Three-dimensional computed tomographic reconstruction of the carotid artery: identifying high bifurcation.

OBJECTIVE: To investigate variability in the level of bifurcation relative to other anatomical landmarks on computed tomography (CT) and to develop an objective and reproducible technique for identifying patients with a high carotid bifurcation who might therefore be at greater risk of operative complications. METHODS: This was a retrospective cross-sectional, imaging study. A series of 86 nonselected consecutive CT carotid angiograms (172 arteries) were analysed. Using three-dimensional reconstructive software, the curved length (CL) of the internal carotid artery (ICA) and the straight-line distance (SLD) from the bifurcation to the base of skull was measured for 140 carotid arteries. The tortuosity index (TI) of each ICA was calculated by dividing CL by SLD. The relationship of the bifurcation to eight anatomical landmarks in the neck was assessed in order to identify a landmark that could act as a surrogate marker of high carotid bifurcation. The landmarks examined were the angle of mandible, greater horn of hyoid, body of hyoid, upper margin of thyroid cartilage, cervical vertebrae, mastoid process, sternoclavicular joint, and sternal notch. RESULTS: The median curved length of the ICA was 80.4 mm (range 58.0-129.0 mm). The median distance of bifurcation from the base of the skull was 72.7 mm (range 58.1-98.1 mm). There was excellent interobserver agreement in measuring SLD, with an intraclass correlation coefficient of 0.993 (p = .00). The median tortuosity index was 1.12 (range 1.01-1.64). Distance from the mastoid process had the greatest correlation with high bifurcation; Pearson's correlation coefficient of 0.894 (two-tailed p = .00). Bifurcations within 5 cm of the mastoid process are likely to be in the highest quartile (82.9% sensitive, 80.1% specific). CONCLUSIONS: Measuring the distance of carotid bifurcation from the base of the skull (SLD), a measure previously not well defined, may be useful in predicting difficult neck dissection and endarterectomy. A distance from mastoid of ≤ 5 cm may also alert the surgeon to potential difficulties.

"Is general surgery still relevant to the subspecialised trainee?" A 10 year comparison of general versus specialty surgical practice.

BACKGROUND: The splintering of general surgery into subspecialties in the past decade has brought into question the relevance of a continued emphasis on traditional general surgical training. With the majority of trainees now expressing a preference to subspecialise early, this study sought to identify if the requirement for proficiency in managing general surgical conditions has reduced over the past decade through comparison of general and specialty surgical admissions at a tertiary referral center. METHODS: A cross-sectional review of all surgical admissions at Cork University Hospital was performed at three individual time points: 2002, 2007 & 2012. Basic demographic details of both elective & emergency admissions were tabulated & analysed. Categorisation of admissions into specialty relevant or general surgery was made using International guidelines. RESULTS: 11,288 surgical admissions were recorded (2002:2773, 2007:3498 & 2012:5017), showing an increase of 81 % over the 10-year period. While growth in overall service provision was seen, the practice of general versus specialty relevant emergency surgery showed no statistically significant change in practice from 2002 to 2012 (p = 0.87). General surgery was mostly practiced in the emergency setting (84 % of all emergency admissions in 2012) with only 28 % elective admissions for general surgery. A reduction in length of stay was seen in both elective (3.62-2.58 bed days, p = 0.342) & emergency admissions (7.36-5.65, p = 0.026). CONCLUSIONS: General surgical emergency work continues to constitute a major part of the specialists practice. These results emphasize the importance of general surgical training even for those trainees committed to sub-specialisation.

ACE up the sleeve - are vascular patients medically optimized?

