Multinuclear nuclear magnetic resonance spectroscopic and high-performance liquid chromatographic characterization of silica, grafted with specifically deuterated 4-((propylamino)methyl)benzoic acid.

Specifically deuterated 4-((propylamino)methyl)benzoic acid-grafted silica (PAMBA-silica) was prepared by benzylation of propylamino-grafted silica (PA-silica) by either in situ reduction by sodium cyanoborodeuteride (NaCNBD3) of the Schiff base, formed by the reaction between PA-silica and 4-formylbenzoic acid, or by NaCNBD3 reduction of the isolated Schiff base. The PAMBA-silicas, which contain amine and carboxylic acid functionalities, were characterized by elemental analysis, (13)C, (29)Si, and (2)H solid state NMR, and HPLC. Solid state (13)C NMR revealed that PAMBA-silica prepared by the in situ method consists of di-benzylated, mono-benzylated, and unreacted amino-groups while PAMBA-silica prepared by the two-step synthesis consists of only mono-benzylated and unreacted amino-groups. (29)Si solid-state NMR spectra indicated that no significant loss of propylamine groups had occurred during benzylation. Nearly ideal uniaxial rigid-limit (2)H NMR spectra of grafted 4-PAMBA ligands indicates that they form a rigid structure, which provides effective electrostatic screening of inner positive charges when the ligands are in zwitterionic form. HPLC columns packed with PAMBA-silica and PA-silica were evaluated for ionic solutes at different pH of the mobile phase. Retention times increased for cations and decreased for anions at increasing pH. These trends show that PAMBA-silicas act as cation and anion exchangers at high and low pH, respectively. The pKa values of grafted carboxylic acid, determined from HPLC of weakly retaining solutes, are close to pKa of the solution PAMBA.

Neuronal Hemoglobin Expression and Its Relevance to Multiple Sclerosis Neuropathology.

Multiple sclerosis (MS) is characterized by demyelination and progressive neurological disability. Previous studies have reported defects to mitochondria in MS including decreased expression of nuclear encoded electron transport chain subunit genes and inhibition of respiratory complexes. We previously reported increased levels of the hemoglobin ß subunit (Hbb) in mitochondrial fractions isolated from postmortem MS cortex compared to controls. In the present study, we performed immunohistochemistry to determine the distribution of Hbb in postmortem MS cortex and identified proteins which interact with Hbb by liquid chromatography tandem mass spectrometry (LC-MS/MS). We found that Hbb was enriched in pyramidal neurons in internal layers of the cortex and interacts with subunits of ATP synthase, histones, and a histone lysine demethylase. We also found that Hbb is present in the nucleus and that expression of Hbb in SH-SY5Y neuroblastoma cells increased trimethylation of histone H3 on lysine 4 (H3K4me3), a histone mark that regulates cellular metabolism. These data suggest that Hbb may be a part of a mechanism linking neuronal energetics with epigenetic changes to histones in the nucleus and may provide neuroprotection in MS by supporting neuronal metabolism.

A SELDI mass spectrometry study of experimental autoimmune encephalomyelitis: sample preparation, reproducibility, and differential protein expression patterns.

BACKGROUND: Experimental autoimmune encephalomyelitis (EAE) is an autoimmune, inflammatory disease of the central nervous system that is widely used as a model of multiple sclerosis (MS). Mitochondrial dysfunction appears to play a role in the development of neuropathology in MS and may also play a role in disease pathology in EAE. Here, surface enhanced laser desorption ionization mass spectrometry (SELDI-MS) has been employed to obtain protein expression profiles from mitochondrially enriched fractions derived from EAE and control mouse brain. To gain insight into experimental variation, the reproducibility of sub-cellular fractionation, anion exchange fractionation as well as spot-to-spot and chip-to-chip variation using pooled samples from brain tissue was examined. RESULTS: Variability of SELDI mass spectral peak intensities indicates a coefficient of variation (CV) of 15.6% and 17.6% between spots on a given chip and between different chips, respectively. Thinly slicing tissue prior to homogenization with a rotor homogenizer showed better reproducibility (CV = 17.0%) than homogenization of blocks of brain tissue with a Teflon® pestle (CV = 27.0%). Fractionation of proteins with anion exchange beads prior to SELDI-MS analysis gave overall CV values from 16.1% to 18.6%. SELDI mass spectra of mitochondrial fractions obtained from brain tissue from EAE mice and controls displayed 39 differentially expressed proteins (p≤ 0.05) out of a total of 241 protein peaks observed in anion exchange fractions. Hierarchical clustering analysis showed that protein fractions from EAE animals with severe disability clearly segregated from controls. Several components of electron transport chain complexes (cytochrome c oxidase subunit 6b1, subunit 6C, and subunit 4; NADH dehydrogenase flavoprotein 3, alpha subcomplex subunit 2, Fe-S protein 4, and Fe-S protein 6; and ATP synthase subunit e) were identified as possible differentially expressed proteins. Myelin Basic Protein isoform 8 (MBP8) (14.2 kDa) levels were lower in EAE samples with advanced disease relative to controls, while an MBP fragment (12. 4kDa), likely due to calpain digestion, was increased in EAE relative to controls. The appearance of MBP in mitochondrially enriched fractions is due to tissue freezing and storage, as MBP was not found associated with mitochondria obtained from fresh tissue. CONCLUSIONS: SELDI mass spectrometry can be employed to explore the proteome of a complex tissue (brain) and obtain protein profiles of differentially expressed proteins from protein fractions. Appropriate homogenization protocols and protein fractionation using anion exchange beads can be employed to reduce sample complexity without introducing significant additional variation into the SELDI mass spectra beyond that inherent in the SELDI- MS method itself. SELDI-MS coupled with principal component analysis and hierarchical cluster analysis provides protein patterns that can clearly distinguish the disease state from controls. However, identification of individual differentially expressed proteins requires a separate purification of the proteins of interest by polyacrylamide electrophoresis prior to trypsin digestion and peptide mass fingerprint analysis, and unambiguous identification of differentially expressed proteins can be difficult if protein bands consist of several proteins with similar molecular weights.

