PECAM-1/IgG attenuates peroxynitrite-mediated extremity reperfusion injury.

OBJECTIVE: Neutrophil transendothelial migration, a key feature of skeletal muscle ischemia and reperfusion (I/R) injury, is mediated by the platelet endothelial cell adhesion molecule-1 (PECAM-1). Peroxynitrite anion, a toxic product of neutrophil superoxide anion and nitric oxide, contributes to oxidative skeletal muscle injury and can be quantified by measurement of protein tyrosine nitration after I/R. This study hypothesizes that administration of the PECAM-1/IgG antibody chimera will inhibit peroxynitrite-mediated injury after I/R. METHODS: The study was composed of five groups: an I/R group (n = 4), a sham treatment group anesthetic control (n = 3), a treatment group receiving the PECAM-1/immunoglobulin G (IgG) antibody chimera with I/R (n = 9), a treatment group receiving human IgG with I/R as an antibody control (n = 6), and a treatment group receiving normal saline solution with I/R as a vehicle control (n = 5). The right hind limb in male New Zealand white rabbits was rendered ischemic by occluding the iliac and femoral arteries for 3 hours, followed by 2 hours of reperfusion (I/R). Sham-treated rabbits underwent arterial dissection without arterial occlusion. PECAM-1/IgG-treated rabbits and IgG-treated rabbits received an infusion of 1 mg/kg in normal saline solution 20 mL via an ear vein catheter during the last 5 minutes of ischemia and the first 15 minutes of reperfusion. Saline solution-treated rabbits similarly received normal saline solution 20 mL. The anterior tibialis muscle was harvested after reperfusion. Immunohistochemical staining for nitrotyrosine was performed with monoclonal antinitrotyrosine antibodies and fluorescently labeled secondary antibodies. Computed morphometric study was performed to calculate relative fluorescence scores for each histologic section. Averaged fluorescence scores were analyzed by one-way analysis of variance with Bonferroni post hoc comparison. RESULTS: The averaged fluorescence scores (mean +/- SEM) for the sham-treated (2.88 +/- 0.78) and PECAM-1/IgG-treated (6.16 +/- 0.43) groups demonstrated a significant reduction in quantitative fluorescence compared with the IgG- (15.17 +/- 2.01) and saline solution-treated (17.46 +/- 3.71) control groups, and the I/R-treated (18.52 +/- 3.00) group, (P <.05). CONCLUSIONS: These results suggest that PECAM-1/IgG diminishes peroxynitrite-mediated oxidative skeletal muscle injury by inhibiting neutrophil transendothelial migration and may therefore prove a useful therapeutic agent in the treatment of reperfusion injury.

Lessons learned in adopting endovascular techniques for treating abdominal aortic aneurysm.

HYPOTHESIS: Endovascular exclusion of abdominal aortic and common iliac aneurysms can be performed safely, and in the short term represents a feasible alternative to traditional, open aneurysm repair. PATIENTS AND METHODS: Forty-one patients were treated with endovascular grafts for 39 abdominal aortic and 2 common iliac artery aneurysms. RESULTS: All devices were successfully deployed. The size of the abdominal aortic aneurysms varied from 4.9 to 11.9 cm (average, 6.13 cm). The median procedure time was 195 minutes. There was one iliac artery rupture, which required celiotomy for repair. The hospital stay varied from 2 to 39 days (average, 6.7 days). The perioperative mortality rate was 2.4%. Sixteen patients (39%) had groin wound complications. Ten patients (24%) had evidence of contrast (endoleak) within the aneurysm sac on completion of the procedure. There were no obvious direct leaks from either the point of proximal or distal fixation. Seven of these endoleaks have resolved spontaneously. Two patients required additional procedures in the postoperative period to treat endoleak. The final patient has evidence of persistent endoleak on 3-month surveillance computed tomography scan. Major late problems occurred in 3 patients. CONCLUSION: Patients with large abdominal aortic aneurysms and considerable cardiac comorbidity can safely undergo endovascular aneurysm repair. Femoral groin wound complications resulting in prolonged hospitalization remain the major cause of perioperative morbidity. In contradistinction to open aneurysm repair, long-term surveillance is essential to detect migration of the device and identify flow within the residual aneurysm sac-complications that could lead to aneurysm rupture following endovascular repair.

