Swine Carotid Artery Research Articles

Basal release and relaxation responses to 6-nitrodopamine in swine carotid, coronary, femoral, and renal arteries

Mammalian and reptilian vascular tissues present basal release of 6-nitrodopamine, which is reduced when the tissues are pre-incubated with the NO synthase inhibitor L-NG-Nitro arginine methyl ester (L-NAME), or when the endothelium is mechanically removed. 6-Nitrodopamine induces vasorelaxation in pre-contracted vascular rings by antagonizing the dopaminergic D2-like receptor. Here it was investigated whether male swine vessels (including carotid, left descendent coronary, renal, and femoral arteries) release 6-nitrodopamine, dopamine, noradrenaline, and adrenaline, as measured by liquid chromatography coupled to tandem mass spectrometry. The in vitro vasorelaxant action of 6-nitrodopamine was evaluated in carotid, coronary, renal, and femoral arteries precontracted by U-46619 (3 nM), and compared to that induced by the dopamine D2-receptor antagonist L-741,626. Expression of tyrosine hydroxylase and the neuromaker calretinin was investigated by immunohistochemistry. All vascular tissues presented basal release of endothelium-derived catecholamines. The relaxation induced by 6-nitrodopamine was not affected by preincubation of the tissues with either L-NAME (100 μM, 30-min preincubation) or the heme-site inhibitor of soluble guanylyl cyclase ODQ (100 μM, 30-min preincubation). Electrical field stimulation (EFS)-induced contractions were significantly potentiated by previous incubation with L-NAME, but unaffected by ODQ preincubation. The contractions induced by EFS were reduced by preincubation with either 6-nitrodopamine or L-741,626. Immunohistochemistry in all arteries revealed the presence of tyrosine hydroxylase in the endothelium, whereas immunoreactivity for calretinin was negative. Swine vessels present basal release of endothelium-derived catecholamines and expression of tyrosine hydroxylase in the endothelium. The vasodilation induced by 6-nitrodopamine is due to blockade of dopaminergic D2-like receptors.

P11: Endoscopic Procurement of Saphenous Vein: a Hydraulic Conductivity and Microscopic Assessment

Background: Endoscopic saphenous vein harvest (ESVH) technique is an alternate and widely used conduit in coronary artery bypass grafting CABG in leu of the internal mammary artery (IMA). Unfortunately, multiple studies have shown low-patency rate in saphenous vein (SV) than IMA and prone to early atherosclerotic changes leading to obstruction. Thus, our goal was to explore the effects of physical distention on the endothelial integrity of SV by using hydraulic conductivity (HC) and scanning electron microscopy (SEM). Method: Saphenous vein conduits were collected intra-operatively from consented patients and procured at 37 °C. Vessel wall filtration rate was determined over 30-minute period at 100 mm Hg pressure and compared with baseline and negative control by using dialysis tubing, and isolated fresh swine carotid arteries. Conduits were stored in saline after harvest. Longitudinal sections of IMAs were fixed and dehydrated in various ethanol concentrations in deionized water for 15-minutes. Samples were then analyzed under SEM imaging. Result: Hydraulic conductivity of ESHV is tenfold higher than the pig carotid and dialysis tubing. Furthermore, SEM analyses of ESHV showed intimal injuries on the luminal surface. Conclusion: ESHV technique may increase the risk of traumatic handling by creating physical gaps in endothelial barrier. Alteration in vascular integrity may impact on permeability of larger molecules enhancing pathologic occlusive state and graft failure.

