Margaret D. Allen

Vascular cell adhesion molecule-1 is expressed in human coronary atherosclerotic plaques. Implications for the mode of progression of advanced coronary atherosclerosis.

Endothelial attachment is the initial step in leukocyte recruitment into developing atherosclerotic lesions. To determine whether vascular cell adhesion molecule-1 (VCAM-1) expression may play a role in inflammatory cell recruitment into human atherosclerotic lesions, immunohistochemistry was performed with a polyclonal rabbit antisera, raised against recombinant human VCAM-1, on 24 atherosclerotic coronary plaques and 11 control coronary segments with nonatherosclerotic diffuse intimal thickening from 10 patients. Immunophenotyping was performed on adjacent sections to identify smooth muscle cells, macrophages, and endothelial cells. To confirm VCAM-1-expressing cell types, double immunostaining with VCAM-1 antisera and each of the cell-specific markers and in situ hybridization were performed. All atherosclerotic plaques contained some VCAM-1, compared to 45% of control segments. VCAM-1 was found infrequently on endothelial cells at the arterial lumen din both plaques (21%) and in control segments (27%), but was prevalent in areas of neovascularization and inflammatory infiltrate in the base of plaques. Double immunostaining and in situ hybridization confirmed that most VCAM-1 was expressed by subsets of plaque smooth muscle cells and macrophages. The results document the presence of VCAM-1 in human atherosclerosis, demonstrate VCAM-1 expression by human smooth muscle cells in vivo, and suggest that intimal neovasculature may be an important site of inflammatory cell recruitment into advanced coronary lesions.

Neovascular Expression of E-Selectin, Intercellular Adhesion Molecule-1, and Vascular Cell Adhesion Molecule-1 in Human Atherosclerosis and Their Relation to Intimal Leukocyte Content

BACKGROUND Leukocyte recruitment is an early event in atherogenesis, and the leukocyte adhesion molecules E-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) recently have been detected in human atherosclerosis. However, no previous study has evaluated either the distribution of these three molecules at different sites within the arterial intima or their relation to plaque leukocyte content. METHODS AND RESULTS Immunohistochemistry was performed on 99 coronary artery segments (34 controls and 65 with atherosclerotic plaque) to identify E-selectin, ICAM-1, VCAM-1, macrophages, smooth muscle cells, and T lymphocytes. For each segment, the presence or absence of adhesion molecule was determined at the arterial lumen, on intimal neovasculature, and on intimal nonendothelial cells. Each segment was scored for intimal macrophage and T-lymphocyte densities on a semiquantitative scale of 0 to 3. In atherosclerotic plaques, the prevalences of E-selectin, ICAM-1, and VCAM-1 on plaque neovasculature were twofold higher than their prevalences on arterial luminal endothelium. E-selectin was the only adhesion molecule for which expression on arterial luminal endothelial cells was more prevalent in plaques than in control segments. Increased plaque intimal macrophage density was associated with expression of VCAM-1 on neovasculature (P < .01) and on nonendothelial cells (P < .01). Increased plaque intimal T-lymphocyte density was associated with the presence of both ICAM-1 and VCAM-1 on neovasculature (both P < .01) and on nonendothelial cells (both P < .01). CONCLUSIONS In atherosclerotic plaques, the expression of all three leukocyte adhesion molecules was more prevalent on intimal neovasculature than on arterial luminal endothelium. Further, the presence on neovasculature and nonendothelial cells of VCAM-1 and ICAM-1 was strongly associated with increased intimal leukocyte accumulation. These findings suggest that leukocyte recruitment through and/or activation of intimal neovasculature may play important roles in the pathogenesis of human atherosclerosis.

A prospective randomized evaluation of biphasic versus monophasic waveform pulses on defibrillation efficacy in humans