Determinates of functional disability after complex upper extremity trauma.

This is a retrospective chart review of 71 patients who were operated on for presumed upper extremity arterial trauma between June 1992 and June 1998. Penetrating trauma occurred in 50 (70%) patients, and blunt trauma in 21 (30%). There were 2 innominate, 6 subclavian, 13 axillary, 26 brachial, 5 radial, 6 ulnar, and 6 multiple arterial injuries. There were 7 negative explorations (4 venous injuries, 2 false-positive angiograms, and 1 branch artery injury). In addition to the vascular injury, 44 patients (69%) had another injury in the extremity, including 8 (12.5%) orthopedic injuries, 12 (19%) nerve injuries, and 24 (37.5%) combination nerve and orthopedic injuries. There were three arterial thromboses, one arterial disruption, and four amputations, resulting in a patency rate and limb salvage rate of 94%. Persistent disability was more common in those patients with blunt injury (p = 0.02) and in those patients with associated neurologic and orthopedic injuries (p < 0.05). Full functional recovery was seen in 21 (33%) patients, while some form of disability was noted in the remaining 67%. The magnitude of the concomitant neurologic injury was the major determinate of functional outcome in this patient population.

Shear force regulates matrix metalloproteinase activity in human saphenous vein organ culture.

BACKGROUND: Development of vein graft intimal hyperplasia has been related both to shear force and to the activity of matrix metalloproteinases (MMPs). Little data are available regarding the effects of shear on MMP expression and activity. The aim of this study was to examine the relationship among shear force, metalloproteinase activity, and intimal thickening in human saphenous vein segments maintained in organ culture. MATERIALS AND METHODS: Segments of human saphenous vein were cultured under static conditions, or perfused under low-flow and high-flow conditions in a perfusion apparatus for 7 days. Metalloproteinase levels and activities were measured using ELISA and substrate gel zymography, respectively. Intimal thickening was determined by morphometric analysis. Results were compared with control vein tissue, which was not subjected to organ culture, using a one-way ANOVA. RESULTS: A 13% increase in proteolytic activity was noted on substrate gel zymography at 68-72 kDa in high-flow vein tissue. The protein content of MMP-2, MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), and TIMP-2 was increased in high-flow vein tissue by 21%, 126%, more than 100-fold, and 86%, respectively. In culture media bathing the outside of the vein, TIMP-2 was increased in high-flow specimens, while TIMP-1 was inversely related to flow rate. Intimal thickening was directly related to flow rates, and was progressively increased in the low-flow and high-flow groups by 3-fold and 4-fold, respectively. CONCLUSIONS: Metalloproteinase levels in human saphenous vein cultures are related to shear force. MMP levels and activity correlate with the degree of intimal thickening. This model may provide a valuable tool for the analysis of physical forces and their influence on intimal thickening in human saphenous vein.

Valuing educational scholarship at the Medical College of Wisconsin.

Since the late 1980s faculty and staff at the Medical College of Wisconsin (MCW) have actively sought to align their school's academic culture and promotional process with its mission of educational excellence and innovation. As one of the top 50 medical schools receiving NIH funds, MCW has well-established mechanisms to evaluate and recognize the scholarship of discovery. Understanding, evaluating, and recognizing the value of individuals engaged in the scholarship of teaching, however, required changes in individuals' beliefs and in the MCW's promotion processes and organizational infrastructure. Building on the successful introduction of the MCW's Educator's PortfolioCopyright, a tool for documenting educational scholarship, a multifaceted change strategy was implemented to influence underlying beliefs and values about clinician-educators. Retrospectively, this strategy was consistent with John Kotter's eight-step change model, which the authors apply as an organizing framework for this case report of educational evolution at the MCW. Through creating a guiding coalition, developing vision and strategy, generating short-term wins, and anchoring new approaches in the MCW's culture, the MCW has made substantive progress in recognizing and rewarding educational scholarship. Changing academic cultures to value education is itself an educational process, requiring persistence and the ability to teach others about educational scholarship and its associated criteria.

