Proteomic profile of circulating immune complexes in chronic Chagas disease.

Immune complexes (ICs) are the direct and real-time products of humoral immune responses. The identification of constituent foreign or autoantigens within ICs might bring new insights into the pathology of infectious diseases. We applied immune complexome analysis of plasma to the study of Chagas disease caused by Trypanosoma cruzi. Twenty seropositive plasma samples including cardiac and/or megacolon determinate patients (n = 11) and indeterminate (n = 9) were analysed along with 10 seronegative individuals to characterize the antigens bound to circulating ICs. We identified 39 T. cruzi antigens and 114 human autoantigens specific to patients with Chagas. Among those antigens, two T. cruzi antigens (surface protease GP63, glucose-6-isomerase) and six human autoantigens (CD180 antigen, ceruloplasmin, fibrinogen beta chain, fibrinogen beta chain isoform 2 preprotein, isoform gamma-A of fibrinogen γ-chain, serum paraoxonase) were detected in more than 50% of the patients tested. Human isoform short of complement factor H-related protein 2 and trans-sialidase of T. cruzi were more frequently found in the indeterminate (5/9 for both) compared with in the determinate Chagas (0/11, P = 0·046 for human, 1/11, P = 0·0498 for T. cruzi). The immune complexome could illustrate the difference of immune status between clinical forms of chronic Chagas disease.

Impact on physician workload and revenue following the creation of a specialty vein clinic within an academic vascular practice.

OBJECTIVE: To evaluate the impact of creating a new specialty vein clinic within an academic-based vascular practice on clinical volume, physician workload and financial parameters. METHODS: All patients evaluated and treated for varicose vein related problems within an academic vascular surgery practice were identified from institutional billing databases. Data were stratified according to the time period prior to establishing a vein clinic (PRE-VC) (1999-2001) and after creation of a vein clinic (POST-VC) (2002-2004). Clinical volume, physician workload and financial parameters were evaluated. Comparisons were made between vein (VEIN) and overall vascular (VASC) practice trends. RESULTS: Comparison of clinical volume, physician workload and financial parameters in both the clinic and operative settings showed larger and more rapid expansion of the VEIN practice than VASC practice between PRE-VC and POST-VC time periods (VEIN vs.VASC growth, respectively: new patient clinic volume +162 vs. +18%; clinic relative value units (RVUs) +131 vs. +1%, clinic revenue +201 vs. +44%; procedure volume +348 vs. +19%; procedure RVUs +129 vs. +11%; procedure revenue +93 vs. +10%). Comparing the beginning of PRE-VC to the end of POST-VC time periods, an increasing trend was also present for the percentage of VEIN practice accounting for the total VASC practice (%VEIN PRE-VC to POST-VC, respectively: new patient clinic volume 11.6-30.2%; clinic RVUs 3.2-48.2%; clinic revenue 17.6-31.2%; procedure volume 3.1-14.3%; procedure RVUs 2.8-9.8%; procedure revenue 3.3-11.7%). CONCLUSION: Establishing a specialty vein clinic within an academic vascular practice can lead to a rapid expansion of clinical volume with associated increase in physician workload and reimbursement at a rate greater than that for the overall vascular practice.

Impact of Medicare denials on noninvasive vascular diagnostic testing.

PURPOSE: The purpose of this study was to evaluate the impact of Medicare coverage limitations and claim denials on noninvasive vascular diagnostic testing. METHODS: All Medicare claims for noninvasive vascular diagnostic studies from January 1, 1999, to December 31, 1999, were identified from the hospital billing database according to Current Procedural Terminology codes for carotid artery duplex ultrasound scan, venous duplex ultrasound scan, and lower-extremity arterial Doppler scan. Reasons for Medicare denial of payment for these tests were reviewed and a cost analysis was performed. RESULTS: During the 1-year period, there were 1096 noninvasive vascular diagnostic studies performed on Medicare patients. Of these 1096 tests, 176 (16.1%) were denied by Medicare (19.6% of 408 carotid duplex ultrasound scans, 16.8% of 345 venous duplex ultrasound scans, and 11.1% of 343 lower-extremity arterial Doppler scans). Of the noninvasive vascular tests denied by Medicare, an abnormal result was present in 72.5% of carotid duplex ultrasound scans, 32.8% of venous duplex ultrasound scans, and 78.9% of lower-extremity arterial Doppler scans. Overall, 88.1% of all initially denied claims (N = 176) were ultimately reimbursed by Medicare after resubmission, including 77.1% of the 118 claims denied based on compliance rules for "medical necessity." CONCLUSION: Because of coverage limitations, Medicare denials of noninvasive vascular diagnostic tests can lead to potential uncompensated physician and hospital technical fees if denied claims are unrecognized. Vascular laboratories performing these tests need to review compliance with Medicare guidelines. Improvements may need to be made at both the provider and Medicare carrier levels in obtaining reimbursement for appropriately ordered noninvasive vascular diagnostic studies.