All-Optical Rotational Ultrasound Imaging

Miniaturised high-resolution imaging devices are valuable for guiding minimally invasive procedures such as vascular stent placements. Here, we present all-optical rotational B-mode pulse-echo ultrasound imaging. With this device, ultrasound transmission and reception are performed with light. The all-optical transducer in the probe comprised an optical fibre that delivered pulsed excitation light to an optical head at the distal end with a multi-walled carbon nanotube and polydimethylsiloxane composite coating. This coating was photoacoustically excited to generate a highly directional ultrasound beam perpendicular to the optical fibre axis. A concave Fabry-Pérot cavity at the distal end of an optical fibre, which was interrogated with a tuneable continuous-wave laser, served as an omnidirectional ultrasound receiver. The transmitted ultrasound had a −6 dB bandwidth of 31.3 MHz and a peak-to-peak pressure of 1.87 MPa, as measured at 1.5 mm from the probe. The receiver had a noise equivalent pressure <100 Pa over a 20 MHz bandwidth. With a maximum outer probe diameter of 1.25 mm, the probe provided imaging with an axial resolution better than 50 µm, and a real-time imaging rate of 5 frames per second. To investigate the capabilities of the probe, intraluminal imaging was performed in healthy swine carotid arteries. The results demonstrate that the all-optical probe is viable for clinical rotational ultrasound imaging.

Safety of carotid artery stent in repetitive transcranial magnetic stimulation-The histopathological proof from swine carotid artery.

Repetitive transcranial magnetic stimulation (rTMS), including theta burst stimulation (TBS), is considered helpful for functional recovery in post-stroke patients. However, the safety is a common concern forusingr TMS for neuro-rehabilitation and research in patients with stents. Prolonged continuous theta burst stimulation (cTBS) with 1200 pulses at 50% of maximum output of Magstim Super Rapid2 was delivered in three different angles to a carotid stent placed in an isolated fresh swine carotid artery or on a table at a distance of 5cm. The possible migration and temperature change of the stent caused by cTBS was monitored by video recording and a digital thermometer, respectively. Histopathological study with hematoxylin and eosin stain were done on the vessel wall to identify possible micro thermal injury. Stents in vessels did not cause any significant morphology change, such as thermal damage, after cTBS was given at three different angles. Neither visible migration nor significant temperature elevation was induced by cTBS. There was no temperature change, thermal injury or migration after prolonged TBS at a high intensity, suggesting TBS is safe for clinical neuro-rehabilitation and physiological assessments in stroke patients with vascular stents.

Sustained Contraction in Vascular Smooth Muscle by Activation of L-type Ca2+ Channels Does Not Involve Ca2+ Sensitization or Caldesmon.

Vascular smooth muscle (VSM) is unique in its ability to maintain an intrinsic level of contractile force, known as tone. Vascular tone is believed to arise from the constitutive activity of membrane-bound L-type Ca2+ channels (LTCC). This study used a pharmacological agonist of LTCC, Bay K8644, to elicit a sustained, sub-maximal contraction in VSM that mimics tone. Downstream signaling was investigated in order to determine what molecules are responsible for tone. Medial strips of swine carotid artery were stimulated with 100 nM Bay K8644 to induce a sustained level of force. Force and phosphorylation levels of myosin light chain (MLC), MAP kinase, MYPT1, CPI-17, and caldesmon were measured during Bay K8644 stimulation in the presence and absence of nifedipine, ML-7, U0126, bisindolylmaleimide (Bis), and H-1152. Nifedipine and ML-7 inhibited force and MLC phosphorylation in response to Bay K8644. Inhibition of Rho kinase (H-1152) but not PKC (Bis) inhibited Bay K8644 induced force. U0126 significantly increased Bay K8644-dependent force with no effect on MLC phosphorylation. Neither CPI-17 nor caldesmon phosphorylation were increased during the maintenance of sustained force. Our results suggest that force due to the influx of calcium through LTCCs is partially MLC phosphorylation-dependent but does not involve PKC or caldesmon. Interestingly, inhibition of MLC kinase (MLCK) and PKC significantly increased MAP kinase phosphorylation suggesting that MLCK and PKC may directly or indirectly inhibit MAP kinase activity during prolonged contractions induced by Bay K8544.