All-trans-retinoic acid decreases cell proliferation and increases apoptosis in an animal model of vein bypass grafting.

BACKGROUND: We have previously demonstrated a decrease in intimal hyperplasia in vein bypass grafts from animals treated with all-trans-retinoic acid (atRA). The purpose of this study was to examine the effect of atRA on proliferation and apoptosis rates in healing vein bypass grafts. METHODS: Interposition jugular vein bypass grafts were placed in the carotid artery of 30 New Zealand white rabbits. Animals received either atRA (10 mg/kg/d) or vehicle (corn oil) for a period of 2 weeks. Animals were killed at 3, 7, or 28 days after graft placement after having received 3 doses of 5-bromo-2'-¿Deoxyuridine (BRDU, 35 Mg/KG). Animals Were Perfusion Fixed, And Vein Grafts Were Prepared For Immunohistochemistry By Using Antibodies To Brdu, Proliferating Cell Nuclear Antigen, And Bcl-XL. Apoptosis Was Measured By Using The Tunel Assay. Histologic Sections Were Analyzed By A Pathologist Blinded To The Study, And An Index Of Positively Stained Cells Was Generated For Each Layer Of The Vein Graft Wall. RESULTS: All-trans-retinoic acid reduced the proliferation index in the neointima of vein grafts during the first week after surgery. Apoptotic rates were higher in the intima of vein grafts from animals treated with atRA, which could not be explained by changes in bcl-xl expression. No differences were noted in the media or adventitia between the groups. CONCLUSIONS: atRA decreased cell proliferation and increased apoptosis in the intima of healing vein bypass grafts. These effects contribute to decreased intimal hyperplasia, which has been previously noted.

All-trans-retinoic acid decreases vein graft intimal hyperplasia and matrix metalloproteinase activity in vivo.

BACKGROUND: Development of vein graft intimal hyperplasia has been associated with increased activity of matrix metalloproteinases (MMPs). All-trans-retinoic acid (atRA) decreases expression and activity of MMPs in tissue culture and has decreased intimal hyperplasia following arterial balloon catheter injury. We examined the effect of oral administration of atRA on intimal hyperplasia and MMP expression in an animal model of vein bypass grafting. MATERIALS AND METHODS: Interposition jugular vein bypass grafts were placed in the carotid artery of New Zealand white rabbits. Animals received either atRA (10 mg/kg/day) or vehicle (corn oil) for a period of 2 weeks. Retinoic acid serum levels were determined by HPLC. Intimal and medial areas were measured using morphometric analysis of perfusion-fixed vein graft specimens, and intimal thickness was calculated using circumferential measurements. Expression of MMP-2, MMP-9, and TIMP-1 in vein grafts and unoperated control veins was determined using Northern analysis, and proteolytic activity was determined using substrate gel zymography. RESULTS: Animals treated with atRA had significantly elevated serum levels of this compound and its metabolites. A decrease in intimal to medial ratio was noted after 28 days in vein grafts from treated animals (0.63 vs 0.88, P < 0.01), and a decrease in calculated intimal thickness was noted at 7 and 28 days. Expression of MMP-2 was decreased in treated animals 7 days following surgery, and expression of both MMP-2 and MMP-9 was decreased at 28 days. A decrease in proteolytic activity was noted on zymography at 68 kDa, 7 and 28 days following surgery in vein grafts from animals treated with atRA, corresponding with a decrease in the active form of MMP-2. Increased expression of TIMP-1 was noted in vein grafts from both the treated and the control groups, 7 and 28 days following graft placement. CONCLUSIONS: Oral administration of all-trans-retinoic acid resulted in decreased intimal hyperplasia in an animal model of vein bypass grafting. This was associated with decreased expression and activity of MMP-2 in treated animals.

Intraoperative carotid evaluation.

