Brigitta C. Brott

A Nitric Oxide Releasing, Self Assembled Peptide Amphiphile Matrix that Mimics Native Endothelium for Coating Implantable Cardiovascular Devices

Cardiovascular disease is the number one cause of death in the United States. Deployment of stents and vascular grafts has been a major therapeutic method for treatment. However, restenosis, incomplete endothelialization, and thrombosis hamper the long term clinical success. As a solution to meet these current challenges, we have developed a native endothelial ECM mimicking self-assembled nanofibrous matrix to serve as a new treatment model. The nanofibrous matrix is formed by self-assembly of peptide amphiphiles (PAs), which contain nitric oxide (NO) donating residues, endothelial cell adhesive ligands composed of YIGSR peptide sequence, and enzyme-mediated degradable sites. NO was successfully released from the nanofibrous matrix rapidly within 48 h, followed by sustained release over period of 30 days. The NO releasing nanofibrous matrix demonstrated a significantly enhanced proliferation of endothelial cells (51+/-3% to 67+/-2%) but reduced proliferation of smooth muscle cells (35+/-2% to 16+/-3%) after 48 h of incubation. There was also a 150-fold decrease in platelet attachment on the NO releasing nanofibrous matrix (470+/-220 platelets/cm(2)) compared to the collagen-I (73+/-22 x 10(3)platelets/cm(2)) coated surface. The nanofibrous matrix has the potential to be applied to various cardiovascular implants as a self-assembled coating, thereby providing a native endothelial extracellular matrix (ECM) mimicking environment.

A hybrid biomimetic nanomatrix composed of electrospun polycaprolactone and bioactive peptide amphiphiles for cardiovascular implants

Current cardiovascular therapies are limited by the loss of endothelium, restenosis and thrombosis. The goal of this study was to develop a biomimetic hybrid nanomatrix that combined the unique properties of electrospun polycaprolactone (ePCL) nanofibers with self-assembled peptide amphiphiles (PAs). ePCL nanofibers have interconnected nanoporous structures, but are hampered by a lack of surface bioactivity to control cellular behavior. It has been hypothesized that PAs could self-assemble onto the surface of ePCL nanofibers and endow them with the characteristic properties of native endothelium. The PAs, which comprised hydrophobic alkyl tails attached to functional hydrophilic peptide sequences, contained enzyme-mediated degradable sites coupled to either endothelial cell-adhesive ligands (YIGSR) or polylysine (KKKKK) nitric oxide (NO) donors. Two different PAs (PA-YIGSR and PA-KKKKK) were successfully synthesized and mixed in a 90:10 (YK) ratio to obtain PA-YK. PA-YK was reacted with pure NO to develop PA-YK-NO, which was then self-assembled onto ePCL nanofibers to generate a hybrid nanomatrix, ePCL-PA-YK-NO. Uniform coating of self-assembled PA nanofibers on ePCL was confirmed by transmission electron microscopy. Successful NO release from ePCL-PA-YK-NO was observed. ePCL-YK and ePCL-PA-YK-NO showed significantly increased adhesion of human umbilical vein endothelial cells (HUVECs). ePCL-PA-YK-NO also showed significantly increased proliferation of HUVECs and reduced smooth muscle cell proliferation. ePCL-PA-YK-NO also displayed significantly reduced platelet adhesion compared with ePCL, ePCL-PA-YK and a collagen control. These results indicate that this hybrid nanomatrix has great potential application in cardiovascular implants.

Turbulent flow evaluation of the venous needle during hemodialysis.

Arteriovenous (AV) grafts and fistulas used for hemodialysis frequently develop intimal hyperplasia (IH) at the venous anastomosis of the graft, leading to flow-limiting stenosis, and ultimately to graft failure due to thrombosis. Although the high AV access blood flow has been implicated in the pathogenesis of graft stenosis, the potential role of needle turbulence during hemodialysis is relatively unexplored. High turbulent stresses from the needle jet that reach the venous anastomosis may contribute to endothelial denudation and vessel wall injury. This may trigger the molecular and cellular cascade involving platelet activation and IH, leading to eventual graft failure. In an in-vitro graft/needle model dye injection flow visualization was used for qualitative study of flow patterns, whereas laser Doppler velocimetry was used to compare the levels of turbulence at the venous anastomosis in the presence and absence of a venous needle jet. Considerably higher turbulence was observed downstream of the venous needle, in comparison to graft flow alone without the needle. While turbulent RMS remained around 0.1 m/s for the graft flow alone, turbulent RMS fluctuations downstream of the needle soared to 0.4-0.7 m/s at 2 cm from the tip of the needle and maintained values higher than 0.1 m/s up to 7-8 cm downstream. Turbulent intensities were 5-6 times greater in the presence of the needle, in comparison with graft flow alone. Since hemodialysis patients are exposed to needle turbulence for four hours three times a week, the role of post-venous needle turbulence may be important in the pathogenesis of AV graft complications. A better understanding of the role of needle turbulence in the mechanisms of AV graft failure may lead to improved design of AV grafts and venous needles associated with reduced turbulence, and to pharmacological interventions that attenuate IH and graft failure resulting from turbulence.

