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It is increasingly appreciated that the properties of a biomaterial used in intramyocardial injection therapy influence the outcomes of infarcted hearts that are treated. In this report the extended in vivo efficacy of a thermally responsive material that can deliver dual growth factors while providing a slow degradation time and high mechanical stiffness is examined. Copolymers consisting of N-isopropyla-crylamide, 2-hydroxyethyl methacrylate, and degradable methacrylate polylactide were synthesized. The release of bioactive basic fibroblast growth factor (bFGF) and insulin-like growth factor 1 (IGF1) from the gel and loaded poly(lactide-co-glycolide) microparticles was assessed. Hydrogel with or without loaded growth factors was injected into 2 week-old infarcts in Lewis rats and animals were followed for 16 weeks. The hydrogel released bioactive bFGF and IGF1 as shown by mitogenic effects on rat smooth muscle cells in vitro. Cardiac function and geometry were improved for 16 weeks after hydrogel injection compared to saline injection. Despite demonstrating that left ventricular levels of bFGF and IGF1 were elevated for two weeks after injection of growth factor loaded gels, both functional and histological assessment showed no added benefit to inclusion of these proteins. This result points to the complexity of designing appropriate materials for this application and suggests that the nature of the material alone, without exogenous growth factors, has a direct ability to influence cardiac remodeling.

Intramyocardial hydrogel injection is an innovative and promising treatment for myocardial infarction (MI) and has recently entered clinical trials. By providing mechanical support to the ventricular wall, hydrogel injectate may act to preserve cardiac function and slow the remodeling process that leads to heart failure. However, improved outcomes will likely depend on the use of hydrogels specifically designed for this unique application, and better understanding of the mechanisms affected by the intervention. In this work, we present the first large animal study achieving functional and geometrical improvements in treating MI using a relatively stiff, fully synthetic hydrogel designed for intramyocardial injection. In addition, the renin-angiotensin system coincided with the mechanical effects of hydrogel injection and attenuated left ventricular remodeling, even after significant hydrogel degradation had occurred in vivo. These results may inspire further optimization of hydrogel materials used in intramyocardial hydrogel injection therapy and a better description of physiologic pathways affected by its implementation to facilitate successful clinical translation.

Tissue engineering has gained attention as an alternative approach for developing small diameter tissue-engineered vascular grafts intended for bypass surgery, as an option to treat coronary heart disease. To promote the formation of a healthy endothelial cell monolayer in the lumen of the graft, polycaprolactone/gelatin/fibrinogen scaffolds were developed, and the surface was modified using thermoforming and coating with collagen IV and fibronectin. Human cord blood-derived endothelial cells (hCB-ECs) were seeded onto the scaffolds and the important characteristics of a healthy endothelial cell layer were evaluated under static conditions using human umbilical vein endothelial cells as a control. We found that polycaprolactone/gelatin/fibrinogen scaffolds that were thermoformed and coated are the most suitable for endothelial cell growth. hCB-ECs can proliferate, produce endothelial nitric oxide synthase, respond to interleukin 1 beta, and reduce platelet deposition.

Current extracellular matrix (ECM) derived scaffolds offer promising regenerative responses in many settings, however in some applications there may be a desire for more robust and long lasting mechanical properties. A biohybrid composite material that offers both strength and bioactivity for optimal healing towards native tissue behavior may offer a solution to this problem. A regionally distinct biocomposite scaffold composed of a biodegradable elastomer (poly(ester urethane)urea) and porcine dermal ECM gel was generated to meet this need by a concurrent polymer electrospinning/ECM gel electrospraying technique where the electrosprayed component was varied temporally during the processing. A sandwich structure was achieved with polymer fiber rich upper and lower layers for structural support and an ECM-rich inner layer to encourage cell ingrowth. Increasing the upper and lower layer fiber content predictably increased tensile strength. In a rat full thickness abdominal wall defect model, the sandwich scaffold design maintained its thickness whereas control biohybrid scaffolds lacking the upper and lower fiber-rich regions failed at 8 weeks. Sandwich scaffold implants also showed higher collagen content 4 and 8 weeks after implantation, exhibited an increased M2 macrophage phenotype response at later times and developed biaxial mechanical properties better approximating native tissue. By employing a processing approach that creates a sheet-form scaffold with regionally distinct zones, it was possible to improve biological outcomes in body wall repair and provide the means for further tuning scaffold mechanical parameters when targeting other applications. Copyright © 2013 John Wiley & Sons, Ltd.

Objective Tissue-engineered vascular grafts containing adipose-derived mesenchymal stem cells offer an alternative to small-diameter vascular grafts currently used in cardiac and lower-extremity revascularization procedures. Adipose-derived, mesenchymal stem cell–infused, tissue-engineered vascular grafts have been shown to promote remodeling and vascular homeostasis in vivo and offer a possible treatment solution for those with cardiovascular disease. Unfortunately, the time needed to cultivate adipose-derived mesenchymal stem cells remains a large hurdle for tissue-engineered vascular grafts as a treatment option. The purpose of this study was to determine if stromal vascular fraction (known to contain progenitor cells) seeded tissue-engineered vascular grafts would remain patent in vivo and remodel, allowing for a “same-day” process for tissue-engineered vascular graft fabrication and implantation. Methods Stromal vascular fraction, obtained from adult human adipose tissue, was seeded within 4 hours after acquisition from the patient onto poly(ester urethane)urea bilayered scaffolds using a customized rotational vacuum seeding device. Constructs were then surgically implanted as abdominal aortic interposition grafts in Lewis rats. Results Findings revealed patency in 5 of 7 implanted scaffolds at 8 weeks, along with neotissue formation and remodeling occurring in patent tissue-engineered vascular grafts. Patency was documented using angiography and gross inspection, and remodeling and vascular components were detected using immunofluorescent chemistry. Conclusions A “same-day” cell-seeded, tissue-engineered vascular graft can remain patent after implantation in vivo, with neotissue formation and remodeling occurring by 8 weeks.

Individual ventricular assist device (VAD) design may affect leukocytes and impact immunity. Few studies have presented leukocyte and infection profiles in VAD patients over the course of the implant period. CD11b (MAC-1) expression on granulocytes is an indicator of activation during inflammation, mediating extravasation and the release of reactive oxygen species in tissue. No reported studies have presented MAC-1 expression on circulating granulocytes in VAD patients. Fifty-six patients implanted at a single center with a HeartMate II (HMII; n = 32), HeartWare (HW; n = 12), or Thoratec pneumatic VAD (PVAD; n = 12) between 1999 and 2011 were followed for 120 days of support. The leukocyte profiles and infectious events of all patients were evaluated; additionally, a subset had MAC-1 expression on circulating granulocytes was measured (HMII n = 9; HW n = 7; PVAD n = 4). All groups exhibited a significant peak in leukocyte numbers at postoperative day (POD) 14 while simultaneously experiencing a significant decrease in hematocrit. HMII patients exhibited a 3.2-fold increase in granulocyte MAC-1 expression at POD 14, and the temporal trend over the implant period differed from that experienced by HW patients. Further, HW patients experienced significantly fewer infection events. Alterations in leukocyte profiles and granulocyte activation experienced by VAD patients appear to be device-specific. Elevations in leukocyte activation may be related to an increased risk for infection, although the specific relationship between these phenomena in this patient group is not known.