HYPOTHESIS: Intraoperative duplex scanning can identify technical defects and increase the quality of carotid artery repair. DESIGN: We evaluated 100 consecutive carotid operations in 96 patients (60 men and 36 women) from 1995 to 1998. Spectral-derived peak systolic flow velocities (PSV) were graded (PSV < 100 cm/s, normal laminar flow; PSV 100-150 cm/s, mild or moderate flow disturbance; PSV > 150 cm/s, severe flow disturbance). Prospective criteria for intraoperative revision included PSV greater than 150 cm/s, spectral broadening, and B-mode imaging of intimal flaps or intraluminal debris. Preoperative, intraoperative, and 6-week follow-up duplex scan results were analyzed. SETTING: All patients were evaluated and treated at a single academic institution. INTERVENTIONS: All procedures were performed with the patient under general endotracheal anesthesia; 86% underwent shunting and 70% underwent patching. MAIN OUTCOME MEASURE: Number and type of revisions, patency of repair, residual and recurrent stenosis, and ipsilateral neurologic events. RESULTS: There were 33 intraoperative duplex studies with abnormal findings. Seven involved the common carotid artery and resulted in intraoperative revision of 5 intimal flaps at the site of the proximal clamp. In 11 patients, incomplete eversion endarterectomy resulted in elevated distal intimal flaps in the external carotid artery that were repaired through a separate arteriotomy. There were 15 abnormalities in the internal carotid artery prompting 5 revisions. Five studies with PSV of 100 to 150 cm/s had no defects on B-mode imaging and were observed without treatment. Five false-positive studies were attributed to increased flow velocity due to contralateral occlusive discase. At 6 weeks' follow-up, 4 of 5 repaired common carotid arteries were normal on duplex scan and 1 had a mild residual stenosis. Ten of the 11 external carotid repairs were patent and 1 was occluded. Four of the 5 internal carotid artery repairs were normal on postoperative evaluation and 1 had a mild residual stenosis. Of the 10 abnormal internal carotid arteries that were observed, 9 were normal on postoperative duplex and 1 had a mild residual stenosis. One perioperative stroke occurred in a patient with a normal, patent carotid repair. CONCLUSIONS: Intraoperative duplex evaluation of carotid reconstruction is an efficient, sensitive tool that can detect technical lesions that will jeopardize surgical reconstruction. Interpretive judgment is required because all flow disturbances do not dictate surgical intervention. This technique enables the surgeon to maximize the quality of the arterial reconstruction during carotid artery surgery.

Metalloproteinase levels are decreased in symptomatic carotid plaques.

BACKGROUND: Matrix metalloproteinase enzymes (MMP) have been identified in carotid atherosclerotic plaques, but their role in the development of clinical symptoms remains ill defined. We correlated the activity and levels of metalloproteinase enzymes and their inhibitors in human carotid plaques to ischemic neurologic events. METHODS: Carotid plaques were collected at the time of endarterectomy from 23 patients with carotid stenosis. Sixteen patients were asymptomatic and 7 patients had symptoms of stroke or transient ischemic attack within 6 weeks of surgery. Protein was extracted from the plaques, proteolytic activity was determined by gelatin zymography, and pro-MMP and tissue inhibitor of metalloproteinase (TIMP) enzyme content were measured by ELISA assay. Macrophage accumulation in the plaque was determined using immunohistochemistry. RESULTS: Plaques from symptomatic patients had decreased proteolytic activity on substrate gel zymography at the 62- and 92-kDa regions (corresponding to active MMP-2 and pro-MMP-9). A decrease in pro-MMP-9 (8.21 +/- 2.35 vs 17.42 +/- 3.14 ng, P < 0. 05) and an increase in TIMP-2 protein (12.62 +/- 0.58 vs 10.56 +/- 0. 77 ng, P < 0.05) were noted on ELISA in plaques from symptomatic patients. No difference was noted in macrophage accumulation in the plaques between the two groups. CONCLUSIONS: Plaques from patients who present with ischemic neurologic symptoms have decreased proteolytic activity associated with decreased pro-MMP-9 and increased TIMP-2 protein levels. These data suggest that metalloproteinase enzymes are not responsible for plaque instability in the carotid circulation and may in fact promote plaque stability.

Endovascular therapy of iliac arteries: routine application of intraluminal stents does not improve clinical patency.