Regulation of mitogen-activated protein kinase by protein kinase C and mitogen-activated protein kinase phosphatase-1 in vascular smooth muscle

Vascular smooth muscle contraction is primarily regulated by phosphorylation of myosin light chain. There are also modulatory pathways that control the final level of force development. We tested the hypothesis that protein kinase C (PKC) and mitogen-activated protein (MAP) kinase modulate vascular smooth muscle activity via effects on MAP kinase phosphatase-1 (MKP-1). Swine carotid arteries were mounted for isometric force recording and subjected to histamine stimulation in the presence and absence of inhibitors of PKC [bisindolylmaleimide-1 (Bis)], MAP kinase kinase (MEK) (U0126), and MKP-1 (sanguinarine) and flash frozen for measurement of MAP kinase, PKC-potentiated myosin phosphatase inhibitor 17 (CPI-17), and caldesmon phosphorylation levels. CPI-17 was phosphorylated in response to histamine and was inhibited in the presence of Bis. Caldesmon phosphorylation levels increased in response to histamine stimulation and were decreased in response to MEK inhibition but were not affected by the addition of Bis. Inhibition of PKC significantly increased p42 MAP kinase, but not p44 MAP kinase. Inhibition of MEK with U0126 inhibited both p42 and p44 MAP kinase activity. Inhibition of MKP-1 with sanguinarine blocked the Bis-dependent increase of MAP kinase activity. Sanguinarine alone increased MAP kinase activity due to its effects on MKP-1. Sanguinarine increased MKP-1 phosphorylation, which was inhibited by inhibition of MAP kinase. This suggests that MAP kinase has a negative feedback role in inhibiting MKP-1 activity. Therefore, PKC catalyzes MKP-1 phosphorylation, which is reversed by MAP kinase. Thus the fine tuning of vascular contraction is due to the concerted effort of PKC, MAP kinase, and MKP-1.

Microstructural quantification of collagen fiber orientations and its integration in constitutive modeling of the porcine carotid artery

BackgroundMechanical characteristics of vascular tissue may play a role in different arterial pathologies, which, amongst others, requires robust constitutive descriptions to capture the vessel wall’s anisotropic and non-linear properties.Specifically, the complex 3D network of collagen and its interaction with other structural elements has a dominating effect of arterial properties at higher stress levels.The aim of this study is to collect quantitative collagen organization as well as mechanical properties to facilitate structural constitutive models for the porcine carotid artery.This helps the understanding of the mechanics of swine carotid arteries, being a standard in clinical hypothesis testing, in endovascular preclinical trials for example. MethodPorcine common carotid arteries (n=10) were harvested and used to (i) characterize the collagen fiber organization with polarized light microscopy, and (ii) the biaxial mechanical properties by inflation testing.The collagen organization was quantified by the Bingham orientation density function (ODF), which in turn was integrated in a structural constitutive model of the vessel wall.A one-layered and thick-walled model was used to estimate mechanical constitutive parameters by least-square fitting the recorded in vitro inflation test results.Finally, uniaxial data published elsewhere were used to validate the mean collagen organization described by the Bingham ODF. ResultsThick collagen fibers, i.e.the most mechanically relevant structure, in the common carotid artery are dispersed around the circumferential direction.In addition, almost all samples showed two distinct families of collagen fibers at different elevation, but not azimuthal, angles.Collagen fiber organization could be accurately represented by the Bingham ODF (κ1,2,3=[13.5,0.0,25.2] and κ1,2,3=[14.7,0.0,26.6]; average error of about 5%), and their integration into a structural constitutive model captured the inflation characteristics of individual carotid artery samples.Specifically, only four mechanical parameters were required to reasonably (average error from 14% to 38%) cover the experimental data over a wide range of axial and circumferential stretches.However, it was critical to account for fibrilar links between thick collagen fibers.Finally, the mean Bingham ODF provide also good approximation to uniaxial experimental data. ConclusionsThe applied structural constitutive model, based on individually measured collagen orientation densities, was able to capture the biaxial properties of the common carotid artery. Since the model required coupling amongst thick collagen fibers, the collagen fiber orientations measured from polarized light microscopy, alone, seem to be insufficient structural information. Alternatively, a larger dispersion of collagen fiber orientations, that is likely to arise from analyzing larger wall sections, could have had a similar effect, i.e. could have avoided coupling amongst thick collagen fibers. Statement of SignificanceThe applied structural constitutive model, based on individually measured collagen orientation densities, was able to capture the biaxial and uniaxial properties of the common carotid artery. Since the model required coupling amongst thick collagen fibers, an effective orientation density that accounts for cross-links between the main collagen fibers has been porposed. The model provides a good approximation to the experimental data.

