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Biomedical subjects

David J Mooney

Publications and source records attributed to David J Mooney.

At least 19 recordsLinked to original sources
PubMed2003

Elastic biodegradable poly(glycolide-co-caprolactone) scaffold for tissue engineering.

Cyclic mechanical strain has been demonstrated to enhance the development and function of engineered smooth muscle (SM) tissues, and it would be necessary for the development of the elastic scaffolds if one wishes to engineer SM tissues under cyclic mechanical loading. This study reports on the development of an elastic scaffold fabricated from a biodegradable polymer. Biodegradable poly(glycolide-co-caprolactone) (PGCL) copolymer was synthesized from glycolide and epsilon-caprolactone in the presence of stannous octoate as catalyst. The copolymer was characterized by (1)H-NMR, gel permeation chromatography and differential scanning calorimetry. Scaffolds for tissue engineering applications were fabricated from PGCL copolymer using the solvent-casting and particle-leaching technique. The PGCL scaffolds produced in this fashion had open pore structures (average pore size = 250 microm) without the usual nonporous skin layer on external surfaces. Mechanical testing revealed that PGCL scaffolds were far more elastic than poly(lactic-co-glycolic acid) (PLGA) scaffolds fabricated using the same method. Tensile mechanical tests indicated that PGCL scaffolds could withstand an extension of 250% without cracking, which was much higher than withstood by PLGA scaffolds (10-15%). In addition, PGCL scaffolds achieved recoveries exceeding 96% at applied extensions of up to 230%, whereas PLGA scaffolds failed (cracked) at an applied strain of 20%. Dynamic mechanical tests showed that the permanent deformation of the PGCL scaffolds in a dry condition produced was less than 4% of the applied strain, when an elongation of 20% at a frequency of 1 Hz (1 cycle per second) was applied for 6 days. Moreover, PGCL scaffolds in a buffer solution also had permanent deformations less than 5% of the applied strain when an elongation of 10% at a frequency of 1 Hz was applied for 2 days. The usefulness of the PGCL scaffolds was demonstrated by engineering SM tissues in vivo. This study shows that the elastic PGCL scaffolds produced in this study could be used to engineer SM-containing tissues (e.g. blood vessels and bladders) in mechanically dynamic environments.

Absorbable Implants↗
PubMed2003

Comparison of vascular endothelial growth factor and basic fibroblast growth factor on angiogenesis in SCID mice.

Therapeutic angiogenesis is a promising approach to treat patients with cardiovascular disease, and will likely be critical to engineering large tissues. Many growth factors have been found to play significant roles in angiogenesis, and vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are the most extensively investigated angiogenic factors to date. However, the appropriate dose to obtain a desired response and the effectiveness of each factor, relative to the other, in promoting angiogenesis at a specific site in the body remains unclear. We have used alginate hydrogels as localized delivery vehicles for VEGF and bFGF, and compared the ability of these factors to promote new blood vessel formation in the subcutaneous tissue of severe combined immunodeficient (SCID) mice. We have found that the thickness of a granulation tissue layer formed around the gel and the number of blood vessels in the layer increased with the dose of VEGF in the gel, but the density of new blood vessels remained relatively constant. Sustained and localized delivery of bFGF from the gels, while similarly leading to an increase in the density of blood vessels in the granulation tissue, did not lead to as high of a blood vessel density as VEGF. The results of this study support previous studies demonstrating the utility of both VEGF and bFGF in promoting angiogenesis, and suggest VEGF is more appropriate for creating a dense bed of new blood vessels in this model.

Angiogenesis Inducing Agents↗
PubMed2003

Cyclic strain inhibits switching of smooth muscle cells to an osteoblast-like phenotype.

