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Biomedical subjects

D J Mooney

Publications and source records attributed to D J Mooney.

At least 19 recordsLinked to original sources
PubMed2007

Spatiotemporal control of vascular endothelial growth factor delivery from injectable hydrogels enhances angiogenesis.

Therapeutic angiogenesis with vascular endothelial growth factor (VEGF) delivery may provide a new approach for the treatment of ischemic diseases, but current strategies to deliver VEGF rely on either bolus delivery or systemic administration, resulting in limited clinical utility, because of the short half-life of VEGF in vivo and its resultant low and transient levels at sites of ischemia. We hypothesize that an injectable hydrogel system can be utilized to provide temporal control and appropriate spatial biodistribution of VEGF in ischemic hindlimbs. A sustained local delivery of relatively low amounts of bioactive VEGF (3 mug) with this system led to physiologic levels of bioactive VEGF in ischemic murine (ApoE(-/-)) hindlimbs for 15 days after injection of the gel, as contrasted with complete VEGF deprivation after 72 h with bolus injection. The gel delivery system resulted in significantly greater angiogenesis in these limbs as compared to bolus (266 vs. 161 blood vessels mm(-2)). Laser Doppler perfusion imaging showed return of tissue perfusion to normal levels by day 28 with the gel system, whereas normal levels of perfusion were never achieved with saline delivery of VEGF or in control mice. The system described in this article could represent an attractive new generation of therapeutic delivery vehicle for treatment of cardiovascular diseases, as it combines long-term in vivo therapeutic benefit (localized bioactive VEGF for 1-2 weeks) with minimally invasive delivery.

Alginates↗
PubMed2006

Transplanted endothelial cells enhance orthotopic bone regeneration.

The aim of this study was to determine if endothelial cells could enhance bone marrow stromal-cell-mediated bone regeneration in an osseous defect. Using poly-lactide-co-glycolide scaffolds as cell carriers, we transplanted bone marrow stromal cells alone or with endothelial cells into 8.5-mm calvarial defects created in nude rats. Histological analyses of blood vessel and bone formation were performed, and microcomputed tomography (muCT) was used to assess mineralized bone matrix. Though the magnitude of the angiogenic response between groups was the same, muCT analysis revealed earlier mineralization of bone in the co-transplantation condition. Ultimately, there was a significant increase (40%) in bone formation in the co-transplantation group (33 +/- 2%), compared with the transplantation of bone marrow stromal cells alone (23 +/- 3%). Analysis of these data demonstrates that, in an orthotopic site, transplanted endothelial cells can influence the bone-regenerative capacity of bone marrow stromal cells.

Absorbable Implants↗
PubMed2005

Bone regeneration in a rat cranial defect with delivery of PEI-condensed plasmid DNA encoding for bone morphogenetic protein-4 (BMP-4).

Gene therapy approaches to bone tissue engineering have been widely explored. While localized delivery of plasmid DNA encoding for osteogenic factors is attractive for promoting bone regeneration, the low transfection efficiency inherent with plasmid delivery may limit this approach. We hypothesized that this limitation could be overcome by condensing plasmid DNA with nonviral vectors such as poly(ethylenimine) (PEI), and delivering the plasmid DNA in a sustained and localized manner from poly(lactic-co-glycolic acid) (PLGA) scaffolds. To address this possibility, scaffolds delivering plasmid DNA encoding for bone morphogenetic protein-4 (BMP-4) were implanted into a cranial critical-sized defect for time periods up to 15 weeks. The control conditions included no scaffold (defect left empty), blank scaffolds (no delivered DNA), and scaffolds encapsulating plasmid DNA (non-condensed). Histological and microcomputed tomography analysis of the defect sites over time demonstrated that bone regeneration was significant at the defect edges and within the defect site when scaffolds encapsulating condensed DNA were placed in the defect. In contrast, bone formation was mainly confined to the defect edges within scaffolds encapsulating plasmid DNA, and when blank scaffolds were used to fill the defect. Histomorphometric analysis revealed a significant increase in total bone formation (at least 4.5-fold) within scaffolds incorporating condensed DNA, relative to blank scaffolds and scaffolds incorporating uncondensed DNA at each time point. In addition, there was a significant increase both in osteoid and mineralized tissue density within scaffolds incorporating condensed DNA, when compared with blank scaffolds and scaffolds incorporating uncondensed DNA, suggesting that delivery of condensed DNA led to more complete mineralized tissue regeneration within the defect area. This study demonstrated that the scaffold delivery system encapsulating PEI-condensed DNA encoding for BMP-4 was capable of enhancing bone formation and may find applications in other tissue types.