Abstract 500: Vitamin D Supplementation Attenuates ADAM-12-mediated Proliferation of Carotid Artery Smooth Muscle Cells of Hypercholesterolemic Swine

Introduction: Risk of hypertension, peripheral artery disease, myocardial infarction and development of human atherosclerosis has been linked to vitamin D deficiency. Atheromatous cytokines, including IL-6, TNF-α and epidermal growth factor receptor (EGFR) family growth factors are released at the site of atherosclerosis and boost the activity of proteolytic enzymes such as ADAMs (a disintegrin and metalloproteinases). ADAM-12 cleaves proHB-EGF (Heparin-binding EGF-like growth factor) activating EGFR, resulting in increased proliferation of smooth muscle cells (SMCs). The aim of this study was to examine the effect of vitamin D on IL-6 and TNF-α-induced ADAM-12, pEGFR expression, HB-EGF release and SMC proliferation. Methods: Micro-swine were fed with vitamin D-deficient high cholesterol diet, high cholesterol diet containing 900 IU of vitamin D, and high cholesterol diet containing 3000 IU of vitamin D for total of 12 months. After six months, serum cholesterol levels of 500-600 mg/dL were achieved in all the three groups. The protein expression of ADAM-12 &amp; pEGFR, and HB-EGF release, in presence or absence of IL-6, TNF-α and Calcitriol, in SMCs was quantified by western Blot. HB-EGF release was measured by ELISA. The proliferation was assayed by [3H]-Thymidine incorporation and cell counting method. Results: The protein expression of ADAM-12 &amp; pEGFR, HB-EGF release were significantly reduced in carotid artery SMCs isolated from Vitamin D-supplemented swine. IL-6 and TNF-α treatment increased the protein expression of ADAM-12 &amp; pEGFR and HB-EGF release in carotid artery SMCs. Proliferation capacity was higher in SMCs isolated from Vitamin D-deficient swine carotid artery, potentiated by IL-6 and TNF-α. Calcitriol inhibited the ADAM-12, pEGFR expression and HB-EGF released in SMCs of hypercholesterolemic swine. Calcitriol also inhibited the proliferation of carotid artery SMCs isolated from Vit D-deficient, D-sufficient and D-supplemented swine. Conclusion: Together, these results suggest that vitamin D deficiency enhances proliferation of SMCs, which is potentiated by atheromatous cytokines. Whereas, vitamin D supplementation regulates ADAM-12-mediated cleavage of proHB-EGF and activation of EGFR inhibiting SMC proliferation.

Evaluation of coronary adventitial vasa vasorum using 3D optical coherence tomography – Animal and human studies

ObjectivesThis study sought to evaluate adventitial vasa vasorum (VV) in vivo with novel imaging technique of optical coherence tomography (OCT). MethodsTo verify OCT methods for quantification of VV, we first studied 2 swine carotid arteries in a model of focal angiogenesis by autologous blood injection, and compared microchannel volume (MCV) by OCT and VV by m-CT, and counts of those. In OCT images, adventitial MC was identified as signal-voiding areas which were located within 1 mm from the lumen-intima border. After manually tracing microchannel areas and the boundaries of lumen-intima and media-adventitial in all slices, we reconstructed 3D images. Moreover, we performed with OCT imaging in 8 recipients referred for evaluation of cardiac allograft vasculopathy at 1 year after heart transplantation. MCV and plaque volume (PV) were assessed with 3D images in each 10-mm-segment. ResultsIn the animal study, among the 16 corresponding 1-mm-segments, there were significant correlations of count and volume between both the modalities (count r2 = 0.80, P < 0.01; volume r2 = 0.50, P < 0.01) and a good agreement with a systemic bias toward underestimation with m-CT. In the human study, there was a significant positive correlation between MCV and PV (segment number = 24, r2 = 0.63, P < 0.01). ConclusionOur results suggest that evaluation of MCV with 3D OCT imaging might be a novel method to estimate the amount of adventitial VV in vivo, and further has the potential to provide a pathophysiological insight into a role of the VV in allograft vasculopathy.