Recapitulating the microstructure of the native human corneal stromal tissue is believed to be a key feature in successfully engineering the corneal tissue. The stratified multilayered collagen fibril lamellae with orthogonal orientation determine the robust biomechanical properties of this tissue, and the uniform collagen fibril size and interfibrillar spacing are critical to its optical transparency. The objective of this investigation was to develop a highly organized collagen-fibril construct secreted by human corneal stromal stem cells (hCSSCs) to mimic the human corneal stromal tissue. In culture on a highly aligned fibrous substrate made from poly(ester urethane) urea, the fibroblast growth factor-2 (FGF-2, 10 ng/mL) and transforming growth factor-beta 3 (TGF-β3, 0.1 ng/mL) impacted the organization and abundance of the secreted collagen fibril matrix. hCSSCs differentiated into keratocytes with significant upregulation of the typical gene markers, including KERA, B3GnT7, and CHST6. FGF-2 treatment stimulated hCSSCs to secrete collagen fibrils strongly aligned in a single direction, whereas TGF-β3 induced collagenous layers with orthogonal fibril orientation. The combination of FGF-2 and TGF-β3 induced multilayered lamellae with orthogonally oriented collagen fibrils, in a pattern mimicking the human corneal stromal tissue. The constructs were 60-70 μm thick and had an increased content of cornea-specific extracellular matrix components, including keratan sulfate, lumican, and keratocan. The approach of combining substrate cues with growth factor augmentation offers a new means to engineer well-organized, collagen-based constructs with an appropriate nanoscale structure for corneal repair and regeneration.

Extracorporeal CO2 removal from circulating blood is a promising therapeutic modality for the treatment of acute respiratory failure. The enzyme carbonic anhydrase accelerates CO2 removal within gas exchange devices by locally catalyzing HCO3 (-) into gaseous CO2 within the blood. In this work, we covalently immobilized carbonic anhydrase on the surface of polypropylene hollow fiber membranes using glutaraldehyde activated chitosan tethering to amplify the density of reactive amine functional groups for enzyme immobilization. XPS and a colorimetric amine assay confirmed higher amine densities on the chitosan coated fiber compared to control fiber. Chitosan/CA coated fibers exhibited accelerated CO2 removal in scaled-down gas exchange devices in buffer and blood (115% enhancement vs. control, 37% enhancement vs. control, respectively). Carbonic anhydrase immobilized directly on hollow fiber membranes without chitosan tethering resulted in no enhancement in CO2 removal. Additionally, fibers coated with chitosan/carbonic anhydrase demonstrated reduced platelet adhesion when exposed to blood compared to control and heparin coated fibers.

The current prevalence and severity of heart defects requiring functional replacement of cardiac tissue pose a serious clinical challenge. Biologic scaffolds are an attractive tissue engineering approach to cardiac repair because they avoid sensitization associated with homograft materials and theoretically possess the potential for growth in similar patterns as surrounding native tissue. Both urinary bladder matrix (UBM) and cardiac ECM (C-ECM) have been previously investigated as scaffolds for cardiac repair with modest success, but have not been compared directly. In other tissue locations, bone marrow derived cells have been shown to play a role in the remodeling process, but this has not been investigated for UBM in the cardiac location, and has never been studied for C-ECM. The objectives of the present study were to compare the effectiveness of an organ-specific C-ECM patch with a commonly used ECM scaffold for myocardial tissue repair of the right ventricle outflow tract (RVOT), and to examine the role of bone marrow derived cells in the remodeling response. A chimeric rat model in which all bone marrow cells express green fluorescent protein (GFP) was generated and used to show the ability of ECM scaffolds derived from the heart and bladder to support cardiac function and cellular growth in the RVOT. The results from this study suggest that urinary bladder matrix may provide a more appropriate substrate for myocardial repair than cardiac derived matrices, as shown by differences in the remodeling responses following implantation, as well as the presence of site appropriate cells and the formation of immature, myocardial tissue.

Donation after cardiac death has been adopted to address the critical shortage of donor organs for transplant. Recovery of these organs is hindered by low blood flow that leads to permanent organ injury. We propose a novel approach to isolate the perfusion of the abdominal organs from the systemic malperfusion of the dying donor. We reasoned that this design could improve blood flow to organs without open surgery, while respecting the ethical principle that cardiac stress not be increased during organ recovery. Conditions within the stent were analysed using a computational fluid dynamics (CFD) method and validated on two prototypes in vitro. The hydrodynamic pressure drop across the stent was measured as 0.14-0.22 mmHg, which is a negligible influence. Device placement studies were also conducted on swine model fluoroscopically. All these results demonstrated the feasibility of rapidly isolating the perfusion to abdominal organs using a compartmentalised stent graft design.

Many preclinical evaluations of autologous small-diameter tissue-engineered vascular grafts (TEVGs) utilize cells from healthy humans or animals. However, these models hold minimal relevance for clinical translation, as the main targeted demographic is patients at high cardiovascular risk such as individuals with diabetes mellitus or the elderly. Stem cells such as adipose-derived mesenchymal stem cells (AD-MSCs) represent a clinically ideal cell type for TEVGs, as these can be easily and plentifully harvested and offer regenerative potential. To understand whether AD-MSCs sourced from diabetic and elderly donors are as effective as those from young nondiabetics (i.e., healthy) in the context of TEVG therapy, we implanted TEVGs constructed with human AD-MSCs from each donor type as an aortic interposition graft in a rat model. The key failure mechanism observed was thrombosis, and this was most prevalent in grafts using cells from diabetic patients. The remainder of the TEVGs was able to generate robust vascular-like tissue consisting of smooth muscle cells, endothelial cells, collagen, and elastin. We further investigated a potential mechanism for the thrombotic failure of AD-MSCs from diabetic donors; we found that these cells have a diminished potential to promote fibrinolysis compared to those from healthy donors. Together, this study served as proof of concept for the development of a TEVG based on human AD-MSCs, illustrated the importance of testing cells from realistic patient populations, and highlighted one possible mechanistic explanation as to the observed thrombotic failure of our diabetic AD-MSC-based TEVGs.

Objectives The PediaFlow (HeartWare International, Inc, Framingham, Mass) is a miniature, implantable, rotodynamic, fully magnetically levitated, continuous-flow pediatric ventricular assist device. The fourth-generation PediaFlow was evaluated in vitro and in vivo to characterize performance and biocompatibility. Methods Supported by 2 National Heart, Lung, and Blood Institute contract initiatives to address the limited options available for pediatric patients with congenital or acquired cardiac disease, the PediaFlow was developed with the intent to provide chronic cardiac support for infants as small as 3 kg. The University of Pittsburgh–led Consortium evaluated fourth-generation PediaFlow prototypes both in vitro and within a preclinical ovine model (n = 11). The latter experiments led to multiple redesigns of the inflow cannula and outflow graft, resulting in the implantable design represented in the most recent implants (n = 2). Results With more than a decade of extensive computational and experimental efforts spanning 4 device iterations, the AA battery–sized fourth-generation PediaFlow has an operating range of 0.5 to 1.5 L/min with minimal hemolysis in vitro and excellent hemocompatibility (eg, minimal hemolysis and platelet activation) in vivo. The pump and finalized accompanying implantable components demonstrated preclinical hemodynamics suitable for the intended pediatric application for up to 60 days. Conclusions Designated a Humanitarian Use Device for “mechanical circulatory support in neonates, infants, and toddlers weighing up to 20 kg as a bridge to transplant, a bridge to other therapeutic intervention such as surgery, or as a bridge to recovery” by the Food and Drug Administration, these initial results document the biocompatibility and potential of the fourth-generation PediaFlow design to provide chronic pediatric cardiac support.