Animals↗
PubMed2005

Actively regulating bioengineered tissue and organ formation.

OBJECTIVES: Describe current and future approaches to tissue engineering, specifically in the area of bone regeneration. These approaches will allow one to actively regulate the cellular populations participating in this process. DESIGN: Many approaches to actively regulate cellular phenotype are under exploration, and these typically exploit known signal transduction pathways via presentation of specific receptor-binding ligands, and may also deliver mechanical information via the physical bridge formed by the receptor-ligand interactions. Cellular gene expression may also be directly modulated utilizing gene therapy approaches to control tissue regeneration. CONCLUSIONS: Significant progress has been made to date in bone regeneration using inductive molecules and transplanted cells, and FDA approved therapies have resulted. While approaches to date have focused on delivery of single stimuli (e.g. one growth factor), future efforts will likely attempt to more closely mimic developmental processes by the delivery of multiple inputs to the cells in spatially and temporally regulated fashions.

Biocompatible Materials↗
PubMed2004

Bone regeneration via a mineral substrate and induced angiogenesis.

Angiogenesis and biomineral substrates play major roles in bone development and regeneration. We hypothesized that macroporous scaffolds of biomineralized 85:15 poly(lactide-co-glycolide), which locally release vascular endothelial growth factor-165 (VEGF), would direct simultaneous regeneration of bone and vascular tissue. The presence of a bone-like biomineral substrate significantly increased regeneration of osteoid matrix (32 +/- 7% of total tissue area; mean +/- SD; p < 0.05) and mineralized tissue (14 +/- 2%; P < 0.05) within a rat cranium critical defect compared with a non-mineralized polymer scaffold (19 +/- 8% osteoid and 10 +/- 2% mineralized tissue). Further, the addition of VEGF to a mineralized substrate significantly increased the generation of mineralized tissue (19 +/- 4%; P < 0.05) compared with mineralized substrate alone. This appeared to be due to a significant increase in vascularization throughout VEGF-releasing scaffolds (52 +/- 9 vessels/mm(2); P < 0.05) compared with mineralized scaffolds without VEGF (34 +/- 4 vessels/mm(2)). Surprisingly, there was no significant difference in total osteoid between the two samples, suggesting that increased vascularization enhances mineralized tissue generation, but not necessarily osteoid formation. These results indicate that induced angiogenesis can enhance tissue regeneration, supporting the concept of therapeutic angiogenesis in tissue-engineering strategies.

Analysis of Variance↗
PubMed2003

Regulating bone formation via controlled scaffold degradation.

It is widely assumed that coupling the degradation rate of polymers used as cell transplantation carriers to the growth rate of the developing tissue will improve its quantity or quality. To test this hypothesis, we developed alginate hydrogels with a range of degradation rates by gamma-irradiating high-molecular-weight alginate to yield polymers of various molecular weights and structures. Decreasing the size of the polymer chains increased the degradation rate in vivo, as measured by implant retrieval rates, masses, and elastic moduli. Rapidly and slowly degrading alginates, covalently modified with RGD-containing peptides to control cell behavior, were then used to investigate the effect of biodegradation rate on bone tissue development in vivo. The more rapidly degrading gels led to dramatic increases in the extent and quality of bone formation. These results indicate that biomaterial degradability is a critical design criterion for achieving optimal tissue regeneration with cell transplantation.