Novel findings in membrane depolarization‐induced force development in vascular smooth muscle (865.11)

A unique phenomenon of vascular smooth muscle (VSM) is its ability to maintain force for a prolonged period of time. The exact mechanism responsible is not known but studies suggest a critical role for L‐type Ca2+ channels (LTCC). The purpose of this study was to investigate the consequences of prolonged LTCC opening with an agonist, Bay K8644, to elucidate the intracellular mechanisms responsible for maintaining VSM tone. Smooth muscle tissue isolated from swine carotid arteries was mounted for isometric force recording. Tissues were stimulated with Bay K8644 alone or following pretreatment with one of the following inhibitors: nifedipine (LTCC), ML‐7 (MLC kinase), bisindolylmaleimide (PKC), U0126 (MAP kinase), or H‐1152 (Rho kinase). The following results were obtained: • Bay K8644 increased MLC phosphorylation, PKC activity, MAP kinase activity, and force development but partially inhibited ROCK activity; CPI‐17 phosphorylation was not changed.• Bay K8644‐dependent activation of PKC inhibited MAP kinase activity; the Bay K8644 partial inhibition of ROCK activity was reversed by inhibition of LTCC with nifedipine .• Inhibition of MAP kinase activity significantly increased MLC kinase activity and inhibition of MLC kinase activity greatly increased MAP kinase activity.These results indicate that the maintenance of force in VSM involves an intricate signaling network with multiple modulatory steps.

Slack Length Reduces the Contractile Phenotype of the Swine Carotid Artery

Contraction is the primary function of adult arterial smooth muscle. However, in response to vessel injury or inflammation, arterial smooth muscle is able to phenotypically modulate from the contractile state to several ‘synthetic' states characterized by proliferation, migration and/or increased cytokine secretion. We examined the effect of tissue length (L) on the phenotype of intact, isometrically held, initially contractile swine carotid artery tissues. Tissues were studied (1) without prolonged incubation at the optimal length for force generation (1.0 Lo, control), (2) with prolonged incubation for 17 h at 1.0 Lo, or (3) with prolonged incubation at slack length (0.6 Lo) for 16 h and then restoration to 1.0 Lo for 1 h. Prolonged incubation at 1.0 Lo minimally reduced the contractile force without substantially altering the mediators of contraction (crossbridge phosphorylation, shortening velocity or stimulated actin polymerization). Prolonged incubation of tissues at slack length (0.6 Lo), despite return of length to 1.0 Lo, substantially reduced contractile force, reduced crossbridge phosphorylation, nearly abolished crossbridge cycling (shortening velocity) and abolished stimulated actin polymerization. These data suggest that (1) slack length treatment significantly alters the contractile phenotype of arterial tissue, and (2) slack length treatment is a model to study acute phenotypic modulation of intact arterial smooth muscle.

