequency. Oesophagitis is observed in 30 to 50% of examined patients and only erosive and ulcerative lesions must be considered. Distinction is made between non-severe oesophagitis (isolated loss of substance), severe oesophagitis (circonferential loss of substance) and complicated oesophagitis (stenosis, ulcerations, brachyoesophagus). 24-hour pH-monitoring analyses reflux duration and relations between symptoms and reflux specially in unusual extraoesophageal presentations. Symptoms and quality of life are the main criteria for staging. In few patients, oesophagitis is severe. Complications (stenosis, ulcerations, bleeding, endobrachyoesophagus) are observed in 10 to 15% of cases. Endobrachyoesophagus with intestinal metaplasia is a risk for neoplasia. The consensus conference proposes this initial therapeutic strategy. In cases of time-spaced symptoms: antiacids, alginic acid or low doses of anti-H2 with life style changes. In cases of typical frequent symptoms, in patients younger than 50 years: 4-weeks treatment with half dosed proton pump inhibitors (PPI) or standard doses of anti-H2 or prokinetics. Nowadays, the majority of the experts propose empiric full-dose treatment. This attitude is more logical as total symptoms suppression with full dose PPI brings positive clues for exact GOR diagnostic without endoscopy. In patients older than 50 years or with alarming symptoms (weight loss, dysplagia, bleeding, anemia): endoscopy must be performed. Patients with non severe oesophagitis: PPI without checking endoscopy. In patients with severe or complicated oesophagitis: 8-weeks treatment following by endoscopy; in non relieved patients: doses are increased. In cases of extraoesophageal presentations: standard PPI treatment during 4 to 8 weeks if GER is well established. In long term strategy, if recidives are rare: intermittent treatment. In early and frequent recidives: long term adapted PPI or surgery. Stenosis are treated by PPI, pneumatic dilatation or surgery if unsuccessful. Brachyoesophagus must be checked by endoscopy every 2 years (malignancy risk).

Effects of combination of low doses of angiotensin-converting enzyme inhibitor and diuretics on renal function in spontaneously hypertensive rats: comparison between acute and chronic treatment.

The goal of this study was to assess the effect of acute or chronic treatment with S5590, a combination of the angiotensin-converting enzyme inhibitor perindopril (0.76 mg/kg/day) and the diuretic indapamide (0.24 mg/kg/day) on renal function in spontaneously hypertensive rats with moderate renal injury. Renal function was evaluated in conscious rats by clearance methods using labelled inulin and PAH, after catheterisation of the carotid artery, jugular vein and bladder. Both acute and chronic treatment normalised renal vascular resistance, although the effect on blood pressure was more marked after chronic than after acute treatment. Although acute treatment with S5590 increased glomerular filtration rate and renal blood flow, chronic treatment did not affect these parameters. Diuresis and natriuresis were only slightly modified and the results suggest a marked renal vasodilatation. In conclusion, the maintenance of renal function after chronic treatment, in a setting of normalisation of arterial pressure, suggest that such a combined treatment may exert marked renal functional protective effects in hypertension.

N-Terminal domain of annexin 2 regulates Ca(2+)-dependent membrane aggregation by the core domain: a site directed mutagenesis study.

Annexin 2 binds and aggregates biological membranes in a Ca(2+)-dependent manner. This protein exists as a monomer (p36) or as a heterotetramer (p90) in which two p36 chains are associated with a dimer of p11, a member of the S100 protein family. Protein kinase C phosphorylates the protein at the level of the N-terminal tail on serines 11 and 25, thereby modifying its oligomeric structure and its properties of membrane aggregation. To analyze these effects, the properties of a series of mutants in which serines 11 and 25 were replaced by alanine and/or glutamic acid were investigated. The affinity for p11 light chain was decreased in the S11E mutants. Glutamic acid residues in positions 11 or 25 did not change membrane binding, either in the tetrameric or in the monomeric form. On the other hand, these mutations affected the aggregation properties of the two forms. For the tetramer, the aggregation efficiency was decreased but not the Ca(2+) sensitivity, whereas the latter was affected in the case of the monomer. The effects were stronger in the S11E mutants, and they were cumulative in the double mutant. They suggest a different conformation of the N-terminal domain in the mutants (and in the phosphorylated protein), a hypothesis which is supported by proteolysis experiments. This conformational change would affect aggregation by the monomer through a dimerization step.