Objective One of the rate-limiting barriers within the field of vascular tissue engineering is the lengthy fabrication time associated with expanding appropriate cell types in culture. One particularly attractive cell type for this purpose is the adipose-derived mesenchymal stem cell (AD-MSC), which is abundant and easily harvested from liposuction procedures. Even this cell type has its drawbacks, however, including the required culture period for expansion, which could pose risks of cellular transformation or contamination. Eliminating culture entirely would be ideal to avoid these concerns. In this study, we used the raw population of cells obtained after digestion of human liposuction aspirates, known as the stromal vascular fraction (SVF), as an abundant, culture-free cell source for tissue-engineered vascular grafts (TEVGs). Methods SVF cells and donor-paired cultured AD-MSCs were first assessed for their abilities to differentiate into vascular smooth muscle cells (SMCs) after angiotensin II stimulation and to secrete factors (eg, conditioned media) that promote SMC migration. Next, both cell types were incorporated into TEVG scaffolds, implanted as an aortic graft in a Lewis rat model, and assessed for their patency and composition. Results In general, the human SVF cells were able to perform the same functions as AD-MSCs isolated from the same donor by culture expansion. Specifically, cells within the SVF performed two important functions; namely, they were able to differentiate into SMCs (SVF calponin expression: 16.4% ± 7.7% vs AD-MSC: 19.9%% ± 1.7%) and could secrete promigratory factors (SVF migration rate relative to control: 3.1 ± 0.3 vs AD-MSC: 2.5 ± 0.5). The SVF cells were also capable of being seeded within biodegradable, elastomeric, porous scaffolds that, when implanted in vivo for 8 weeks, generated patent TEVGs (SVF: 83% patency vs AD-MSC: 100% patency) populated with primary vascular components (eg, SMCs, endothelial cells, collagen, and elastin). Conclusions Human adipose tissue can be used as a culture-free cell source to create TEVGs, laying the groundwork for the rapid production of cell-seeded grafts.

As an intervention to abrogate ischemic cardiomyopathy, the concept of applying a temporary, local patch to the surface of the recently infarcted ventricle has been explored from a number of design perspectives. Two important features considered for such a cardiac patch include the provision of appropriate mechanical support and the capacity to influence the remodeling pathway by providing cellular or biomolecule delivery. The objective of this report was to focus on these two features by first evaluating the incorporation of a cardiac extracellular matrix (ECM) component, and second by evaluating the impact of patch anisotropy on the pathological remodeling process initiated by myocardial infarction. The functional outcomes of microfibrous, elastomeric, biodegradable cardiac patches have been evaluated in a rat chronic infarction model. Ten weeks after infarction and 8 wk after patch epicardial placement, echocardiographic function, tissue-level structural remodeling (e.g., biaxial mechanical response and microstructural analysis), and cellular level remodeling were assessed. The results showed that the incorporation of a cardiac ECM altered the progression of several keys aspects of maladaptive remodeling following myocardial infarction. This included decreasing LV global mechanical compliance, inhibiting echocardiographically-measured functional deterioration, mitigating scar formation and LV wall thinning, and promoting angiogenesis. In evaluating the impact of patch anisotropy, no effects from the altered patch mechanics were detected after 8 wk, possibly due to patch fibrous encapsulation. Overall, this study demonstrates the benefit of a cardiac patch design that combines both ventricle mechanical support, through a biodegradable, fibrillary elastomeric component, and the incorporation of ECM-based hydrogel components.

Mechanical conditioning of engineered tissue constructs is widely recognized as one of the most relevant methods to enhance tissue accretion and microstructure, leading to improved mechanical behaviors. The understanding of the underlying mechanisms remains rather limited, restricting the development of in silico models of these phenomena, and the translation of engineered tissues into clinical application. In the present study, we examined the role of large strip-biaxial strains (up to 50%) on ECM synthesis by vascular smooth muscle cells (VSMCs) micro-integrated into electrospun polyester urethane urea (PEUU) constructs over the course of 3 weeks. Experimental results indicated that VSMC biosynthetic behavior was quite sensitive to tissue strain maximum level, and that collagen was the primary ECM component synthesized. Moreover, we found that while a 30% peak strain level achieved maximum ECM synthesis rate, further increases in strain level lead to a reduction in ECM biosynthesis. Subsequent mechanical analysis of the formed collagen fiber network was performed by removing the scaffold mechanical responses using a strain-energy based approach, showing that the de-novo collagen also demonstrated mechanical behaviors substantially better than previously obtained with small strain training and comparable to mature collagenous tissues. We conclude that the application of large deformations can play a critical role not only in the quantity of ECM synthesis (i.e. the rate of mass production), but also on the modulation of the stiffness of the newly formed ECM constituents. The improved understanding of the process of growth and development of ECM in these mechano-sensitive cell-scaffold systems will lead to more rational design and manufacturing of engineered tissues operating under highly demanding mechanical environments.

Current artificial lungs and respiratory assist devices designed for carbon dioxide removal (CO(2)R) are limited in their efficiency due to the relatively small partial pressure difference across gas exchange membranes. To offset this underlying diffusional challenge, bioactive hollow fiber membranes (HFMs) increase the carbon dioxide diffusional gradient through the immobilized enzyme carbonic anhydrase (CA), which converts bicarbonate to CO(2) directly at the HFM surface. In this study, we tested the impact of CA-immobilization on HFM CO(2) removal efficiency and thromboresistance in blood. Fiber surface modification with radio frequency glow discharge (RFGD) introduced hydroxyl groups, which were activated by 1M CNBr while 1.5M TEA was added drop wise over the activation time course, then incubation with a CA solution covalently linked the enzyme to the surface. The bioactive HFMs were then potted in a model gas exchange device (0.0084 m(2)) and tested in a recirculation loop with a CO(2) inlet of 50mmHg under steady blood flow. Using an esterase activity assay, CNBr chemistry with TEA resulted in 0.99U of enzyme activity, a 3.3 fold increase in immobilized CA activity compared to our previous method. These bioactive HFMs demonstrated 108 ml/min/m(2) CO(2) removal rate, marking a 36% increase compared to unmodified HFMs (p < 0.001). Thromboresistance of CA-modified HFMs was assessed in terms of adherent platelets on surfaces by using lactate dehydrogenase (LDH) assay as well as scanning electron microscopy (SEM) analysis. Results indicated HFMs with CA modification had 95% less platelet deposition compared to unmodified HFM (p < 0.01). Overall these findings revealed increased CO(2) removal can be realized through bioactive HFMs, enabling a next generation of more efficient CO(2) removal intravascular and paracorporeal respiratory assist devices.

A biodegradable elastomeric scaffold was created by electrospinning a mixed solution of poly(ester urethane)urea (PEUU) and porcine dermal extracellular matrix (dECM) digest, with PEUU included to provide elasticity, flexibility, and mechanical support and dECM used to enhance bioactivity and biocompatibility. Micrographs and differential scanning calorimetry demonstrated partial miscibility between PEUU and dECM. With greater dECM content, scaffolds were found to possess lower breaking strains and suture retention strength, although initial modulus was greater with higher dECM concentrations. The hybrid scaffolds containing 0% to 50% dECM had tensile strengths of 5 to 7 MPa, breaking strains of 138% to 611%, initial moduli of 3 to 11 Mpa, and suture retention strengths of 35 to 59 MPa. When hydrated, scaffolds were found to contract markedly with 50% dECM content. When used in a rat full-thickness abdominal wall replacement model, no herniation, infection, or tissue adhesion was observed after 4 and 8 weeks with a scaffold containing 25% dECM or a control 100% PEUU scaffold. Scaffolds incorporating dECM were significantly thicker at the time of explant, with greater numbers of associated smooth muscle actin-positive staining cells than in the control, but minimal cellular infiltration and remodeling of the scaffold were detected regardless of dECM addition. The processing of dECM and PEUU from a mixed solution thus provided a scaffold with evidence of better bioactivity and with mechanical properties not achievable with digested dECM alone.