Correlative evaluation of variable pressure-scanning electron microscopy to histology methods for vascular tissues

For pathological investigation, vascular tissues implanted with biomaterials or medical devices require microscopic histology and scanning electron microscopy (SEM) assessment to evaluate biocompatibility with the vascular luminal surface. However, these valuable specimens are often susceptible to SEM and histology tissue preparation artifacts. Correlative variable pressure-scanning electron microscopy (VP-SEM) and histology methods were developed and evaluated, using current SEM technology, allowing direct imaging of hydrated samples without lengthy tissue processing, labor, equipment, and their sequela artifacts. Unstained and silver-stained, fixed swine carotid arteries were assessed using a cooling stage and an environmental secondary electron detector. After VP-SEM imaging, arteries were processed for histology evaluation. Assessment of luminal endothelial surface SEM images and tissue histology of the underlying mural wall was made to determine if low vacuum, cryo-freezing, electron beam exposure, and/or staining affected tissue morphology. Conventional SEM requires lengthy tissue processing with dedicated equipment for solvent dehydration, critical point drying, and metallic sputter coating in order to image under high vacuum for secondary electron imaging. This process causes notable artifacts when samples are later processed for microscopic histology. In contrast, silver staining of swine carotid arteries, followed by aldehyde fixation, facilitated VP-SEM imaging and histology preparation with results that were comparable to conventional SEM and had acceptable tissue morphology. VP-SEM with a cooling stage and an environmental secondary electron detector allows efficient SEM imaging and correlative histological assessment of vascular tissue samples. This correlative method may be further developed and implemented for research applications with biomaterial and medical device implanted tissues.

Determination and Modeling of the Inelasticity Over the Length of the Porcine Carotid Artery

The study of the mechanical properties of swine carotids has clinical relevance because it is important for the appropriate design of intravascular devices in the animal trial phases. The inelastic properties of porcine carotid tissue were investigated. Experimental uniaxial cyclic tests were performed along the longitudinal and circumferential directions of vessels. The work focused on the determination, comparison, and constitutive modeling of the softening properties and residual stretch set of the swine carotid artery over long stretches and stress levels in both proximal and distal regions. It was observed that the residual strain depends on the maximum stretch in the previous load cycle. The strain was higher for distal than for proximal samples and for circumferential than for longitudinal samples. In addition, a pseudoelastic model was used to reproduce the residual stretch and softening behavior of the carotid artery. The model presented a good approximation of the experimental data. The results demonstrate that the final results in animal trial studies could be affected by the location studied along the length of the porcine carotid.

Multimodality image fusion to guide peripheral artery chronic total arterial occlusion recanalization in a swine carotid artery occlusion model: Unblinding the interventionalist

To demonstrate the feasibility of magnetic resonance imaging (MRI) to X-ray fluoroscopy (XRF) image fusion to guide peripheral artery chronic total occlusion (CTO) recanalization. Endovascular peripheral artery CTO revascularization is minimally invasive, but challenging, because the occlusion is poorly visualized under XRF. Devices may steer out of the artery, which can lead to severe perforation. Merging preacquired MRI of the CTO to the live XRF display may permit upfront use of aggressive devices and improve procedural outcomes. Swine carotid artery CTOs were created using a balloon injury model. Up to 8 weeks later, MRI of the carotid arteries was acquired and segmented to create three-dimensional surface models, which were then registered onto live XRF. CTO recanalization was performed using incrementally aggressive CTO devices (group A) or an upfront aggressive directed laser approach (group B). Procedural success was defined as luminal or subintimal device position without severe perforation. In this swine model, MRI to XRF fusion guidance resulted in a procedural success of 57% in group A and 100% in group B, which compared favorably to 33% using XRF alone. Fluoroscopy time was significantly less for group B (8.5 ± 2.6 min) compared to group A (48.7 ± 23.9 min), P < 0.01. Contrast dose used was similar between groups A and B. MRI to XRF fusion-guided peripheral artery CTO recanalization is feasible. Multimodality image fusion may permit upfront use of aggressive CTO devices with improved procedural outcomes compared to XRF-guided procedures.

