New methods for time-resolved fluorescence spectroscopy data analysis based on the Laguerre expansion technique--applications in tissue diagnosis.

OBJECTIVES: A new deconvolution method for the analysis of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data is introduced and applied for tissue diagnosis. METHOD: The intrinsic TR-LIFS decays are expanded on a Laguerre basis, and the computed Laguerre expansion coefficients (LEC) are used to characterize the sample fluorescence emission. The method was applied for the diagnosis of atherosclerotic vulnerable plaques. RESULTS: At a first stage, using a rabbit atherosclerotic model, 73 TR-LIFS in-vivo measurements from the normal and atherosclerotic aorta segments of eight rabbits were taken. The Laguerre deconvolution technique was able to accurately deconvolve the TR-LIFS measurements. More interesting, the LEC reflected the changes in the arterial biochemical composition and provided discrimination of lesions rich in macrophages/foam-cells with high sensitivity (> 85%) and specificity (> 95%). At a second stage, 348 TR-LIFS measurements were obtained from the explanted carotid arteries of 30 patients. Lesions with significant inflammatory cells (macrophages/foam-cells and lymphocytes) were detected with high sensitivity (> 80%) and specificity (> 90%), using LEC-based classifiers. CONCLUSION: This study has demonstrated the potential of using TR-LIFS information by means of LEC for in vivo tissue diagnosis, and specifically for detecting inflammation in atherosclerotic lesions, a key marker of plaque vulnerability.

Diagnosis of vulnerable atherosclerotic plaques by time-resolved fluorescence spectroscopy and ultrasound imaging.

In this study, time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) and ultrasonography were applied to detect vulnerable (high-risk) atherosclerotic plaque. A total of 813 TR-LIFS measurements were taken from carotid plaques of 65 patients, and subsequently analyzed using the Laguerre deconvolution technique. The investigated spots were classified by histopathology as thin, fibrotic, calcified, low-inflamed, inflamed and necrotic lesions. Spectral and time-resolved parameters (normalized intensity values and Laguerre expansion coefficients) were extracted from the TR-LIFS data. Feature selection for classification was performed by either analysis of variance (ANOVA) or principal component analysis (PCA). A stepwise linear discriminant analysis algorithm was developed for detecting inflamed and necrotic lesion, representing the most vulnerable plaques. These vulnerable plaques were detected with high sensitivity (>80%) and specificity (>90%). Ultrasound (US) imaging was obtained in 4 carotid plaques in addition to TR-LIFS examination. Preliminary results indicate that US provides important structural information of the plaques that could be combined with the compositional information obtained by TR-LIFS, to obtain a more accurate diagnosis of vulnerable atherosclerotic plaque.

Application of the laguerre deconvolution method for time-resolved fluorescence spectroscopy to the characterization of atherosclerotic plaques.

This study investigates the ability of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) to detect inflammation in atherosclerotic lesion, a key feature of plaque vulnerability. A total of 348 TR-LIFS measurements were taken from carotid plaques of 30 patients, and subsequently analyzed using the Laguerre deconvolution technique. The investigated spots were classified as Early, Fibrotic/Calcified or Inflamed lesions. A stepwise linear discriminant analysis algorithm was developed using spectral and TR features (normalized intensity values and Laguerre expansion coefficients at discrete emission wavelengths, respectively). Features from only three emission wavelengths (390, 450 and 500 nm) were used in the classifier. The Inflamed lesions were discriminated with sensitivity > 80% and specificity > 90 %, when the Laguerre expansion coefficients were included in the feature space. These results indicate that TR-LIFS information derived from the Laguerre expansion coefficients at few selected emission wavelengths can discriminate inflammation in atherosclerotic plaques. We believe that TR-LIFS derived Laguerre expansion coefficients can provide a valuable additional dimension for the detection of vulnerable plaques.

Novel methods of time-resolved fluorescence data analysis for in-vivo tissue characterization: application to atherosclerosis.

This study investigates the ability of new analytical methods of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data to characterize tissue in-vivo, such as the composition of atherosclerotic vulnerable plaques. A total of 73 TR-LIFS measurements were taken in-vivo from the aorta of 8 rabbits, and subsequently analyzed using the Laguerre deconvolution technique. The investigated spots were classified as normal aorta, thin or thick lesions, and lesions rich in either collagen or macrophages/foam-cells. Different linear and nonlinear classification algorithms (linear discriminant analysis, stepwise linear discriminant analysis, principal component analysis, and feedforward neural networks) were developed using spectral and TR features (ratios of intensity values and Laguerre expansion coefficients, respectively). Normal intima and thin lesions were discriminated from thick lesions (sensitivity >90%, specificity 100%) using only spectral features. However, both spectral and time-resolved features were necessary to discriminate thick lesions rich in collagen from thick lesions rich in foam cells (sensitivity >85%, specificity >93%), and thin lesions rich in foam cells from normal aorta and thin lesions rich in collagen (sensitivity >85%, specificity >94%). Based on these findings, we believe that TR-LIFS information derived from the Laguerre expansion coefficients can provide a valuable additional dimension for in-vivo tissue characterization.