Absorbable Implants↗
PubMed2002

Tissue compatibility of two biodegradable tubular scaffolds implanted adjacent to skin or buccal mucosa in mice.

Radiation therapy for cancer in the head and neck region leads to a marked loss of salivary gland parenchyma, resulting in a severe reduction of salivary secretions. Currently, there is no satisfactory treatment for these patients. To address this problem, we are using both tissue engineering and gene transfer principles to develop an orally implantable, artificial fluid-secreting device. In the present study, we examined the tissue compatibility of two biodegradable substrata potentially useful in fabricating such a device. We implanted in Balb/c mice tubular scaffolds of poly-L-lactic acid (PLLA), poly-glycolic acid coated with PLLA (PGA/PLLA), or nothing (sham-operated controls) either beneath the skin on the back, a site widely used in earlier toxicity and biocompatibility studies, or adjacent to the buccal mucosa, a site quite different functionally and immunologically. At 1, 3, 7, 14, and 28 days postimplantation, implant sites were examined histologically, and systemic responses were assessed by conventional clinical chemistry and hematology analyses. Inflammatory responses in the connective tissue were similar regardless of site or type of polymer implant used. However, inflammatory reactions were shorter and without epithelioid and giant cells in sham-operated controls. Also, biodegradation proceeded more slowly with the PLLA tubules than with the PGA/PLLA tubules. No significant changes in clinical chemistry and hematology were seen due to the implantation of tubular scaffolds. These results indicate that the tissue responses to PLLA and PGA/PLLA scaffolds are generally similar in areas subjacent to skin in the back and oral cavity. However, these studies also identified several potentially significant concerns that must be addressed prior to initiating any clinical applications of this device.

Animals↗
PubMed2001

Injection molding of chondrocyte/alginate constructs in the shape of facial implants.

Over one million patients per year undergo some type of procedure involving cartilage reconstruction. Polymer hydrogels, such as alginate, have been shown to be effective carriers for chondrocytes in subcutaneous cartilage formation. The goal of our current study was to develop a method to create complex structures (nose bridge, chin, etc.) with good dimensional tolerance to form cartilage in specific shapes. Molds of facial implants were prepared using Silastic ERTV. Suspensions of chondrocytes in 2% alginate were gelled by mixing with CaSO(4) (0.2 g/mL) and injected into the molds. Constructs of various cell concentrations (10, 25, and 50 million/mL) were implanted in the dorsal aspect of nude mice and harvested at times up to 30 weeks. Analysis of implanted constructs indicated progressive cartilage formation with time. Proteoglycan and collagen constructs increased with time to approximately 60% that of native tissue. Equilibrium modulus likewise increased with time to 15% that of normal tissue, whereas hydraulic permeability decreased to 20 times that of native tissue. Implants seeded with greater concentrations of cells increased proteoglycan content and collagen content and equilibrium and decreased permeability. Production of shaped cartilage implants by this technique presents several advantages, including good dimensional tolerance, high sample-to-sample reproducibility, and high cell viability. This system may be useful in the large-scale production of precisely shaped cartilage implants.

Absorbable Implants↗
PubMed2001

Up-Regulation of Bcl-2 in microvascular endothelial cells enhances intratumoral angiogenesis and accelerates tumor growth.