Constitutive phosphorylation of the vesicular inhibitory amino acid transporter in rat central nervous system.

gamma-Aminobutyric acid (GABA) and glycine are stored into synaptic vesicles by a recently identified vesicular inhibitory amino acid transporter [VIAAT, also called vesicular GABA transporter (VGAT)]. Immunoblotting analysis revealed that rat brain VIAAT migrated as a doublet during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with a predominant slower band in all areas examined except olfactory bulb and retina. The slower band corresponded to a phosphorylated form of VIAAT as it was converted to the faster one by treating brain homogenates with alkaline phosphatase or with an endogenous phosphatase identified as type 2A protein-serine/threonine phosphatase using okadaic acid. In contrast, the recombinant protein expressed in COS-7 or PC12 cells co-migrated with the faster band of the brain doublet and was insensitive to alkaline phosphatase. To investigate the influence of VIAAT phosphorylation on vesicular neurotransmitter loading, purified synaptic vesicles were treated with alkaline phosphatase and assayed for amino acid uptake. However, neither GABA nor glycine uptake was affected by VIAAT phosphorylation. These results indicate that VIAAT is constitutively phosphorylated on cytosolic serine or threonine residues in most, but not all, regions of the rat brain. This phosphorylation does not regulate the vesicular loading of GABA or glycine, suggesting that it is involved at other stages of the synaptic vesicle life cycle.

Selective endothelin-A versus combined endothelin-A/endothelin-B receptor blockade in rat chronic heart failure.

BACKGROUND: The relative efficacy of endothelin-A (ET(A)) receptor blockade versus combined ET(A)-ET(B) receptor blockade in chronic heart failure (CHF) is still largely unknown. METHODS AND RESULTS: We compared, in a rat model of CHF (coronary ligation), the hemodynamic and structural effects of 1 month of treatment with the ET(A) antagonist ABT-627 (5 mg x kg(-1) x d(-1)), the ET(B) antagonist A-192621 (30 mg x kg(-1) x d(-1)) or a combination of the 2 drugs. Doses were chosen for their capacity to block the pressor response to ET-1 (for ET(A) blockade) or the depressor responses to sarafotoxin S6c or ET-1 (for ET(B) blockade). ET(A) and combined ET(A)-ET(B) blockade reduced systolic blood pressure to the same extent, whereas ET(B) blockade had no effect. In contrast, only combined ET(A)-ET(B) blockade significantly reduced heart rate. Both ET(A) and combined ET(A)-ET(B) blockade, but not ET(B) blockade alone, increased left ventricular (LV) fractional shortening and wall thickening and reduced LV end-diastolic pressure, as well as LV end-diastolic and end-systolic volumes. However, all treatments (including ET(B) blockade) decreased LV collagen accumulation. CONCLUSIONS: The chronic blockade of both ET(A) and ET(B) receptors improved systemic hemodynamics, as well as LV function and remodeling, to the same extent as ET(A) receptor blockade alone. However, only combined ET(A)-ET(B) receptor blockade decreased heart rate. Whether this differential effect on heart rate affects the long-term outcome after treatment with ET(A) or mixed ET(A)-ET(B) antagonists in CHF remains to be determined.

Evidence against a role of inducible nitric oxide synthase in the endothelial protective effects of delayed preconditioning.