Corneal stroma is an avascular connective tissue characterized by layers of highly organized parallel collagen fibrils, mono-disperse in diameter with uniform local interfibrillar spacing. Reproducing this level of structure on a nano- and micro-scale may be essential to engineer corneal tissue with strength and transparency similar to that of native cornea. A substrate of aligned poly(ester urethane) urea (PEUU) fibers, 165 ± 55 nm in diameter, induced alignment of cultured human corneal stromal stem cells (hCSSCs) which elaborated a dense collagenous matrix, 8-10 μm in thickness, deposited on the PEUU substratum. This matrix contained collagen fibrils with uniform diameter and regular interfibrillar spacing, exhibiting global parallel alignment similar to that of native stroma. The cells expressed high levels of gene products unique to keratocytes. hCSSCs cultured on PEUU fibers of random orientation or on a cast film of PEUU also differentiated to keratocytes and produced abundant matrix, but lacked matrix organization. These results demonstrate the importance of topographic cues in instructing organization of the transparent connective tissue of the corneal stroma by differentiated keratocytes. This important information will help with design of biomaterials for a bottom-up strategy to bioengineer spatially complex, collagen-based nano-structured constructs for corneal repair and regeneration.

ObjectiveAortic aneurysm (AA) is associated with loss of elastin and structural integrity, accompanied by increased matrix metalloproteinase (MMP) expression. These processes are supported by inflammatory macrophages, with mediators such as tumor necrosis factor-α (TNF-α). Mesenchymal stem cells (MSCs) contribute to aortic remodeling. Therefore, to clarify whether MSCs might be useful for AA cell therapy, we examined the effect of MSCs on vascular smooth muscle cells (SMCs) and macrophages in vitro, on aortic tissue ex vivo, and on aorta in vivo.MethodsMurine macrophages and SMCs were cultured, with or without bone marrow-derived murine MSCs, for 96 hours in vitro. Gene expression of MMPs and TNF-α from macrophages and that of elastin from SMCs were measured. The murine aortic tissues were cultured with or without MSCs for up to 14 days, followed by measurement of MMP enzyme activity and elastin content. The in vivo aneurysm model used apolipoprotein E-deficient (apoE−/−) male mice receiving angiotensin II (Ang II) infusion for 28 days. MSCs were implanted by laparotomy to the abdominal aortic adventitial surface from the superior mesenteric artery origin to the left renal artery. Age-matched apoE−/− mice with or without Ang II infusion were used for control groups. At the end point, aortic diameter, elastin content, MMPs' activity, and cytokines expressed, including interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), TNF-α, insulin-like growth factor-1 (IGF-1), and tissue inhibitor of metalloproteinases-1 (TIMP-1) were quantified.ResultsMSCs suppressed MMP-2 with or without MSCs (2.59 vs 3.94, P < .05), MMP-9 (5.83 vs 9.70, P < .05), and TNF-α (2.79 vs 3.38, P < .05) expression in macrophages, and promoted elastin expression in SMCs (19.35 vs 3.23, P < .05) in vitro. MSCs also decreased active MMP-2 activity (0.310 vs 0.0609 U/μL, P < .05) and preserved elastin content (68.05 vs 40.29 μg/mg, P < .05) ex vivo. AA development was site-specifically inhibited (0.73 vs 1.04 mm aortic diameter, P < .05) and elastin content was preserved (46.9 vs 25.6 μg/mg, P < .05) at 4 weeks. Downregulation of MMPs and IL-6, MCP-1, and TNF-α, and upregulation of IGF-1 and TIMP-1 were demonstrated with MSC implantation in vivo.ConclusionsMSC implantation inhibits Ang II–induced AA development in apoE−/− mice through elastin preservation in the aortic wall and is associated with attenuated levels of MMPs and inflammatory cytokines.Clinical RelevanceAortic aneurysm (AA) rupture is a leading cause of sudden death. Current treatment options are conventional replacement surgery with a prosthetic conduit or endovascular stent grafting. Procedures on patients diagnosed with AA, particularly for those with thoracic lesions, are relatively invasive and often occur in an elderly, high-risk population. Although several pharmacologic strategies have been proposed, mesenchymal stem cell therapy has not been considered as a therapeutic approach for AA treatment. This report evaluates the efficacy of bone marrow-derived mesenchymal stem cells in an AA animal model in attenuation of aneurysmal growth by immunosuppression and elastin preservation.

The Levitronix PediVAS is an extracorporeal magnetically levitated pediatric ventricular assist system with an optimal flow rate range of 0.3 – 1.5 L/min. The system is being tested in preclinical studies to assess hemodynamic performance and biocompatibility. The PediVAS was implanted in 9 ovines for 30 days duration using either commercially available cannulae (N = 3) or customized Levitronix cannulae (N = 6). Blood biocompatibility in terms of circulating activated platelets was measured by flow cytometric assays to detect P-selectin. Platelet activation was further examined following exogenous agonist stimulation. Platelet activation rose following surgery and eventually returned to baseline in animal studies where minimal kidney infarcts were observed. Platelet activation remained elevated for the duration of the study in animals where a moderate number of kidney infarcts with or without thrombotic deposition in the cannulae were observed. When platelet activation did return to baseline, platelets appropriately responded to agonist stimulation signifying conserved platelet function following PediVAS implant. Platelet activation returned to baseline in the majority of studies, representing a promising biocompatibility result for the Levitronix PediVAS.

Ventricular assist devices (VADs) have significantly impacted the treatment of adult cardiac failure, but few options exist for pediatric patients. This has motivated our group to develop an implantable magnetically levitated rotodynamic VAD (PediaFlow®) for 3–20 kg patients. The second prototype design of the PediaFlow (PF2) is 56% smaller than earlier prototypes, and achieves 0.5–1.5 L/min blood flow rates. In vitro hemodynamic performance and hemolysis testing were performed with analog blood and whole ovine blood, respectively. In vivo evaluation was performed in an ovine model to evaluate hemocompatibility and end-organ function. The in vitro normalized index of hemolysis was 0.05–0.14 g/L over the specified operating range. In vivo performance was satisfactory for two of the three implanted animals. A mechanical defect caused early termination at 17 days of the first in vivo study, but two subsequent implants proceeded without complication and electively terminated at 30 and 70 days. Serum chemistries and plasma free hemoglobin were within normal limits. Gross necropsy revealed small, subclinical infarctions in the kidneys of the 30 and 70 day animals (confirmed by histopathology). The results of these experiments, particularly the biocompatibility demonstrated in vivo encourage further development of a miniature magnetically levitated VAD for the pediatric population. Ongoing work including further reduction of size will lead to a design freeze in preparation for of clinical trials.

Restoration of functional endothelium is a requirement for preventing late stent thrombosis. We propose a novel method for targeted delivery of stem cells to a site of arterial injury using ultrasound-generated acoustic radiation force. Mesenchymal stem cells (MSCs) were surface-coated electrostatically with cationic gas-filled lipid microbubbles (mb-MSC). mb-MSC was characterized microscopically and by flow cytometry. The effect of ultrasound (5 MHz) on directing mb-MSC movement toward the vessel wall under physiologic flow conditions was tested in vitro in a vessel phantom. In vivo testing of acoustic radiation force-mediated delivery of mb-MSCs to balloon-injured aorta was performed in rabbits using intravascular ultrasound (1.7 MHz) during intra-aortic infusion of mb-MSCs. Application of ultrasound led to marginalization and adhesion of mb-MSCs to the vessel phantom wall, whereas no effect was observed on mb-MSCs in the absence of ultrasound. The effect was maximal when there were 7±1 microbubbles/cell (n=6). In rabbits (n=6), adherent MSCs were observed in the ultrasound-treated aortic segment 20 min after the injection (334±137 MSCs/cm(2)), whereas minimal adhesion was observed in control segments not exposed to ultrasound (2±1 MSCs/cm(2), p0.05). At 24 h after mb-MSC injection and ultrasound treatment, the engrafted MSCs persisted and spread out on the luminal surface of the artery. The data demonstrate proof of principle that acoustic radiation force can target delivery of therapeutic cells to a specific endovascular treatment site. This approach may be used for endoluminal cellular paving and could provide a powerful tool for cell-based re-endothelialization of injured arterial segments.