Rat and human aortic smooth muscle cells display differing migration and matrix metalloproteinase activities in response to dexamethasone.

OBJECTIVE: The steroid dexamethasone inhibits neointimal hyperplasia development in rats but not in humans. This study investigates the differential effects of dexamethasone on rat and human smooth muscle cell migration and matrix metalloproteinase (MMP) activity. METHODS: Rat aortic smooth muscle cells were harvested from Sprague-Dawley rats. Human aortic smooth muscle cells were obtained from Clonetics. Boyden chamber migration assays were performed with chemoattractant (platelet-derived growth factor) and varying concentrations of dexamethasone (10(-9) to 10(-5) mol/L). Zymography of culture media was used to assess MMP activity, and Western blot analysis was used for quantification of MMP-2 and tissue inhibitor of MMP-2 (TIMP-2) secretion. RESULTS: Dexamethasone inhibits rat aortic smooth muscle cell migration in a dose-dependent fashion. An increase in concentrations of dexamethasone does not effect human aortic smooth muscle cell migration. Rat aortic smooth muscle cell MMP-2 activity is inhibited with dexamethasone in a dose-dependent fashion, and human aortic smooth muscle cell MMP-2 activity is unchanged with dexamethasone. MMP-2 secretion is inhibited with dexamethasone in rat aortic smooth muscle cells but remains unaltered in human aortic smooth muscle cells. Dexamethasone increases rat aortic smooth muscle cell TIMP-2 secretion, and human aortic smooth muscle cell TIMP-2 secretion remains constant. CONCLUSION: Dexamethasone inhibits rat aortic smooth muscle cell migration, MMP-2 activity, and MMP-2 secretion and increases TIMP-2 secretion. These effects are not observed in human aortic smooth muscle cells. These findings may explain why dexamethasone inhibits neointimal hyperplasia in animal models but is ineffective in humans. Inhibition of human smooth muscle cell migration in vitro may be useful in predicting the effectiveness of future therapeutic agents for treatment of neointimal hyperplasia in humans.

Dexamethasone inhibits vascular smooth muscle cell migration via modulation of matrix metalloproteinase activity.

BACKGROUND: Dexamethasone (DEX) has been shown to inhibit development of neointimal hyperplasia in rats. We hypothesize that DEX inhibits neointimal hyperplasia by altering matrix metalloproteinase (MMP) activity, resulting in inhibition of smooth muscle cell migration. METHODS: Rat aortic smooth muscle cells (RASMC) were harvested and cultured for two to four passages. A migration assay was performed in a Boyden chamber with chemoattractant (platelet-derived growth factor) and varying concentrations of DEX (10(-9) to 10(-5) M). The number of migrated cells was counted under light microscopy. Zymography was performed on culture media to assess MMP activity, and Western blotting was performed to assay MMP and levels of tissue inhibitors of MMPs (TIMPs). RESULTS: DEX progressively inhibited RASMC migration in a dose-dependent fashion. This effect was statistically significant for concentrations of 10(-7) to 10(-5) M (P < 0.0005). Zymography showed that DEX inhibits MMP-2 activity in a dose-dependent manner. Western blots indicated that total MMP-2 secretion was inhibited and that TIMP-2 secretion was increased by DEX. CONCLUSIONS: DEX inhibits platelet-derived growth factor-induced migration of RASMCs and MMP-2 activity in vitro. Our data suggest that DEX suppresses MMP activity and secretion, resulting in the inhibition of smooth muscle cell migration. This may explain the mechanism by which DEX inhibits neointimal hyperplasia.

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.

Acute arterial thrombosis causes endothelial dysfunction: a new paradigm for thrombolytic therapy.