Biphasic waveforms have been suggested as a superior waveform for ventricular defibrillation. To test this premise, a prospective randomized intraoperative evaluation of defibrillation efficacy of monophasic and biphasic waveform pulses was performed in 22 survivors of out of hospital ventricular fibrillation who were undergoing cardiac surgery for implantation of an automatic defibrillator. The initial waveform used in a patient for defibrillation testing, either monophasic or biphasic, was randomly selected. Subsequently, each patient served as his or her own control for defibrillation testing of the other waveform. The defibrillation threshold was defined as the lowest pulse amplitude that would effectively terminate ventricular fibrillation with a single discharge delivered 10 s after initiation of an episode of ventricular fibrillation induced with alternating current. Each defibrillation pulse was recorded oscilloscopically, and defibrillation pulse voltage, current, resistance and stored energy were measured. Fifteen (68%) of the 22 patients had a lower defibrillation threshold with the biphasic pulse, 3 (14%) had a lower threshold with the monophasic pulse and 4 (18%) had equal defibrillation thresholds (within 1.0 J) regardless of waveform. The mean leading edge defibrillation threshold voltage was 317 +/- 105 V when the monophasic pulse was used and 267 +/- 102 V (16% less) when the biphasic pulse was used (p = 0.008). Mean leading edge defibrillation threshold current was 7.9 +/- 3.7 A when the monophasic pulse was used and 6.8 +/- 3.8 A (14% less) when the biphasic pulse was used (p = 0.051).(ABSTRACT TRUNCATED AT 250 WORDS)

Prospective Trial of Catheter Irrigation and Muscle Flaps for Sternal Wound Infection

BACKGROUND Sternal wound infection is a relatively rare but potentially devastating complication of open heart operations. The most common treatments after debridement are rewiring with antibiotic irrigation and muscle flaps. Here we present the results of a prospective trial to determine the appropriate roles of closed-chest catheter irrigation and muscle flap closure for sternotomy infection and to assess the effect of internal mammary artery bypass grafting on the outcome of each treatment modality. METHODS Between 1990 and 1994, 5,658 sternotomies were performed at the University of Washington Medical Center. Sternal dehiscence occurred in 43 patients, 25 of whom had infection (overall incidence, 0.44%). Because of the infrequency of this complication, a prospective, randomized trial was developed in which the initial approach to sternal dehiscence was rewiring and catheter irrigation. Muscle flaps were used as the primary treatment if the sternum could not be restabilized or as secondary treatment if catheter irrigation failed. Wound resolution, length of hospital stay, and complications were evaluated. RESULTS Sterile dehiscences were successfully closed with irrigation in 17 of 18 patients; the other patient required flap closure. Of the 25 patients with infection, 19 had irrigation and 6, closure with flaps primarily. In the group of infected patients, 17 of the 19 who received irrigation also had internal mammary artery bypass grafting. Irrigation failed in 15 (88.2%) of these 17 patients, and salvage was accomplished with muscle flap closure. All 6 patients with infection who were closed primarily with muscle flaps had a successful outcome. Hospitalization averaged 10.2 days when muscle flaps were used primarily and 14.3 additional days for unsuccessful irrigation. When irrigation was successful, the hospital stay averaged 11.2 days. CONCLUSIONS Catheter irrigation should be reserved for patients without infection or patients with infection but without internal mammary artery bypass grafts in whom dehiscence occurs less than 1 month after sternotomy. All others should have closure with muscle flaps.

Evaluation of electrode polarity on defibrillation efficacy.

The effect of electrode polarity on defibrillation thresholds in humans is unknown. This prospective, randomized evaluation of electrode polarity on defibrillation thresholds was performed in 21 survivors of ventricular fibrillation (VF) undergoing cardiac surgery. Defibrillation was always performed with 2 identical large rectangular, wire mesh electrodes positioned over the anterior wall of the right ventricle and the posterolateral wall of the left ventricle. The initial electrode polarity for the left ventricular (LV) electrode was chosen randomly for determination of the defibrillation threshold. Subsequently, electrode polarity was reversed. The defibrillation threshold was defined as the lowest pulse amplitude that would effectively terminate VF with a single discharge delivered 10 seconds after initiation of an episode of VF with alternating current. For each defibrillation pulse, voltage, current, resistance and delivered energy were recorded. Of the 21 patients, 15 (71%) had a lower defibrillation threshold when the LV electrode was positive, 2 patients (10%) had a lower defibrillation threshold when the LV electrode was negative and 4 patients (19%) had equal defibrillation thresholds (within 0.5 J) regardless of polarity. The mean leading edge defibrillation threshold voltage was 370 +/- 88 volts when the LV electrode was negative and 320 +/- 109 volts (14% less) when the LV electrode was positive (p = 0.014). Mean leading edge defibrillation threshold current was 9.3 +/- 3.1 amps when the LV electrode was negative compared to 7.7 +/- 3.1 amps (17% less) when the LV electrode was positive (p = 0.0033). There were no differences in resistance with the 2 configurations.(ABSTRACT TRUNCATED AT 250 WORDS)

Transvenous defibrillation in humans via the coronary sinus.

