Genetic risk score associations for myocardial infarction are comparable in persons with and without rheumatoid arthritis: the population-based HUNT study.

Persons with rheumatoid arthritis (RA) have increased risk of myocardial infarction (MI). Overlapping associations with MI of weighted genetic risk scores (wGRS) for coronary artery disease (CAD) and RA is unknown in a population-based setting. Data from the prospective Nord-Trøndelag Health Study (HUNT2: 1995-1997 and HUNT3: 2006-2008) were used. wGRS added each participant's carriage of all risk variants weighted by the coefficient from published association studies. Published wGRS for CAD and RA were analysed in Cox regression with MI as outcome, age as analysis time, and censoring at the first MI, death, or 31.12.2017. 2609 of 61,465 participants developed MI during follow-up (mean 17.7 years). The best-fitting wGRS for CAD and RA included 157 and 27 single-nucleotide polymorphisms, respectively. In multivariable analysis including traditional CAD risk factors, the CAD wGRS was associated with MI [hazard ratio = 1.23 (95% CI 1.18-1.27) for each SD increase, p < 0.0001] in RA patients (n = 433) and controls. The RA wGRS was not significant (p = 0.06). Independently from traditional risk factors, a CAD wGRS was significantly associated with the risk for MI in RA patients and controls, whereas an RA wGRS was not. The captured genetic risk for RA contributed little to the risk of MI.

Autoimmune diabetes in adults and risk of myocardial infarction: the HUNT study in Norway.

BACKGROUND: The long-term consequences of autoimmune diabetes in adults (AIDA) are largely unexplored. OBJECTIVE: To investigate the risk of myocardial infarction (MI) in AIDA compared to type 2 diabetes, taking into consideration the effects of socio-economic and lifestyle factors, the metabolic syndrome and glycaemic control. METHODS: A total of 62 995 participants including 207 individuals with AIDA (onset ≥35 years and anti-GAD positive) and 2322 individuals with type 2 diabetes (onset ≥35 years and anti-GAD negative), from the population-based Norwegian HUNT study, were followed for a first MI during the period 1995-2008. We identified 2614 MIs by hospital records or the National Cause of Death Registry. Cox proportional hazard models were used to estimate the risk of MI by diabetes subgroups after adjustment for age and socio-economic and lifestyle factors. RESULTS: AIDA amongst women was associated with a nearly fourfold increased risk of MI [hazard ratio (HR) 3.63, 95% confidence interval (CI) 2.21-5.96) compared to nondiabetic participants, whereas no excess risk was found in men with AIDA (HR 1.30, 95% CI 0.70-2.52). By contrast, type 2 diabetes was associated with an increased MI risk in both men (HR 1.92, 95% CI 1.62-2.26) and women (HR 2.39, 95% CI 1.98-2.89). The metabolic profile was more favourable in patients with AIDA than in those with type 2 diabetes, but glycaemic control was worse. Multivariable models and sensitivity analyses suggest that these results were robust. CONCLUSIONS: Women with AIDA were more likely to develop MI, compared to men with AIDA and both men and women with type 2 diabetes. Further investigations are warranted to confirm this gender difference.

Cardiac function associated with previous, current and repeated depression and anxiety symptoms in a healthy population: the HUNT study.

OBJECTIVE: Symptoms of anxiety and depression often co-exist with cardiovascular disease (CVD), yet little is known about the association with left ventricular (LV) subclinical dysfunction. We aimed to study the cross-sectional associations of previous, current and repeated depression or anxiety symptoms, with sensitive indices of LV systolic and diastolic function, based on tissue Doppler (TD) and speckle tracking (ST) imaging methods. METHODS: A random selection of 1296 individuals free from known CVD, hypertension and diabetes were examined with echocardiography at baseline of the third Nord-Trøndelag Health Study, (HUNT3, 2006-2008). The primary outcomes were LV diastolic function (e') and LV systolic function (longitudinal global strain). The primary exposures were self-report on the Hospital Anxiety and Depression Scale (HADS). Associations between outcomes and baseline exposures were available for 1034 (80%), and with previous and repeated exposures for 700 participants who also participated in HUNT2 (1995-1997). RESULTS: Previous and repeated depression symptoms, but not current depression, were linearly associated with a reduction in e'. The average sum of two repeated HADS-D scores 10 years apart had the strongest effect on e' (-8.3%; 95% CI -13.9% to -2.7%) per 5 units. We observed a sex difference between depression symptoms and longitudinal global strain (p for interaction 0.019), where women had a marginal negative effect. Anxiety symptoms, neither previous, current nor repeated were associated with subclinical LV dysfunction. CONCLUSIONS: In a healthy sample, confirmed free of CVD, past and repeated depression symptoms were associated with subclinical LV dysfunction. Thus, depression symptoms might represent a modifiable risk factor for future CVD.