Ectopic calcification is commonly associated with cardiovascular disease, injury, aging, and biomaterial implantation. We hypothesized that the normal mechanical environment of smooth muscle cells (SMCs) inhibits a phenotypic switch to a pattern of gene expression more typical for bone and inducive for calcification. This hypothesis was tested using a 3-D engineered smooth muscle tissue model subjected to cyclic mechanical strain. This simplified model maintained a 3-D tissue architecture while eliminating systemic effects as can be seen with in vivo models. All engineered tissues were found to express bone-associated genes (osteopontin, matrix gla protein, alkaline phosphatase, and the transcription factor CBFA-1). Strikingly, however, expression of these genes was down-regulated in tissues exposed to cyclic strain at all time points ranging from 5 to 150 days. Furthermore, long-term strain played a protective role in regard to calcification, as unstrained tissues exhibited increased calcium deposition with respect to strained tissues. The results of this study suggest that without an appropriate mechanical environment, SMCs in 3-D culture undergo a phenotypic conversion to an osteoblast-like pattern of gene expression. This finding has significant implications for the mechanisms underlying a variety of cardiovascular diseases and indicates the broad utility of engineered tissue models in basic biology studies.

Animals↗
PubMed2003

Cyclic strain enhances matrix mineralization by adult human mesenchymal stem cells via the extracellular signal-regulated kinase (ERK1/2) signaling pathway.

Physical stimuli play critical roles in the development, regeneration, and pathology of many mesenchymal tissues, most notably bone. While mature bone cells, such as osteoblasts and osteocytes, are clearly involved in these processes, the role of their progenitors in mechanically mediated tissue responses is unknown. In this study, we investigated the effect of cyclic substrate deformation on the proliferation and osteogenic differentiation of human mesenchymal stem cells (hMSCs). Application of equibiaxial cyclic strain (3%, 0.25Hz) to hMSCs cultured in osteogenic media inhibited proliferation and stimulated a 2.3-fold increase in matrix mineralization over unstrained cells. The strain stimulus activated the extracellular signal-regulated kinase (ERK1/2) and p38 mitogen-activated protein kinase pathways, but had no effect on c-Jun N-terminal kinase phosphorylation or activity. Strain-induced mineralization was largely mediated by ERK1/2 signaling, as inhibition of ERK1/2 attenuated calcium deposition by 55%. Inhibition of the p38 pathway resulted in a more mature osteogenic phenotype, suggesting an inhibitory role for p38 signaling in the modulation of strain-induced osteogenic differentiation. These results demonstrate that mechanical signals regulate hMSC function, suggesting a critical role for physical stimulation of this specific cell population in mesenchymal tissue formation.

Adult↗
PubMed2003

Designing alginate hydrogels to maintain viability of immobilized cells.

Hydrogel-forming materials have been widely utilized as an immobilization matrix and transport vehicle for cells. Success in these applications is dependent upon maintaining cell viability through the gel preparation process. We hypothesized that the high viscosity of pre-gelled solutions typically used in these applications may decrease cell viability due to the high shear forces required to mix cells with these solutions. Further, we proposed this harmful effect could be mediated by decreasing the molecular weight (Mw) of the polymer used to form the gel, while maintaining its gel-forming ability. To investigate this hypothesis, alginate was used as model system, as this copolymer consists of cross-linkable guluronic acid (G) blocks and non-cross-linkable blocks. Decreasing the Mw of alginate using irradiation (e.g., irradiating at dose of 2 Mrad) decreased the low shear viscosity of 2% (w/w) pre-gelled solutions from 1000 to 4 cP, while maintaining high elastic moduli, once cross-linked to form a gel. Importantly, the immobilization of cells with these polymer hydrogels increased cell viability from 40% to 70%, as compared to using high Mw polymer chains to form the gels. Furthermore, the solids concentration of gels formed with the low Mw alginate could be raised to further increase the moduli of gels without significantly deteriorating the viability of immobilized cells. This was likely due to the limited increase in the viscosity of these solutions. This material design approach may be useful with a variety of synthetic or naturally occurring block copolymers used to immobilize cells.

Alginates↗
PubMed2003

Hydrogels for tissue engineering: scaffold design variables and applications.

Polymer scaffolds have many different functions in the field of tissue engineering. They are applied as space filling agents, as delivery vehicles for bioactive molecules, and as three-dimensional structures that organize cells and present stimuli to direct the formation of a desired tissue. Much of the success of scaffolds in these roles hinges on finding an appropriate material to address the critical physical, mass transport, and biological design variables inherent to each application. Hydrogels are an appealing scaffold material because they are structurally similar to the extracellular matrix of many tissues, can often be processed under relatively mild conditions, and may be delivered in a minimally invasive manner. Consequently, hydrogels have been utilized as scaffold materials for drug and growth factor delivery, engineering tissue replacements, and a variety of other applications.

