Prospective comparison of intraoperative angiography with duplex scanning in evaluating lower-extremity bypass grafts in a community hospital.

The gold standard for intraoperative evaluation of lower-extremity bypass grafts has been angiography. Limitations of this technique include inability to measure flow dynamics, violation of graft integrity, cost, and length of assessment time. The goal of this study was to evaluate duplex scanning as an alternative modality for intraoperative graft assessment. Our study group comprised of 19 consecutive patients undergoing infrainguinal bypass procedures at our institution between March 1999 and March 2000. Intraoperative angiography was compared with duplex scanning by evaluating parameters of assessment time, graft flow velocities, serum creatinine levels, and 30-day graft patency rates. Mean study times were the following: cut-film angiography, 22 +/- 1.8 minutes; real-time fluoroscopy, 17 +/- 2.5 minutes; and duplex imaging, 10.4 +/- 1.1 minutes. As noted duplex imaging times as compared with radiographic modalities were significantly shorter (P < 0.05). There was a substantial cost difference between angiography ($650) and duplex scanning ($350). A 100 per cent correlation of study findings was noted between angiography and duplex scanning. No significant change in pre- versus postoperative creatinine levels was found. We conclude that duplex scanning is an effective modality and provides reliable intraoperative vascular graft assessment data in a community hospital setting. Advantages include a shorter study time, lower cost, flow dynamic data acquisition, and avoidance of mechanical graft trauma.

Molecular signals required for type III secretion and translocation of the Xanthomonas campestris AvrBs2 protein to pepper plants.

Strains of Xanthomonas campestris pv. vesicatoria (Xcv) carrying avrBs2 are specifically recognized by Bs2 pepper plants, resulting in localized cell death and plant resistance. Agrobacterium-mediated transient expression of the Xcv avrBs2 gene in plant cells results in Bs2-dependent cell death, indicating that the AvrBs2 protein alone is sufficient for the activation of disease resistance-mediated cell death in planta. We now provide evidence that AvrBs2 is secreted from Xcv and that secretion is type III (hrp) dependent. N- and C-terminal deletion analysis of AvrBs2 has identified the effector domain of AvrBs2 recognized by Bs2 pepper plants. By using a truncated Pseudomonas syringae AvrRpt2 effector reporter devoid of type III signal sequences, we have localized the minimal region of AvrBs2 required for type III secretion in Xcv. Furthermore, we have identified the region of AvrBs2 required for both type III secretion and translocation to host plants. The mapping of AvrBs2 sequences sufficient for type III delivery also revealed the presence of a potential mRNA secretion signal.

Expression of the Bs2 pepper gene confers resistance to bacterial spot disease in tomato.

The Bs2 resistance gene of pepper specifically recognizes and confers resistance to strains of Xanthomonas campestris pv. vesicatoria that contain the corresponding bacterial avirulence gene, avrBs2. The involvement of avrBs2 in pathogen fitness and its prevalence in many X. campestris pathovars suggests that the Bs2 gene may be durable in the field and provide resistance when introduced into other plant species. Employing a positional cloning strategy, the Bs2 locus was isolated and the gene was identified by coexpression with avrBs2 in an Agrobacterium-mediated transient assay. A single candidate gene, predicted to encode motifs characteristic of the nucleotide binding site-leucine-rich repeat class of resistance genes, was identified. This gene specifically controlled the hypersensitive response when transiently expressed in susceptible pepper and tomato lines and in a nonhost species, Nicotiana benthamiana, and was designated as Bs2. Functional expression of Bs2 in stable transgenic tomatoes supports its use as a source of resistance in other Solanaceous plant species.

Serum proteins facilitate neutrophil induction of endothelial leukocyte adhesion molecule 1.

BACKGROUND: Although the individual actions of neutrophils and serum proteins such as complement in acute inflammation are well characterized, less is known about their effects in combination. We investigated the combined effects of neutrophil contact and active serum proteins on the expression of endothelial leukocyte adhesion molecule 1 (ELAM-1). METHODS: Confluent monolayers of human umbilical vein endothelial cells were incubated with neutrophils in the presence and absence of fresh human serum. Flow cytometry was used to assess expression of endothelial intercellular adhesion molecule 1 (ICAM-1) and ELAM-1. In addition, neutrophils were retained in a semipermeable insert, which allowed their secretions to contact the endothelium but restricted neutrophil-endothelial contact. RESULTS: ELAM-1 expression was significantly increased on the cells coincubated with neutrophils and fresh human serum (25.8%; p < 0.001). There was no significant change in ELAM-1 expression on endothelial cells incubated with fresh human serum alone (3.9%; p > 0.01) or in those incubated with neutrophils and heat-inactivated serum (9.3%; p > 0.01). In the absence of neutrophil contact, ELAM-1 expression was increased only in the presence of fresh human serum (9.6%; p < 0.05). CONCLUSIONS: These findings suggest that serum proteins may potentiate the volume or potency of neutrophil-derived diffusable mediators of ELAM-1 expression. These effects are eliminated with the heat inactivation of serum proteins, implicating a heat sensitive mediator such as the complement cascade.