Vascular endothelial growth factor (VEGF) has been shown to be a potent mediator of angiogenesis that functions as a survival factor for endothelial cells by up-regulating Bcl-2 expression. We have recently reported that human dermal microvascular endothelial cells (HDMECs) seeded in biodegradable sponges and implanted into severe combined immunodeficient (SCID) mice organize into functional human microvessels that transport mouse blood cells. In this study, we implanted sponges seeded with OSCC-3 (oral squamous cell carcinoma) or SLK (Kaposi's sarcoma) together with endothelial cells into SCID mice to generate human tumors vascularized with human microvessels. This model system was used to examine the role of both endothelial cell Bcl-2 and the proangiogenic chemokine interleukin-8 (IL-8) on tumor growth and intratumoral microvascular density. Coimplantation of HDMECs overexpressing Bcl-2 (HDMEC-Bcl-2) and tumor cells resulted in a 3-fold enhancement of tumor growth when compared with the coimplantation of control HDMECs and tumor cells. This was associated with increased intratumoral microvascular density and enhanced endothelial cell survival. To determine whether the enhanced neovascularization mediated by Bcl-2 overexpression in endothelial cells was influenced by the synthesis of endogenous mediators of angiogenesis, we screened these cells for expression of VEGF, basic fibroblast growth factor (bFGF), and IL-8 by ELISA. HDMEC-Bcl-2 cells and VEGF-treated HDMECs exhibited a 15-fold and 4-fold increase, respectively, in the expression of the proangiogenic chemokine IL-8 in vitro, whereas the expression of VEGF and bFGF remained unchanged. Transfection of antisense Bcl-2 into HDMECs blocked VEGF-mediated induction of IL-8. Conditioned media from HDMEC-Bcl-2 induced proliferation and sprouting of endothelial cells in vitro and neovascularization in rat corneas. Anti-IL-8 antibody added to HDMEC-Bcl-2 conditioned media markedly reduced the potency of these responses. SCID mice bearing VEGF-producing tumor implants that were treated with anti-lL-8 antibody exhibited a 43% reduction in microvessel density and a 50% reduction in tumor weight compared with treatment with a nonspecific antibody. These results demonstrate that the up-regulation of Bcl-2 expression in endothelial cells that constitute tumor microvessels enhances intratumoral microvascular survival and density and accelerates tumor growth. Furthermore, endothelial cells that overexpress Bcl-2 have more angiogenic potential than control cells, and IL-8-neutralizing antibodies attenuate their angiogenic activity in vitro and in vivo.

Animals↗
PubMed2001

Degradable and injectable poly(aldehyde guluronate) hydrogels for bone tissue engineering.

Degradable and injectable hydrogels may be ideal for bone-tissue engineering, especially in the craniofacial region because of the ease of access for injection. Alginate hydrogels potentially could be used as injectable cell delivery vehicles, but they exhibit a limited range of mechanical properties and uncontrollable disintegration time. Therefore we synthesized new hydrogels, composed of poly(aldehyde guluronate) (PAG) and adipic acid dihydrazide, that have a wide range of mechanical stiffness and controllable degradation rate. MC3T3-E1 cells adhered and multiplied on PAG hydrogels in vitro. When primary rat calvarial osteoblasts were mixed with PAG hydrogels and subcutaneously injected into the backs of mice, mineralized bone tissues were formed 9 weeks following implantation. These hydrogels may find wide utility as an injectable delivery system for bone precursor cells as well as for other applications in tissue engineering.

3T3 Cells↗
PubMed2001

Hydrogels for combination delivery of antineoplastic agents.

The systemic delivery of anticancer agents has been widely investigated during the past decade but localized delivery may offer a safer and more effective delivery approach. We have designed and synthesized a novel hydrogel to locally deliver antineoplastic agents, and demonstrate the different types of release that can be achieved from these hydrogels using three model drugs: methotrexate, doxorubicin, and mitoxantrone. Alginate was chemically modified into low molecular weight oligomers and cross-linked with a biodegradable spacer (adipic dihydrazide) to form biodegradable hydrogels. The model antineoplastic agents were loaded into the hydrogel via three different mechanisms. Methotrexate was incorporated within the pores of the hydrogel and was released by diffusion into the surrounding medium. Doxorubicin was covalently attached to the polymer backbone via a hydrolytically labile linker and was released following the chemical hydrolysis of the linker. Mitoxantrone was ionically complexed to the polymer and was released after the dissociation of this complex. These three release mechanisms could potentially be used to deliver a wide selection of antineoplastic agents, based on their chemical structure. This novel delivery system allows for the release of single or combinations of antineoplastic agents, and may find utility in localized antineoplastic agent delivery.