Biocompatible Materials↗
PubMed2003

Polymeric growth factor delivery strategies for tissue engineering.

PURPOSE: Tissue engineering seeks to replace and regrow damaged or diseased tissues and organs from either cells resident in the surrounding tissue or cells transplanted to the tissue site. The purpose of this review is to present the application of polymeric delivery systems for growth factor delivery in tissue engineering. METHODS: Growth factors direct the phenotype of both differentiated and stem cells, and methods used to deliver these molecules include the development of systems to deliver the protein itself, genes encoding the factor, or cells secreting the factor. RESULTS: Results in animal models and clinical trials indicate that these approaches may be successfully used to promote the regeneration of numerous tissue types. CONCLUSIONS: Controlling the dose, location, and duration of these factors through polymeric delivery strategies will dictate their utility in tissue regeneration.

Animals↗
PubMed2003

Role of vascular endothelial growth factor in bone marrow stromal cell modulation of endothelial cells.

One of the fundamental principles that underlies tissue-engineering strategies using cell transplantation is that a newly formed tissue must acquire and maintain sufficient vascularization in order to support its growth. Enhancing angiogenesis through delivery of growth factors is one approach to establishing a vascular network to these tissues. In this study, we tested the potential of bone marrow stromal cells (BMSCs) to modulate the growth and differentiation activities of blood vessel precursors, endothelial cells (ECs), by their secretion of soluble angiogenic factors. The growth and differentiation of cultured ECs were enhanced in response to exposure to BMSC conditioned medium (CM). Enzyme-linked immunosorbent assays demonstrated that both mouse and human BMSCs secreted significant quantities of vascular endothelial growth factor (VEGF) (2.4-3.1 ng/10(6) cells per day). Furthermore, eliminating the activity of BMSC-secreted VEGF with blocking antibodies completely blocked the CM effects on cultured ECs. These data demonstrate that human BMSCs secrete sufficient quantities of VEGF to enhance survival and differentiation of endothelial cells in vitro, and suggest they may be capable of directly orchestrating angiogenesis in vivo.

Bone Marrow Cells↗
PubMed2003

Selective adipose tissue ablation by localized, sustained drug delivery.

The reduction of adipose depots is widely considered to be the optimal approach to limit pathologies associated with obesity. While many current antiobesity strategies are centered on regulating satiety, these approaches typically attempt an overall weight loss and are unable to target distinct adipose depots specifically associated with disease risk. The authors report a novel therapeutic modality utilizing localized and sustained delivery of drugs to provide for the selective ablation of adipose tissue. Using the epididymal fat pad of Sprague-Dawley rats as a model, they injected into the tissue poly(lactide-co-glycolide) microspheres encapsulating tumor necrosis factor-alpha, a well-known regulator of adipose tissue mass. The utility of this approach was investigated in vivo by measuring the fat pad mass relative to the contralateral control within the same animal (n = 4 at each time point) and in vitro by measuring apoptosis in adipose organ cultures. The authors demonstrated control over the localization of tumor necrosis factor-alpha by performing blood analysis. This is the first report of localized drug delivery for adipose tissue ablation, and these results indicate the potential utility of the general tissue ablation approach for treatment of numerous pathologies.

Adipose Tissue↗
PubMed2002

External mechanical strain regulates membrane targeting of Rho GTPases by controlling microtubule assembly.

