Adenoviral-mediated gene transfer in human and animal vein grafts using clinically relevant exposure times, pressures, and viral concentrations.

This study examined the efficiency of adenoviral-mediated gene transfer in experimental vein grafts and cultured human saphenous vein under physiologic conditions using clinically relevant exposure times, pressures, and viral concentrations. The external jugular veins of 25 male New Zealand White rabbits were exposed to 0.5 mL of replication-deficient adenovirus vectors encoding beta-galactosidase (AdlacZ), control adenovirus (AdBg/II), or vehicle at pressures ranging from 0 to 120 mmHg for 10 min. Veins were excised and grafted into the carotid circulation. After 5 days, the vessels were reexposed, excised, and stained with X-gal chromagen for beta-galactosidase (beta-gal) activity. Gene transfer was also performed in 13 segments of human saphenous vein discarded at the time of bypass grafting. The veins were cultured for 0-21 days and assayed for beta-gal activity as above. Rabbit vein grafts exposed to high-pressure AdlacZ transfection showed significant transgene expression in 100% of grafts (39 +/- 2% positive cells/hpf) while only 60% of those transfected at low pressure expressed beta-gal (9 +/- 3% positive cells/hpf). All human veins exposed to AdlacZ expressed beta-gal to a variable degree (range 10-50% positive cells/hpf). No control grafts or veins expressed the transgene. Efficient adenoviral-mediated gene transfer in experimental vein grafts and human saphenous vein segments can be achieved using clinically feasible parameters of exposure time, pressure, and viral concentration.

Effective hemodynamic diameter: an intrinsic property of vein grafts with predictive value for patency.

INTRODUCTION: Conduit size and quality are major determinants of the long-term success of infrainguinal autologous vein grafting. However, accurate measurement of the internal diameter of vein grafts is difficult given their variable wall thickness and taper. The purpose of this study was to define the "effective" internal diameter of a vein graft according to its hemodynamic properties and to determine its significance for graft patency. METHODS: Sixty infrainguinal bypass grafts performed on 57 patients were evaluated intraoperatively. Proximal and distal graft pressure and blood flow (Q(meas)) were measured with fluid-filled catheter transduction and ultrasonic transit-time flowimetry, respectively, after unclamping. Waveforms were recorded digitally at 200 Hz under baseline conditions and after stimulation with 60 mg of papaverine. According to Fourier transformation of the measured pressure gradient (DeltaP), the Womersley solution for fluid flow in a straight rigid tube was used to calculate theoretical flow waveforms (Q(calc)) for a range of graft diameters. The theoretical waveforms were then compared with the measured flow waveforms and the best-fit diameter chosen as the "effective hemodynamic diameter" (EHD). Only grafts in which the correlation coefficient of Q(calc) versus Q(meas) was more than 0.90 were accepted (n = 47) to assure validity of the hemodynamic model. After a mean follow-up of 12.5 months (range, 0.1-43.9 months), patency was determined by the life table method. Hemodynamic and clinical variables were tabulated, and their effect on patency determined the use of univariate and multivariate Cox regression. RESULTS: Mean EHD was 4.1 +/- 0.1 mm with a range of 2.5 to 5.7 mm. Administration of papaverine caused profound changes in DeltaP (+78% +/- 17%) and Q(meas) (+71% +/- 12%) as expected, but had no effect on EHD (+0.05% +/- 0.1%). Univariate regression identified five variables associated with decreased secondary patency (P <.10): low EHD, conduit source other than the greater saphenous vein, high baseline DeltaP(mean), female sex, and redo operation. Of these, only low EHD was significant after multivariate analysis (P =.03). Patency of small diameter grafts (EHD < 3.6 mm; n = 11) was compared with patency of larger grafts (EHD > 3.6 mm; n = 36) to test a frequently espoused clinical guideline. Grafts with an EHD less than 3.6 mm exhibited significantly lower secondary patency compared with larger grafts (P =.0001). The positive and negative predictive values for an EHD less than 3.6 mm for secondary graft failure for grafts with at least 1 year follow-up were 86% and 88%, respectively. CONCLUSION: An EHD is a unique parameter that quantifies conduit size and has a significant impact on vein graft patency. An EHD less than 3.6 mm portends graft failure.

The effect of 'non-critical' (<50%) stenosis on vein graft longitudinal resistance and impedance.