OBJECTIVE: To examine the current medical management of arteriopathic patients attending a vascular surgical service at a university teaching hospital over a 6-month period. The prescribing of antiplatelets, statins, angiotensin-converting enzyme (ACE) inhibitors, or angiotensin receptor blockers and beta-blockers was specifically examined. Vascular patients are often under the care of multiple specialties, and therefore the influence of different medical specialties on the patients' medical management was also examined. DESIGN: Between January and June 2009, data were recorded on sequential patients with arterial disease attending the vascular surgical service. Patients' demographics, type of arterial disease, medical consultations within the previous 12 months, and current medications were recorded. RESULTS: The study included 180 patients with a mean age of 69 years (39-88 years). All but 4% were taking an antiplatelet or anticoagulant, predominantly aspirin. There were 86% taking a statin, 44% taking a beta-blocker, and 51% taking an ACE inhibitor. Suboptimal prescription of ACE inhibitors and beta-blockers was evident regardless of the type of medical consultations in the previous year. No specialty group differed significantly from vascular surgeons in their prescribing pattern. CONCLUSIONS: While almost all arteriopaths receive some form of antiplatelet and statin in line with clinical evidence, ACE inhibitors and beta-blockers appear to be under-prescribed in this arteriopathic population. We conclude that opportunity exists for vascular surgeons to embrace recent guidelines and lead the way in both surgical and medical optimization of arteriopathic patients through improving links with primary care physicians or taking greater responsibility themselves for the medical as well as the surgical care of their arteriopathic patients.

Atmospheric pressure variations and abdominal aortic aneurysm rupture.

BACKGROUND: Ruptured abdominal aortic aneurysm (RAAA) presents with increased frequency in the winter and spring months. Seasonal changes in atmospheric pressure mirrors this pattern. AIM: To establish if there was a seasonal variation in the occurrence of RAAA and to determine if there was any association with atmospheric pressure changes. METHODS: A retrospective cohort-based study was performed. Daily atmospheric pressure readings for the region were obtained. RESULTS: There was a statistically significant monthly variation in RAAA presentation with 107 cases (52.5%) occurring from November to March. The monthly number of RAAA and the mean atmospheric pressure in the previous month were inversely related (r = -0.752, r (2) = 0.566, P = 0.03), and there was significantly greater daily atmospheric pressure variability on days when patients with RAAA were admitted. CONCLUSION: These findings suggest a relationship between atmospheric pressure and RAAA.

Combination therapy of cholesterol reduction and L-arginine supplementation controls accelerated vein graft atheroma.

Hyperlipidemia contributes to the development of intimal hyperplasia and accelerated atheroma in vein bypass grafts. Dietary cholesterol reduction and oral supplementation with L-arginine have been shown to reduce accelerated atheroma in experimental vein grafts. This study extends these observations by examining the effect of the combination therapy of cholesterol reduction and L-arginine supplementation on the development of intimal hyperplasia in vein grafts in hypercholesterolemic animals. Thirty New Zealand White rabbits had a carotid vein bypass graft performed and were sacrificed at 28 days postoperatively either for morphology (light and electron microscopy) and videomorphometry, or for in vitro contractile studies. Twenty animals received a 1% cholesterol diet for 4 weeks prior to surgery. This diet was continued until harvest in ten animals. Ten cholesterol-fed animals received L-arginine supplementation (2 g/kg/day, p.o.) for 7 days preoperatively and thereafter until harvest and in addition were returned to a normal diet on the day of surgery. The last ten animals were controls (normal diet). Combined cholesterol reduction and L-arginine supplementation prevented accelerated atheroma in vein grafts, halted the change in enhanced smooth muscle cell contractility, and improved endothelial cell function. Early postoperative therapy targeting atheroma development in the high-risk patient could offer significant morphological and functional benefits.

Early morphology of accelerated vein graft atheroma in experimental vein grafts.

Vein grafts fail because of the development of intimal hyperplasia and atheroma. Recent experimental evidence suggests that the presence of hypercholesterolemia induces a three-fold increase in intimal hyperplasia with early atheroma development within 4 weeks of implantation. We have previously demonstrated endothelial cell preservation and a short-lived (3-day) polymorphonuclear leukocyte infiltrate in vein grafts. The aim of this study is to define the early morphology and ultrastructure of vein grafts implanted into a hyperlipidemic environment to provide a pathological foundation on which to examine the cellular and molecular events that determine this accelerated response. Twenty-one male New Zealand White rabbits underwent a right carotid interposition bypass graft using the ipsilateral external jugular vein; all animals received a 1% cholesterol diet for 4 weeks prior to surgery and continuing postoperatively until harvest. Animals (n = 3 per time point) were sacrificed at 60 min, 1 day, 3 days, 5 days, 7 days, 14 days, and 28 days postoperatively for scanning and transmission electron microscopy of the vein grafts. No concurrent controls were employed. The results of this study suggest that in the presence of hypercholesterolemia, the pathophysiological processes involved in the vein graft are similar to those reported for noncholesterol-fed animals. There is a sustained subendothelial response with the prolonged presence of macrophages and cellular debris and the accumulation of foam cells.