Interviews with pioneers of vascular surgery.

The purpose of the SVS is not just to hold an annual meeting, but also to pursue social, financial, and political responsibilities. In addition, the Society leads in research, training, education, and practice (ie, patient care). The current leadership of the SVS is dedicated to preserving the history of the Society. The History Project Work Group will execute the orders of the Society to develop a series of digital videography recorded interviews with leaders in vascular surgery to be made available to our membership. The information collected will form the basis for a book on the history of the SVS.

Analysis of the mitochondrial proteome in multiple sclerosis cortex.

Mitochondrial dysfunction has been proposed to play a role in the neuropathology of multiple sclerosis (MS). Previously, we reported significant alterations in the transcription of nuclear-encoded electron transport chain genes in MS and confirmed translational alterations for components of Complexes I and III that resulted in reductions in their activity. To more thoroughly and efficiently elucidate potential alterations in the expression of mitochondrial and related proteins, we have characterized the mitochondrial proteome in postmortem MS and control cortex using Surface-Enhanced Laser Desorption Ionization Time of Flight Mass Spectrometry (SELDI-TOF-MS). Using principal component analysis (PCA) and hierarchical clustering techniques we were able to analyze the differential patterns of SELDI-TOF spectra to reveal clusters of peaks which distinguished MS from control samples. Four proteins in particular were responsible for distinguishing disease from control. Peptide fingerprint mapping unambiguously identified these differentially expressed proteins. Three proteins identified are involved in respiration including cytochrome c oxidase subunit 5b (COX5b), the brain specific isozyme of creatine kinase, and hemoglobin ß-chain. The fourth protein identified was myelin basic protein (MBP). We then investigated whether these alterations were consistent in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. We found that MBP was similarly altered in EAE but the respiratory proteins were not. These data indicate that while the EAE mouse model may mimic aspects of MS neuropathology which result from inflammatory demyelinating events, there is another distinct mechanism involved in mitochondrial dysfunction in gray matter in MS which is not modeled in EAE.

Significant reductions in methicillin-resistant Staphylococcus aureus bacteraemia and clinical isolates associated with a multisite, hand hygiene culture-change program and subsequent successful statewide roll-out.

OBJECTIVE: To assess the efficacy of a multimodal, centrally coordinated, multisite hand hygiene culture-change program (HHCCP) for reducing rates of methicillin-resistant Staphylococcus aureus (MRSA) bacteraemia and disease in Victorian hospitals. DESIGN, PARTICIPANTS AND SETTING: A pilot HHCCP was conducted over a 24-month period (October 2004 to September 2006) in six Victorian health care institutions (4 urban, 2 rural; total beds, 2379). Subsequently, we assessed the efficacy of an identical program implemented throughout Victorian public hospitals over a 12-month period (beginning between March 2006 and July 2006). MAIN OUTCOME MEASURES: Rates of hand hygiene (HH) compliance; rates of MRSA disease (patients with bacteraemia and number of clinical isolates per 100 patient discharges [PD]). RESULTS: Mean HH compliance improved significantly at all pilot program sites, from 21% (95% CI, 20%-22%) at baseline to 48% (95% CI, 47%-49%) at 12 months and 47% (95% CI, 46%-48%; range, 31%-75%) at 24 months. Mean baseline rates for the number of patients with MRSA bacteraemia and the number of clinical MRSA isolates were 0.05/100 PD per month (range, 0.00-0.13) and 1.39/100 PD per month (range, 0.16-2.39), respectively. These were significantly reduced after 24 months to 0.02/100 PD per month for bacteraemia (P = 0.035 for trend; 65 fewer patients with bacteraemia) and 0.73/100 PD per month for MRSA isolates (P = 0.003; 716 fewer isolates). Similar findings were noted 12 months after the statewide roll-out, with an increase in mean HH compliance (from 20% to 53%; P < 0.001) and reductions in the rates of MRSA isolates (P = 0.043) and bacteraemias (P = 0.09). CONCLUSIONS: Pilot and subsequent statewide implementation of a multimodal HHCCP was effective in significantly improving HH compliance and reducing rates of MRSA infection.

