Safety and efficacy of ultraviolet-a light-activated gene transduction for gene therapy of articular cartilage defects.

Background: Gene therapies for articular cartilage defects are limited by the absence of an in vivo delivery system that can mediate site-specific transduction restricted to within the margins of the defect during routine arthroscopy. We have proposed the use of ultraviolet light to stimulate gene expression following infection by recombinant adeno-associated virus (rAAV). However, research has demonstrated that short-wavelength ultraviolet light (ultraviolet C), while effective, is neither safe nor practical for this purpose.

Remodeling of cortical bone allografts mediated by adherent rAAV-RANKL and VEGF gene therapy.

Structural allograft healing is limited because of a lack of vascularization and remodeling. To study this we developed a mouse model that recapitulates the clinical aspects of live autograft and processed allograft healing. Gene expression analyses showed that there is a substantial decrease in the genes encoding RANKL and VEGF during allograft healing.

Primary murine limb bud mesenchymal cells in long-term culture complete chondrocyte differentiation: TGF-beta delays hypertrophy and PGE2 inhibits terminal differentiation.

In vitro models of endochondral bone formation using both primary and immortalized cells have provided insight regarding factors and signaling pathways involved in chondrocyte maturation and endochondral bone formation. However, primary murine cell culture models of chondrocyte differentiation have not been established but have enormous potential due to the possible use of cells from transgenic and knockout animals. Here, we show that stage E11.

Light-activated gene transduction of recombinant adeno-associated virus in human mesenchymal stem cells.

Deficiencies in skeletal tissue repair and regeneration lead to conditions like osteoarthritis, osteoporosis and degenerative disc disease. While no cure for these conditions is available, the use of human bone marrow derived-mesenchymal stem cells (HuMSCs) has been shown to have potential for cell-based therapy. Furthermore, recombinant adeno-associated viruses (rAAV) could be used together with HuMSCs for in vivo or ex vivo gene therapy.

NFkappaB: a pivotal transcription factor in prostate cancer metastasis to bone.

Prostate cancers frequently metastasize to bone and this accounts for substantial morbidity. We investigated the potential role of the transcription factor NFkappaB as a central regulator of prostate cancer metastasis using the prostate adenocarcinoma cell line, PC-3, in a series of in vitro studies. Wild type PC-3 cells (PC-3.

Adeno-associated virus-mediated osteoprotegerin gene transfer protects against particulate polyethylene-induced osteolysis in a murine model.

Objective: Osteoprotegerin (OPG), a natural negative regulator of osteoclastogenesis and bone resorption, may be a potential therapeutic agent for treatment of osteolysis-associated prosthetic joint loosening. Using an in vivo adeno-associated virus (AAV)-mediated gene transfer technique, this study was designed to evaluate the protective effects of OPG transgene against orthopedic wear debris-induced bone loss in a murine model of osteolysis.

Methods: Bone tissue was implanted into established pouches on BALB/c mice, followed by the introduction of ultra-high-molecular-weight polyethylene (UHMWPE) particles to provoke inflammation and osteolysis.

Light-activated gene transduction enhances adeno-associated virus vector-mediated gene expression in human articular chondrocytes.

Objective: To evaluate the effects of ultraviolet (UV) light as an adjuvant for recombinant adeno-associated virus (rAAV) transduction in human articular chondrocytes.

Methods: Primary articular chondrocytes and immortalized chondrocytes (tsT/AC62) were exposed to various doses of UV light (0-1,000 J/m(2)) and infected at various multiplicities of infection (MOIs) with rAAV containing the enhanced green fluorescent protein (EGFP) gene. Cells were analyzed for viability and EGFP expression by fluorescence-activated cell sorting on days 2, 4, and 8 following infection.

Recombinant adeno-associated virus-mediated osteoprotegerin gene therapy inhibits wear debris-induced osteolysis.

Background: Aseptic loosening of orthopaedic implants secondary to wear debris-induced osteolysis is a serious problem. Osteoprotegerin (OPG) is a natural decoy protein that inhibits osteoclast activation and bone resorption. This study investigated whether gene therapy using a recombinant adeno-associated viral vector that expresses OPG can inhibit wear debris-induced osteolysis.

Efficacy of ex vivo OPG gene therapy in preventing wear debris induced osteolysis.

Aseptic loosening of prosthetic implants remains a serious orthopaedic problem and the greatest limitation to total joint arthroplasty. Central to the etiology of aseptic loosening is periprosthetic osteolysis at the bone-implant interface, which is caused by wear debris-induced inflammation. This inflammation produces the critical osteoclast differentiation factor RANKL, which directly stimulates osteoclastogenesis and osteoclastic bone resorption.

Effect of anti-tumor necrosis factor-alpha gene therapy on wear debris-induced osteolysis.

Background: Particle phagocytosis by macrophages induces the secretion of tumor necrosis factor-alpha, which is involved in the development of an osteolytic response. Therefore, we aimed to determine whether gene delivery of a soluble inhibitor of tumor necrosis factor-alpha (sTNFR:Fc) could prevent wear debris-induced osteolysis in a mouse model. sTNFR:Fc is a fusion protein containing the extracellular domain of the human type-I tumor necrosis factor receptor fused to the Fc region of mouse immunoglobulin.

Efficacy of etanercept for wear debris-induced osteolysis.

A major limitation of total joint arthroplasty is that up to 20% of patients require revision surgery to correct prosthetic loosening. Aseptic loosening is believed to result from the phagocytosis of wear debris particles by macrophages, which secrete proinflammatory cytokines that stimulate osteolysis. Tumor necrosis factor alpha (TNF-alpha) has been shown to be one of the prominent cytokines in this cascade and to be involved critically in the generation of particle-induced osteolysis.

Quantitative small-animal surrogate to evaluate drug efficacy in preventing wear debris-induced osteolysis.

Individuals who suffer from severe joint destruction caused by the various arthritidies often undergo total joint arthroplasty. A major limitation of this treatment is the development of aseptic loosening of the prosthesis in as many as 20% of patients. The current paradigm to explain aseptic loosening proposes that wear debris generated from the prosthesis initiates a macrophage-mediated inflammatory response by resident macrophages, leading to osteoclast activation and bone resorption at the implant interface.

Tumor necrosis factor-alpha/nuclear transcription factor-kappaB signaling in periprosthetic osteolysis.

Due to irreversible joint destruction caused by the various arthritides, more than 400,000 total joint arthroplasties are performed each year in the United States. As many as 20% of these require revision surgery because of aseptic loosening. The current paradigm to explain aseptic loosening is that wear debris generated from the prosthesis stimulates the release of proinflammatory cytokines (i.

Empirical advantages of adeno associated viral vectors in vivo gene therapy for arthritis.

Objective: To evaluate the utility of the adeno associated viral (AAV) vector for gene delivery to joint cells in vivo and in vitro, and to assess its potential as a vector for arthritis gene therapy.

Methods: A recombinant AAV (rAAV) vector expressing the bacterial beta-galactosidase (beta-gal) gene (rAAV-CMV-LacZ) was directly introduced into healthy-normal mouse knees, or arthritic knees in mice overexpressing tumor necrosis factor-alpha (hTNFalpha-Tg). Beta-gal expression levels were determined by immunohistochemistry and chemiluminescence.