A consistently effective transvenous defibrillation system for use in automatic defibrillators could significantly alter the approach to patients at risk of sudden death. Transvenous defibrillation systems that use a right ventricular (RV) electrode only or an RV electrode in combination with a chest patch are relatively inefficient at applying current to the posterolateral left ventricle. An RV electrode combined with a coronary sinus (CS) electrode, however, may improve current distribution to the posterolateral left ventricle. The purpose of this investigation, therefore, was to evaluate the effectiveness and safety of a specially designed transvenous lead system with a CS electrode capable of current delivery to this relatively inaccessible region of the heart. In 20 survivors of cardiac arrest, we determined defibrillation efficacy immediately before defibrillator surgery for monophasic pulses delivered between an RV catheter electrode and a CS catheter electrode system and compared these findings with an RV catheter electrode-thoracic patch defibrillation system. Subsequently, we referenced the efficacy of both transvenous systems to an epicardial patch electrode system at the time of defibrillator implantation. The mean delivered-energy defibrillation threshold for the CS-RV electrode system was 17.5 +/- 7.9 J, which was substantially lower than the RV electrode-thoracic patch system (25.6 +/- 11.4 J, p = 0.0016 [46% more]). Defibrillation threshold voltage was 529 +/- 123 V for the CS-RV electrode system and 647 +/- 164 V (22% more) for the RV electrode-thoracic patch system (p = 0.0013).(ABSTRACT TRUNCATED AT 250 WORDS)

Liposome-Mediated Gene Transfection of Endothelial Nitric Oxide Synthase Reduces Endothelial Activation and Leukocyte Infiltration in Transplanted Hearts

Background—During cardiac ischemia-reperfusion injury, neutrophilic infiltration of the myocardium is mediated by adhesion molecule expression on activated coronary endothelium. Nitric oxide inhibits neutrophil adhesion to endothelium in vitro by blocking the nuclear factor (NF)-&kgr;B-dependent transcription of adhesion molecules. We investigated whether intraoperative gene delivery of endothelial nitric oxide synthase (eNOS) into donor hearts before transplantation would have a similar effect on an entire organ. Methods and Results—In an allogeneic rabbit heart transplant model, liposomes complexed to the gene encoding eNOS were infused into the donor coronary circulation before transplantation. By 24 hours after transplantation, calcium-dependent nitrite production was significantly higher in eNOS-transfected donor hearts than in the 3 control groups: donor hearts transfected with empty plasmids alone, donor hearts treated with diluent only, and untransplanted native hearts. Intramyocardial neutrophil and T-lymphocyte populations were halved in eNOS-transfected hearts compared with control donor hearts (P <0.05). Moreover, the prevalence of NF-&kgr;B activation in microvascular endothelial cells and surrounding cardiac myocytes as well as endothelial vascular cell adhesion molecule-1 and intracellular adhesion molecule-1 expression were all significantly reduced in eNOS-transfected hearts compared with control donor hearts (P <0.01). Without immunosuppression, eNOS-transfected hearts survived longer than controls. Conclusions—Intraoperative liposome-mediated gene delivery of eNOS to donor hearts can result in early gene expression sufficient to reduce ischemia-reperfusion injury by inhibiting NF-&kgr;B activation, adhesion molecule expression, and the early infiltration of leukocytes, all of which may improve graft survival.

Gene therapy for donor hearts: ex vivo liposome-mediated transfection.