Alginates↗
PubMed2001

Engineering and characterization of functional human microvessels in immunodeficient mice.

SUMMARY: Current model systems used to investigate angiogenesis in vivo rely on the interpretation of results obtained with nonhuman endothelial cells. Recent advances in tissue engineering and molecular biology suggest the possibility of engineering human microvessels in vivo. Here we show that human dermal microvascular endothelial cells (HDMEC) transplanted into severe combined immunodeficient (SCID) mice on biodegradable polymer matrices differentiate into functional human microvessels that anastomose with the mouse vasculature. HDMEC were stably transduced with Flag epitope or alkaline phosphatase to confirm the human origin of the microvessels. Endothelial cells appeared dispersed throughout the sponge 1 day after transplantation, became organized into empty tubular structures by Day 5, and differentiated into functional microvessels within 7 to 10 days. Human microvessels in SCID mice expressed the physiological markers of angiogenesis: CD31, CD34, vascular cellular adhesion molecule 1 (VCAM-1), and intercellular adhesion molecule 1 (ICAM-1). Human endothelial cells became invested by perivascular smooth muscle alpha-actin-expressing mouse cells 21 days after implantation. This model was used previously to demonstrate that overexpression of the antiapoptotic protein Bcl-2 in HDMEC enhances neovascularization, and that apoptotic disruption of tumor microvessels is associated with apoptosis of surrounding tumor cells. The proposed SCID mouse model of human angiogenesis is ideally suited for the study of the physiology of microvessel development, pathologic neovascular responses such as tumor angiogenesis, and for the development and investigation of strategies designed to enhance the neovascularization of engineered human tissues and organs.

Absorbable Implants↗
PubMed2001

Polymeric system for dual growth factor delivery.

The development of tissues and organs is typically driven by the action of a number of growth factors. However, efforts to regenerate tissues (e.g., bone, blood vessels) typically rely on the delivery of single factors, and this may partially explain the limited clinical utility of many current approaches. One constraint on delivering appropriate combinations of factors is a lack of delivery vehicles that allow for a localized and controlled delivery of more than a single factor. We report a new polymeric system that allows for the tissue-specific delivery of two or more growth factors, with controlled dose and rate of delivery. The utility of this system was investigated in the context of therapeutic angiogenesis. We now demonstrate that dual delivery of vascular endothelial growth factor (VEGF)-165 and platelet-derived growth factor (PDGF)-BB, each with distinct kinetics, from a single, structural polymer scaffold results in the rapid formation of a mature vascular network. This is the first report of a vehicle capable of delivery of multiple angiogenic factors with distinct kinetics, and these results clearly indicate the importance of multiple growth factor action in tissue regeneration and engineering.

Actins↗
PubMed2001

Using HSV-thymidine kinase for safety in an allogeneic salivary graft cell line.