When is supervised exercise therapy considered useful in peripheral arterial occlusive disease? A nationwide survey among vascular surgeons.

OBJECTIVES: Although international guidelines state that supervised exercise therapy (SET) should be offered to all patients with intermittent claudication (IC), SET appears to be underutilised in clinical practice. The aim of this study was to document current opinions of Dutch vascular surgeons on SET as treatment option for peripheral arterial occlusive disease (PAOD). MATERIALS AND METHODS: Vascular surgeons and fellows in vascular surgery were asked to complete a 24-question survey either at the 2011 Annual Meeting of the Dutch Society for Vascular Surgery or online. RESULTS: Ninety-one participants, including 83 vascular surgeons (51% of all Dutch vascular surgeons), completed the survey. The respondents would refer 75.4% of newly diagnosed patients with IC for SET. SET was considered less useful in patients with IC and major (cardiopulmonary) co-morbidity or a significant iliac artery stenosis. In critical limb ischaemia, the combination of SET and angioplasty was considered useful in 71.9%. Respondents regarded patient satisfaction (63.3%) and improvement in pain-free or maximal walking ability (26.6%) as clinically most relevant goals of SET. Most (84.4%) agreed that SET should also include lifestyle management. CONCLUSION: Although the vast majority of Dutch vascular surgeons consider SET as an important treatment option for PAOD, SET should receive more emphasis in clinical practice since arguments not to refer for SET are outdated. Furthermore, vascular surgeons agree that lifestyle management should be integrated in SET.

Alpha-tocopheryl succinate sensitises a T lymphoma cell line to TRAIL-induced apoptosis by suppressing NF-kappaB activation.

Activation of nuclear factor-kappaB (NF-kappaB) can interfere with induction of apoptosis triggered by the tumour necrosis factor-related apoptosis-inducing ligand (TRAIL; Apo2L). Therefore, agents that suppress NF-kappaB activation may sensitise cells to TRAIL-dependent apoptosis. Exposure of Jurkat cells to TRAIL resulted in massive and saturable apoptosis induction, following an initial lag time. This lag was abolished by pretreatment of the cells with subapoptotic doses of alpha-tocopheryl succinate (alpha-TOS) or the proteasome inhibitor MG132. Exposure of the cells to TRAIL led to a rapid, transient activation of NF-kappaB, a process that was suppressed by cell pretreatment with alpha-TOS or MG132. Activation of NF-kappaB by TNF-alpha prior to TRAIL exposure increased resistance of the cells to TRAIL-mediated apoptosis. We conclude that alpha-TOS sensitises cells to TRAIL killing, at least in some cases, through inhibition of NF-kappaB activation. This further supports the possibility that this semisynthetic analogue of vitamin E is a potential adjuvant in cancer treatment, such as in the case of TRAIL-mediated inhibition of cancer.

Apoptotic death of inflammatory cells in human atheroma.

Although the accumulation of cholesterol and other lipidic material is unquestionably important in atherogenesis, the reasons why this material progressively accumulates, rather than being effectively cleared by phagocytic cells such as macrophages, are not completely understood. We hypothesize that atheromatous lesions may represent "death zones" that contain toxic materials such as oxysterols and in which monocytes/macrophages become dysfunctional and apoptotic. Indeed, cathepsins B and L, normally confined to the lysosomal compartment, are present in the cytoplasm and nuclei of apoptotic (caspase-3-positive) macrophages within human atheroma. The possible involvement of oxysterols is suggested by experiments in which cultured U937 and THP-1 cells exposed to 7-oxysterols similarly undergo marked lysosomal destabilization, caspase-3 activation, and apoptosis. Like macrophages within atheroma, intralysosomal cathepsins B and L are normally present in the cytoplasm and nuclei of these oxysterol-exposed cells. Lysosomal destabilization, cathepsin release, and apoptosis may be causally related, because inhibitors of cathepsins B and L suppress oxysterol-induced apoptosis. Thus, toxic materials such as 7-oxysterols in atheroma may impair the clearance of cholesterol and other lipidic material by fostering the apoptotic death of phagocytic cells, thereby contributing to further development of atherosclerotic lesions.