The PediaFlow™ pediatric ventricular assist device (VAD) is a magnetically levitated turbodynamic pump under development for circulatory support of small children with a targeted flow rate range of 0.3 - 1.5 L/min. As the design of this device is refined, ensuring high levels of blood biocompatibility is essential. In this study we characterized platelet activation during the implantation and operation of a second generation prototype of the PediaFlow VAD (PF2) and also performed a series of surgical sham studies to examine purely surgical effects on platelet activation. In addition, a newly available monoclonal antibody was characterized and shown to be capable of quantifying ovine platelet activation. The PF2 was implanted in 3 chronic ovine experiments of 16, 30, and 70 days, while surgical sham procedures were performed in 5 ovines with 30 d monitoring. Blood biocompatibility in terms of circulating activated platelets was measured by flow cytometric assays with and without exogenous agonist stimulation. Platelet activation following sham surgery returned to baseline in approximately 2 weeks. Platelets in PF2 implanted ovines returned to baseline activation levels in all three animals, and showed an ability to respond to agonist stimulation. Late term platelet activation was observed in one animal corresponding with unexpected pump stoppages related to a manufacturing defect in the percutaneous cable. The results demonstrated encouraging platelet biocompatibility for the PF2 in that basal platelet activation was achieved early in the pump implant period. Furthermore, this first characterization of the effect of a major cardiothoracic procedure on temporal ovine platelet activation provides comparative data for future cardiovascular device evaluation in the ovine model.

One hundred thirty-seven spring-born yearling beef heifers of British breed types were used to determine the relationships between residual feed intake (RFI) and growth rate, body composition, mature size, and fertility. Heifers were housed in a dry lot facility during the experimental period, and data were collected over a 2-yr period (yr 1, n = 67; yr 2, n = 70). Individual feed intake, BW, BCS, hip height, and ultrasonic measurements [subcutaneous rib fat (UBF), rump fat (URF), LM area (LMA), and intramuscular fat (IMF)] of body composition were recorded. Individual feed intakes (kg of TDN consumed/d) were used to calculate RFI combining both years of data. Heifers averaged 387.0 ± 19.4 d of age and 337.1 ± 29.9 kg of BW at initiation of the experiment. Mean ADG was 1.14 ± 0.21 kg/d during the trial. Based on RFI, with year of test and farm of origin included in the model as covariates, heifers were classified into groups: positive (POS; 0.74 kg of TDN/d) or negative (NEG; -0.73 kg TDN/d) for first analysis and high (HI), medium (MED), or low (LO; mean RFI = 1.06, -0.01, and -1.13 kg of TDN/d, respectively) subsequently. An initial phenotypic relationship (P 0.10) from MED heifers at either the beginning or the end of test. Additionally, a negative linear relationship was observed between RFI and age at puberty (P 0.10). Differences in body fat and rate of metabolism associated with RFI could delay reproductive maturity.

The PediaFlow pediatric ventricular assist device is a miniature magnetically levitated mixed flow pump under development for circulatory support of newborns and infants (3-15 kg) with a targeted flow range of 0.3-1.5 L/min. The first generation design of the PediaFlow (PF1) was manufactured with a weight of approximately 100 g, priming volume less than 2 mL, length of 51 mm, outer diameter of 28 mm, and with 5-mm blood ports. PF1 was evaluated in an in vitro flow loop for 6 h and implanted in ovines for three chronic experiments of 6, 17, and 10 days. In the in vitro test, normalized index of hemolysis was 0.0087 ± 0.0024 g/100L. Hemodynamic performance and blood biocompatibility of PF1 were characterized in vivo by measurements of plasma free hemoglobin, plasma fibrinogen, total plasma protein, and with novel flow cytometric assays to quantify circulating activated ovine platelets. The mean plasma free hemoglobin values for the three chronic studies were 4.6 ± 2.7, 13.3 ± 7.9, and 8.8 ± 3.3 mg/dL, respectively. Platelet activation was low for portions of several studies but consistently rose along with observed animal and pump complications. The PF1 prototype generated promising results in terms of low hemolysis and platelet activation in the absence of complications. Hemodynamic results validated the magnetic bearing design and provided the platform for design iterations to meet the objective of providing circulatory support for young children with exceptional biocompatibility.

Heart failure initiated by coronary artery disease and myocardial infarction (MI) is a widespread, debilitating condition for which there are a limited number of options to prevent disease progression. Intra-myocardial biomaterial injection following MI theoretically provides a means to reduce the stresses experienced by the infarcted ventricular wall, which may alter the pathological remodeling process in a positive manner. Furthermore, biomaterial injection provides an opportunity to concurrently introduce cellular components and depots of bioactive agents. Biologically derived, synthetic and hybrid materials have been applied, as well as materials designed expressly for this purpose, although optimal design parameters, including degradation rate and profile, injectability, elastic modulus and various possible bioactivities, largely remain to be elucidated. This review seeks to summarize the current body of growing literature where biomaterial injection, with and without concurrent pharmaceutical or cellular delivery, has been pursued to improve functional outcomes following MI. The literature to date generally demonstrates acute functional benefits associated with biomaterial injection therapy across a broad variety of animal models and material compositions. Further functional improvements have been reported when cellular or pharmaceutical agents have been incorporated into the delivery system. Despite these encouraging early results, the specific mechanisms behind the observed functional improvements remain to be fully explored and future studies employing hypothesis-driven material design and selection may increase the potential of this approach to alleviate the morbidity and mortality of heart failure.

Thrombosis and thromboembolism remain problematic for a large number of blood contacting medical devices and limit broader application of some technologies due to this surface bioincompatibility. In this study we focused on the covalent attachment of zwitterionic phosphorylcholine (PC) or sulfobetaine (SB) moieties onto a TiAl 6 V 4 surface with a single step modification method to obtain a stable blood compatible interface. Silanated PC or SB modifiers (PCSi or SBSi) which contain an alkoxy silane group and either PC or SB groups were prepared respectively from trimethoxysilane and 2-methacryloyloxyethyl phosphorylcholine (MPC) or N -(3-sulfopropyl)- N -(methacryloxyethyl)- N , N -dimethylammonium betaine (SMDAB) monomers by a hydrosilylation reaction. A cleaned and oxidized TiAl 6 V 4 surface was then modified with the PCSi or SBSi modifiers by a simple surface silanization reaction. The surface was assessed with X-ray photoelectron spectroscopy (XPS), attenuated total reflection-Fourier transform infrared spectroscopy (ATR-FTIR) and contact angle goniometry. Platelet deposition and bulk phase activation were evaluated following contact with anticoagulated ovine blood. XPS results verified successful modification of the PCSi or SBSi modifiers onto TiAl 6 V 4 based on increases in surface phosphorous or sulfur respectively. Surface contact angles in water decreased with the addition of hydrophilic PC or SB moieties. Both the PCSi and SBSi modified TiAl 6 V 4 surfaces showed decreased platelet deposition and bulk phase platelet activation compared to unmodified TiAl 6 V 4 and control surfaces. This single step modification with PCSi or SBSi modifiers offers promise for improving the surface hemocompatibility of TiAl 6 V 4 and is attractive for its ease of application to geometrically complex metallic blood contacting devices.

Replacement or regeneration of load-bearing soft tissues has long been the impetus for the development of bioactive materials. While maturing, current efforts continue to be confounded by our lack of understanding of the intricate multi-scale hierarchical arrangements and interactions typically found in native tissues. The current state of the art in biomaterial processing enables a degree of controllable microstructure that can be used for the development of model systems to deduce fundamental biological implications of matrix morphologies on cell function. Furthermore, the development of computational frameworks which allow for the simulation of experimentally derived observations represents a positive departure from what has mostly been an empirically driven field, enabling a deeper understanding of the highly complex biological mechanisms we wish to ultimately emulate. Ongoing research is actively pursuing new materials and processing methods to control material structure down to the micro-scale to sustain or improve cell viability, guide tissue growth, and provide mechanical integrity, all while exhibiting the capacity to degrade in a controlled manner. The purpose of this review is not to focus solely on material processing but to assess the ability of these techniques to produce mechanically sound tissue surrogates, highlight the unique structural characteristics produced in these materials, and discuss how this translates to distinct macroscopic biomechanical behaviors.