PURPOSE: The goals of this study were to delineate the time course of endothelial dysfunction after arterial thrombosis, to determine the cause of endothelial dysfunction in this setting, and to determine whether modulating standard thrombolytic therapy would ameliorate the thrombosis-mediated endothelial dysfunction. METHODS: Male adult rats underwent infrarenal aortic occlusion by means of clip ligature to induce arterial thrombosis. After 30 minutes, 1, 2, and 3 hours, ring segments from the infrarenal aorta were harvested and placed into physiologic buffer baths. With the use of a force transducer, both endothelial-dependent relaxation (EDR) and endothelial-independent relaxation (EIR) were measured. Endothelial function and presence were determined by means of factor VIII immunohistochemical staining. Endothelial morphology was evaluated with scanning electron microscopy (SEM). Nitric oxide (NO) levels were determined with a chemiluminescent assay of its nitrite/nitrate metabolites (NO(x)). Standard thrombolytic therapy with urokinase (UK) was infused into thrombosed aortic ring segments and compared with UK supplemented with both low-dose L -arginine (2 mmol) and high-dose L -arginine (20 mmol). RESULTS: Arterial thrombosis decreases EDR. The nadir of EDR occurs 1 hour after thrombosis (mean +/- SE, 13% +/- 6.4% vs 94% +/- 2.6% for controls, P <.005), with persistent lowering of EDR as long as 3 hours after thrombosis. EIR is preserved, and vasoconstriction with norepinephrine or potassium buffer is unaltered. Both endothelial function and presence (n = 6 per group) were documented by means of factor VIII immunohistochemistry. An intact monolayer of endothelium at all time intervals after thrombosis was revealed by means of SEM analysis. No differences between control and thrombosed specimens were revealed by means of the grading of SEM images. Local NO(x) levels were lower after 1 hour of thrombosis, with an increase higher than baseline values at 3 hours. The addition of low-dose L -arginine resulted in a minor increase in EDR. However, high-dose L -arginine resulted in a significant increase in EDR versus controls receiving UK alone (64% +/- 6.3% vs 38% +/- 4.4%, P <.05). Correspondingly, local NO(x) levels were 20-fold higher after the high-dose L -arginine supplementation when compared with UK thrombolysis alone (2.8 +/- 0.52 micromol/L vs 0.133 +/- 0.02 micromol/L, n = 6 samples/group, P <.005). CONCLUSION: Acute arterial thrombosis causes endothelial dysfunction, without causing endothelial cell loss. Endothelial function reaches a nadir after 1 hour of thrombosis. EIR and vasoconstriction remain unaffected, indicating normal smooth muscle cell function. NO(x) levels suggest that NO levels are decreased acutely after thrombosis. Supplementing standard thrombolytic therapy with the NO precursor, l-arginine, ameliorates the endothelial dysfunction seen after acute thrombosis by increasing local NO production.

Combined carotid endarterectomy and coronary bypass: a decade experience at UCLA.

PURPOSE: The purpose of this review was to determine outcomes for combined carotid endarterectomy (CEA) and coronary revascularization (CABG) in patients with asymptomatic carotid stenosis. METHODS: We reviewed the medical records of consecutive combined procedures (CEA and CABG), performed at UCLA Medical Center from October, 1989 to January, 1999. FINDINGS: There were 43 patients, 27 men and 16 women, with a mean age of 71 yr (range 51-87). Thirty-four patients 79% (34/43) had asymptomatic carotid stenosis. Stroke occurred in three patients (3/43 = 6.9%). Stroke ipsilateral to the CEA occurred in two patients: one asymptomatic (1/34 = 2.9%) and one symptomatic (1/9 = 11.1%). CONCLUSIONS: The majority of patients undergoing combined CEA/CABG have asymptomatic carotid stenosis identified in preparation for elective CABG. The asymptomatic carotid subset stroke rate of 2.9% resulting from a combined CEA/CABG is higher than our reported rate for CEA performed alone. In patients with asymptomatic carotid stenosis, the combined procedure should be selectively performed.

Carotid endarterectomy with normal findings from a completion study: Is there need for early duplex scan?

OBJECTIVE: The objective of this study was to determine the value of early (< 6 months) duplex scanning after carotid endarterectomy (CEA) with an intraoperative completion study with normal results. Attention was paid to restenosis rates and reoperation for recurrent stenosis within the first 6 months. METHODS: A retrospective review was performed on 380 CEAs (338 patients) with intraoperative completion studies and duplex surveillance within the first 6 months. Results of completion studies, restenosis rates, and recurrent symptoms were evaluated for each operation. Studies were performed from 0 to 200 days postoperatively (median, 28). RESULTS: Intraoperative completion studies included 333 angiograms, 26 duplex scans, and 21 angiograms with duplex scans. Of the 380 intraoperative completion studies, 28 (7.5%) had abnormal findings, including 14 abnormal internal carotid arteries (ICAs). Twenty-four procedures were revised, and the findings of all repeat completion studies were normal. Of the initial completion studies, in four cases, abnormalities (3 ICAs) were insignificant and did not warrant further intervention. Follow-up ICA duplex scans had normal results after 364 (95.8%) CEAs. There were 14 mild recurrent ICA stenoses and two moderate recurrent ICA stenoses; neither had abnormal findings from the completion study. There were no severe recurrent ICA stenoses. External carotid artery (ECA) recurrent stenosis included 7 mild, 15 moderate, and 9 severe restenoses. CONCLUSIONS: Only 0.5% of CEAs developed moderate restenosis. No procedures had severe recurrent stenosis on duplex scan within the first 6 months, and none required intervention. Duplex surveillance in the first 6 months is relatively unproductive, providing that there were normal results from an intraoperative completion study for each patient. Routine surveillance can be started at 1 year.