Physiologic concentrations of TNFalpha and IL-1beta released from reperfused human intestine upregulate E-selectin and ICAM-1.

Intestinal ischemia-reperfusion (I/R) causes local and distant tissue injury via neutrophil (PMN) activation and adhesion. Endothelial cell adhesion molecules (E-selectin, ICAM-1) mediate the adhesion and transmigration of PMN in the microcirculation. Expression of these receptors is influenced by cytokines. To determine the physiologic concentrations of two specific cytokines involved in I/R, tumor necrosis factor (TNF) and interleukin-1 (IL-1), human intestinal segments were exposed to 30 min of ischemia followed by reperfusion. Venous effluent samples were obtained; enzyme immunoassays measured maximum concentrations of TNF (30.5 +/ 1.0 pg/ml) and IL-1 (59.0 +/- 6.0 pg/ml). Cultured human endothelial cells were then exposed to physiologic concentrations of human recombinant TNF (10 pg/ml) and IL-1 (10 pg/ml), individually and in combination. Flow cytometric analysis of receptor expression demonstrated upregulation of E-selectin as early as 2 hr (P < 0.05) with maximum effects at 4 hr. At 4 hr, E-selectin expression (% shift from baseline) was greater with TNF and IL-1 combined (50.9 +/- 2.9, P < 0.01) than with either cytokine alone (TNF 34.6 +/- 4.0; IL-1 23.5 +/- 4.0, P < 0.01). ICAM-1 receptor expression began at 4 hr with maximum effects at 24 hr. ICAM-1 expression after TNF and IL-1 exposure (15.4 +/- 1.3, P < 0.001) was also greater than TNF (10.9 +/- 0.3, P < 0.01) or IL-1 (3.1 +/- 1.5) alone. TNF and IL-1 are present in venous effluent in concentrations capable of increasing PMN adhesion in the microcirculation. These findings support a role for these cytokines in local and distant organ injury from I/R. Since combined effects are greater than either cytokine alone, antagonism of both TNF and IL-1 may be required for a therapeutic benefit in clinical applications.

E-selectin gene induction by ionizing radiation is independent of cytokine induction.

The mechanism of the x-ray-mediated inflammatory response in normal tissues is unknown. To determine whether leukocyte infiltration into irradiated tissue is regulated by adhesion molecule expression, we quantified the synthesis of glycoproteins that participate in inflammation. We found that E-selectin is synthesized in a time-dependent manner following exposure to doses as low as 0.5 Gy. Northern blot analysis demonstrated that E-selectin mRNA expression increased at 2 h after x-irradiation and increased expression required no de novo protein synthesis. Transcription of the promoter region of E-selectin (-578 to +35) was transiently induced following x-irradiation, whereas deletion of the NFkB binding site eliminated x-ray induction. Electrophoretic mobility gel shift analysis confirmed increased binding of nuclear proteins from irradiated endothelial cells to the NFkB binding sequence from the E-selectin promoter. Nuclear protein binding to the NFkB binding sequence was altered by antibodies to the p50 and p65 components of NFkB. These data demonstrate that E-selectin expression does not require cytokine synthesis, but involves NFkB activation.

Aortofemoral bypass via transperitoneal approach.

The transperitoneal route remains the most widely used approach to the infrarenal abdominal aorta. The specific indications and techniques are reviewed in full detail. Special circumstances that require modifications of the standard operative approach are presented. Finally, current and expected results of aortofemoral bypass are discussed.

Endothelial cell response to hypoxia-reoxygenation is mediated by IL-1.