Controlling transplant vasculopathy in cryopreserved vein grafts with polyethylene glycol and glutathione during transport.

BACKGROUND: the biological characteristics of cryopreserved allografts are poorly understood, although many factors are known to influence their outcome. This study examines the development of transplant vasculopathy in both fresh and cryopreserved vein allografts and specifically assesses the efficacy of a transport solution containing 10% polyethylene glycol and 10 microM glutathione (PEG/GSH). METHODS: jugular veins were harvested from control donor rabbits and transplanted as interposition carotid bypass grafts in 30 New Zealand White (NZW) rabbits. Ten received the fresh jugular veins (fresh). Ten animals received jugular veins which had been harvested, transported in a physiological solution, cryopreserved and stored in a standard fashion (cryopreserved). Ten animals received jugular veins which had been harvested, transported in the same solution with the addition of PEG/GSH, cryopreserved and stored in a standard fashion (PEG/GSH). Cryopreserved jugular veins were stored for 6 weeks before transplantation. All animals were sacrificed 28 days postoperatively. Vein grafts were perfusion-fixed and wall dimensions were determined by planimetry. RESULTS: all transplanted grafts were patent at harvest. The control cryopreserved vein grafts showed a 54% increase in mean intimal thickness (63+/-10 micron vs. 41+/-3 micron p<0.05) but no change in mean medial thickness (125+/-9 micron vs. 119+/-13 micron; p = N.S. ) compared to the fresh allograft. Transport of the grafts in PEG/GSH solution resulted in the abolition of the increase in intimal thickness (41+/-4 micron; p <0.01) associated with cryopreservation without a change in medial thickness (140+/-15 micron; p = N.S.) compared to the cryopreserved allograft. CONCLUSION: cryopreserved vein grafts develop significant intimal hyperplasia compared to freshly transplanted grafts. The use of PEG/GSH in the transport solution significantly reduces this transplant graft intimal hyperplasia to that which develops in fresh grafts and may lead to improvements in the clinical use of cryopreserved veins.

Time course of the regression of intimal hyperplasia in experimental vein grafts.

A previous study in which vein grafts were removed from the arterial circulation and reimplanted into the venous circulation of the same animal demonstrated regression of vein graft intimal hyperplasia and medial thickening within 14 days. The present study was designed to characterize the kinetics of the morphological and ultrastructural changes over this 14-day period. Twenty-one male New Zealand White rabbits received a reversed vein interposition bypass graft of the right common carotid artery. Fourteen days after the procedure, 21 vein grafts were isolated, removed, and reimplanted into the contralateral external jugular venous system as veno-venous interposition bypass grafts (reversal grafts). The grafts were harvested at 60 minutes, 1 day, 3 days, 5 days, 7 days, and 14 days after reversal. Before insertion into the venous circulation, the vein graft had a confluent endothelial cell surface with multiple layers of smooth muscle cells representing intimal hyperplasia. After 1 hour, the reversal graft retained an intact endothelial cell layer with no evidence of tissue edema or cellular disruption. By 24 hours, there were a few blood cells on the endothelial cell surface. There was no inflammatory infiltrate seen in the subendothelium, and the smooth muscle cells were unaltered. At 3 days, the endothelial cell lining remained intact with no polymorphonucleocytes in the subendothelium or within the graft wall. Underlying smooth muscle cells at this time were noted to contain cytoplasmic vacuoles. At 5 days, there were no inflammatory cells seen on the surface or within the vein graft wall, but many of the underlying smooth muscle cells within the intimal hyperplasia were noted to be fragmented and to have clumping of chromatin. After 7 days, the endothelial cells remained intact and there was widespread evidence of apoptosis beneath the subendothelium with highly fragmented smooth muscle cells, some of which were histologically in the process of breaking up. At 14 days, the grafts retained uniform endothelial cell surfaces. Most of the smooth muscle cells that composed the intimal hyperplasia seen before implantation as a reversal graft were gone. Areas of newly laid down collagen could be observed. There were no acute inflammatory cells but for some mast cells seen in the graft wall. This study demonstrates that in this model, regression of intimal hyperplasia was associated with apoptosis of the smooth muscle cells and the deposition of collagen. There was no evidence that this process is mediated by an acute inflammatory response. Regression therefore appears to be due to induction of smooth muscle cell apoptosis by either a reduction in pressure or flow or a combination of both factors. The findings will enable a systematic cellular and molecular analysis of the biology of regression, which may afford clues to better understand the biology of the developing intimal hyperplasia.