Injectable thermoresponsive hydrogels are of interest for a variety of biomedical applications, including regional tissue mechanical support as well as drug and cell delivery. Within this class of materials there is a need to provide options for gels with stronger mechanical properties as well as variable degradation profiles. To address this need, the hydrolytically labile monomer, methacrylate-polylactide (MAPLA), with an average 2.8 lactic acid units, was synthesized and copolymerized with N-isopropylacrylamide (NIPAAm) and 2-hydroxyethyl methacrylate (HEMA) to obtain bioabsorbable thermally responsive hydrogels. Poly(NIPAAm-co-HEMA-co-MAPLA) with three monomer feed ratios (84/10/6, 82/10/8, and 80/10/10) was synthesized and characterized with NMR, FTIR, and GPC. The copolymers were soluble in saline at reduced temperature (10 degrees C), forming clear solutions that increased in viscosity with the MAPLA feed ratio. The copolymers underwent sol-gel transition at lower critical solution temperatures of 12.4, 14.0, and 16.2 degrees C, respectively, and solidified immediately upon being placed in a 37 degrees C water bath. The warmed hydrogels gradually excluded water to reach final water contents of approximately 45%. The hydrogels as formed were mechanically strong, with tensile strengths as high as 100 kPa and shear moduli of 60 kPa. All three hydrogels were completely degraded (solubilized) in PBS over a 6-7 month period at 37 degrees C, with a higher MAPLA feed ratio resulting in a faster degradation period. Culture of primary vascular smooth muscle cells with degradation solutions demonstrated a lack of cytotoxicity. The synthesized hydrogels provide new options for biomaterial injection therapy where increased mechanical strength and relatively slow resorption rates would be attractive.

Biodegradable elastomeric scaffolds are of increasing interest for applications in soft tissue repair and regeneration, particularly in mechanically active settings. The rate at which such a scaffold should degrade for optimal outcomes, however, is not generally known and the ability to select from similar scaffolds that vary in degradation behavior to allow such optimization is limited. Our objective was to synthesize a family of biodegradable polyurethane elastomers where partial substitution of polyester segments with polycarbonate segments in the polymer backbone would lead to slower degradation behavior. Specifically, we synthesized poly(ester carbonate)urethane ureas (PECUUs) using a blended soft segment of poly(caprolactone) (PCL) and poly(1,6-hexamethylene carbonate) (PHC), a 1,4-diisocyanatobutane hard segment and chain extension with putrescine. Soft segment PCL/PHC molar ratios of 100/0, 75/25, 50/50, 25/75, and 0/100 were investigated. Polymer tensile strengths varied from 14 to 34 MPa with breaking strains of 660-875%, initial moduli of 8-24 MPa and 100% recovery after 10% strain. Increased PHC content was associated with softer, more distensible films. Scaffolds produced by salt leaching supported smooth muscle cell adhesion and growth in vitro. PECUU in aqueous buffer in vitro and subcutaneous implants in rats of PECUU scaffolds showed degradation slower than comparable poly(ester urethane)urea and faster than poly(carbonate urethane)urea. These slower degrading thermoplastic polyurethanes provide opportunities to investigate the role of relative degradation rates for mechanically supportive scaffolds in a variety of soft tissue repair and reconstructive procedures.

Artificial lungs composed of bundles of micro-porous hollow fiber membranes (HFMs), which are made from polymeric materials such as poly (methylpentene) and poly(propylene), are routinely employed to both oxygenate blood and remove carbon dioxide (CO2) in an extraluminal blood flow format. The efficiency of CO2 and oxygen (O2) gas exchange in the current artificial lung model, which is based on passive diffusion, is limited by the fiber surface area to blood volume ratio with devices requiring approximately 1–2 m2 of HFM surface to provide adequate gas exchange (1,2). Such large blood-contacting surfaces present significant challenges to hemocompatibility, necessitating aggressive anticoagulation and stimulating research into devices with improved efficiency (i.e., smaller) and hemocompatibility. A wide variety of surface modification techniques have been evaluated to reduce the thrombogenicity of blood-contacting biomaterials, although there have been relatively few reports specifically focused on the HFMs utilized in artificial lung applications. Siloxane-grafted HFMs prepared by a plasma polymerization process with 1,3,3,7-tetramethyhydrocyclosiloxane (TMCTS)-coated fibers have shown reduced thrombogenesis relative to unmodified fibers (3). Plasma polymerization techniques have a number of advantages with regard to surface modification including: facile preparation of a thin, conformal, and pinhole-free coating; amenability to a wide variety of substrates; achievement of good adhesion between coating and substrate; and the ability to generate coatings that present excellent thermal and chemical resistance (4,5). Another approach to improving artificial lung biocompatibility is to effectively reduce the required HFM surface area by increasing the gas exchange rate of HFM-based devices. Increasing the efficiency of CO2 removal is especially important because the natural concentration gradient for CO2 diffusion is much smaller than that for O2 addition. Furthermore, in many patients with respiratory failure, the need for CO2 removal is more important clinically, as oxygenation can be provided by nasal cannula or lung-protective ventilation (1,6–8). In a previous study, we reported the development of a bioactive HFM that could improve CO2 removal rates in lung failure patients (9). Carbonic anhydrase (CA) was covalently immobilized to the surface of a conventional HFM, and by catalyzing the dehydration of bicarbonate in blood was shown to facilitate diffusion of CO2 toward the fiber membranes, essentially mimicking the function of the enzyme on lung capillary surfaces. Results indicated that CO2 exchange rates from buffer were increased by as much as 75% in the model device. In this brief report, we addressed whether the attachment of CA onto a HFM surface increased fiber thrombogenicity, which would negate the biocompatibility benefits associated with a smaller device. A commercial poly(propylene) HFM (Celgard, Charlotte, NC,USA) was used as the base material and for control purposes. Fibers representing the intermediate modification steps of siloxane coating and amine grafting on siloxane (Alung Technologies, Pittsburgh, PA, USA) as well as the final CA-modified fiber were evaluated. A second control of a commercial poly (methyl pentene) HFM (Oxyplus, Membrana, Wuppertal, Germany) was also included.

Limited autologous vascular graft availability and poor patency rates of synthetic grafts for bypass or replacement of small-diameter arteries remain a concern in the surgical community. These limitations could potentially be improved by a tissue engineering approach. We report here our progress in the development and in vivo testing of a stem-cell-based tissue-engineered vascular graft for arterial applications. Poly(ester urethane)urea scaffolds (length = 10 mm; inner diameter = 1.2 mm) were created by thermally induced phase separation (TIPS). Compound scaffolds were generated by reinforcing TIPS scaffolds with an outer electrospun layer of the same biomaterial (ES-TIPS). Both TIPS and ES-TIPS scaffolds were bulk-seeded with 10 x 10(6) allogeneic, LacZ-transfected, muscle-derived stem cells (MDSCs), and then placed in spinner flask culture for 48 h. Constructs were implanted as interposition grafts in the abdominal aorta of rats for 8 weeks. Angiograms and histological assessment were performed at the time of explant. Cell-seeded constructs showed a higher patency rate than the unseeded controls: 65% (ES-TIPS) and 53% (TIPS) versus 10% (acellular TIPS). TIPS scaffolds had a 50% mechanical failure rate with aneurysmal formation, whereas no dilation was observed in the hybrid scaffolds. A smooth-muscle-like layer of cells was observed near the luminal surface of the constructs that stained positive for smooth muscle alpha-actin and calponin. LacZ+ cells were shown to be engrafted in the remodeled construct. A confluent layer of von Willebrand Factor-positive cells was observed in the lumen of MDSC-seeded constructs, whereas acellular controls showed platelet and fibrin deposition. This is the first evidence that MDSCs improve patency and contribute to the remodeling of a tissue-engineered vascular graft for arterial applications.