Extreme salivary hypofunction is a result of tissue damage caused by irradiation therapy for cancer in the head and neck region. Unfortunately, there is no currently satisfactory treatment for this condition that affects up to 40,000 people in the United States every year. As a novel approach to managing this problem, we are attempting to develop an orally implantable, fluid-secreting device (an artificial salivary gland). We are using the well-studied HSG salivary cell line as a potential allogeneic graft cell for this device. One drawback of using a cell line is the potential for malignant transformation. If such an untoward response occurred, the device could be removed. However, in the event that any HSG cells escaped, we wished to provide additional patient protection. Accordingly, we have engineered HSG cells with a hybrid adeno-retroviral vector, AdLTR.CMV-tk, to express the herpes simplex virus thymidine kinase (HSV-tk) suicide gene as a novel safety factor. Cells were grown on plastic plates or on poly-L-lactic acid disks and then transduced with different multiplicities of infection (MOIs) of the hybrid vector. Thereafter, various concentrations of ganciclovir (GCV) were added, and cell viability was tested. Transduced HSG cells expressed HSV-tk and were sensitive to GCV treatment. Maximal effects were seen at a MOI of 10 with 50 microM of GCV, achieving 95% cell killing on the poly-L-lactic acid substrate. These results suggest that engineering the expression of a suicide gene in an allogeneic graft cell may provide additional safety for use in an artificial salivary gland device.

Artificial Organs↗
PubMed2001

Control of microtubule assembly by extracellular matrix and externally applied strain.

A number of studies have suggested that externally applied mechanical forces and alterations in the intrinsic cell-extracellular matrix (ECM) force balance equivalently induce changes in cell phenotype. However, this possibility has never been directly tested. To test this hypothesis, we directly investigated the response of the microtubule (MT) cytoskeleton in smooth muscle cells to both mechanical signals and alterations in the ECM. A tensile force that resulted in a positive 10% step change in substrate strain increased MT mass by 34 +/- 10% over static controls, independent of the cell adhesion ligand and tyrosine phosphorylation. Conversely, a compressive force that resulted in a negative 10% step change in substrate strain decreased MT mass by 40 +/- 6% over static controls. In parallel, increasing the density of the ECM ligand fibronectin from 50 to 1,000 ng/cm(2) in the absence of any applied force increased the amount of polymeric tubulin in the cell from 59 +/- 11% to 81 +/- 13% of the total cellular tubulin. These data are consistent with a model in which MT assembly is, in part, controlled by forces imposed on these structures, and they suggest a novel control point for MT assembly by altering the intrinsic cell-ECM force balance and applying external mechanical forces.

Animals↗
PubMed2001

Cell-interactive alginate hydrogels for bone tissue engineering.

There is significant interest in the development of injectable carriers for cell transplantation to engineer bony tissues. In this study, we hypothesized that adhesion ligands covalently coupled to hydrogel carriers would allow one to control pre-osteoblast cell attachment, proliferation, and differentiation. Modification of alginate with an RGD-containing peptide promoted osteoblast adhesion and spreading, whereas minimal cell adhesion was observed on unmodified hydrogels. Raising the adhesion ligand density increased osteoblast proliferation, and a minimum ligand density (1.5-15 femtomoles/cm2) was needed to elicit this effect. MC3T3-E1 cells demonstrated increased osteoblast differentiation with the peptide-modified hydrogels, as confirmed by the up-regulation of bone-specific differentiation markers. Further, transplantation of primary rat calvarial osteoblasts revealed statistically significant increases of in vivo bone formation at 16 and 24 weeks with G4RGDY-modified alginate compared with unmodified alginate. These findings demonstrate that biomaterials may be designed to control bone development from transplanted cells.

3T3 Cells↗
PubMed2001

Craniofacial tissue engineering.

There is substantial need for the replacement of tissues in the craniofacial complex due to congenital defects, disease, and injury. The field of tissue engineering, through the application of engineering and biological principles, has the potential to create functional replacements for damaged or pathologic tissues. Three main approaches to tissue engineering have been pursued: conduction, induction by bioactive factors, and cell transplantation. These approaches will be reviewed as they have been applied to key tissues in the craniofacial region. While many obstacles must still be overcome prior to the successful clinical restoration of tissues such as skeletal muscle and the salivary glands, significant progress has been achieved in the development of several tissue equivalents, including skin, bone, and cartilage. The combined technologies of gene therapy and drug delivery with cell transplantation will continue to increase treatment options for craniofacial cosmetic and functional restoration.

Adipose Tissue↗