Laparoscopic-assisted and hand-access laparoscopic surgery.

Therapeutic laparoscopy has developed rapidly since the first report of laparoscopic cholecystectomy in 1986. Several other reports followed before the end of that decade. Recent developments have resulted from a combination of the many technical improvements in laparoscopic equipment, together with the innovation and increasing experience of those performing laparoscopic surgery. Cholecystectomy, appendectomy, gastric fundoplication, and preperitoneal hernia repair are all now widely practiced and often laparoscopy is the favored approach.

Locally applied antisense oligonucleotide to proliferating cell nuclear antigen inhibits intimal thickening in experimental vein grafts.

This study examines the effect of antisense oligonucleotide to proliferating cell nuclear antigen (PCNA) on the formation of vein graft intimal hyperplasia in vivo, using localized administration. Twenty-four New Zealand white rabbits had a right carotid interposition bypass graft using the external jugular vein and were sacrificed on the 28th postoperative day. To determine the effect of PCNA on the development of intimal hyperplasia, 6 animals had their grafts coated with a pluronic gel containing 18 base antisense oligonucleotide to PCNA (1 mg/ml), 6 received a pluronic gel containing an 18 base nonsense oligonucleotide (1 mg/ml), and 12 animals were controls (6 with and 6 without pluronic gel). These grafts were harvested for morphology and videomorphometry. There was no change in the intimal thickness between the control and gel-treated groups. (70 +/- 4 microm versus 72 +/- 4 microm; mean +/- s.e.m.; p = ns). The presence of nonsense oligonucleotide had no further effect. Antisense PCNA produced a 26% decrease in intimal thickness to 50 +/- 4 microm in the treated vein grafts (p < 0.03) without a change in medial thickness. This study shows that a local single application of antisense oligonucleotide to PCNA will reduce the intimal hyperplasia in experimental vein grafts over 28 days.

G protein signaling and vein graft intimal hyperplasia: reduction of intimal hyperplasia in vein grafts by a Gbetagamma inhibitor suggests a major role of G protein signaling in lesion development.

Vein grafting results in the development of intimal hyperplasia with accompanying changes in guanine nucleotide-binding (G) protein expression and function. Several serum mitogens that act through G protein-coupled receptors, such as lysophosphatidic acid, stimulate proliferative pathways that are dependent on the G protein betagamma subunit (Gbetagamma)-mediated activation of p21ras. This study examines the role of Gbetagamma signaling in intimal hyperplasia by targeting a gene encoding a specific Gbetagamma inhibitor in an experimental rabbit vein graft model. This inhibitor, the carboxyl terminus of the beta-adrenergic receptor kinase (betaARK(CT)), contains a Gbetagamma-binding domain. Vein graft intimal hyperplasia was significantly reduced by 37% (P<0.01), and physiological studies demonstrated that the normal alterations in G protein coupling phenotypically seen in this model were blocked by betaARK(CT) treatment. Thus, it appears that Gbetagamma-mediated pathways play a major role in intimal hyperplasia and that targeting inhibitors of Gbetagamma signaling offers novel intraoperative therapeutic modalities to inhibit the development of vein graft intimal hyperplasia and subsequent vein graft failure.

Alterations in collagen subtype III and IV protein in experimental venous bypass grafting.

