Regional Angiogenesis with Vascular Endothelial Growth Factor (VEGF) in peripheral arterial disease: Design of the RAVE trial

Background

Patients with intermittent claudication caused by infrainguinal atherosclerosis have limited pharmacologic options “Therapeutic angiogenesis” is a novel treatment approach that seeks to improve perfusion of ischemic limbs by the induction of collateral vessel formation. This trial is a phase 2 randomized double-blind placebo-controlled proof of concept trial that will use an intramuscular adenoviral gene transfer approach of vascular endothelial growth factor, 121 isoform (Ad_GVVEGF_121.10) to patients with severe IC caused by infrainguinal disease.

Methods

This is a phase 2, double-blind, randomized, placebo-controlled, dose-finding, multicenter study. Patients with severe intermittent claudication caused by infrainguinal atherosclerosis predominantly involving the superficial femoral artery confirmed with imaging studies that meet inclusion criteria will be stratified on the basis of the presence or absence of diabetes mellitus and randomized in a 1:1:1 fashion to low dose (4 × 10⁹ particle units), high dose (4 × 10¹⁰ particle units), or placebo arms (35-36 patients per group). Subjects are required to have exercise-limiting IC in the index extremity during 2 qualifying exercise treadmill tests, with peak walking times between 1 and 10 minutes. A single dose of Ad_GVVEGF_121.10 will be administered as 20 intramuscular injections throughout the area of the lower limb requiring collateralization.

Results

The primary efficacy parameter for the Regional Angiogenesis With Vascular Endothelial Growth Factor (RAVE) trial is the change in peak walking time at 12 weeks compared with baseline. The sample size is expected to provide an 80% power to detect a difference of 1.5 minutes between any of the 2 treatment groups and the placebo group. Secondary efficacy parameters include claudication onset time, hemodynamic effects of therapy assessed with ankle-brachial index, assessment of physical impairment, and health-related quality of life as measured with the Walking Impairment Questionnaire and SF-36 Health Survey. All randomized patients will also be evaluated for safety.     

Cholesteatoma epithelium is characterized by increased expression of Ki-67, p53 and p21, with minimal apoptosis

OBJECTIVE: To investigate differences in cell proliferation, cell cycle arrest and apoptosis between cholesteatoma and control skin. MATERIAL AND METHODS: Immunohistochemical sections of 15 cholesteatoma and 15 paired control retro-auricular skin samples were examined for Ki-67, p53, p21 and active caspase 3, using image analysis, as well as for DNA fragmentation. RESULTS: The retro-auricular skin samples contained 5.7% +/- 3.6%, Ki-67-positive cells and showed a normal expression pattern. In the cholesteatoma epithelium 11.7% +/- 9.5% of the cells were Ki-67-positive and these cells were dominantly expressed in the basal and parabasal cell layers. Retro-auricular skin contained 5.8% +/- 5.4% p53-positive cells and 1.0% +/- 0.9%, p21-positive cells. In the cholesteatoma epithelium 17.8% +/- 12.3% of the cells were p53-positive and 14.3% +/- 11.6% were p21-positive The expression of Ki-67, p53 and p21 differed significantly between the two groups (p < 0.05). In the cholesteatoma epithelium a positive correlation was found between p53 and p21 expression (p = 0.016). Active caspase 3 positivity and DNA fragmentation were rarely seen in the cholesteatoma epithelium. CONCLUSION: Our results indicate that increased cell proliferation in cholesteatoma epithelium is accompanied by an increase in p53 and p21 protein levels, whilst apoptosis is minimal.     

Validation of the screening strategy in the NIAAA "Physicians' Guide to Helping Patients with Alcohol Problems"

OBJECTIVE: This study was undertaken to determine the diagnostic test characteristics of the alcohol screening strategy recommended in the National Institute on Alcoholism and Alcohol Abuse (NIAAA) "Physicians' Guide to Helping Patients with Alcohol Problems." METHOD: A research interview was performed on patients who presented to one urban emergency department (N = 395; 61% women). It asked three alcohol consumption questions, the CAGE questionnaire, and about past alcohol problems. The NIAAA-recommended screen was considered positive for alcohol consumption in excess of 14 drinks per week or 4 drinks per occasion for men, or 7 drinks per week or 3 drinks per occasion for women, or a CAGE score of 1 or greater. A sample of patients (n = 250) received the Composite International Diagnostic Interview substance abuse module, a gold standard interview, to determine lifetime or prior 12-month alcohol abuse or dependence; results were adjusted for verification bias. RESULTS: The prevalence of lifetime:alcohol abuse or dependence was 13%, for which the NIAAA strategy was 81% sensitive and 80% specific. The prevalence of alcohol abuse or dependence in the prior 12 months was 10%, for which the strategy was 83% sensitive and 84% specific. Its positive likelihood ratio exceeded that of the CAGE, augmented CAGE or consumption questions alone, and its negative likelihood ratio was the lowest. CONCLUSIONS: The screening strategy combining alcohol consumption and CAGE questions recommended in the NIAAA "Physicians' Guide" is valid, and has superior test characteristics compared to the CAGE alone, in this predominantly black (86%) emergency department population. Its brevity and simple interpretation recommend wider dissemination of the NIAAA "Physicians' Guide," although future research should examine its test characteristics in other clinical settings and with other populations.     