Cytoskeletal involvement during hypo-osmotic swelling and volume regulation in cultured chick cardiac myocytes.

The membrane skeleton in spherical cardiac myocytes subjected to hypo-osmotic challenge was examined by laser scanning confocal microscopy. A distinct cortical layer intimately localized under the plasmalemma was revealed for spectrin and actin (including filamentous actin and alpha-sarcomeric actin). Desmin filaments were abundant and in close contact with the plasmalemma. During swelling and subsequent regulatory volume decrease (RVD) the structural integrity of these cytoskeletal elements remained intact, and the close association between actin and plasmalemma persisted as confirmed by double immunolabeling. Subplasmalemmal beta-tubulin labeling was sparse. Hypo-osmotic conditions disrupted the microtubules and depolymerized tubulin. Neither pretreatment with taxol nor with colchicine, resulted in any effect on cell volume regulation. The present results show that actin, desmin, and spectrin contribute to a subplasmalemmal cytoskeletal network in spherical cardiac myocytes, and that this membrane skeleton remains structurally intact during swelling and RVD. It is suggested that the integrity of this membrane skeleton is important for stabilization of the plasmalemma and the membrane-integrated proteins during hypo-osmotic challenge, and that it may participate in the regulation of the cell volume.

N-Glycans influence the in vitro adhesive and invasive behaviour of three metastatic cell lines.

Alterations in cellular glycosylation may play a key role in metastatic behaviour of tumour cells. We studied three metastatic cell lines, LOX (malignant melanoma), FEMX (malignant melanoma) and MA-11 (mammary carcinoma). These cell lines have a very different metastatic behaviour in vivo, and different glycans have been postulated to be partly responsible for these differences. To further investigate the functional role of carbohydrates, these three cell lines have been treated with tunicamycin, an inhibitor of the biosynthesis of N-glycans and benzyl- alpha-N-acetylgalactosamine (benzyl-alpha-GalNAc; BnGalNAc), an inhibitor of mature O-linked glycans. Various in vitro adhesion and invasion assays were undertaken for functional studies. Tunicamycin significantly inhibited adhesion to laminin, but only slightly affected cell adhesion to collagen IV. The same compound significantly decreased cellular invasiveness through a Matrigel invasion chamber. Moreover, tunicamycin reduced homotypic aggregation of cells. BnGalNAc had generally little effect on cell behaviour in in vitro assay. The effects of the inhibitors were, however, to some extent cell line-specific. We conclude that N-glycans, but probably not mature O-glycans have important in vitro functions in cell adhesion to laminin, cell invasion through Matrigel and cellular aggregation in the studied cell lines. These results support the view that carbohydrates are functionally involved in several steps of the metastatic process.

Induction of cell death by the lysosomotropic detergent MSDH.

Controlled lysosomal rupture was initiated in lysosome-rich, macrophage-like cells by the synthetic lysosomotropic detergent, O-methyl-serine dodecylamide hydrochloride (MSDH). When MSDH was applied at low concentrations, resulting in partial lysosomal rupture, activation of pro-caspase-3-like proteases and apoptosis followed after some hours. Early during apoptosis, but clearly secondary to lysosomal destabilization, the mitochondrial transmembrane potential declined. At high concentrations, MSDH caused extensive lysosomal rupture and necrosis. It is suggested that lysosomal proteases, if released to the cytosol, may cause apoptosis directly by pro-caspase activation and/or indirectly by mitochondrial attack with ensuing discharge of pro-apoptotic factors.

Lysosomal destabilization during macrophage damage induced by cholesterol oxidation products.

We have previously shown that oxidized low-density lipoprotein (LDL) induces damage to the macrophage lysosomal membranes, with ensuing leakage of lysosomal contents and macrophage cell death. Cholesterol oxidation products (ChOx) have been reported to be the major cytotoxic components of oxidized LDL/LDL- and also to stimulate cholesterol accumulation in vascular cells. In the present study, we characterized the initial events during macrophage damage induced by cholesterol oxidation products (ChOx). Within 24 h of exposure, ChOx caused lysosomal destabilization, release to the cytosol of the lysosomal marker-enzyme cathepsin D, apoptosis, and postapoptotic necrosis. Enhanced autophagocytosis and chromatin margination was found 12 h after the exposure to ChOx, whereas apoptosis and postapoptotic necrosis was pronounced 24 and 48 h after the exposure. Some lysosomal vacuoles were then filled with degraded cellular organelles, indicating phagocytosis of apoptotic bodies by surviving cells. Because caspase-3 activation was detected in the ChOx-exposed cells, lysosomal destabilization may associate with the leakage of lysosomal enzymes, and activation of the caspase cascade. MnSOD mRNA levels were markedly increased after 24 h of exposure to ChOx, suggesting associated induction of mitochondrial protection repair or turnover. We conclude that ChOx-induced damage to lysosomes and mitochondria are sequelae to the cascade of oxysterol cytotoxic events. The early disruption of lysosomes induced by ChOx, with resultant autophagocytosis may be a critical event in apoptosis and/or necrosis of macrophages/foam cells during the development of atherosclerotic lesions.