Utility of basilic vein transposition for dialysis access.

Autologous arteriovenous access is the key to long-term success with hemodialysis and is strongly supported by the National Kidney Foundation's Dialysis Outcomes Quality Initiative guidelines. Basilic vein transposition (BVT) fulfills the need for a durable conduit with high patency and maturation rates. This retrospective review examines a single group's experience with this procedure. All patients undergoing BVT for hemodialysis with available follow-up data were reviewed. Telephone interviews were used to supplement clinical data where needed. Functional assisted patency was used as the end point for this procedure, and if the access was never used for dialysis, then the patency was considered zero. Secondary interventions performed while the access remained patent and in use were not considered detrimental to the patency reported. One hundred seventy BVTs in 162 patients were performed between November 1992 and October 2001. There were 87 women (53.7%) and 112 black patients (69.1%); hypertension was present in 138 patients (85.2%) and diabetes in 89 patients (54.9%). Each year, an increasing incidence of BVT was performed in our dialysis population. The BVT was performed as the first access in that extremity in 73 of the procedures (42.9%). Functional patency (primary assisted) was achieved in 40.0% at 2 years and 15.2% at 5 years. The mean assisted patency was 14.6 months. To maintain BVT patency, 40 percutaneous secondary interventions (69.0%) and 18 surgical revisions (31.0%) occurred in 32 patients (19.0%). Ligation for swelling was necessary in 4 patients (2.5%), and steal syndrome occurred in 3 patients (1.9%). BVT is a useful autologous procedure for hemodialysis and the preferred access alternative in patients without an adequate cephalic vein. Although patencies remain poor relative to other conventional arterial vascular procedures, BVT is our most durable hemodialysis access procedure and is often the only available autologous conduit for hemodialysis.

Endograft limb occlusion and stenosis after ANCURE endovascular abdominal aneurysm repair.

OBJECTIVE: The purpose of this study was to define the incidence and treatment of endograft limb stenosis or occlusion (endograft limb dysfunction [ELD]) in a single center with the ANCURE unsupported bifurcated or aortouniiliac endograft by using intraoperative completion angiography and postoperative color duplex ultrasound scanning (CDU). METHODS: Sixty-seven endografts (58 bifurcated, 9 uniiliac) were implanted between February 1996 and July 2000. Intraoperative completion aortography was performed in every patient. Postoperative assessment of the endograft consisted of CDU and computed tomography scanning and kidney, ureter and bladder radiographs within 7 days of implantation, at 3 and 6 months after the operation, and every 6 months thereafter. RESULTS: At the time of endograft implantation, widely patent normal-appearing endograft limbs were revealed by means of the initial completion angiogram in 58 of 67 patients (group 1). ELD subsequently developed in seven of these 58 patients (13.4%). The results of the completion angiogram were not normal in the remaining nine patients (group 2), leading to the deployment of a self-expanding stent within the endograft limbs. The results of subsequent angiography were normal. No ELD has occurred in any patient in group 2 to date. The primary assisted patency rate at 30 months was 88% +/- 5.2% for group 1 versus 100% +/- 0% for group 2 (P = not significant, Log-rank test). Postoperative ELD occurred in seven patients (10.4%). Endovascular graft thrombosis occurred in three patients (3 endograft limbs). In each case, an endovascular approach was attempted; however, the guidewire would not traverse the occluded endovascular graft limb. Revascularization was accomplished by means of femorofemoral bypass grafting. Endovascular graft stenosis occurred in four patients (4 endograft limbs). Three patients with bifurcated endografts and limb stenosis who had no symptoms diagnosed by means of CDU were successfully treated by means of balloon angioplasty with self-expanding stent implantation, and the endograft limbs remained patent at 3, 5, and 26 months follow-up. The remaining patient who had an aortouniiliac endograft with recurrent severe stenoses underwent endograft explantation and aortobifemoral bypass grafting. The overall incidence of ELD during or after endovascular abdominal aortic aneurysm repair was 23.8% (16 of 67 patients). CONCLUSION: Unsupported endografts are at risk for developing ELD. The use of stents for limb support at the time of the initial endograft implantation may prevent subsequent ELD and bears further study. Endograft limb occlusion usually presents with acute severe ischemic symptoms, and the failure of operative thrombectomy necessitates femorofemoral artery bypass grafting. Endograft limb stenosis is identified by means of CDU surveillance in the postoperative period. Prompt treatment with percutaneous transluminal angioplasty/stent yields satisfactory primary assisted patency. Intraoperative intravenous ultrasound scanning, oblique angiograms, pressure gradients, and completion angiography may be necessary to detect and treat ELD.