Hemodynamic Analysis of a Compliant Femoral Artery Bifurcation Model using a Fluid Structure Interaction Framework

The influence of wall motion on the hemodynamic characteristics of the human femoral bifurcation and its effects on the development of peripheral artery disease has not been previously investigated. This study aimed in investigating the hemodynamics of a compliant patient-specific femoral artery bifurcation model by a fluid structure interaction (FSI) scheme. The complex physiological geometry of the femoral artery bifurcation was reproduced from sequentially obtained transverse CT scan images. Velocity waveforms derived from phase contrast MR images were extracted and mapped to define boundary conditions. Equations governing blood flow and wall motion were solved using an FSI framework that utilizes commercial codes: FLUENT for computational fluid dynamics and ANSYS for computational structural dynamics. The results showed that wall compliance decreased flow velocities at the relatively high curvature geometries including common and superficial femoral artery (SFA), and it created strong recirculation in the profunda femoris artery close to the bifurcation. In the SFA region near the apex, time averaged wall shear stress (TAWSS) differences up to 25% between compliant and rigid models were observed. The compliant model also exhibited lower TAWSS and oscillatory shear at the superior section of the common femoral artery close to the bifurcation. The presence of wall motion, however, created minor differences in the general flow-field characteristics. We conclude that wall motion does not have significant influence on the global fluid dynamic characteristics of the femoral artery bifurcation. Longer arterial segments need to be simulated to see the effect of wall motion on tortuousity which was previously cited as an important factor in the development of atherosclerosis at the femoral artery.

Clinical Device-Related Article Surface characterization of explanted endovascular stents: Evidence of in vivo corrosion

Limited information exists regarding the in vivo stability of endovascular stents. Nine excised human vascular segments with implanted stents (n = 16) manufactured from stainless steel, nickel-titanium, tantalum, and cobalt-based alloys were analyzed. The stent/tissue components were separated using an established tissue dissolution protocol and control and explanted stents were evaluated by digital optical and electron microscopy. Metallic content in surrounding tissues was measured by mass spectroscopy. Surface alterations, consistent with corrosion mediated by electrochemical and mechanical factors, were observed in 9 of the 16 explanted stents and were absent from control stents. Tissue dissolved from around corroded stents corresponded with a higher metallic content. The effect of these changes in the microtopography of stents on their mechanical properties (fatigue strength and fracture limit) in addition to the potential for released metallic debris contributing to the biological mechanisms of in-stent restenosis supports the need for further investigations.

Stainless Steel Ions Stimulate Increased Thrombospondin-1-Dependent TGF-Beta Activation by Vascular Smooth Muscle Cells: Implications for In-Stent Restenosis

Background/Aims: Despite advances in stent design, in-stent restenosis (ISR) remains a significant clinical problem. All implant metals exhibit corrosion, which results in release of metal ions. Stainless steel (SS), a metal alloy widely used in stents, releases ions to the vessel wall and induces reactive oxygen species, inflammation and fibroproliferative responses. The molecular mechanisms are unknown. TGF-β is known to be involved in the fibroproliferative responses of vascular smooth muscle cells (VSMCs) in restenosis, and TGF-β antagonists attenuate ISR. We hypothesized that SS ions induce the latent TGF-β activator, thrombospondin-1 (TSP1), through altered oxidative signaling to stimulate increased TGF-β activation and VSMC phenotype change. Methods: VSMCs were treated with SS metal ion cocktails, and morphology, TSP1, extracellular matrix production, desmin and TGF-β activity were assessed by immunoblotting. Results: SS ions stimulate the synthetic phenotype, increased TGF-β activity, TSP1, increased extracellular matrix and downregulation of desmin in VSMCs. Furthermore, SS ions increase hydrogen peroxide and decrease cGMP-dependent protein kinase (PKG) signaling, a known repressor of TSP1 transcription. Catalase blocks SS ion attenuation of PKG signaling and increased TSP1 expression. Conclusions: These data suggest that ions from stent alloy corrosion contribute to ISR through stimulation of TSP1-dependent TGF-β activation.