OBJECTIVE Liposomes may be an appropriate transfection vehicle for transplanted hearts, avoiding the use of viruses in immunosuppressed hosts and allowing transfection of nondividing cells. To study whether liposome-mediated transfection could be accomplished during transplantation, we used a liposome-reporter gene system in a rabbit model of allograft cardiac transplantation. METHODS After aortic crossclamping, Stauffland donor hearts were injected with 10 ml Stanford cardioplegic solution; then a 1.3 to 2.0 mg/kg dose of chloramphenicol acetyl transferase in 1:1 deoxyribonucleic acid-liposome complexes was injected proximal to the aortic crossclamp for coronary artery perfusion. The hearts were transplanted into New Zealand White rabbit recipients in the heterotopic cervical position (n = 11 transplants). Recipients were sacrificed at 24 hours. Myocardial specimens (right and left ventricles) and vascular specimens (epicardial coronary artery, aortic root, and coronary sinus) from both the transplanted and native hearts were analyzed for chloramphenicol acetyl transferase protein by means of the enzymatic liquid scintillation assay (counts per minute per milligram of total protein). RESULTS In the recipient, myocardial chloramphenicol acetyl transferase activity was significantly greater in treated donor hearts (mean 4.6 x 10(5) cpm/mg +/- 1.1 x 10(5) [standard error]) than in native hearts (mean 4.1 x 10(2) cpm/mg +/- 72 [standard error], p < 0.01, Mann-Whitney U test). In treated donor hearts, right and left ventricular specimens, as well as apical and basal myocardial specimens, were transfected equally. Chloramphenicol acetyl transferase activity in vascular specimens also indicated transfection (mean 5.4 x 10(5) cpm/mg +/- 2.5 x 10(5) [standard error]). Chloramphenicol acetyl transferase activity in the coronary sinus was comparable with that in the coronary arteries, which suggests that liposomes can transverse the coronary capillary beds. CONCLUSIONS These findings demonstrate that ex vivo transfection of donor hearts with a liposome-reporter gene system results in significant in vivo expression of the transfected gene product after cardiac transplantation. Genetic alteration of myocardium and cardiac vasculature has potential clinical applications in the prevention of posttransplantation rejection, ischemia-reperfusion injury, and both transplant and nontransplant coronary artery disease.

Knowledge and opinions about organ donation among urban high school students: pilot test of a health education program

Background: Increasing the diversity of the organ donor pool might improve the opportunities for people of color on organ transplant waiting lists to receive donated organs. We report on the results of a pilot classroom health education program to improve knowledge about organ donation and transplantation among a diverse student body at an urban high school. 
Methods: The effectiveness of the educational program was evaluated with baseline and follow‐up questionnaires which examined: 1) whether the program increased knowledge about organ donation; 2) whether the students’ opinions about organ donation changed; and 3) whether the program was related to any changes in opinion. 
Results: On the follow‐up questionnaire, correct answers on 15 factual questions increased by 18% for the treatment group, compared to 5% for the control group (p=0.00). Regarding opinions, at baseline 92% of white students had positive opinions about donation, compared to 48% of the students of color (p=0.00). In the follow‐up survey, the increase in positive opinions among the students of color was significantly greater than among white students (p=0.04). In this pilot study, however, changes in opinions occurred with equal frequency among students in the treatment and control groups. 
In regression analysis, both knowledge of the subject and discussing donation with ones family were significantly associated with positive opinions about donation. 
Conclusions: Overall, this pilot study provided encouraging evidence that the classroom health education program affected knowledge about organ donation, and that opinions about organ donation are responsive to increases in knowledge.

Prospective comparison of sequential pulse and single pulse defibrillation with use of two different clinically available systems

Sixteen out-of-hospital survivors of ventricular fibrillation underwent a prospective, randomized, intraoperative comparison of sequential pulse and single pulse defibrillation with use of two distinct electrode systems and waveform shapes currently available for clinical use. Defibrillation was tested alternately with either the single pulse or the sequential pulse system 10 s into an episode of ventricular fibrillation. Sequential pulse defibrillation was performed with two 4 ms truncated exponential pulses of constant duration delivered to three equally spaced oval epicardial patch electrodes composed of concentric coils. The posterior left ventricular electrode served as the common cathode. The first anode was over the anterior right ventricle and the second anode was over the anterior left ventricle. Single pulse defibrillation was performed with the standard intracardiac defibrillation system with use of a single truncated exponential pulse with a fixed 65% tilt delivered across two rectangular, wire mesh epicardial patch electrodes positioned over the anterior right ventricle and posterolateral left ventricle. During defibrillation threshold determination, voltage and current waveforms were recorded and used to determine pulsing resistance and delivered and stored energy. Average defibrillation threshold leading edge voltage for the single pulse technique was 273 +/- 101 V compared with 246 +/- 67 V (11% less) for the sequential pulse technique (p = 0.136). Defibrillation threshold leading edge current for the single pulse technique was 6.7 +/- 2.5 A compared with 5.2 +/- 1.7 A (29% less) for the sequential pulse method (p = 0.005). The defibrillation threshold delivered energy was 5.6 +/- 4.0 J for the single pulse technique and 3.5 +/- 1.8 J (38% less) for the sequential pulse technique (p = 0.021).(ABSTRACT TRUNCATED AT 250 WORDS)