The incidence and natural history of type I and II endoleak: a 5-year follow-up assessment with color duplex ultrasound scan.

PURPOSE: The purpose of this study was the demonstration of the value of color duplex ultrasound (CDU) scanning in the detection of type I endoleak (T1EL) and type II endoleak (T2EL), the correlation of Doppler scan waveform pattern to endoleak persistence or seal, and the description of the natural history of endoleak. METHODS: The study was a retrospective review of 83 patients who underwent periodic CDU scan and computed tomographic (CT) scan surveillance of the endograft and aneurysm sac after insertion of an aortic endograft for abdominal aortic aneurysm (AAA). Forty-one patients (49%) with an endoleak at anytime in the follow-up period form the basis of this report. RESULTS: T1EL was detected in all five patients with CDU and CT scans. T2EL was detected in 36 patients with CDU scan as compared with 18 patients with CT scan. With CT scan, endoleak was not detected when CDU scan showed no endoleak. Conversely, all CT scan--detected endoleaks were found with CDU scanning. The T2EL source artery was identified with CT scan in seven patients, whereas the source was identified in all 36 patients with CDU scan. Endoleak source did not correlate with outcome (seal or persistence). However, a to/fro Doppler scan waveform pattern was associated with spontaneous T2EL seal in seven of 12 patients, and a monophasic or biphasic waveform was associated with endoleak persistence in 14 of 17 patients (P =.023, with chi(2) test). Thirteen of 36 T2ELs underwent spontaneous seal by 6.2 +/- 2.8 months. T2ELs without increasing AAA diameter were observed. Eight patients with persistent T2EL present for more than 12 months did not undergo treatment. However, two patients underwent T2EL obliteration with coils because of AAA sac enlargement. T1EL of the distal attachment site was the initial endoleak identified in five patients, but seven patients harboring T2ELs had subsequent T1ELs develop. For the entire 83 patients, the combined T1EL and T2EL prevalence rate was 20% of patients at a 6-month follow-up period, but this rate increased to 50% after 24 months. The incidence rate of newly detected endoleaks and of spontaneous sealing was 24.4% at 12 months and 12.5% in longer-term follow-up period. CONCLUSION: CDU scan is effective in the identification of the type of endoleak, the delineation of the vessel involved, and the hemodynamic information not available with any other testing method. Endoleaks have a dynamic natural history characterized by a variable onset with changing branch vessel involvement and spectral flow patterns. Periodic long-term endograft surveillance with CDU scanning is necessary for following existing endoleaks and for detecting new ones. Corroboration of these findings in larger multicenter prospective trials will be needed to determine whether CDU scan analysis of endoleaks would be predictive of long-term success in endovascular AAA repair.

Recommended standards for reports dealing with arteriovenous hemodialysis accesses.

The incidence rate of treated end-stage renal disease in the united states is 180 per million and continues to rise at a rate of 7.8% per year. Arteriovenous hemodialysis access (AV access) creation and maintenance are two of the most difficult issues associated with the management of patients on hemodialysis. The 1-year complication rate of a primary prosthetic AV access for hemodialysis ranges from 33% to 99%. Various investigators report on patency and complications of AV access. However, it is rather difficult to compare outcomes because of the wide variety of access materials, configurations, locations, risk factors, and quality of inflow and outflow vessels. Although there have been reporting standards for dialysis access endovascular interventions and for central venous access placement, standards regarding surgical access placement and its revision are lacking. The "Dialysis Outcome Quality Initiative," published by the National Kidney Foundation, provides recommendations for optimal clinical practices aimed at improving dialysis outcome and patient survival. This reporting standards document is not meant to be a "practice guidelines" or "best practices" document. Rather, the purpose of this document is to provide standardized definitions related to AV access procedures and to recommend reporting standards for patency and complications, to be used by surgeons, nephrologists, and interventional radiologists, that will permit meaningful comparisons among AV access procedures. The terms, definitions, and categories featured in this article have been approved by the Committee on Reporting Standards of the Society for Vascular Surgery and the American Association for Vascular Surgery and should be observed in preparing manuscripts on AV accesses for submission to the Journal Of Vascular Surgery.