OBJECTIVE: To examine changes in collagen III and IV protein during the development of intimal hyperplasia in experimental vein grafts. METHODS: Sixteen New Zealand White rabbits underwent reversed, jugular vein, interposition grafting of the carotid artery. Vessels were harvested 3, 7 or 28 days after operation and subjected to immunohistochemical examination and gelatinase assays. RESULTS: In control vein, collagen IV was expressed around adventitial blood vessels and throughout the endothelium. Compared with its presence in control veins, collagen IV protein was decreased in the endothelium in all 3-day vein grafts and undetectable in the endothelium and intima in 7-day vein grafts, but was present in the endothelium and intimal hyperplasia in 28-day vein grafts. In contrast, collagen III was absent from the endothelium of control vein and 3-day vein grafts, was present at low levels in the intima of 7-day vein grafts, but was absent from the endothelium and intimal hyperplasia in 28-day vein grafts. In 3-day vein grafts, areas of collagen IV loss colocalized to areas of leukocyte infiltration. Protein extracts from 3-day vein grafts contained a 72 kDa gelatinase. CONCLUSIONS: The presence and alterations of collagen protein in veins and vein grafts are subtype specific. Collagen III does not appear to be a normal component of intimal hyperplasia in vein grafts. The decrease in collagen IV protein in the endothelium of veins may be a component of the endothelial changes that follow bypass grafting, mediated by leukocytes, the induction of gelatinase activity, or both.

Local effects of nitric oxide supplementation and suppression in the development of intimal hyperplasia in experimental vein grafts.

OBJECTIVES: The universal response of vein grafts after insertion into the arterial circulation is the development of intimal hyperplasia; smooth muscle cell proliferation and connective tissue deposition, which may be modulated in part by dysfunctional endothelial nitric oxide (NO) metabolism. This study examines the effects of single dose, local application by pluronic gel of a NO donor, S-nitroso-N-acetylpenicillamine (SNAP) and an NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME) on the formation of intimal hyperplasia. MATERIALS: Forty New Zealand white rabbits underwent jugular vein interposition grafting of the common carotid artery. DESIGN: Ten animals were controls, 10 animals had the outer surface of the vein graft coated with 30% pluronic gel (2.5 ml), and 10 each were immersed for 15 min prior to insertion in Ringer lactate containing 10(-3) M of SNAP or L-NAME and then had their vein grafts coated with 2.5 ml of gel containing either SNAP (10(-3) M) or L-NAME (10(-3) M), which allows for sustained delivery for up to 6 h. On the 28th post operative day, the animals were sacrificed and vein grafts were harvested for morphology by electron microscopy (SEM and TEM) and dimensional analysis by videomorphometry. RESULTS: All vein grafts developed intimal hyperplasia. On SEM the vein grafts had a confluent layer of endothelial cells with multiple layers of smooth muscle cells representing intimal hyperplasia in TEM. There were no demonstrable morphological differences between the four groups. Local treatment with SNAP produced a significant 36% decrease in mean intimal thickness (72 +/- 4 microns vs. 45 +/- 4 microns; mean +/- S.E.M.; p < 0.01) without a change in medial thickness compared to gel-only treated groups (58 +/- 6 microns vs. 61 +/- 7 microns; p = ns). Inhibition of NO synthase by L-NAME had no effect on the development of intimal hyperplasia (72 +/- 4 microns vs. 79 +/- 10 microns; p = ns); medial thickness was also unchanged. CONCLUSION: These data confirm the protective effect of NO in vascular injury and suggest that NO synthase activity is either absent or reduced to such a level that further inhibition in this short time course is not relevant to the pathophysiology of vein graft intimal hyperplasia.

Localized versus systemic angiotensin II receptor inhibition of intimal hyperplasia in experimental vein grafts by the specific angiotensin II receptor inhibitor L158,809.

BACKGROUND: This study examines the effect of the angiotensin II receptor (type 1) antagonist (L158,809) on the formation of vein graft intimal hyperplasia in vivo, by both localized and systemic administration. METHODS: Forty New Zealand White rabbits underwent carotid interposition bypass grafting with the external jugular vein and were killed on postoperative day 28. To determine the effect of L158,809 on the development of intimal hyperplasia, 10 animals received long-term oral therapy with L158,809 (10 mg/kg/day, begun 5 days before operation and continued until harvest), 10 animals underwent coating of the grafts with a pluronic gel containing L158,809 (10(-5) mol/L), and 20 animals were controls (10 with and 10 without pluronic gel). These grafts were harvested for either histologic analysis (n = 6 per group) or in vitro isometric tension studies to angiotensin II (n = 4 per group). RESULTS: Long-term oral treatment with L158,809 produced a 43% decrease in intimal thickness from 76 +/- 6 microns (mean +/- SEM) in the control animals to 43 +/- 7 microns in the treated vein grafts (p = 0.002). There was also a significant decrease (44%) in the medial thicknesses between the control (75 +/- 4 microns) and L158,809-treated (42 +/- 6 microns) vein grafts (p = 0.007). The contractile responses to angiotensin II were abolished in the vein grafts by long-term L158,809 therapy. Local application by gel of L158,809 produced a significant decrease (33%) in the intimal thickness (48 +/- 3 microns) but no change in medical thicknesses (76 +/- 6 microns) compared with control grafts. The contractile responses to angiotensin II were unchanged in the vein grafts by local L158,809 therapy. CONCLUSIONS: This study shows that a local single application of L158,809 will reduce the intimal response but not the medial response in vein grafts, whereas long-term treatment will reduce intimal hyperplasia and the medial response in experimental vein grafts. Therefore angiotensin II acting through AT1 receptors mediates a significant part of the intimal hyperplastic response in vein grafts that appears to involve two phases: an acute intimal response requiring short-term therapy and a long-term medial response that requires prolonged therapy.