Membrane skeleton in cultured chick cardiac myocytes revealed by high resolution immunocytochemistry.

Distribution of cytoskeletal proteins with emphasis on the membrane-cytoskeleton interface was examined in cultured cardiac myocytes. Using specific antibodies recognizing alpha-sarcomeric actin, desmin, beta-tubulin, spectrin/alpha-fodrin and ankyrin, respectively, the cellular localization of these cytoskeletal proteins was detected by laser scanning confocal microscopy. In addition, the fine filamentous structure of these proteins was identified by combining silver-enhanced immunogold labelling with electron microscopy. The latter technique employed the sequence of quick-freezing, deep-etching and rotary shadowing of the specimens. Conventional transmission electron microscopy of the spherical cardiac myocytes revealed a filamentous submembranous layer, approximately 100 nm thick. Specific immunolabelling of alpha-sarcomeric actin and spectrin/alpha-fodrin as well as ankyrin was seen beneath the plasmalemma. A three-dimensional meshwork of spectrin/alpha-fodrin was shown. Numerous desmin filaments that exhibited a tortuous course throughout the cells were also observed running in parallel with the surface in the submembranous area, whereas beta-tubulin was infrequently detected in these areas. In conclusion, the present study shows that spherical cardiac myocytes contain a distinct and complex three-dimensional membrane skeleton. Major constituents of this distinct submembranous layer were spectrin/alpha-fodrin fibres as well as actin and desmin filaments.

Minute oxidative stress is sufficient to induce apoptotic death of NIT-1 insulinoma cells.

When cultured NIT-1 cells were subjected to a low level of oxidative stress (30 microM hydrogen peroxide for 15 min at 37 degrees C) several of their lysosomes ruptured, as demonstrated by intravital staining with the lysosomotropic weak base acridine orange. Such rupture is due to intralysosomal, iron-catalyzed oxidative reactions, since it was largely prevented by previous endocytotic uptake of desferrioxamine. The resultant limited leakage of lysosomal hydrolytic enzymes into the cytosol could be important for an apoptotic-type degradation/fragmentation process within initially intact plasma membranes. In contrast, extensive lysosomal rupture leads to necrosis. The development of the damage process was followed by light- and electron microscopy; and by the TUNEL-reaction. As a result of the applied oxidative stress, which is comparable to that expected to occur within the microenvironment surrounding activated macrophages under oxidative burst (e.g. during autoimmune insulitis), about 90% of the cells eventually died due to post-apoptotic secondary necrosis. The few surviving cells phagocytosed the debris from their fragmented neighbours and began to divide about 24 h after the insult. Thus the sensitivity to oxidative stress varies, perhaps as a consequence of varying amounts of intralysosomal redox-active iron, as we have found to be the case in several other cellular systems. Since the NIT-1 cells are highly differentiated, and in many ways like beta cells, we consider our result to be of value for the understanding of beta-cell death during the development of insulin-dependent (Type I) diabetes mellitus (IDDM).

A Norwegian nosocomial outbreak of methicillin-resistant Staphylococcus aureus resistant to fusidic acid and susceptible to other antistaphylococcal agents.

In Norway, infections caused by methicillin resistant Staphylococcus aureus (MRSA) are still uncommon. From December 1993 to January 1997, MRSA was isolated from 22 people in Oslo county; 17 patients and five carriers (healthcare workers). A cluster of ten people (five patients and five healthcare workers) were associated with an outbreak at two hospitals in Oslo. The five patients were all admitted to the same intensive care unit (ICU) at Ullevål University Hospital between May-July 1995 (they were not transferred from abroad) and treated for acute neurological lesions. After surgery, four of them (one died) were transferred to another hospital for rehabilitation and training. The presence of MRSA was discovered in the patients and the five healthcare workers during the 10 months June 1995-March 1996. All cluster strains showed an unusual antibiotic resistance pattern in vitro, with a relatively low degree of methicillin resistance, resistance to fusidic acid, but sensitivity to all other anti-staphylococcal agents. A clonal spread of this fusidic acid resistant MRSA was supported by strain typing using pulsed-field gel electrophoresis (PFGE), which showed that all ten cluster strains belonged to one type or its subtype.