Carotid body tumors: the role of preoperative embolization.

Resection of carotid body tumors (neck paragangliomas) carries inherent risks of injury to the cranial nerves and other structures as well excessive blood loss. Preoperative embolization has been used to lessen the morbidity in tumors that are larger than 2 cm in diameter. Two female patients presented for treatment with large asymptomatic carotid body tumors-one 4 cm and one 5 cm in diameter. Both patients had preoperative angiography the day before surgery that revealed the feeding arterial vessels so that successful embolization could be accomplished with gel. Success was judged by diminution of the angiographic blush. Both patients had an uneventful surgical excision the following day with the carotid body tumors being able to be resected periadventitially without damage to either the external or internal carotid artery. The cranial nerves were preserved in both patients and blood loss was only 200 cc in both cases. We conclude that preoperative embolization is an important adjunct in treating patients with large carotid body tumors. The surgical exploration proceeds much smoother, the blood loss is minimal, and patients have minimal morbidity.

Penetrating ulceration of the infrarenal aorta: case reports of an embolic and an asymptomatic lesion.

Penetrating aortic ulceration is uncommon in the infrarenal aorta. We describe a patient with a penetrating infrarenal aortic ulcer manifesting as blue toe syndrome, and a second patient with a similar lesion identified as an incidental finding. These two patients were treated for penetrating infrarenal aortic ulceration within the past 9 months at two university-affiliated hospitals, a regional Veterans Administration Medical Center, and a County Medical Center. Both lesions demonstrated aneurysm changes with varying degrees of mural thrombus. The lesion filled with fresh thrombus proved labile, with embolization manifesting as blue toe syndrome. We support the aggressive treatment of aneurysmal penetrating aortic ulcer with aortic graft replacement to eliminate the potential for distal embolization and to obviate the risk of rupture and death.

Safety and efficacy of early surgical decompression of the thoracic outlet for Paget-Schroetter syndrome.

The surgical treatment of Paget-Schroetter syndrome has evolved to include early thrombolytic therapy and an interval period of anticoagulation, followed by late surgical decompression of the thoracic outlet. More recently, we have developed an abbreviated course of therapy in which the thrombolytic therapy is followed by early surgical decompression during the same admission, then a period of anticoagulation. We compared early surgical decompression with the standard management protocol to determine safety and efficacy of the early treatment algorithm. Nine patients were treated with lysis and early operation. These were compared with the preceding nine consecutive patients treated with lysis and staged operation. Demographic data, risk factors, duration of thrombosis, lytic therapy, time to surgery, operative variables, and postoperative complications were analyzed. Our results showed that thrombolysis followed by early operation does not result in increased perioperative morbidity or mortality. Early surgical decompression of the thoracic outlet during the same admission as lysis is as safe and efficacious as the traditional (staged decompression) approach to Paget-Schroetter syndrome. Lysis followed by early surgical decompression should be considered a new standard of care in the management of Paget-Schroetter syndrome.

Selective botulinum chemodenervation of the scalene muscles for treatment of neurogenic thoracic outlet syndrome.

Patients with thoracic outlet syndrome (TOS) who improve temporarily after anesthetic blockade of the anterior scalene muscles have been shown to improve after ultimate surgical decompressions at the interscalene triangle. Anesthetic blockade of the scalene muscles, even with the addition of steroids, however, rarely produces any prolonged relief as patients are awaiting definitive surgery. The present study was undertaken to determine if more effective and prolonged relief might be obtained with electrophysiologically and fluoroscopically guided selective injection of the scalene muscles with botulinum toxin, which has been used in the past for treating conditions associated with spasm of cervical muscles. In 14 of 22 patients (64%) with a clinical diagnosis of TOS, there was more than a 50% reduction of symptoms measured by a 101-point scale for at least 1 month after botulinum chemodenervation of the scalene muscles. Only 4 of the 22 patients (18%) had a 50% reduction of symptoms for at least 1 month after injection with lidocaine and steroids. In no patient were the results of lidocaine and steroid injection superior to botulinum chemodenervation. Chemodenervation had a mean duration of effect of 88 days. No significant side effects were encountered with botulinum chemodenervation except for mild transient dysphagia in two cases. These results appear to demonstrate that botulinum chemodenervation of the scalene muscles may be helpful in alleviating symptoms in patients with TOS awaiting definitive surgical decompression.