OBJECTIVE: vein graft stenoses <50% cause minimal flow impairment, velocity elevation, or symptomatology and are therefore usually assumed to be "non-critical". The purpose of this study was to assess the effect of <50% vein graft stenosis on vein graft longitudinal impedance, as elevated impedance has been found to correlate with clinical graft failure. METHODS: eight segments of non-reversed cryopreserved vein (mean length 23+/-1 cm; mean outer diameter 4.7+/-0.2 mm) were saline-perfused in vitro utilising a variable pulsatile perfusion pump, Windkessel, and clamp resistor simulating the haemodynamic conditions of arterial bypass. Proximal (Pprox) and distal (Pdist) pressure were continuously measured by fluid-filled catheter transduction, and flow (Q) by ultrasonic transit-time flowmetry. Waveforms were digitally recorded at 200 Hz at pulse rates ranging from 60-180 b.p.m. with mean flow (Q) of 154 ml/min and mean proximal pressure (Pprox) of 100 mmHg (max/min 120/90). Graded mid-graft stenoses of <50% were created using an inflatable vascular occluder and measured by the corresponding changes in mean pressure gradient (DeltaP=Pprox-Pdist) and Q (%stenosis=1-{DeltaPbaselineQstenosis/Delta PstenosisQbaseline}1/4). Vein graft longitudinal resistance (RL) was calculated as DeltaP/Q. After Fourier transformation, vein graft longitudinal impedance (ZL) was calculated as DeltaP/Q at each harmonic, with ZL determined by integration over 0-4 Hz. Results are reported as mean+/-S.E.M. RESULTS: the desired levels of pressure and flow were established in all vein segments. Graded inflation of the occluder resulted in vein graft stenosis of 23+/-3% and 39+/-3%. This was accompanied by a mild reduction in Q (12% and 30%) and considerable increases in both RL (180% and 710%) and ZL (140% and 430%). CONCLUSIONS: "non-critical" vein graft stenosis (<50%) causes minimal change in mean flow, but substantial elevations in longitudinal resistance and impedance. The contribution of "non-critical" stenosis to vein graft failure may be under-appreciated.

Adenoviral-mediated gene transfer of a constitutively active form of the retinoblastoma gene product attenuates neointimal thickening in experimental vein grafts.

PURPOSE: Inappropriate or excessive vascular smooth muscle cell proliferation leads to the development of occlusive lesions in up to 50% of vein grafts. The purpose of this study was to test the hypothesis that induced overexpression of a cytostatic nonphosphorylatable form of the retinoblastoma protein (DeltaRb) would attenuate neointimal thickening in experimental vein grafts. METHODS: A replication-deficient adenovirus vector that encoded a nonphosphorylatable, constitutively active form of DeltaRb was constructed (AdDeltaRb) and contained an NH2-terminal epitope tag from the influenza hemagglutinin molecule (HA). Forty-eight male New Zealand white rabbits underwent surgical exposure of the external jugular vein for transfection with either 3 x 10(10) plaque-forming units/mL AdDeltaRb (n = 16), 3 x 10(10) plaque-forming units/mL control adenovirus (AdBglII, n = 15), or vehicle (n = 17) for 10 minutes at 120 mm Hg. After vector exposure, the vein was excised and interposed end-to-end into the carotid circulation. After 5 days, 12 grafts (four from each group) were excised and assayed for genomic DeltaRb DNA with the polymerase chain reaction or for hemagglutinin molecule expression and localization with immunohistochemistry. The remainder of the grafts (n = 36) were perfusion-fixed after 4 weeks, and 5 microm sections prepared for digital planimetric analysis. RESULTS: Polymerase chain reaction results identified the DeltaRb gene only in the grafts that were transfected with AdDeltaRb. Immunohistochemical analysis results revealed transgene expression in most of the endothelial cells and in many of the smooth muscle cells. After 4 weeks, the grafts that were exposed to AdDeltaRb exhibited a 22% reduction in neointimal thickness (vehicle, 77 +/- 7 microm; AdBglII, 75 +/- 5 microm; AdDeltaRb, 60 +/- 5 microm; P =.05), and medial thickness, luminal diameter, and other parameters were unchanged (medial thickness: vehicle, 72 +/- 10 microm; AdBglII, 85 +/- 7 microm; AdDeltaRb, 69 +/- 9 microm; P = NS; luminal diameter: vehicle, 4.5 +/- 0.2 mm; AdBglII, 4.4 +/- 0.2 mm; AdDeltaRb, 4.7 +/- 0.1 mm; P = NS). CONCLUSION: With this delivery system, adenoviral-mediated gene transfer is highly efficient and induced overexpression of DeltaRb leads to a reduction in vein graft neointimal thickening.

Current results of surgical therapy for chronic mesenteric ischemia.