Ceroid/lipofuscin-loaded human fibroblasts show decreased survival time and diminished autophagocytosis during amino acid starvation.

To test whether heavy accumulation of ceroid/lipofuscin can disturb important functions of the lysosomal system, AG-1518 human fibroblasts, ceroid/lipofuscin-loaded (following prolonged culture at normobaric hyperoxia) or not, were exposed to amino acid starvation. Ceroid/lipofuscin-loading resulted in decreased cellular survival. Also, there was an inverse relationship between amounts of ceroid/lipofuscin and the survival time of individual cells within the same cultures. Ceroid/lipofuscin-loaded fibroblasts displayed diminished autophagocytotic capacity, as demonstrated by electron microscopy and by treatment of cell cultures with NH4Cl (which inhibits autophagocytotic degradation by increasing intralysosomal pH) for 1 week before ensuing starvation. The latter treatment increased survival of control cells (due to deposition of nondegraded autophagocytosed material before start of starvation), but not that of ceroid/lipofuscin-loaded cells. Moreover, when NH4Cl treatment was combined with starvation, both groups of cells showed approximately the same shortened survival times, testifying to the causal relationship between diminished autophagocytosis and decreased survival of starving ceroid/lipofuscin-loaded cells. We hypothesize that large amounts of undegradable ceroid/lipofuscin within the acidic vacuolar compartment may interfere with lysosomal function, resulting in poor renewal of long-lived proteins and worn-out/damaged organelles, decreased adaptability, and cell death.

Beta-cells, oxidative stress, lysosomal stability, and apoptotic/necrotic cell death.

Reactive oxygen intermediates (ROI) may be involved in the destruction of pancreatic beta-cells during the development of insulin-dependent diabetes mellitus (IDDM). To investigate the possible role of lysosomes in this process, normal mouse beta-cells were cultured as monolayers at D-glucose concentrations of 1.6 (pronounced crinophagy), 11 or 28 mM (minimal crinophagy), subjected to a low level of oxidative stress and returned to standard culture conditions. Some cultures were exposed to desferrioxamine (Des) before the oxidative stress. As a result of such stress, many of the cells' lysosomes ruptured with consequent apoptosis or necrosis. Cells kept at 1.6 mM glucose were rich in secretory granules, showed crinophagy/autophagy, were very sensitive to oxidative stress, and had the least stable lysosomes. Cells kept at 28 mM glucose did not show crinophagy, contained fewer secretory granules, were less sensitive to oxidative stress, and had more stable lysosomes. Des-treated cells behaved almost as cells not exposed to oxidative stress at all. The findings suggest that iron may occur together with zinc within the secretory granules and that it sensitizes crinophagic lysosomes to oxidative stress. The stress that was applied in this study may be comparable to what occurs within the vicinity of activated macrophages during autoimmune insulitis.

Effect of collagenase on surface expression of immunoreactive fibronectin and laminin in freshly isolated cardiac myocytes.

The extracellular glycoproteins fibronectin (FN) and laminin (LMN) are ubiquitously expressed in myocardial tissue. These glycoproteins are important for cellular attachment and differentiation of the cardiac myocytes. Utilizing specific antibodies for the detection of FN and LMN, respectively, the distribution of these extracellular proteins was examined in enzymatically isolated adult cardiac myocytes. Immunofluorescence staining of rod-shaped cardiac myocytes revealed only remnants of immunoreactive FN on the cellular surface and in the transverse tubular membrane system. LMN expression, however, was preserved in a raster-like pattern in the cardiac myocytes. In order to study the distribution of these glycoproteins at high resolution, scanning electron microscopy using the backscattered electron mode was combined with immunogold staining and silver-enhancement. In addition, to confirm the immunofluorescence microscopic observations it was shown that FN labelling was restricted to ill-defined extracellular material and that LMN was absent from the intercalated discs of the cardiac myocytes. The hypercontracted cells were characterized by numerous surface protrusions devoid of immunoreactive LMN. Thus, these results indicate that FN and LMN are differently affected by collagenase treatment, and that these changes of glycoprotein expression may influence the normal function of the cardiac myocytes as well as the membrane stability during the development of irreversible cellular lesions.