Local delivery of a tissue factor antibody reduces early leukocyte infiltration but fails to limit intimal hyperplasia in experimental vein grafts.

BACKGROUND: Tissue Factor-mediated thrombin generation involves the generation of VIIa and Xa and has been implicated in the pathogenesis of intimal hyperplasia. In experimental vein grafts, Tissue Factor protein is increased over the first 3 days and colocalized with CD18-positive leukocytes; this increase in Tissue Factor precedes the development of intimal hyperplasia. This study further evaluates the potential role of Tissue Factor in vein graft intimal hyperplasia by directly inhibiting Tissue Factor protein. METHODS: New Zealand white rabbits underwent interpositional bypass grafting of the common carotid artery using the external jugular vein. Perioperatively, murine anti-rabbit Tissue Factor antibody (109 microg/ml gel, 12,500x IC50 of Tissue Factor activity) was applied to the adventitial surface of the graft, using a pluronic gel (30% soln.). Tissue Factor antibody treated vein grafts were compared to control and empty gel-treated vein grafts. Vein grafts were examined at 3 days to assess CD18-positive leukocyte infiltration and the presence of residual antibody by Western blotting. At 28 days, intimal and medial dimensions were quantified using videomorphometry. RESULTS: At day 3, there was marked reduction in CD18-positive leukocytes in the Tissue Factor antibody versus control vein grafts (6.3 +/- 4.7 vs 20.8 +/- 7.4 per 200x field, P < 0.05). At 28 days, intimal hyperplasia was similar for the control (70 +/- 4 microm, mean +/- SEM), gel (73 +/- 4 microm), and Tissue Factor antibody (75 +/- 4 microm) vein grafts. However, medial thickness (76 +/- 4 microm;, P < 0.05) was significantly increased compared to the gel treated vein graft (61 +/- 5 microm). CONCLUSION: Local delivery of pharmacologic doses of an anti-rabbit Tissue Factor antibody decreased CD18-positive leukocyte infiltration but failed to limit intimal hyperplasia in experimental vein grafts. The results suggest that inhibition of Tissue Factor protein modulates polymorphonuclear leukocyte-endothelial interactions but not in the subsequent development of intimal hyperplasia. It implies that the relationship between the extrinsic coagulation cascade and intimal hyperplasia in vein grafts is complex.

Modulation of tissue factor protein expression in experimental venous bypass grafts.

Vein graft failure is a major limitation of coronary artery and peripheral vascular surgery. Tissue factor (TF), a transmembrane glycoprotein, generates thrombin by initiating the extrinsic coagulation cascade and plays a major role in the response to arterial injury. This study was designed to examine changes in TF protein expression in response to venous bypass grafting. New Zealand White rabbits underwent interposition bypass grafting of the common carotid artery via the ipsilateral external jugular vein. The contralateral control jugular veins (n = 6), early vein grafts (1 or 3 days after grafting, n = 18), and late vein grafts (14 or 28 days after grafting, n = 8) were examined by immunohistochemistry. The presence or absence of TF immunostaining in the intima was assessed in each vessel quadrant. In control veins, intimal TF staining was present in 5 of 24 vessel quadrants. In early vein grafts, TF staining was markedly increased in the intima (72 of 72 quadrants, P < .001 vs control veins), and TF immunostaining colocalized with CD18-positive leukocytes but not with endothelial cells, vascular smooth muscle cells, or RAM11-positive macrophages. In late vein grafts with intimal hyperplasia, TF expression was low or absent in the intima (6 of 32 quadrants, P < .001 vs early vein grafts; P = NS vs control veins), although medial smooth muscle cells expressed TF. Marked changes in TF expression occur in vein grafts. In early vein grafts, TF protein was greatly increased in the intima for at least 3 days and was associated with CD18-positive leukocytes. In late vein grafts with intimal hyperplasia, however, TF protein was not seen in the intima. These findings may have important implications for the development of therapeutic strategies to limit vein graft failure.