Reperfusion injury involves the adhesion and activation of neutrophils (PMN) both in affected tissues and distant organs. Cell adhesion molecules (CAM) such as endothelial-leukocyte adhesion molecule-1 (ELAM-1) and intercellular adhesion molecule-1 (ICAM-1) are known to mediate, at least in part, the adherence of activated PMN to the endothelium. To characterize the cellular mechanisms of this phenomenon, we exposed cultured human umbilical vein endothelial cells (HU-VEC) to hypoxia and reoxygenation (H/R) using an incubator chamber purged of oxygen with 100% nitrogen. Confluent monolayers of HUVEC were subjected to 60 min of hypoxia followed by variable periods of reoxygenation (120 and 240 min). Flow cytometry was utilized to assess the expression of ELAM-1 and ICAM-1, expressed as percent shift from baseline expression. To determine what role endothelium-derived cytokines such as IL-1 play in the expression of CAM after H/R, we performed additional experiments in the presence of recombinant IL-1 receptor antagonist (IL-1RA). ICAM-1 was present on unstimulated HUVEC while ELAM-1 was not constitutively expressed. Following exposure of cells to hypoxia and reoxygenation, significant increases in ELAM expression were seen (8.4 +/- 2.4% at 120 min; 19.1 +/- 7.4%, P < 0.05). While there was similar trend in ICAM expression, this did not achieve statistical significance (0.10 < P < 0.05). The addition of IL-1RA (10 ng/ml) to hypoxic HUVEC consistently attenuated ELAM-1 upregulation during reoxygenation (0.8 +/- 0.7% at 120 min and 5.9 +/- 4.1% at 240 min) such that expression was not significantly greater than baseline.(ABSTRACT TRUNCATED AT 250 WORDS)

Limiting oxygen delivery attenuates intestinal reperfusion injury.

Since free radical-mediated injury is dependent on the reintroduction of oxygen into ischemic tissues, restriction of oxygen content in the initial reperfusate has therapeutic potential. The degree to which oxygen must be restricted is crucial since hypoxic injury would continue if reperfusion O2 delivery remained below the ischemic threshold of the tissue. We examined this treatment strategy in 20 pump-perfused intestinal preparations subjected to 30 min of flow interruption. The oxygen content of the reperfusate was varied by utilizing arterial (A) or venous (V) blood; as a further modification, we also performed experiments in which hemodiluted arterial blood (HD) was the reperfusate at normal (NHD) and high (HHD) flow rates. The flow rates and O2 contents of the reperfusates were adjusted to produce either high (approximately 12 ml O2/min/100 g) or low (approximately 8 ml O2/min/100 g) levels of O2 delivery. Histologic sections, obtained after ischemia and after 1 hr of reperfusion, were blindly evaluated for mucosal injury (1 = normal to 5 = severe injury). Immediately after 30 min of ischemia, all groups had comparable histologic grades (A 2.0 +/- 0.3, V 1.8 +/- 0.3, NHD 1.6 +/- 0.3, HHD 2.3 +/- 0.3). One hour after reperfusion, intestines reperfused with blood with high O2 content and hence high O2 delivery showed significantly more damage (P < 0.001) than those with exposed to low O2 delivery during reperfusion: A 3.9 +/- 0.5 and HHD 4.4 +/- 0.4 versus V 2.7 +/- 0.5 and NHD 2.9 +/- 0.3.(ABSTRACT TRUNCATED AT 250 WORDS)

True aneurysmal disease in the hand and upper extremity.

Ten patients with true aneurysmal disease of the hand and forearm vessels were treated at our institution between 1981 and 1990. Pseudoaneurysms resulting from penetrating trauma or mycotic aneurysms were specifically excluded. Eight patients were male, two were female; mean patient age was 38 years (range 26 to 72 years). A history of repetitive occupational or recreational trauma was elicited in five patients. All patients presented with painful masses or neurologic symptoms due to nerve compression. Ischemic changes were evident in five patients due to thrombosis or distal embolization. Arteriography and transcutaneous Doppler ultrasound aided in documentation of flow characteristics and planning for operative intervention. Three patients underwent excision and ligation once collateral flow was demonstrated to be adequate and reconstruction was not felt to be feasible. Seven patients underwent resection with vein graft reconstruction. Immediate postoperative and interval patency rates were 100%. No digital amputations were required even in those patients presenting with severe distal ischemia.

Intermittent ischemia potentiates intestinal reperfusion injury.