Percent infarct mapping: An R1‐map‐based CE‐MRI method for determining myocardial viability distribution

Viability detection is crucial for the management of myocardial infarction (MI). Signal intensity (SI)‐based MRI methods may overestimate infarct size in vivo. In contrast to SI, the longitudinal relaxation‐rate enhancement (ΔR1) is an intrinsic parameter that is linearly proportional to the concentration of contrast agent (CA). Determining ΔR1 in the presence of an infarct‐avid persistent CA (PCA) allows determination of the per‐voxel percentage of infarcted tissue. Introduced here is a ΔR1‐based CE‐MRI method, termed percent infarct mapping (PIM), for quantifying myocardial viability following delayed PCA accumulation. In a canine MI model (N = 6), PIMs were generated using a persistent CA (PCA) and validated using triphenyltetrazolium‐chloride (TTC) histochemistry. Voxel‐by‐voxel R1 maps of the entire left ventricle (LV) were generated 24 and 48 hr after PCA administration using inversion recovery (IR) with multiple inversion times (TIs). PI values were calculated voxel by voxel. Significant correlations (P < 0.01, R = 0.97) were obtained for PI per slice (PIS) determined using PIM vs. corresponding TTC‐based values. Median deviations of PIS with PIM from that with TTC were only 1.01% and –0.53%, at 24 hr and 48 hr. Median deviations from the true infarction fraction (IF) were 1.23% and 0.49% of LV at 24 hr and 48 hr, respectively. No significant difference was found between PIM24 hr and PIM48 hr. ΔR1‐based PIM is an accurate and reproducible method for quantifying myocardial viability distribution, and thus enhances the clinical utility of CE‐MRI. Magn Reson Med, 2006.

Stent overlapping and geometric curvature influence the structural integrity and surface characteristics of coronary nitinol stents.

Preliminary studies have revealed that some stents undergo corrosion and fatigue-induced fracture in vivo, with significant release of metallic ions into surrounding tissues. A direct link between corrosion and in-stent restenosis has not been clearly established; nonetheless in vitro studies have shown that relatively high concentrations of heavy metal ions can stimulate both inflammatory and fibrotic reactions, which are the main steps in the process of restenosis. To isolate the mechanical effects from the local biochemical effects, accelerated biomechanical testing was performed on single and overlapping Nickel-Titanium (NiTi) stents subjected to various degrees of curvature. Post testing, stents were evaluated using Scanning Electron Microscopy (SEM) to identify the type of surface alterations. Fretting wear was observed in overlapping cases, in both straight and curved configurations. Stent strut fractures occurred in the presence of geometric curvature. Fretting wear and fatigue fractures observed on stents following mechanical simulation were similar to those from previously reported human stent explants. It has been shown that biomechanical factors such as arterial curvature combined with stent overlapping enhance the incidence and degree of wear and fatigue fracture when compared to single stents in a straight tube configuration.

The role of vascular calcification in inducing fatigue and fracture of coronary stents.

Traditional approaches for in-vitro pulsatile and fatigue testing of endovascular stents do not take into consideration the pathologies of the stented vessel and their associated biomechanical effects. One important pathology is calcification, which may be capable of inducing changes in the vessel wall leading to inhomogeneous distribution of stresses combined with wall motion during the cardiac cycle. These local property changes in the region adjacent to stents could directly influence in-vivo stent performance. Seven cases containing a total of 18 stents were obtained from autopsy. Radiographs were evaluated and vessels were sectioned for histology and stent topographical analysis. Stents were retrieved by chemical removal of surrounding tissue and surfaces were evaluated using 3D digital optical and scanning electron microscopy for biomechanical abrasion and fracture features. Pathologic complications such as restenosis and thrombus formation were assessed from histological sections. Direct evidence of fracture was found in 6 of the 7 cases (in 12 out of 18 stents; 9 drug eluting and 3 bare metal). The degree of stent alterations was variable, where separation of segments due to fracture occurred mostly in drug-eluting stents. All fracture surfaces were representative of a high cycle fatigue mechanism. These fractures occurred in complex lesions involving the presence of diffuse calcification alone, or in combination with vessel angulations and multiple overlapping stents. Morphologic analysis of tissue at or near some fracture sites showed evidence of thrombus formation and/or neointimal tissue growth.

Effect of Posture Change on the Geometric Features of the Healthy Carotid Bifurcation

Segmented cross-sectional MRI images were used to construct 3-D virtual models of the carotid bifurcation in ten healthy volunteers. Geometric features, such as bifurcation angle, internal carotid artery (ICA) angle, planarity angle, asymmetry angle, tortuosity, curvature, bifurcation area ratio, ICA/common carotid artery (CCA), external carotid artery (ECA)/CCA, and ECA/ICA diameter ratios, were calculated for both carotids in two head postures: 1) the supine neutral position; and 2) the prone sleeping position with head rotation to the right (~80°). The results obtained have shown that head rotation causes 1) significant variations in bifurcation angle [32% mean increase for the right carotid (RC) and 21% mean decrease for the left carotid (LC)] and internal carotid artery angle (97% mean increase for the RC, 43% mean decrease for the LC); 2) a slight increase in planarity and asymmetry angles for both RC and LC; 3) minor and variable curvature changes for the CCA and for the branches; 4) slight tortuosity changes for the braches but not for the CCA; and 5) unsubstantial alterations in area and diameter ratios (percentage changes <;10%). The significant geometric changes observed in most subjects with head posture may also cause significant changes in bifurcation hemodynamics and warrant future investigation of the hemodynamic parameters related to the development of atherosclerotic disease such as low oscillating wall shear stress and particle residence times.