BACKGROUND: Although recognition of chronic mesenteric ischemia has increased in recent years, this disorder has continued to present diagnostic and therapeutic challenges. OBJECTIVE: To examine the modern results of surgical revascularization for chronic mesenteric ischemia. DESIGN: Retrospective review. SETTING: University medical center. PATIENTS: The management of 24 consecutive patients (mean +/- SEM age, 58 +/- 3 years; 5 men, 19 women) who were undergoing surgical treatment of chronic mesenteric ischemia between 1986 and 1996 was reviewed. INTERVENTION: Surgical mesenteric revascularization. MAIN OUTCOME MEASURES: Postoperative course, long-term graft patency rate, and long-term symptom-free survival rate. RESULTS: The most frequent presenting symptoms were postprandial abdominal pain (18 patients [75%]) and weight loss (14 patients [58%]). Less specific complaints included nausea and vomiting (8 patients [33%]), diarrhea (7 patients [29%]), and constipation (4 patients [17%]). Atherosclerotic risk factors were common, including tobacco use (20 patients [83%]), coronary artery disease (10 patients [42%]), and hypertension (10 patients [42%]). The cause was identified as atherosclerosis in 21 patients, median arcuate ligament compression in 2 patients who were monozygotic twins, and Takayasu arteritis in 1 patient. Lesions were localized to all 3 major visceral vessels (celiac artery, superior mesenteric artery [SMA], and inferior mesenteric artery) in 8 patients, celiac artery and SMA in 13, SMA alone in 2, and SMA and inferior mesenteric artery in 1. Seventeen patients underwent antegrade reconstructions from the supraceliac aorta to the SMA and/or celiac artery; 7 patients underwent revascularization by use of a retrograde bypass that originated from the infrarenal aorta or a prosthetic graft. There were no perioperative deaths although 1 patient died in the hospital 6 weeks after early graft failure and sepsis (overall in-hospital mortality, 4%). Follow-up ranged from 3 months to 10 years (median, 2.4 years). The mean +/- SEM 5-year primary graft patency rate, as objectively documented by use of contrast angiography or duplex scanning in 19 of 24 patients, was 78% +/- 11%. Primary failure was documented in 3 patients (at 3 weeks, 5 months, and 7 months). Two patients required a thrombectomy; 1 of these patients subsequently died of an intestinal infarction. The mean +/- SEM 5-year survival rate by use of life-table analysis was 71% +/- 11%. No patient with a patent graft experienced a symptomatic recurrence. CONCLUSIONS: Chronic mesenteric ischemia is usually a manifestation of advanced systemic atherosclerosis. Symptoms almost always reflect midgut ischemia in the distribution of the SMA. An antegrade bypass from the supraceliac aorta can be performed with acceptable operative morbidity and is currently the preferred reconstructive technique. Surgical revascularization affords long-term symptom-free survival in a majority of patients with chronic mesenteric ischemia.

Validation of a new and specific intraoperative measurement of vein graft resistance.

PURPOSE: Clinical studies have revealed that the most important predictor of successful bypass grafting is the origin and quality of the bypass conduit. Attempts at intraoperative evaluation of the hemodynamic properties of the conduit, including assessment of blood flow (Q), pressure gradients (delta P), and resistance (R), have not been useful. This is because each of these parameters measures the characteristics of the graft plus the outflow bed. To date, no specific measurement of the resistive properties of the conduit only is available. The purpose of this investigation was to evaluate longitudinal impedance (ZL) as a measure of conduit-specific resistance and to evaluate its potential in predicting the outcome of infrainguinal vascular reconstructions. METHODS: ZL was measured during surgery in 73 infrainguinal autologous vein reconstructions performed in 68 patients in two separate institutions over a 21-month period. Vein graft ultrasonic transit time Q and delta P (from proximal to distal anastomosis) were measured at baseline and after maximal peripheral vasodilatation with an intraarterial injection of papaverine 30 mg. Waveforms were recorded for 10 seconds at 200 Hz using a digital acquisition system. R was calculated as proximal mean pressure divided by mean blood flow (Q). After Fourier transformation, ZL was calculated as delta P/Q at each harmonic and total ZL (integral of ZL) was defined as the integral of moduli from 0 to 4 Hz. RESULTS: All hemodynamic variables were significantly affected by papaverine vasodilatation (delta P, 3.9 +/- 0.5 vs 6.3 +/- 0.8 mm Hg; Q, 78.2 +/- 7.0 vs 126 +/- 11 ml/min; R, 134 +/- 17 vs 72.7 +/- 6.2 x 10(3) dyne.sec.cm-5; p < 0.0001), except integral of ZL, which remained constant (31.1 +/- 2.8 vs 30.8 +/- 2.8 x 10(3) dyne.cm-5; p = NS). After follow-up of 1 week to 17 months (median, 5 months), the 1-year primary, primary-assisted, and secondary patency rates were 72% +/- 7%, 77% +/- 6%, and 81% +/- 6%, respectively. Using Cox analysis, primary patency was significantly associated with decreased integral of ZL (p = 0.0001), but not with baseline or papaverine-stimulated delta P, Q, delta P/Q, or R integral of ZL > 47 x 10(3) dyne.cm-5 predicted primary failure with 90% positive and negative predictive value. CONCLUSIONS: Intraoperative measurement of integral of ZL in infrainguinal vein grafts is independent of outflow conditions (that is, does not change with papaverine), and hence describes the resistive properties of the conduit only. In addition, these preliminary data suggest that integral of ZL is predictive of short-term primary patency. integral of ZL is the first available hemodynamic measurement that is conduit-specific and may therefore be a better predictor of graft patency than currently available methods.