Durability of percutaneous angioplasty and stent implantation for the treatment of abdominal aortic coarctation: a case report.

Abdominal aortic coarctation (AAC) is an uncommon vascular lesion with serious sequelae related to uncontrolled hypertension. Balloon-expandable stents have recently been utilized in the treatment of AAC as an alternative to surgical intervention. A 17-year-old female presented with hypertension uncontrolled by beta blockade. She underwent angiography, which revealed an isolated supraceliac aortic coarctation without visceral or renal artery involvement. Balloon angioplasty with stent placement was performed. At 2-year follow-up, a restenosis was identified and was treated with repeat balloon-expandable stent placement. Implantation of balloon-expandable stents is a safe and technically feasible treatment modality for abdominal aortic coarctation not involving the renal and mesenteric arteries. However, it is currently unknown whether the long-term durability of this approach may limit its effectiveness when compared to traditional surgical interventions.

Matrix metalloproteinase inhibition limits arterial enlargements in a rodent arteriovenous fistula model.

BACKGROUND: We administered a specific, nonselective matrix metalloproteinase (MMP) inhibitor (RS-113,456) to examine the effect of MMP inhibition on flow-mediated arterial enlargement in a rodent arteriovenous fistula (AVF) model. METHODS: Four groups of male Sprague-Dawley rats were created: sham (sham operated; n = 10), control (2.0 mm left common femoral AVF alone; n = 16), vehicle (AVF plus 0.5 mL vehicle orally twice a day; n = 20), and treatment (AVF plus 25 mg/kg RS-113,456 in 0.5 mL vehicle orally twice a day; n = 16). Heart rate, mean arterial pressure, and body weight were recorded on postoperative days 0, 7, 14, and 21. On day 21, AVF patency was confirmed, the infrarenal aorta and common iliac arteries were exposed, blood flow velocity and external diameter were measured, and wall shear stress (WSS) was calculated. Analysis was performed by paired, two-tailed Student t test, one-way analysis of variance, and the Bonferroni/Dunn procedure for post hoc testing. RESULTS: Heat rate, mean arterial pressure, and weight did not vary at any time between groups. Aortic and left iliac diameter was larger in the AVF groups than in sham groups (P < .001), and control and vehicle groups were larger than treatment groups (P < .0001). Changes in aortic and left iliac flow were also significant (AVF was more than sham and control, and vehicle was more than treatment). No difference in aortic and left iliac artery velocity and WSS or right iliac diameter, velocity, flow, or WSS was observed between groups. CONCLUSIONS: MMP inhibition diminishes flow-mediated arterial enlargement in the rat AVF model.

Flow-induced arterial enlargement is inhibited by suppression of nitric oxide synthase activity in vivo.

BACKGROUND: Acute flow-induced arterial dilation is mediated by nitric oxide (NO). The role of NO in chronic flow-induced adaptive enlargement is unknown. We assessed the role of NO in arterial adaptation to increased blood flow (BF). METHODS: Iliac artery BF was increased in adult male rats by creating a left femoral arteriovenous fistula. Left iliac BF and diameter were measured, and wall shear stress was calculated. The effect of the NO synthase inhibitor N omega-nitro-L-arginine-methyl ester (L-NAME) was studied in arteriovenous fistula rats divided into three groups (group 1, vehicle, group 2, 0.5 mg/ml; group 3, 2 mg/ml) in drinking water. Arterial diameter, blood pressure, and medial cell density were assessed after 21 days. Left iliac cyclic guanosine monophosphate content was measured in an additional group of animals. RESULTS: BF and wall shear stress in the left iliac artery increased fourfold immediately after arteriovenous fistula. Arterial enlargement was evident after 7 days, and wall shear stress normalized after 42 days. Flow-induced arterial enlargement was inhibited by both low- and high-dose L-NAME compared with control (analysis of variance p < 0.05). Blood pressure was elevated only in animals treated with high-dose L-NAME. Left iliac cyclic guanosine monophosphate content was lower in rats treated with L-NAME than in the control group (p < 0.05). CONCLUSIONS: NO suppression by L-NAME inhibits flow-induced iliac artery enlargement in rats. This finding suggests that NO plays a role in flow-induced arterial remodeling.