Our objective in this study was to review our experience with endovascular therapy of iliac artery occlusive disease over the past decade, and to compare the results of angioplasty alone with the addition of endovascular stents to these procedures. This report details a retrospective analysis of clinical data on 141 consecutive patients with iliac artery occlusive disease, treated by balloon angioplasty alone, or with the addition of intraluminal stents. The procedures analyzed included 58 common iliac artery interventions (26 angioplasties and 32 stent insertions) and 83 external iliac artery procedures (43 angioplasties and 40 stent insertions). Early and continued success, and their components, are reported and compared according to published standards. While endovascular therapy of iliac artery occlusive disease is effective in relieving symptoms, clinical patency rates are lower than those reported for direct reconstruction. Primary stent placement has not enhanced clinical patency in the iliac arteries, and the selective insertion of these devices for more complicated angioplasty procedures seems warranted.

cGMP is decreased after acute ischemia in chronically ischemic canine limbs.

BACKGROUND: A chronic partially ischemic state may alter the skeletal muscle response to acute ischemia and free radical formation. METHODS: In order to investigate this hypothesis, a chronic ischemic state was established by ligating the right femoral artery of four mongrel dogs. ABIs were decreased from 1.05 +/- 0.25 preligation to 0.54 +/- 0.14 at 6 weeks (P = 0.04). At the end of 8 weeks, the hindlimb was subjected to 3 h of acute ischemia by clamping the iliac artery. The clamp was then released for 2 h of reperfusion. Plasma samples from the right iliac vein were taken during the ischemia-reperfusion period for analysis of cGMP. Tibialis anterior biopsies for Western analysis of eNOS and iNOS were taken upon completion of reperfusion. Comparisons to control dogs subjected to the acute ischemia and reperfusion without prior femoral artery ligation were made. RESULTS: cGMP levels were increased in the controls at 3 h of ischemia (3539 +/- 350) and 2 h of reperfusion (2880 +/- 269). The chronic ischemia group did not develop a corresponding increase in cGMP at 3 h of ischemia (2762 +/- 251) or after 2 h of reperfusion (2102 +/- 130). Western analysis of eNOS and iNOS revealed similar levels in both groups. Analysis of eNOS revealed 0.6429 +/- 0.086 and 0.5916 +/- 0.072 (densitometric units +/- SEM) for study and control dogs, respectively. Analysis of iNOS revealed 0.3401 +/- 0.067 and 0.2475 +/- 0.066 for study and control dogs, respectively. CONCLUSION: Previous ligation of the femoral artery resulting in chronic partial ischemia in this model demonstrated no increase in cGMP following acute ischemia that was not accompanied by a change in eNOS or iNOS levels. Nitric oxide activity is reflected by cGMP levels, which may increase in response to free radicals in the acute setting of complete ischemia.

Cholesterol, but not cigarette smoke, decreases rabbit carotid artery relaxation.

The purpose of this study was to determine the physiologic effects of cigarette smoke exposure and dietary cholesterol on the availability of nitric oxide in carotid vascular rings. New Zealand white rabbits were placed in an airflow chamber for 3 hr/day over an 8-week period and were exposed to smoke from 600 cigarettes/per day added to the chamber inflow by a robotic smoke generator. New Zealand white rabbits, made hypercholesterolemic, and one group fed a normal diet, were similarly placed in the chamber without exposure to cigarette smoke. In those exposed groups, serum cotinine and cholesterol levels were consistently elevated. After the 8-week period, the carotid arteries were harvested. The vessels were cut into 3-mm rings which were suspended from pressure transducers. The rings were contracted with potassium chloride (KCl) to determine vessel integrity. One ring from each carotid was maximally contracted with 1 x 10(-3) molar norepinephrine (NE) while the experimental ring was contracted to 50% of maximum. Relaxation of the rings was achieved by adding incremental doses of acetylcholine. Our results showed that endothelial dysfunction, as measured by acetylcholine-mediated vasorelaxation, occurs in the rabbit carotid artery when exposed to high dietary cholesterol. Cigarette exposure alone in this particular vessel did not result in significant alteration in acetylcholine-mediated vasorelaxation.

Health-related quality of life and functional outcome following arterial reconstruction for limb salvage.