Chronic mesenteric ischemia. Clinical presentation and diagnosis.

Owing to a heightened awareness of the disease as well as improved diagnostic tests, chronic mesenteric ischemia is now recognized as a more common cause of abdominal pain. The classic symptoms of postprandial abdominal pain with weight loss are evident in the majority of proven cases; most patients also have other evidence of advanced atherosclerotic vascular disease. Several new diagnostic techniques are being developed and tested, most notably color duplex imaging, although angiography still remains the diagnostic gold standard. It is hoped that better noninvasive testing may eventually eliminate the need for angiography, as well as lead to a more expedient diagnosis of chronic mesenteric ischemia.

Gene therapy for vascular disease.

Atherosclerosis is a degenerative process characterized by endothelial cell dysfunction, inflammatory cell adhesion and infiltration, and the accumulation of cellular and matrix elements leading to the formation of fibrocellular plaques. In the end stages, advanced occlusive plaques limit blood flow and oxygen delivery resulting in the well-known ischemic syndromes of the coronary, skeletal muscle, mesenteric, and cerebrovascular circulation. Moreover, sudden critical ischemic events may be precipitated by plaque disturbance, rupture, hemorrhage, and/or thrombosis. Traditional pharmacologic approaches have been effective in reducing serum cholesterol and controlling thrombosis but, in the main, have had little impact on the treatment of advanced lesions. The purpose of this review is to examine the current status of gene therapy for vascular proliferation, aberrant endothelial function, thrombosis, peripheral ischemia, and modification of the blood/biomaterial interface.

Measurement of vascular input impedance in infrainguinal vein grafts.

The purpose of this investigation was to measure vascular input impedance in infrainguinal vein grafts and assess the importance of clinical and hemodynamic parameters in predicting graft patency. Fifty-seven patients undergoing infrainguinal vein bypass grafting for limb salvage (n = 40) or claudication (n = 17) were prospectively studied. At the time of revascularization, simultaneously acquired intraluminal pressure and blood flow waveforms were digitized at 200 Hz and subjected to Fourier transformation in near real-time. Input impedance was calculated at baseline (immediately after unclamping) and after stimulation with either papaverine or completion arteriography. Resistance (Rin) was calculated as mean pressure divided by mean blood flow (Q). Characteristic impedance (Z0) was calculated as the mean of harmonics 3-10. Intraoperative data acquisition required approximately 5 min, utilized the completion angiography cannula already in place, and was uncomplicated in all patients. Stimulation with either papaverine or arteriography resulted in increased Q (72 +/- 7 to 146 +/- 11 ml/min, p < 0.001), decreased Rin (126 +/- 13 to 52 +/- 4 x 10(3) dyne.s.cm-5, p < 0.001), and slightly decreased Z0 (18 +/- 2 to 15 +/- 1, p = 0.002). After a mean follow-up of 20 months, the 2-year primary patency, secondary patency, limb salvage, and survival rates were 61 +/- 8%, 74 +/- 7%, 76 +/- 6%, and 86 +/- 6%, respectively. Primary patency was not associated with any of the clinical variables studied including age, sex, smoking history, history of previous vascular surgery, hypertension, coronary artery disease, diabetes mellitus, creatinine, indication for revascularization (claudication versus limb salvage), anesthesia (general versus regional), or level of distal anastomosis (popliteal versus infrapopliteal). Furthermore, there was no association between primary patency and baseline Q, baseline Rin, or stimulated Z0. However, using univariate analysis, patency was positively associated with decreased stimulated Rin (p = 0.002), elevated stimulated Q (p = 0.006), and decreased baseline Z0 (p = 0.02). Multiple regression analysis identified stimulated Rin as the only independent predictor of primary patency (p = 0.002). Stimulated Rin > or = 50 x 10(3) dyne.s.cm-5 was 71% sensitive and 65% specific for graft failure. It is concluded that 1) vascular input impedance can be simply and reliably measured in the operating room, and 2) elevated stimulated Rin is an independent predictor of primary patency.