Inhibition of vascular smooth muscle cell proliferation and neointimal accumulation by adenovirus-mediated gene transfer of cytosine deaminase.

BACKGROUND: Restenosis remains a significant problem after balloon angioplasty. Previous studies have demonstrated that recombinant adenoviruses are efficient vectors for gene transfer to the arterial wall and can be used to inhibit the proliferative aspect of restenosis. We sought to extend these observations using AdCMV.CD, an adenovirus that encodes cytosine deaminase (CD) and is capable of metabolizing 5-fluorocytosine (5-FC) to 5-fluorouracil. METHODS AND RESULTS: Infection of vascular smooth muscle cells (VSMC) with AdCMV.CD increases by two to three orders of magnitude the growth-inhibitory effects of 5-FC. The degree of VSMC inhibition in vitro was a function of 5-FC concentration and the level of CD expression. Cells infected with AdCMV.CD exhibited a profound bystander effect on the growth of neighboring cells, which did not require direct cell-to-cell contact. The predominant effect of AdCMV.CD on growth of VSMC appeared to be cytostatic, not cytotoxic. Assessment of this strategy in a rabbit femoral artery model of balloon-induced injury demonstrated that compared with animals in either of two control groups, animals treated with the active combination of infection with AdCMV.CD and 1-week treatment with parenteral 5-FC had a significant reduction at 30 days in the neointimal-to-medial ratio. CONCLUSIONS: Our results suggest that adenovirus-mediated gene transfer of CD along with 5-FC administration may be a useful strategy to treat the proliferative aspects of restenosis.

In vivo suppression of injury-induced vascular smooth muscle cell accumulation using adenovirus-mediated transfer of the herpes simplex virus thymidine kinase gene.

Restenosis, a process characterized in part by excessive smooth muscle cell (SMC) proliferation in areas of vascular injury, occurs in up to 50% of patients undergoing balloon angioplasty. In an effort to develop a treatment strategy for restenosis, we constructed a replication-deficient recombinant adenovirus (AdMLP.HSTK) containing the herpes simplex virus thymidine kinase gene (HSV tk). This viral gene product phosphorylates the prodrug ganciclovir to form a nucleoside analog that inhibits DNA synthesis. Cultured primary rat SMCs infected with AdMLP.HSTK were completely growth-inhibited by incubation in ganciclovir-containing medium. In addition, when only a portion of the SMC population received the HSV tk transgene, an inhibitory effect on neighboring SMCs was evident. Evaluation of this strategy in vivo using a rat carotid balloon injury model demonstrated that local infection of injured arteries with AdMLP.-HSTK followed by 2 weeks of systemic ganciclovir treatment significantly (P < 0.01) reduced injury-induced SMC accumulation. In contrast, there was no suppression of injury-induced SMC accumulation in animals infected with AdMLP.HSTK but not receiving ganciclovir or in those animals infected with a control adenovirus and either treated or not treated with ganciclovir. These results demonstrate the potential utility of adenovirus-mediated gene transfer for treatment of restenosis after balloon injury.

The basis of molecular strategies for treating coronary restenosis after angioplasty.

Excessive smooth muscle cell proliferation significantly contributes to restenosis, which occurs in 25% to 50% of patients within 6 months of coronary angioplasty. Because successful treatment will probably depend on our acquiring a comprehensive knowledge of the molecular and cellular mechanisms involved, this report reviews 1) information relevant to the molecular and cellular mechanisms responsible for the smooth muscle cell(s) response to vascular injury, and 2) several molecular-based therapeutic strategies currently being explored as possible approaches to the control of restenosis, including recombinant DNA technology to target delivery of cytotoxic molecules to proliferating smooth muscle cell(s), antisense strategies to inhibit expression of gene products necessary for cell proliferation and gene therapy.