We hypothesized that even brief periods of reperfusion interjected between ischemic episodes would increase tissue injury. Studies were performed in a rat small intestine preparation in which metabolic, hemodynamic, and histologic responses to ischemia have been well characterized. Animals were subjected to a total of 30 or 45 minutes of complete intestinal ischemia. Flow interruption was continuous (C, single episode) or intermittent (I, two or three episodes of 15-minute ischemia separated by 5 minutes of reperfusion). In some experiments 5-minute reperfusions were performed with arterial blood depleted of leukocytes (IL). This additional perturbation was included to determine the role of neutrophils that have been strongly implicated in reperfusion injury. In all three protocols histologic sections were obtained after each ischemic insult and after 1 hour of reperfusion with arterial blood. Villous histology was graded in a blinded fashion with 1 = normal and 5 = severe injury. No significant differences were found between groups in immediate postischemic histologies before reperfusion. After 1 hour of reperfusion, intermittent episodes of ischemia were associated with significantly worse histologic injury than that seen with comparable durations of continuous ischemia (30 min: I, 4.4 +/- 0.5 vs C, 2.7 +/- 0.4; 45 min: I, 4.9 +/- 0.2 vs C, 2.8 +/- 0.3). However, if 5-minute reperfusions were with leukopenic blood, this effect was markedly reduced (30 min IL, 3.4 +/- 0.3; 45 min IL, 3.6 +/- 0.2). Even short periods of reperfusion during an ischemic insult greatly increased mucosal injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Current results of elective aortic reconstruction for aneurysmal and occlusive disease.

Decisions to resect small aortic aneurysms or employ non-operative treatment for aorto-iliac occlusive disease must depend on current rather than historical surgical results. To assess current morbidity and mortality, we reviewed 200 consecutive aortic resections in two groups of patients treated from 1981 to 1989: those undergoing elective aortofemoral bypass for occlusive disease (AFB, no. 100) or resection of infrarenal abdominal aortic aneurysms (AAA, no. 100). Indications for AFB included claudication (54%), rest pain (32%), and gangrene (13%). AAA size ranged from 3 to 14 cm (mean 6.5 +/- 2.4 cm); 45% presented with abdominal or back pain. Patients undergoing AFB were younger (AFB 61.5 +/- 10 years vs AAA 68.7 +/- 8.9 years) with a higher incidence of some atherosclerotic risk factors, diabetes mellitus 30% vs 10%, tobacco use 77% vs 49%, hyperlipidemia 21% vs 7%; p less than 0.001). Coronary artery disease (CAD) was more prevalent in AAA patients (49% vs 34%; p less than 0.001). Postoperative mortality was not different in occlusive or aneurysmal disease (3% AFB vs 2% AAA), nor was the occurrence of serious complications such as myocardial infarction (2% vs 1%) or pulmonary embolism (2% vs 3%). Improvements in patient selection, perioperative care and surgical technique have lowered the mortality of elective aortic surgery. Given the current standard of care, an aggressive approach to AAA even in high risk patients is appropriate. The low morbidity of AFB for occlusive disease mandates a critical appraisal of less effective nonoperative therapies.

Glucagon potentiates intestinal reperfusion injury.

Vasoactive agents, including glucagon, have been used in treatment of mesenteric ischemia. Such drugs change both intestinal blood flow and metabolism. Since reperfusion injury reflects the metabolic state of an organ as well as the duration and severity of ischemia, we investigated the effect of glucagon in a standard model of intestinal ischemia. Data were generated from denervated isoperfused rat small intestinal preparations (n = 39). Arterial and venous pressures, intestinal blood flow, and oxygen consumption were monitored. Animals were subjected to 15, 30, or 45 minutes of ischemia followed by 1 hour reperfusion. Experiments were performed without drug infusion or during intravenous glucagon administration (0.1, 0.2, or 0.4 micrograms/kg/min). After the rats were killed, histologic sections of intestine were graded 1 through 5 in a blinded fashion with 1 = normal villi and 5 = severe injury. Results (mean +/- SD) were analyzed by analysis of variance (*p less than 0.05). Glucagon at all concentrations increased intestinal blood flow and oxygen consumption before ischemia. For example, with 0.2 micrograms/kg/min glucagon, intestinal blood flow increased from 80.78 +/- 13.5 to 114.79 +/- 21.02 ml/min.100 gm* and oxygen consumption increased from 3.65 +/- 0.73 to 5.73 +/- 1.37 ml/min.100 gm.* Mucosal injury after ischemia reflected duration of ischemia and glucagon infusion rate. At all ischemic intervals, increased glucagon concentrations were associated with greater mucosal injury. In fact the histologic injury with 15 minutes of ischemia + 0.2 microgram/kg/min glucagon (3.04 +/- 0.49) exceeded that of 30 minutes of ischemia (2.87 +/- 0.06).(ABSTRACT TRUNCATED AT 250 WORDS)