Abstract 279: Vitamin D3 Attenuates ADAM-12--Mediated Shedding of EGFR in Carotid Artery Smooth Muscle Cells of Hypercholesterolemic Swine

Deficiency of Vitamin D is linked to an increased risk of hypertension, peripheral artery disease, and myocardial infarction and is a major risk factor for the development of human atherosclerosis. Atheromatous cytokines, including TNF-α, IL-6 and IFN-γ, and EGF receptor family growth factors are released at the site of atherosclerosis and act on proteolytic enzymes, MMPs (matrix metalloproteinases), ADAMs (a disintegrin and metalloproteinases), and ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs). ADAM-12 activates EGFR resulting in increased migration and proliferation of smooth muscle cells (SMCs). The aim of this study was to examine the effect of vitamin D on IL-6-induced ADAM-12 expression and SMC migration and proliferation. Micro-swine were fed with either vitamin D-deficient high cholesterol diet or high cholesterol diet containing 900 IU of vitamin D for 6 months. After six months when serum cholesterol levels ranged from 500-600 mg/dL, vitamin D-deficient group continued on the same deficient diet, whereas the other group received supplementation of vitamin D (1,000 IU/d) for 6 months. The mRNA expression of ADAM-12 and EGFR in whole carotid artery and in IL-6-treated SMCs was quantified by qPCR. The proliferation was assayed by CyQuant NF cell proliferation assay. The mRNA transcripts of ADAM-12 and EGFR were significantly increased in carotid arteries from Vitamin D-deficient than in vitamin D- supplemented swine. Treatment of SMCs with IL-6 also increased the mRNA transcripts of ADAM-12 and EGFR in vitamin D-deficient swine SMCs compared to vitamin D-supplemented swine SMCs. The cell proliferation was higher in SMCs isolated from Vitamin D-deficient swine carotid artery compared to vitamin D- supplemented swine carotid artery. Together, these results suggest that Vitamin D regulates ADAM-12-mediated activation of EGFR and vitamin D deficiency further enhances proliferation of SMCs, which is potentiated by atheromatous cytokines.

Experimental study and constitutive modelling of the passive mechanical properties of the porcine carotid artery and its relation to histological analysis: Implications in animal cardiovascular device trials

The present study focusses on the determination, comparison and constitutive modelling of the passive mechanical properties of the swine carotid artery over very long stretches in both proximal and distal regions. Special attention is paid to the histological and mechanical variations of these properties depending on the proximity to the heart. The results can have clinical relevance, especially in the research field of intravascular device design. Before the final clinical trials on humans, research in the vascular area is conducted on animal models, swine being the most common due to the similarities between the human and swine cardiovascular systems as well as the fact that the swine size is suitable for testing devices, in this case endovascular carotid systems. The design of devices usually involves numerical techniques, and an important feature is the appropriate modelling of the mechanical properties of the vessel. Fourteen carotid swine arteries were harvested just after sacrifice and cyclic uniaxial tension tests in longitudinal and circumferential directions were performed for distal and proximal samples. The stress–stretch curves obtained were fitted with a hyperelastic anisotropic model. Stress-free configuration states were also analyzed. Finally, human and swine samples were processed in a histological laboratory and images were used to quantify their microconstituents. The statistical analysis revealed significant differences between the mechanical behavior of proximal and distal locations in the circumferential but not in the longitudinal direction. Circumferential direction samples show clear differences both in residual stretches and tensile curves between the two locations, while the features of longitudinal specimens are independent of the axial position. The statistical analysis provides significant evidence of changes depending on the position of the sample, mainly in elastin and SMC quantification.

Influence of dispersion in myosin filament orientation and anisotropic filament contractions in smooth muscle

A new constitutive model for the biomechanical behaviour of smooth muscle tissue is proposed. The active muscle contraction is accomplished by the relative sliding between actin and myosin filaments, comprising contractile units in the smooth muscle cells. The orientation of the myosin filaments, and thereby the contractile units, are taken to exhibit a statistical dispersion around a preferred direction. The number of activated cross-bridges between the actin and myosin filaments governs the contractile force generated by the muscle and also the contraction speed. A strain-energy function is used to describe the mechanical behaviour of the smooth muscle tissue. Besides the active contractile apparatus, the mechanical model also incorporates a passive elastic part. The constitutive model was compared to histological and isometric tensile test results for smooth muscle tissue from swine carotid artery. In order to be able to predict the active stress at different muscle lengths, a filament dispersion significantly larger than the one observed experimentally was required. Furthermore, a comparison of the predicted active stress for a case of uniaxially oriented myosin filaments and a case of filaments with a dispersion based on the experimental histological data shows that the difference in generated stress is noticeable but limited. Thus, the results suggest that myosin filament dispersion alone cannot explain the increase in active muscle stress with increasing muscle stretch.