Although a variety of materials are currently used for abdominal wall repair, general complications encountered include herniation, infection, and mechanical mismatch with native tissue. An approach wherein a degradable synthetic material is ultimately replaced by tissue mechanically approximating the native state could obviate these complications. We report here on the generation of biodegradable scaffolds for abdominal wall replacement using a wet electrospinning technique in which fibers of a biodegradable elastomer, poly(ester urethane)urea (PEUU), were concurrently deposited with electrosprayed serum-based culture medium. Wet electrospun PEUU (wet ePEUU) was found to exhibit markedly different mechanical behavior and to possess an altered microstructure relative to dry processed ePEUU. In a rat model for abdominal wall replacement, wet ePEUU scaffolds (1 × 2.5 cm) provided a healing result that developed toward approximating physiologic mechanical behavior at 8 weeks. An extensive cellular infiltrate possessing contractile smooth muscle markers was observed together with extensive extracellular matrix (collagens, elastin) elaboration. Control implants of dry ePEUU and expanded polytetrafluoroethylene did not experience substantial cellular infiltration and did not take on the native mechanical anisotropy of the rat abdominal wall. These results illustrate the markedly different in vivo behavior observed with this newly reported wet electrospinning process, offering a potentially useful refinement of an increasingly common biomaterial processing technique.

A major barrier to the development of a clinically useful small diameter tissue engineered vascular graft (TEVG) is the scaffold component. Scaffold requirements include matching the mechanical and structural properties with those of native vessels and optimizing the microenvironment to foster cell integration, adhesion and growth. We have developed a small diameter, bilayered, biodegradable, elastomeric scaffold based on a synthetic, biodegradable elastomer. The scaffold incorporates a highly porous inner layer, allowing cell integration and growth, and an external, fibrous reinforcing layer deposited by electrospinning. Scaffold morphology and mechanical properties were assessed, quantified and compared with those of native vessels. Scaffolds were then seeded with adult stem cells using a rotational vacuum seeding device to obtain a TEVG, cultured under dynamic conditions for 7 days and evaluated for cellularity. The scaffold showed firm integration of the two polymeric layers with no delamination. Mechanical properties were physiologically consistent, showing anisotropy, an elastic modulus (1.4 + or - 0.4 MPa) and an ultimate tensile stress (8.3 + or - 1.7 MPa) comparable with native vessels. The compliance and suture retention forces were 4.6 + or - 0.5 x 10(-4) mmHg(-1) and 3.4 + or - 0.3N, respectively. Seeding resulted in a rapid, uniform, bulk integration of cells, with a seeding efficiency of 92 + or - 1%. The scaffolds maintained a high level of cellular density throughout dynamic culture. This approach, combining artery-like mechanical properties and a rapid and efficient cellularization, might contribute to the future clinical translation of TEVGs.

To improve the thromboresistance of a titanium alloy (TiAl 6 V 4 ) surface which is currently utilized in several ventricular assist devices (VADs), a plasma-induced graft polymerization of 2-methacryloyloxyethyl phosphorylcholine (MPC) was carried out and poly(MPC) (PMPC) chains were covalently attached onto a TiAl 6 V 4 surface by a plasma induced technique. Cleaned TiAl 6 V 4 surfaces were pretreated with H 2 O-vapor-plasma and silanated with 3-methacryloylpropyltrimethoxysilane (MPS). Next, a plasma-induced graft polymerization with MPC was performed after the surfaces were pretreated with Ar plasma. Surface compositions were verified by X-ray photoelectron spectroscopy (XPS). In vitro blood biocompatibility was evaluated by contacting the modified surfaces with ovine blood under continuous mixing. Bulk phase platelet activation was quantified by flow cytometric analysis, and surfaces were observed with scanning electron microscopy after blood contact. XPS data demonstrated successful modification of the TiAl 6 V 4 surfaces with PMPC as evidenced by increased N and P on modified surfaces. Platelet deposition was markedly reduced on the PMPC grafted surfaces and platelet activation in blood that contacted the PMPC-grafted samples was significantly reduced relative to the unmodified TiAl 6 V 4 and polystyrene control surfaces. Durability studies under continuously mixed water suggested no change in surface modification over a 1-month period. This modification strategy shows promise for further investigation as a means to reduce the thromboembolic risk associated with the metallic blood-contacting surfaces of VADs and other cardiovascular devices under development.

Our objective was to develop a surface modification strategy for a titanium alloy (TiAl6V4) to provide thromboresistance for surfaces in rigorous blood-contacting cardiovascular applications, such as that found in ventricular assist devices. We hypothesized that this could be accomplished by the covalent attachment of a phospholipid polymer, poly(2-methacryloyloxyethylphosphorylcholine (MPC)-co-methacryl acid) (PMA). TiAl6V4 was H2O plasma treated by radio frequency glow discharge, silanated with 3-aminopropyltriethoxysilane (APS), and ammonia plasma treated to increase surface reactivity. The TiAl6V4 surface was then modified with PMA via a condensation reaction between the amino groups on the TiAl6V4 surface and the carboxyl groups on PMA. The surface composition was verified by X-ray photoelectron spectroscopy, confirming successful modification of the TiAl6V4 surfaces with APS and PMA as evidenced by increased Si and P. Plasma treatments with H2O and ammonia were effective at further increasing the surface reactivity of TiAl6V4 as evidenced by increased surface PMA. The adsorption of ovine fibrinogen onto PMA-modified surfaces was reduced relative to unmodified surfaces, and in vitro ovine blood contact through a rocking test revealed marked reductions in platelet deposition and bulk phase platelet activation relative to unmodified TiAl6V4 and polystyrene controls. The results indicate that the PMA-modification scheme for TiAl6V4 surfaces offers a potential pathway to improve the thromboresistance of the blood-contacting surfaces of cardiovascular devices.

Injection of a bulking material into the ventricular wall has been proposed as a therapy to prevent progressive adverse remodeling due to high wall stresses that develop after myocardial infarction. Our objective was to design, synthesize and characterize a biodegradable, thermoresponsive hydrogel for this application based on copolymerization of N-isopropylacrylamide (NIPAAm), acrylic acid (AAc) and hydroxyethyl methacrylate-poly(trimethylene carbonate) (HEMAPTMC). By evaluating a range of monomer ratios, poly(NIPAAm-co-AAc-co-HEMAPTMC) at a feed ratio of 86/4/10 was shown to be ideal since it formed a hydrogel at 37 degrees C, and gradually became soluble over a 5 month period in vitro through hydrolytic cleavage of the PTMC residues. HEMAPTMC, copolymer and degradation product chemical structures were verified by NMR. No degradation product cytotoxicity was observed in vitro. In a rat chronic infarction model, the infarcted left ventricular (LV) wall was injected with the hydrogel or phosphate buffered saline (PBS). In the PBS group, LV cavity area increased and contractility decreased at 8 wk (p

We have utilized soft lithography techniques to create three-dimensional arrays of blood microchannels and gas pathways in poly(dimethylsiloxane) (PDMS) that approach the microvascular scale of the natural lung. The blood microchannels were lined with endothelial cells in an effort to provide a non-thrombogenic surface that might ultimately reduce the need for systemic anticoagulation. A novel design and fabrication method were developed to create prototype modules for gas permeance and cell culture testing. The gas permeance modules contained 6 layers, four gas and two blood, while the modules for cell culture testing contained two layers of blood channels. The gas permeance of the modules was examined and maximum values of 9.16 x 10(-6) and 3.55 x 10(-5) mL/s/cm(2)/cmHg, for O(2) and CO(2) respectively, were obtained. Finally, endothelial cells were seeded and dynamically cultured in prototype cell culture modules. Confluent and viable cell monolayers were achieved after 10 days of perfusion.