Transmission of externally applied mechanical forces to the interior of a cell requires coordination of biochemical signaling pathways with changes in cytoskeletal assembly and organization. In this study, we addressed one potential mechanism for this signal integration by applying uniform single external mechanical strains to aortic smooth muscle cells (SMCs) via their adhesion substrate. A tensile strain applied to the substrate for 15 min significantly increased microtubule (MT) assembly by 32 +/- 7%, with no apparent effect on the cells' focal adhesions as revealed by immunofluorescence and quantitative analysis of Triton X-100-insoluble vinculin levels. A compressive strain decreased MT mass by 24 +/- 9% but did not influence the level of vinculin in focal adhesions. To understand the decoupling of these two cell responses to mechanical strain, we examined a redistribution of the small GTPases RhoA and Rac. Tensile strain was found to decrease the amount of membrane-associated RhoA and Rac by 70 +/- 9% and 45 +/- 11%, respectively, compared with static controls. In contrast, compressive strain increased membrane-associated RhoA and Rac levels by 74 +/- 17% and 36 +/- 13%, respectively. Disruption of the MT network by prolonged treatments with low doses of either nocodazole or paclitaxel before the application of strain abolished the redistribution of RhoA and Rac in response to the applied forces. Combined, these results indicate that the effects of externally applied mechanical strain on the distribution and activation of the Rho family GTPases require changes in the state of MT polymerization.

Animals↗
PubMed2002

Engineering growing tissues.

Regenerating or engineering new tissues and organs may one day allow routine replacement of lost or failing tissues and organs. However, these engineered tissues must not only grow to fill a defect and integrate with the host tissue, but often they must also grow in concert with the changing needs of the body over time. We hypothesized that tissues capable of growing with time could be engineered by supplying growth stimulus signals to cells from the biomaterial used for cell transplantation. In this study, chondrocytes and osteoblasts were cotransplanted on hydrogels modified with an RGD-containing peptide sequence to promote cell multiplication. New bone tissue was formed that grew in mass and cellularity by endochondral ossification in a manner similar to normal long-bone growth. Transplanted cells organized into structures that morphologically and functionally resembled growth plates. These engineered tissues could find utility in treating diseases and injuries of the growth plate, testing the effect of experimental drugs on growth-plate function and development, and investigating the biology of long-bone growth. Furthermore, this concept of promoting the growth of engineered tissues could find great utility in engineering numerous tissue types by way of the transplantation of a small number of precursor cells.

Alginates↗
PubMed2002

Engineering vascular networks in porous polymer matrices.

Enhanced vascularization is critical to the treatment of ischemic tissues and the engineering of new tissues and organs. We have investigated whether sustained and localized delivery of vascular endothelial growth factor (VEGF) combined with transplantation of human microvascular endothelial cells (HMVECs) can be used to engineer new vascular networks. VEGF was incorporated and released in a sustained manner from porous poly(lactic-co-glycolic acid) (PLG) matrices to promote angiogenesis at the transplantation site. VEGF could be incorporated and released in a biologically active form from PLG matrices, with the majority of VEGF release (64%) occurring within 2 weeks. These matrices promoted a 260% increase in the density of host SCID mouse-derived capillaries invading the matrices after 7 days of implantation, confirming the activity of the released VEGF. HMVECs were transplanted into SCID mice on PLG matrices, and organized to form immature human-derived vessels within 3 days. Functional vessels were observed within 7 days. Importantly, when HMVECs were transplanted on VEGF-releasing matrices, a 160% increase in the density of human-derived blood vessels was observed after 14 days. These findings suggest that combining elements of vasculogenesis and angiogenesis provides a viable and novel approach to enhancing local vascularization.

Animals↗
PubMed2002

Bioinspired growth of crystalline carbonate apatite on biodegradable polymer substrata.

Mineralization in biological systems is a widespread, yet incompletely understood phenomenon involving complex interactions at the biomacromolecule-mineral nucleus interface. This study was aimed at understanding and controlling mineral formation in a poly(alpha-hydroxy ester) model system, to gain insight into biological mineralization processes and to develop biomaterials for orthopaedic tissue regeneration. We specifically hypothesized that providing a high surface density of anionic functional groups would enhance nucleation and growth of bonelike mineral following exposure to simulated body fluids (SBF). Polymer surface functionalization was achieved via hydrolysis of 85:15 poly(lactide-co-glycolide) (PLG) films. This treatment led to an increase in surface carboxylic acid and hydroxyl groups, resulting in a substantial increase in polymer surface energy from 42 to 49 dynes/cm2. Treated polymers exhibited a 3-fold increase in heterogeneous mineral grown and growth of a continuous mineral film on the polymer surface. The mineral grown on PLG surfaces is a carbonate apatite, the major mineral component of vertebrate bone tissue. Mineral crystal size and morphology were dependent on the solution characteristics but unaffected by the degree of surface prehydrolysis. The mechanism of heterogeneous carbonate apatite growth was examined via ion binding assays, which indicated that calcium binding is mediated independently by the presence of soluble phosphate counterions and surface functional groups. These findings indicate that poly(alpha-hydroxy ester) materials can be readily mineralized using a biomimetic process, and that the impetus for mineral nucleation in this system appears more complicated than the simple electrostatic interactions proposed in previous biomineralization theory.