Oral monoterpene therapy (perillyl alcohol) reduces vein graft intimal hyperplasia.

The development of intimal hyperplasia is recognized as a major impediment to graft patency. D-Limonenes are monoterpenes with a recognized cytostatic effect on cell proliferation by inhibiting posttranslational isoprenylation of p21ras and other small G-proteins. This study examines the effect of perillyl alcohol, an oral hydroxylated D-limonene, on the development of intimal hyperplasia and its associated smooth muscle cell physiological responses in an experimental model of vein bypass grafting. Twenty New Zealand white rabbits had a right carotid interposition bypass graft using the ipsilateral external jugular vein. Ten animals received chronic oral therapy with a perillyl alcohol (200 mg/kg/day; begun 5 days before surgery and continued until harvest) and 10 control animals received vehicle only. All animals were sacrificed on the 28th postoperative day. Vein grafts were harvested either for morphology/videomorphometry (n = 6 per group) or for in vitro isometric tension studies (n = 4; four 5-mm rings per graft). The cell proliferation and incorporation of [3H]thymidine into the cellular DNA of serum-stimulated rabbit aortic smooth muscle cells was also assessed in the presence of increasing concentrations of perillyl alcohol (10(-9)-10(-4) M). Perillyl alcohol treated vein grafts showed a 22% reduction in overall mean intimal thickness (54 +/- 4 microns vs 69 +/- 3 microns; P = 0.006) but a 25% increase in overall mean medial thickness (86 +/- 4 microns vs 61 +/- 3 microns). The intimal ratio of the perillyl alcohol treated vein grafts decreased by 27% compared to controls. Perillyl alcohol induced norepinephrine and serotonin hypersensitivity in vein grafts compared to controls. The IC50 for perillyl alcohol was 176 nM with maximal inhibition at 5 microM. Incubation of smooth muscle cell cultures with increasing concentrations of perillyl alcohol showed a dose-dependent decrease in in vitro cellular proliferation, maximal at 1 microM. Therapy with perillyl alcohol alters the early development of intimal hyperplasia reducing the intimal response but increasing the medial response without significant changes in the physiological responses of the smooth muscle cells. Modulating G-proteins will affect the intimal hyperplastic response in vein grafts.

Antisense oligonucleotide to proto-oncogene c-myb inhibits the formation of intimal hyperplasia in experimental vein grafts.

BACKGROUND: The development of intimal hyperplasia is a major cause of early vein graft failure. The study examines the effects of locally delivered antisense oligonucleotides to the proto-oncogene c-myb on the development of vein graft intimal hyperplasia. METHODS: Common carotid vein bypass grafting procedures were performed on 60 New Zealand White rabbits. Seventeen grafts were controls, 14 had grafts coated with a commercial gel, 17 had grafts coated with gel containing 200 micrograms of an antisense c-myb oligonucleotide, and 6 rabbits each had grafts coated with gel containing one of two control oligonucleotides. Grafts were harvested 28 days after surgery, and sections were taken for dimensional analysis, morphologic evaluation, and vasomotor function. Grafts were also harvested at 1 day for oligonucleotide uptake/localization analysis and at 3 days for c-myb mRNA analysis. RESULTS: Oligonucleotides were uniformly distributed within the media and adventitia by 1 day. A 38% reduction occurred in mean intimal thickness in the vein grafts coated with antisense to c-myb compared with the other groups. No difference in medial thickness was seen among groups. By scanning and transmission electron microscopy all vein grafts showed a confluent endothelium. In contrast to control vein grafts, which did not relax to acetylcholine, most of the gel and all of the gel/oligonucleotide-coated grafts relaxed by more than 40% of precontracted tension. Responses to a panel of contractile agents were unchanged in the treated groups compared with those in the control group. CONCLUSIONS: Locally delivered antisense oligonucleotides to proto-oncogene c-myb significantly reduces intimal hyperplasia with preservation of acetylcholine-mediated endothelium-dependent relaxation in experimental vein grafts. These findings suggest that targeting a common regulatory pathway of vascular smooth muscle mitogenesis can be successful in reducing the early development of intimal hyperplasia.