Heart failure (HF) is a major public health problem in the United States, and its prevalence is likely to increase with the aging U.S. population. Mechanical circulatory support (MCS) utilizing bladder-based blood pumps generating pulsatile flow has been reserved for patients with severe HF failing medical therapy. As MCS technology has advanced to include rotary blood pumps, so has our understanding of the biological and clinical responses to MCS, which in turn has altered the risk/benefit profile of this therapy. This may lead to paradigm shifts in device usage from support of end-stage HF to temporary support for recovery of cardiac function and earlier usage, to, ultimately, prevention of disease progression. This review serves to explore the current state and future opportunities of MCS within our larger understanding of the epidemiology, pathophysiology, and treatment options for HF.

Myocardial contrast echocardiography utilizes intravenously injected gas-filled microspheres as acoustically active red blood cell tracers. During ultrasound imaging, unimpeded microsphere transit through the intramyocardial microcirculation causes transient myocardial opacification, which can be mapped and quantified as myocardial perfusion. Ultrasound molecular imaging utilizes similar acoustically active microspheres, which are modified to bear a receptor-specific ligand on the surface, conferring microsphere binding to a disease- specific endothelial epitope. Because the microspheres adhere to the endothelium, ultrasound imaging reveals a persistent, rather than transient, contrast effect, indicating the presence and location of the molecule of interest in real time. Molecular contrast echocardiography has been developed to detect upregulated leukocyte adhesion molecules during microvascular inflammation, such as occurs in cardiac transplant rejection and ischemia–reperfusion. Principles of microsphere targeting and ultrasound imaging of microvascular epitopes have been extended to larger vessels to image molecular markers of atherosclerosis. This Article summarizes the current status of cardiovascular ultrasound molecular imaging. Experimental proofs of concept will be outlined and the clinical extension of these concepts to the molecular imaging of cardiovascular disease using clinical ultrasound technology will be discussed.

To develop elastase-sensitive polyurethane scaffolds that would be applicable to the engineering of mechanically active soft tissues.A polyurethane containing an elastase-sensitive peptide sequence was processed into scaffolds by thermally induced phase separation. Processing conditions were manipulated to alter scaffold properties and anisotropy. The scaffold's mechanical properties, degradation, and cytocompatibility using muscle-derived stem cells were characterized. Scaffold in vivo degradation was evaluated by subcutaneous implantation.When heat transfer was multidirectional, scaffolds had randomly oriented pores. Imposition of a heat transfer gradient resulted in oriented pores. Both scaffolds were flexible and relatively strong with mechanical properties dependent upon fabrication conditions such as solvent type, polymer concentration and quenching temperature. Oriented scaffolds exhibited anisotropic mechanical properties with greater tensile strength in the orientation direction. These scaffolds also supported muscle-derived stem cell growth more effectively than random scaffolds. The scaffolds expressed over 40% weight loss after 56 days in elastase containing buffer. Elastase-sensitive scaffolds were complete degraded after 8 weeks subcutaneous implantation in rats, markedly faster than similar polyurethanes that did not contain the peptide sequence.The elastase-sensitive polyurethane scaffolds showed promise for application in soft tissue engineering where controlling scaffold mechanical properties and pore architecture are desirable.

There is a clinical need for a tissue-engineered vascular graft (TEVG), and combining stem cells with biodegradable tubular scaffolds appears to be a promising approach. The goal of this study was to characterize the incorporation of muscle-derived stem cells (MDSCs) within tubular poly(ester urethane) urea (PEUU) scaffolds in vitro to understand their interaction, and to evaluate the mechanical properties of the constructs for vascular applications. Porous PEUU scaffolds were seeded with MDSCs using our recently described rotational vacuum seeding device, and cultured inside a spinner flask for 3 or 7 days. Cell viability, number, distribution and phenotype were assessed along with the suture retention strength and uniaxial mechanical behavior of the TEVGs. The seeding device allowed rapid even distribution of cells within the scaffolds. After 3 days, the constructs appeared completely populated with cells that were spread within the polymer. Cells underwent a population doubling of 2.1-fold, with a population doubling time of 35 h. Stem cell antigen-1 (Sca-1) expression by the cells remained high after 7 days in culture (77+/-20% vs. 66+/-6% at day 0) while CD34 expression was reduced (19+/-12% vs. 61+/-10% at day 0) and myosin heavy chain expression was scarce (not quantified). The estimated burst strength of the TEVG constructs was 2127+/-900 mm Hg and suture retention strength was 1.3+/-0.3N. We conclude from this study that MDSCs can be rapidly seeded within porous biodegradable tubular scaffolds while maintaining cell viability and high proliferation rates and without losing stem cell phenotype for up to 7 days of in-vitro culture. The successful integration of these steps is thought necessary to provide rapid availability of TEVGs, which is essential for clinical translation.

Scaffolds that better approximate the mechanical properties of cardiovascular and other soft tissues might provide a more appropriate mechanical environment for tissue development or healing in vivo. An ability to induce local angiogenesis by controlled release of an angiogenic factor, such as basic fibroblast growth factor (bFGF), from a biodegradable scaffold with mechanical properties more closely approximating soft tissue could find application in a variety of settings. Toward this end biodegradable poly(ester urethane)urea (PEUU) scaffolds loaded with bFGF were fabricated by thermally induced phase separation. Scaffold morphology, mechanical properties, release kinetics, hydrolytic degradation and bioactivity of the released bFGF were assessed. The scaffolds had inter-connected pores with porosities of 90% or greater and pore sizes ranging from 34-173 microm. Scaffolds had tensile strengths of 0.25-2.8 MPa and elongations at break of 81-443%. Incorporation of heparin into the scaffold increased the initial burst release of bFGF, while the initial bFGF loading content did not change release kinetics significantly. The released bFGF remained bioactive over 21 days as assessed by smooth muscle mitogenicity. Scaffolds loaded with bFGF showed slightly higher degradation rates than unloaded control scaffolds. Smooth muscle cells seeded into the scaffolds with bFGF showed higher cell densities than for control scaffolds after 7 days of culture. The bFGF-releasing PEUU scaffolds thus exhibited a combination of mechanical properties and bioactivity that might be attractive for use in cardiovascular and other soft tissue applications.

Biodegradable synthetic matrices that resemble the size scale, architecture and mechanical properties of the native extracellular matrix (ECM) can be fabricated through electrospinning. Tubular conduits may also be fabricated with properties appropriate for vascular tissue engineering. Achieving substantial cellular infiltration within the electrospun matrix in vitro remains time consuming and challenging. This difficulty was overcome by electrospraying smooth muscle cells (SMCs) concurrently with electrospinning of a biodegradable, elastomeric poly(ester urethane) urea (PEUU) small-diameter conduit. Constructs were cultured statically or in spinner flasks. Hematoxylin and eosin (H&E) staining demonstrated qualitatively uniform SMCs integration radially and circumferentially within the conduit after initial static culture. In comparison with static culture, samples cultured in spinner flasks indicated 2.4 times more viable cells present from MTT and significantly larger numbers of SMCs spread within the electrospun fiber networks by H&E image analysis. Conduits were strong and flexible with mechanical behaviors that mimicked those of native arteries, including static compliance of 1.6+/-0.5 x 10(-3)mmHg(-1), dynamic compliance of 8.7+/-1.8 x 10(-4)mmHg(-1), burst strengths of 1750+/-220 mmHg, and suture retention. This method to rapidly and efficiently integrate cells into a strong, compliant biodegradable tubular matrix represents a significant achievement as a tissue engineering approach for blood vessel replacement.