Análise morfométrica da carótida de suínos submetidos a angioplastia com ou sem implante de stent de cromo-cobalto

Contexto: A hiperplasia intimal é a reação tardia mais comum decorrente da angioplastia. O uso de stents de cromo-cobalto é bem estudado na circulação coronariana, porém não há muitos estudos que abordem o uso desses stents nas circulações carotídea e periférica. Objetivo: Analisar mediante morfometria a reação intimal presente na artéria carótida de suínos submetidos a angioplastia isoladamente e a angioplastia seguida de implante de stent de cromo-cobalto. Materiais e métodos: Em oito suínos, foi realizada angioplastia da artéria carótida comum direita e angioplastia seguida de implante de um stent de cromo-cobalto na artéria carótida comum esquerda. Após 4 semanas, os animais foram submetidos a eutanásia para a retirada de amostras de tecido arterial e preparo de lâminas histológicas. As imagens das lâminas foram digitalizadas e analisadas por programa de morfometria digital. A análise estatística foi realizada através da média e desvio padrão das áreas em cada grupo, utilizando-se o Teste t de Student. O valor de p &lt; 0,05 foi considerado significativo. Resultados: O implante do stent provocou maior grau de hiperplasia comparado à angioplastia isolada. A diferença em resposta ao implante de stent foi estatisticamente significativa quando as áreas do lúmen, da lâmina elástica interna e da lâmina elástica externa foram comparadas entre os dois grupos. Não se observou diferença significativa quando se realizou a comparação entre as camadas médias dos dois grupos. Conclusão: O implante de stent de cromo-cobalto gerou um espessamento intimal maior do que o produzido apenas pela angioplastia, porém ele não foi suficiente para reduzir o lúmen arterial.

Adeno‐associated Virus 9 Vector in the Gene Therapy of Occlusive Vascular Diseases

We analyzed the efficiency and duration of transfection of AAV9 vector containing SM22α promoter with either eGFP or LacZ as reporter genes. The vectors (1 × 1012 pfu) were incorporated into coronary and carotid arteries of swine using microporous balloon catheter‐based gene delivery approach. Segments from all vessels were excised at various time points and immediately frozen for mRNA/protein expression or were processed for frozen sections for X‐Gal, GFP or H&amp;E staining. GFP was visualized distinctly in the medial layer. Frozen sections were stained and β‐galactosidase gene transfer was assessed by blue‐stained cell nuclei. The endothelial and adventitial layers were carefully scraped off from the harvested vessel and total RNA was isolated from the remaining medial layer for qPCR. In the medial layer of coronary and femoral arteries LacZ mRNA expression was visualized 7 days after vector administration. The GFP mRNA expression was detected at 7 days and lasted for at least 2 months showing the prolonged expression of the AAV9‐vector. The GFP expression was confirmed by Western blot. Control arteries did not show any expression of GFP or LacZ. These findings support the use of AAV9 as a vector to effectively transduce a gene in coronary arteries.(Supported by LB692 Nebraska Tobacco settlement Funds and NHLBI‐sponsored Penn Vector Core, University of Pennsylvania)

Stimulated Actin Polymerization Induces Force Potentiation in Swine Carotid Artery

The phenomenon of post-tetanic potentiation, in which a single submaximal contraction or series of submaximal contractions strengthens a subsequent contraction, has been observed in both skeletal and cardiac muscle.