[Methicillin resistant yellow staphylococci]. / Meticillinresistente gule stafylokokker.

The incidence of methicillin-resistant Staphylococcus aureus in Norway is extremely low. Isolation of such strains is nearly always associated with import. From December 1993 to January 1997 at the Ullevål University Hospital Department of Medical Microbiology, methicillin-resistant Staphylococcus aureus was isolated from 22 persons in Oslo (17 patients and five healthy carriers). A cluster of ten infected persons was detected (five patients and five carriers (nurses)) who were infected with strains showing an unusual antibiotic resistance pattern. All of the cluster strains except for beta-lactams were resistant to fucidic acid and sensitive to other antistaphylococcal agents. The cluster was associated with two hospitals. The five patients were all admitted to the same intensive care unit during the period May to July 1995. Four of the five patients (one died) were referred to the same department in a long-term care hospital for rehabilitation and training. Problems concerning epidemiological investigation and control are discussed.

Functional and histological differences in autogenous and allogenic vein grafts: two different vasculopathies?

The long-term biological characteristics and the functional and morphological changes that occur in fresh allografts are poorly understood. This study tests the hypothesis that the development of intimal hyperplasia and its associated functional changes are accelerated in an allograft compared to an autograft due to the additional immunological stimuli. Common carotid vein bypass grafts were performed in 40 New Zealand White rabbits: 20 received their ipsilateral jugular veins (autologous) and 20 received the fresh contralateral jugular veins from the control rabbit (allogenic). Electron microscopy was performed and intimal and medial dimensions were determined by videoplanimetry at 7, 14, and 28 days. Contraction and relaxation studies to a panel of agonists were also performed. The EC50's (agonist concentration which produces 50% of the maximal response) were calculated. All grafts remained patent. Allografts showed a 51% decrease in overall mean intimal thickness (41 +/- 3 microns vs. 83 +/- 12 microns; P < 0.01) and a 97% increase in overall mean medial thickness (140 +/- 15 microns vs. 71 +/- 3 microns; P < 0.01) compared to the autografts. The lumen of the allogenic vein grafts was equivalent to the autologous vein grafts. Overall mean total wall thickness only increased by 17%, 181 microns vs. 154 microns for allo- and autografts, respectively. The EC50 for norepinephrine, histamine, and bradykinin were similar in the auto- and allografts, while the EC50 to serotonin was significantly less in the allografts than in the autografts. Neither the precontracted auto- or allografts relaxed to acetylcholine or serotonin (receptor mediated, endothelium dependent). The EC50 for calcium ionophore (nonreceptor mediated, endothelium dependent) was equivalent in the auto- and allografts. The EC50 for the sodium nitroprusside-induced relaxation (endothelium independent) was significantly higher in the allograft than in the autograft. This study demonstrates that there are two different vasculopathies occurring in autografts and allografts: intimal hyperplasia is predominant in the autograft while an exaggerated medial response is predominant in the allograft. Serotonin contractility and endothelial-independent relaxation are enhanced in the allograft compared to the autograft.

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.

The morphology of venovenous bypass graft endothelium.

Venovenous bypass grafts are commonly used in the repair of vascular trauma to large- and small-caliber veins. This study examines the morphology of the venous wall in an experimental model of the venovenous bypass graft. The morphology of the venous endothelium from unmanipulated jugular veins and from jugular veins implanted as a venovenous bypass graft in the external jugular venous system for 10 min, 6 h, and 1, 3, 5, 7 and 28 days was examined. Veins and venovenous grafts were pressure fixed in situ at 80 mmHg and were examined by scanning and transmission electron microscopy. The endothelial cell lining remained confluent and intact over the 28-day period with evidence of endothelial cell contraction (spindle-shaped cells) for the first 72 h. Pinocytotic activity in endothelial cells and underlying smooth muscle cells was observed throughout the study, strongly indicating physiologically active cells. There was some accumulation of blood cells, predominantly polymorphonuclear leukocytes on the endothelial surface. Polymorphonuclear leukocytes were observed to infiltrate into the subendothelium through endothelial cell junctions within 6 h but by day 3, none was noted in the subendothelial space. There was no major disruption of the graft wall at any time point. By day 28, there was evidence of intimal thickening in the venovenous bypass grafts but no well-demarcated intimal hyperplasia. This study shows that there is no significant endothelial injury in the venovenous bypass grafts and that the endothelial cells remain physiologically active. Short-term failure of venovenous bypass grafts, therefore, appears not be due to significant endothelial cell damage in the graft.