Efficient adenoviral gene transfer to early venous bypass grafts: comparison with native vessels.

OBJECTIVES: Gene therapy may provide new approaches to reduce vein graft failure following coronary or peripheral bypass surgery. The aim of this study was to investigate the relative efficacy of intraoperative adenoviral gene transfer to vein grafts, comparing transgene expression in vein grafts with that in matched native vessels in the same animal. In addition, we assessed the impact of bypass grafting on the cellular targets of gene transfer. METHODS: New Zealand White rabbits underwent interposition bypass grafting of the carotid artery, using the ipsilateral external jugular vein, which was infected with an adenovirus expressing beta-galactosidase immediately prior to bypass grafting (n = 16). The contralateral native jugular vein (n = 16) and carotid artery (n = 8) were infected concurrently with the same adenoviral preparation. After 3, 7 or 14 days, beta-galactosidase protein expression was quantified by ELISA, and specific cell types expressing beta-galactosidase were identified by X-Gal staining and by immunohistochemistry. RESULTS: After 3 days, endothelial cells were efficiently transduced in all vessels; medial smooth muscle cells were transduced infrequently. In contrast to jugular veins after gene transfer, endothelium in vein grafts showed expression of VCAM-1 and ICAM-1, and intense inflammation with CD18+ leukocytes. Transgene expression in vein grafts at day 3 was maintained at levels approximately 50% of that in ungrafted jugular veins, but continued to decrease through day 7. CONCLUSIONS: Although vascular injury in early venous bypass grafts reduces gene transfer efficacy, significant transgene expression is maintained for at least 7 days. These findings have important implications for intraoperative gene transfer strategies in vein grafts.

Characterisation of angiotensin II receptor mediated responses and inhibition of intimal hyperplasia in experimental vein grafts by the specific angiotensin II receptor inhibitor, L158,809.

OBJECTIVES: This study characterises pharmacologically the angiotensin II receptor in experimental vein grafts and examines the effect of the angiotensin II receptor (type 1) antagonist (L158,809) on the formation of vein graft intimal hyperplasia in vivo, as well as the in vitro physiological response to angiotensin II of vein grafts after chronic oral L158,809 treatment. MATERIALS: Thirty New Zealand White rabbits had a right carotid interposition bypass graft using the external jugular vein and were killed on the 28th postoperative day. DESIGN: To characterise the angiotensin II receptors, concentration response curves to angiotensin II were obtained in vitro in the presence or absence of L158,809. To determine the effect of L158,809 on the development of intimal hyperplasia, 10 animals received chronic oral therapy with L158,809 (10 mg/kg/day; begun 5 days before surgery and continued until harvest) and 10 animals received vehicle only as controls. These grafts were harvested either for histology (n = 6 per group) or for in vitro isometric tension studies to angiotensin II. RESULTS: The monophasic contractile response to angiotensin II in the untreated vein grafts could be inhibited in a concentration dependent manner by L158,809 with first order kinetics. Chronic oral treatment with L158,809 produced a 48% decrease in intimal thickness from 82 +/- 1 micron (mean +/- S.E.M.) in the controls to 43 +/- 7 microns in the treated vein grafts (p = 0.002). There was also a significant decrease (45%) in the medial thickness between the control (76 +/- 6 microns) and L158,809 treated (42 +/- 6 microns) vein grafts (p = 0.007). The responses to angiotensin II were abolished in the vein grafts by chronic L158,809 therapy. CONCLUSIONS: This study suggests that vein graft angiotensin II responses are mediated through a type 1 receptor and that chronic inhibition with L158,809, significantly reduces intimal hyperplasia and medial hypertrophy in experimental vein grafts and concomitantly abolishes the in vitro responses to angiotensin II. Therefore, angiotensin II acting through AT1 receptors mediates a significant part of the intimal hyperplastic response in vein grafts.