Apatites↗
PubMed2002

Alginate type and RGD density control myoblast phenotype.

Alginates are being increasingly used for cell encapsulation and tissue engineering applications; however, these materials cannot specifically interact with mammalian cells. We have covalently modified alginates of varying monomeric ratio with RGD-containing cell adhesion ligands using carbodiimide chemistry to initiate cell adhesion to these polymers. We hypothesized that we could control the function of cells adherent to RGD-modified alginate hydrogels by varying alginate polymer type and cell adhesion ligand density, and we have addressed this possibility by studying the proliferation and differentiation of C2C12 skeletal myoblasts adherent to these materials. RGD density on alginates of varying monomeric ratio could be controlled over several orders of magnitude, creating a range of surface densities from 1-100 fmol/cm(2). Myoblast adhesion to these materials was specific to the RGD ligand, because adhesion could be competed away with soluble RGD in a dose-dependent manner. Myoblast proliferation and differentiation could be regulated by varying the alginate monomeric ratio and the density of RGD ligands at the substrate surface, and specific combinations of alginate type and RGD density were required to obtain efficient myoblast differentiation on these materials.

Alginates↗
PubMed2002

Salt fusion: an approach to improve pore interconnectivity within tissue engineering scaffolds.

Macroporous scaffolds composed of biodegradable polymers have found extensive use as three-dimensional substrates either for in vitro cell seeding followed by transplantation, or as conductive substrates for direct implantation in vivo. Methods abound for creation of macroporous scaffolds for tissue engineering, and common methods typically employ a solid porogen within a three-dimensional polymer matrix to create a well-defined pore size, pore structure, and total scaffold porosity. This study describes an approach to impart improved pore interconnectivity to polymer scaffolds for tissue engineering by partially fusing the solid porogen together prior to creation of a continuous polymer matrix. Three dimensional, porous scaffolds of the copolymer 85:15 poly(lactide-co-glycolide) were fabricated via either a solvent casting/particulate leaching process, or a gas foaming/particulate leaching process. Prior to creation of a continuous polymer matrix the NaCl crystals, which serve as the solid porogen, are partially fused via treatment in 95% humidity. Scanning electron micrographs clearly display fused salt crystals and an enhancement in pore interconnectivity in the salt fused scaffolds prepared via both solvent casting and gas foaming, and the extent of pore interconnectivity is enhanced with longer treatment times. Fusion of salt crystal for 24 h increased the radius of curvature of salt crystals, and led to a twofold increase in the compressive modulus of solvent cast scaffolds (total porosity of 97 +/- 1%). Fusion of NaCl crystals prior to gas foaming resulted in a decrease in scaffold compressive modulus from 277 +/- 60k Pa to 187 +/- 30k Pa (total porosity of 94 +/- 1%). The resulting highly interconnected scaffolds have implications for facilitated cell migration, abundant cell-cell interaction, and potentially improved neural and vascular growth within tissue engineering scaffolds.

Biocompatible Materials↗
PubMed2002

Tissue engineering strategies for in vivo neovascularisation.

Neovascularisation is a promising alternative therapeutic approach to re-establish blood flow in ischaemic tissues of patients suffering from coronary artery or peripheral artery disease. Often, these patients are not suitable candidates for current revascularisation procedures such as coronary angioplasty or bypass surgery. Several strategies are presently under investigation to induce vascularisation by stimulating the body's natural processes of vasculogenesis, angiogenesis and arteriogenesis. These strategies involve transplantation of various cell types into the ischaemic site and the delivery of recombinant angiogenic agents through direct protein administration or gene transfer. We will examine the basic approaches for these neovacularisation strategies and their therapeutic potential as demonstrated in animal models and human trials to date.

Animals↗