Vascular surgery outcomes have traditionally been measured by limb salvage and graft patency. However, as health care resources are rationed, the patient's functional outcome and quality of life will require assessment. The in situ saphenous vein graft has proven successful in achieving long-term limb salvage for patients with critical ischemia, with the expectation of preserving a life-style and sense of well-being that would be lost with limb amputation. This study was conducted to measure functional capacity and quality of life in these patients. Seventy patients with successful in situ saphenous vein bypass grafts constructed for limb-threatening ischemia, followed for a mean of 45.6 months in a surveillance program with normal graft flow characteristics, were compared with a group of age and gender-matched controls with normal limb pressures and no history of vascular occlusive disease. A questionnaire was designed from standardized health status scales and administered to the two groups to assess symptoms, health perceptions, physical functioning and life quality. When comparing the groups of revascularized and control patients, symptoms and perceptions about their health were similar. However, the revascularized patients had significantly decreased functional capacity in their ability to walk various distances (P< or =0.005), perform household tasks (P< or =0.001) and bathe (P< or =0.001). The patient group with vascular grafts functioned as well as the controls only in activities of dressing and using the toilet. Indicators of life quality that rate independence and mobility, including the ability to procure groceries (P< or = 0.001), prepare meals (P< or =0.005) participate in social activities (P< or =0.001) and drive an automobile (P< or =0.01), were also significantly limited in the patients with successful vascular reconstructions. Although achieving long-term limb salvage and graft patency, the patients in this group of successful vascular reconstructions retain functional disabilities that require significant care. Despite these physical handicaps, these patients have a remarkably similar sense of well-being and lack of somatic complaints compared with the control group. This medical outcome study identifies the functional capacity and lifetime needs for vascular surgery patients that will provide useful data for those responsible for allocating health care resources.

Ascorbic acid prevents cigarette smoke injury to endothelium-dependent arterial relaxation.

BACKGROUND: Arterial endothelium-dependent acetylcholine relaxation is impaired by smoking, and this injury may be mediated by oxygen free radicals. The purpose of this study was to examine the effect of ascorbic acid (AA) treatment on this injury. MATERIALS AND METHODS: New Zealand White rabbits (n = 6, each group) were placed in a 240-ft3 airflow chamber for 3 h per day, 5 days per week over an 8-week period. Animals were divided into four groups and fed rabbit chow and water or rabbit chow and AA (250 ml/kg)-supplemented water. The control-smoke rabbit group and the ascorbic acid-smoke rabbit group were exposed to mainstream cigarette smoke from a robotic smoke generator for the 3-h period, while the control-no smoke rabbit group and ascorbic acid-no smoke rabbit group were similarly placed in the chamber without smoke. At the end of 8 weeks, rabbits were sacrificed and segments of their superficial femoral arteries were suspended from tension transducers and the maximal contraction was determined. The remaining rings were contracted to 50% of the maximum and relaxation was determined by adding acetylcholine. Groups were compared using one-way ANOVA. RESULTS: Rings from control-smoke (5.13 +/- 0.21 g) and AA-smoke rabbits (6. 24 +/- 0.46 g) exhibited increased mean contraction to KCl (P < 0. 05) compared to control-no smoke rabbits (3.86 +/- 0.40 g). Acetylcholine-dependent relaxation was significantly reduced in the rings from the control-smoke rabbits compared to control-no smoke rabbits (acetylcholine, 5 x 10(-7) M: 24.7 +/- 2.7% versus 55.3 +/- 8.0%; acetylcholine, 7 x 10(-7) M: 27.5 +/- 2.3% versus 56.3 +/- 9. 2%). The AA-smoke group (acetylcholine, 5 x 10(-7) M: 61.8 +/- 12. 4%; 7 x 10(-7) M: 67.9 +/- 11.4%) had significantly increased relaxation compared to the control-smoke group (P < 0.05). There was no statistical difference in the mean percentage ring relaxation between the control-no smoke, AA-no smoke, and AA-smoke groups. CONCLUSIONS: Ascorbic acid protected the artery from cigarette smoke-induced endothelial injury.