Bismuth overdosing-induced reversible nephropathy in rats

Overdosing of colloidal bismuth subcitrate (CBS), used to treat peptic ulcers and Helicobacter pylori infections, has been reported to result in serious, though reversible, nephrotoxicity in humans. However, little is known about the nature of the renal damage induced by bismuth (Bi), and no well-described experimental model exists. Single large oral CBS doses (0.75, 1.5, and 3.0 mmol Bi/kg) were administered to three groups of 20 female Wistar rats. A control group (n = 20) received only the vehicle. Standard kidney function parameters, urinary excretion of N-acetyl-beta-D-glucosaminidase (NAG) and the Bi content were monitored in blood, urine, liver, and kidneys for 14 days. A dose of 3.0 mmol Bi/kg, 100 times the daily therapeutic dose, caused kidney damage within 6 h as detected by proteinuria, glucosuria, and elevated plasma urea and plasma creatinine levels. The kidneys of all animals, except two that died, recovered functionally within 10 days. At a dose of 1.5 mmol Bi/kg, clinical parameters changed less and normalized within 48 h, whereas a dose of 0.75 mmol Bi/kg induced no changes. Histological evaluation revealed that the S3 tubular segment necrotized first with additional necrotization of the S1/S2 segment when more Bi was absorbed. The lesions were accompanied by interstitial infiltrates of CD45+ leukocytes. In summary, we developed a rat model for Bi-induced reversible nephropathy. A large single oral overdose of CBS administered to Wistar rats led to damage to the proximal tubule, especially in the last segment.     

Accuracy of color-Doppler in the quantification of proximal vertebral artery stenoses

BACKGROUND: Vertebrobasilar (VB) strokes appear to have the same causes as carotid strokes. Obstructive lesions of proximal vertebral arteries probably occur in about 30% of stroke patients. PURPOSE: Our aim was to assess the validity of color Doppler sonography compared to selective intra-arterial angiography in the quantification of proximal vertebral artery stenoses. MATERIALS AND METHODS: A prospective blind study of 316 vertebral arteries was undertaken between 1996 and 1998. One hundred and fifty-eight patients with cerebrovascular disorders without cerebral hemorrhage were studied consecutively by frequency or amplitude color Doppler flow imaging and intra-arterial angiography. The lesions were quantified by morphological and hemodynamic criteria and classified into 6 groups: 0% 207 arteries; 1-29% 32 arteries; 30-49% 29 arteries; 50-69% 13 arteries; 70-99% 23 arteries; 100% 12 arteries. RESULTS: Ten of the 12 occlusions were identified, the 2 false-negatives were due to 2 revascularized vessels. Moderate stenoses (<50%) were differentiated from tight stenoses (>50%) using hemodynamic criteria. The majority of false-negative stenoses (38) in the different groups were related to intrathoracic or very deep origin of the artery, anechogenic stenosis or a tortuous vessel. Stenoses greater than 70% were diagnosed in 71% of cases with a specificity of 99%. The kappa value was 0.80. CONCLUSION: Duplex sonography should be proposed first in VB attacks or stroke to detect and quantify vertebral artery stenoses for surgery and angioplasty.     

Right Ventricular Function Evolution With Pregnancy in Repaired Tetralogy of Fallot

This case illustrates the evolution of right ventricular (RV) 3-dimensional (3D) area strain during pregnancy in a patient with repaired Tetralogy of Fallot. The report highlights impairment in RV function with pregnancy, suggesting the importance of prepregnancy RV systolic function assessment, especially using 3D echocardiography.     

Large-scale screening of the in vitro susceptibility of Prototheca zopfii towards polyene antibiotics

A large scale screening of the in vitro susceptibility of 105 strains of Prototheca zopfii to a panel of polyene antibiotics (amphotericin B, nystatin, primaricin and filipin) was conducted. Strains studied were isolated from dairy-associated environments in five different localities. Groups 1-4 included strains recovered from four separate regions of Italy, while group 5 included isolates from Belgium. Amphotericin B and primaricin exhibited the highest activity, with th MIC90 ranging from 4 and 8 microg/ml, respectively. On the other hand, the MIC90 of nystatin and filipin were from two to four times higher. Two strains were resistant to all four polyenes tested. The above results are compared with those in the literature and the importance of carrying out large-scale screening surveys to assess polyene susceptibility patterns within the species P. zopfii is discussed.     

Binding of mannose‐binding lectin to fructosamines: a potential link between hyperglycaemia and complement activation in diabetes

Background

Complement activation via the MBL pathway has been proposed to play a role in the pathogenesis of diabetic complications. As protein glycation is increased in diabetes, we tested the possibility that the glycation product fructoselysine is a ligand for MBL and that its interaction with this protein may initiate complement activation.

Methods

We investigated the binding of MBL to fructoselysine by chromatography of human serum on fructoselysine‐Sepharose, followed by Western blot and mass spectrometry analysis. We also performed enzyme‐linked immunosorbent assays using purified MBL and fructoselysine‐derivatized (binding assay) or mannan‐coated plates (inhibition assay). Complement activation was determined by the fixation of C3d following incubation of fructoselysine‐derivatized plates with serum from subjects with different levels of MBL.

Results

MBL and its associated proteases were selectively purified from serum by chromatography on fructoselysine‐Sepharose. Competition experiments indicated that MBL had a similar affinity for mannose, fructose and fructoselysine. MBL bound, in a highly cooperative manner, to fructoselysine‐derivatized plates. This binding was associated with complement activation and was much lower with serum from subjects with low‐MBL genotypes.

Conclusions

MBL binding to fructoselysine and the ensuing complement activation may provide a physiopathological link between enhanced glycation and complement activation in diabetes. The cooperative character of this binding may explain the high sensitivity of diabetic complications to hyperglycaemia. Copyright © 2010 John Wiley & Sons, Ltd.