Preconditioning the heart with brief periods of ischaemia induces delayed endothelial protection against reperfusion injury, but the precise mechanisms involved in this endogenous protein are still unclear. Induction of the type II-nitric oxide synthase (iNOS) acts as a mediator of the preconditioning against myocardial infarction and stunning. The present study was designed to assess whether iNOS also contributes to the delayed endothelial protective effects of preconditioning. Rats were subjected to 20 min ischaemia followed by 60 min reperfusion 24 h after sham surgery or preconditioning (one cycle or 2 min ischaemia/5 min reperfusion and two cycles of 5 min ischaemia/5 min reperfusion). At the end of the reperfusion, coronary segments were removed distal to the site of occlusion and mounted in wire myographs. Ischaemia-reperfusion (I/R) decreased the endothelium-dependent relaxations to acetylcholine (maximal relaxations: sham, 66+/-5%; I/R, 40+/-1%; P<0.05) and this impairment was prevented by preconditioning (maximal relaxation: 61+/-6%). Administration of N-(3-aminomethyl)benzyl)acetaminide (1400W), a highly selective inhibitor for iNOS, 10 min before prolonged ischaemia did not modify the beneficial effect of preconditioning (maximal relaxation: 66+/-5%). These data show that preconditioning induces delayed protection against reperfusion-injury. However, in contrast to the myocytes, these endothelial protective effects of delayed preconditioning do not involve iNOS.

Improvement of endothelial function by chronic angiotensin-converting enzyme inhibition in heart failure : role of nitric oxide, prostanoids, oxidant stress, and bradykinin.

BACKGROUND-Chronic heart failure (CHF) impairs the endothelium-dependent, flow-mediated dilation (FMD) of small arteries. However, whether chronic angiotensin-converting enzyme (ACE) inhibition affects the impairment of FMD in CHF is unknown. We investigated the effects of long-term ACE inhibition on the FMD of peripheral arteries in rats with CHF and the mechanism(s) involved. METHODS AND RESULTS-FMD was assessed in isolated, perfused gracilis muscle arteries from sham-operated, and untreated or ACE inhibitor-treated (perindopril 2 mg. kg(-1). day(-1) for 10 weeks) rats with CHF (coronary artery ligation). The role of nitric oxide (NO), prostaglandins, and free radicals was assessed by pretreating the vessels with the NO synthase inhibitor N(W)-nitro-L-arginine, the cyclooxygenase inhibitor diclofenac, or the free radical scavenger N-2-mercaptopropionyl-glycine (MPG). Endothelial NO synthase mRNA expression was determined by reverse transcriptase polymerase chain reaction. In animals with hemodynamic and echographic signs of CHF, FMD was converted into vasoconstriction, and this was prevented by ACE inhibition. FMD of arteries from sham-operated or ACE inhibitor-treated CHF rats was abolished by N(W)-nitro-L-arginine. In untreated CHF rats, FMD was increased by diclofenac and MPG. In contrast, in arteries from ACE inhibitor-treated rats, neither diclofenac nor MPG affected FMD. In parallel, ACE inhibition prevented the reduction of endothelial NO synthase mRNA by CHF. CONCLUSIONS-In CHF, ACE inhibition normalized NO-dependent dilatation and suppressed the production of vasoconstrictor prostanoid(s), resulting in improved FMD. The improvement of FMD might contribute to the beneficial effects of ACE inhibition during CHF.

Biochemical characterization of Rab3-GTPase-activating protein reveals a mechanism similar to that of Ras-GAP.

Small G proteins of the Rab family are regulators of intracellular vesicle traffic. Their intrinsic rate of GTP hydrolysis is very low but is enhanced by specific GTPase-activating proteins (GAPs) that switch G proteins to their inactive form. We have characterized the activity of recombinant Rab3-GAP on Rab3A in solution. The K(m) and K(d) values (75 microm) indicate a low affinity of Rab3-GAP for its substrate. The affinity is higher for the transition state analog Rab3A:GDP:AlF(x) (15 microm). The k(cat) (1 s(-)(1)) is within the range of values reported for other GAPs. A mutation in the switch I region of Rab3A disrupted the interaction with Rab3-GAP. Furthermore, Rabphilin, a putative target of Rab3, inhibited the activity of Rab3-GAP on Rab3. Therefore, the Rab3-GAP-binding site involves the switch I region of Rab3 and overlaps with the Rabphilin-binding domain. Substitution of a single arginine residue (Arg-728) of Rab3-GAP disrupted its catalytic activity but not its interaction with Rab3A. We propose that Rab3-GAP, like Ras- and Rho-GAPs, stabilizes the transition state of Rab3 and provides a critical arginine residue to accelerate the GTPase reaction.