Yield of repeated screening for abdominal aortic aneurysm after a 4-year interval. Aneurysm Detection and Management Veterans Affairs Cooperative Study Investigators.

BACKGROUND: Little is known about the rate at which new abdominal aortic aneurysms (AAAs) develop or whether screening older men for AAA, if undertaken, should be limited to once in a lifetime or repeated at intervals. METHODS: A large population of veterans, aged 50 through 79 years, completed a questionnaire and underwent ultrasound screening for AAA. Of these, 5151 without AAA on the initial ultrasound (defined as infrarenal aortic diameter of 3.0 cm or larger) were selected randomly to be invited for a second ultrasound screening after an interval of 4 years. Local records and national databases were searched to identify deaths and AAA diagnoses made during the study interval in subjects who did not attend the rescreening. RESULTS: Of the 5151 subjects selected for a second screening, 598 (11.6%) had died (none due to AAA), and 20 (0.4%) had an interim diagnosis of AAA. A second screening was performed on 2622 (50.9%), of whom 58 (2.2%; 95% confidence interval, 1.6%-2.8%) had new AAA. Three new AAAs were 4.0 to 4.9 cm, 10 were 3.5 to 3.9 cm, and 45 were 3.0 to 3.4 cm. Independent predictors of new AAA at the second screening included current smoker (odds ratio, 3.09; 95% confidence, 1.74-5.50), coronary artery disease (odds ratio, 1.81; 95% confidence interval, 1.07-3.07), and, in a separate model using a composite variable, any atherosclerosis (odds ratio, 1.97; 95% confidence interval, 1.16-3.35). Adding the interim and rescreening diagnosis rates suggests a 4-year incidence rate of 2.6%. Rescreening only in subjects with infrarenal aortic diameter of 2.5 cm or greater on the initial ultrasound would have missed more than two thirds of the new AAAs. CONCLUSIONS: A second screening is of little practical value after 4 years, mainly because the AAAs detected are small. However, the incidence that we observed suggests that a second screening after longer intervals (ie, more than 8 years) may provide yields similar to those seen in initial screening and therefore warrants further study.

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.

Gelatin-sealed polyester resists Staphylococcus epidermidis biofilm infection.

BACKGROUND: The purpose of this study was to show that gelatin-impregnated polyester grafts inhibit Staphylococcus epidermidis biofilm infection in a canine model of aortic graft interposition. A clinically native species and two engineered strains, which differed in slime and adhesin antigen components, were compared to determine differential gelatin and slime interactions. METHODS: In vitro bacterial graft colonization was validated by immersion of graft segments in inoculating solutions (10(6) colony forming units/ml) of a clinically native species RP62A and two genetically engineered S. epidermidis species, M187sn3 (SN3: slime and adhesin negative) or M187sp11 (SP11: slime and adhesin positive), for 18 h at 23 degrees C. The grafts were washed, sonicated, and cultured to assess in vitro bacterial graft adherence. Grafts similarly inoculated were placed as aortic interposition grafts in dogs. Three sterile grafts were implanted as controls. Grafts were excised after 6 weeks and cultured for bacterial growth as in the in vitro study. Infection was defined by a positive culture in the excised grafts. Data were analyzed with nonparametric statistical methods. RESULTS: In vitro bacterial graft adherence in colony forming units per milliliter was similar at 18 h postsonication for RP62A (8 x 10(4) +/- 1 x 10(4)), SN3 (7 x 10(4) +/- 2 x 10(4)), and SP11 (6 x 10(4) +/- 2 x 10(4)) (P = NS). Only one of five grafts inoculated with RP62A was culture positive after 6 weeks. No grafts inoculated with the engineered strains SN3 or SP11 were culture positive after explanation. CONCLUSION: In vitro bacterial inoculation of gelatin-impregnated polyester was similar among the species and not dependent upon the presence of slime and adhesin components. Gelatin-impregnated polyester grafts demonstrated in vivo resistance to coagulase-negative staphylococcal biofilm infection.

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.