Efficient gene transfer into myocardium by direct injection of adenovirus vectors.

Previous studies have established that gene transfer into myocardial cells in vivo is detectable after direct injection of plasmid DNA. Recently, adenovirus vectors have been shown to provide an efficient method for gene transfer into a wide range of tissues. Therefore, this study sought to assess the efficiency and stability of adenovirus-mediated gene transfer into myocardium and to compare this method with that using plasmid-based gene transfer techniques. Adult rats underwent myocardial injection via a subdiaphragmatic approach. Gene transfer efficiency was compared using direct injection of an adenovirus vector encoding for the marker gene beta-galactosidase (beta-gal), a control adenovirus vector encoding for the cystic fibrosis transmembrane conductance regulator gene, a plasmid encoding for beta-gal, or a control plasmid. Hearts infected with an adenovirus vector containing the beta-gal gene showed significantly increased beta-gal enzymatic activity compared with hearts injected with beta-gal plasmid. Histological examination revealed that cardiac myocytes were the target of adenovirus-mediated gene transfer. A time course of gene expression showed that beta-gal enzymatic activity peaked during the first week following injection. Adenovirus vectors provide an efficient but transient method for in vivo gene expression in myocardium.

Efficient and selective adenovirus-mediated gene transfer into vascular neointima.

BACKGROUND: Previous attempts to target arterial smooth muscle cells (SMCs) for gene delivery using liposomal or retroviral methods were limited by low transfection efficiency. We therefore evaluated the efficiency of adenovirus-mediated gene delivery in cultured vascular SMCs and in an in vivo model of balloon injury-induced SMC cell proliferation. METHODS AND RESULTS: We used a recombinant adenovirus, Ad.RSV beta gal, which contained the beta-galactosidase (beta-gal) histochemical marker gene. For in vitro studies, rat aortic SMCs were incubated in media containing Ad.RSV beta gal for 5 to 120 minutes. The proportion of SMCs expressing the beta-gal gene product increased from 25% (5-minute exposure) to 80% (120-minute exposure). For in vivo studies, uninjured and injured rat carotid segments were incubated with 0.5 to 1.0 x 10(9) pfu Ad.RSV beta gal for 45 minutes. Uninjured arteries showed adenovirus-mediated gene transfer limited to the endothelium. Injured arteries were exposed to adenovirus 0, 3, 7, or 12 days after injury. In these segments, beta-gal expression was minimal with infection at 0 or 3 days after injury but marked when infection was delayed until 7 or 12 days after injury. Neointimal cells constituted the dominant target of adenovirus gene transfer, with efficiency of gene transfer ranging from 10% to > 75%. Medial SMCs, whether covered or uncovered by neointimal cells, were minimally infected. Infection with a control adenovirus vector showed no beta-gal staining. CONCLUSIONS: Recombinant adenovirus selectively targets neointimal cells with high-efficiency gene transfer. This suggests that adenovirus vectors should be useful in targeting cells for the delivery of genes whose products may be relevant to the treatment of restenosis.

Physics of thin-section MR imaging at low field strength.

Thin-section magnetic resonance imaging at low field strengths requires analysis of the relative merits of data collection techniques for direct three-dimensional versus multisection two-dimensional imaging. This analysis was done with specific emphasis on shortened relaxation times and the use of reduced magnetic field gradients. Three-dimensional Fourier transform techniques can provide thin-section images of good diagnostic quality when combined with partial flip-angle gradient-reversal techniques.

Influence of dosage regimen on responses of the arcuate nucleus to subcutaneous injection of a protein hydrolysate.

The effect of administration of a glutamate-containing protein hydrolysate on the arcuate nucleus of 10-day-old mice was studied by two methods. Arcuate nucleus damage resulted when administration was by a single large subcutaneous dose (100 ml/kg). When the same total dose was administered subcutaneously in five small doses (20 ml/kg) over a period of 8 h, the damage to the arcuate nucleus did not occur. The latter method of administration was to simulate a clinical infusion. The results demonstrate that there is no hazard to the arcuate nucleus w-en glutamate-containing protein hydrolysates are administered by infusion.