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Biomedical subjects

C H Evans

Publications and source records attributed to C H Evans.

At least 73 records · Page 4Linked to original sources
PubMed2000

Nitric oxide inhibits chondrocyte response to IGF-I: inhibition of IGF-IRbeta tyrosine phosphorylation.

Chondrocytes in arthritic cartilage respond poorly to insulin-like growth factor I (IGF-I). Studies with inducible nitric oxide synthase (iNOS) knockout mice suggest that NO is responsible for part of the cartilage insensitivity to IGF-I. These studies characterize the relationship between NO and chondrocyte responses to IGF-I in vitro, and define a mechanism by which NO decreases IGF-I stimulation of chondrocyte proteoglycan synthesis. Lapine cartilage slices, chondrocytes, and cartilage from osteoarthritic (OA) human knees were exposed to NO from the donors S-nitroso-N-acetylpenicillamine (SNAP) or (Z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1- ium-1, 2-diolate] (DETA NONOate), by transduction with adenoviral transfer of iNOS (Ad-iNOS), or by activation with interleukin-1 (IL-1). NO synthesis was estimated from medium nitrite, and proteoglycan synthesis was measured as incorporation of (35)SO(4). IGF-I receptor phosphorylation was evaluated with Western analysis. SNAP, DETA NONOate, endogenously synthesized NO in Ad-iNOS-transduced cells, or IL-1 activation decreased IGF-I-stimulated proteoglycan synthesis in cartilage and monolayer cultures of chondrocytes. OA cartilage responded poorly to IGF-I; however, the response to IGF-I was restored by culture with N(G)-monomethyl-L-arginine (L-NMA). IGF-I receptor phosphotyrosine was diminished in chondrocytes exposed to NO. These studies show that NO is responsible for part of arthritic cartilage/chondrocyte insensitivity to anabolic actions of IGF-I; inhibition of receptor autophosphorylation is potentially responsible for this effect.

Animals↗
PubMed2000

Gene therapy for juvenile rheumatoid arthritis?

Gene therapy shows promise as a means of improving the treatment of a number of rheumatic diseases, including adult rheumatoid arthritis (RA). We summarize these developments and discuss the merits of extending such approaches to the treatment of juvenile RA. The special issues that arise when treating children by experimental new procedures predicate a cautious approach using strategies first shown to be safe in adults. Patients with severe, erosive, nonremitting pauciarticular juvenile RA might be suitable first candidates for gene therapy.

Arthritis, Juvenile↗
PubMed2000

Cartilage injury and repair.

Articular cartilage is a complex structure that, once damaged, has little capacity for permanent repair. The problem lies in the inability of the body to regenerate tissue with the appropriate macromolecular constituents and architecture of normal hyaline cartilage. Although full-thickness defects are capable of stimulating a repair response, the resulting fibrocartilage is inferior and cannot withstand long-term, repetitive use. Numerous surgical approaches that involve penetration of subchondral bone offer short-term to moderate-term relief of symptoms, whereas other approaches have seen significant improvement through transplantation of osteochondral and periosteal tissue and implantation of autologous chondrocytes. Despite these procedures, articular cartilage damage continues to be an unmet clinical problem. Improvements in biochemical and molecular biologic techniques may allow advances in the understanding of chondrocyte and cartilage biology and may provide innovative and novel approaches to stimulating the repair of articular cartilage through biologic means.

Animals↗
PubMed2000

Potential applications of gene therapy in sports medicine.

Gene therapy has the potential to improve greatly the management of sports medicine injuries. By the expeditious introduction of the appropriate genes to sites of injury, it should prove possible to initiate healing responses in those tissues that normally fail to heal, to accelerate healing in tissues which do naturally heal, and to improve the quality of the repair tissue. In addition, genes with anti-inflammatory properties offer novel possibilities for the control of chronic, sports related inflammation. None of these applications of gene therapy in sports medicine is yet in the clinic. It would seem, however, only a matter of time before the first clinical trials emerge. Their acceptance will be facilitated by the two human protocols that are presently underway for the treatment of rheumatoid arthritis. The present status of sports medicine gene therapy is summarized in Table 3.

Athletic Injuries↗
PubMed2000

The Marshall R. Urist Young Investigator Award. Orthopaedic applications of gene therapy. From concept to clinic.

Gene therapy offers new possibilities for the clinical management of orthopaedic conditions that are difficult to treat by traditional surgical or medical means. To bring the potential of this novel technology into the clinic, a research program was initiated that aimed to identify orthopaedically useful genes and develop methods for delivering them to suitable sites under conditions in which gene expression remains at therapeutic levels for the appropriate periods of time; this program is now 10 years old. Rheumatoid arthritis was selected as the lead disease. Preclinical studies evaluating the local and systemic delivery of numerous different genes by in vivo and ex vivo methods in murine and lapin models led to the development of a human gene therapy protocol for arthritis. In this protocol, a gene encoding the human interleukin-1 receptor antagonist protein is transferred to the metacarpophalangeal joints of female patients with rheumatoid arthritis. The first patient was treated this way in July 1996. This is not only the first orthopaedic application of human gene therapy, but also the first use of gene therapy approved for a nonlethal disease. In addition to providing additional therapeutic options for the treatment of rheumatoid arthritis, the experimental data from this study suggest that gene transfer approaches may improve the treatment of osteoarthritis, the repair of cartilage, ligaments, tendons, menisci, intervertebral discs and bone, and the management of disorders such as osteoporosis and osteogenesis imperfecta. They also show promise as a means for developing novel and improved animal models of orthopaedic diseases. If the current rate of progress continues, wide clinical application of gene therapy in various orthopaedic indications should occur within the next 5 to 10 years.

Animals↗
PubMed2000

Interaction of strain and interleukin-1 in articular cartilage: effects on proteoglycan synthesis in chondrocytes.

In temporomandibular joint disorders, the release of proinflammatory cytokines such as interleukin-1 (IL-1) initiates an inflammatory process disrupting cartilage homeostasis, ultimately leading to cartilage destruction. Additionally, mechanical stimuli affect articular chondrocyte metabolism. While articular chondrocytes generate nitric oxide (NO) in the presence of IL-1 proteoglycan synthesis is consecutively suppressed. The purpose of this study was to assess the effects of proinflammatory cytokines and mechanical strain in the form of cyclic tensile stretch on proteoglycan synthesis in chondrocytes, as compared to the NO competitive inhibitor L-N-monomethyl arginine (LMA), and to assess whether this effect is secondarily related to the activity of growth factors such as transforming growth factor beta (TGF-beta). Lapine articular chondrocytes were exposed to one of four different treatment regimens: no cyclic tensile stretch, IL-1, cyclic tensile stretch, or IL-1 plus cyclic tensile stretch. NO production was determined as medium nitrite accumulation. TGF-beta-bioactivity in chondrocyte conditioned medium was measured with the mink-lung epithelial cell bioassay. Proteoglycan synthesis was measured as the incorporation of 35-[S]-sodium sulfate into macromolecules separated from unincorporated label by gel filtration on PD-10 columns. In resting chondrocyte cultures, only baseline levels of NO were measured and the application of stretch for 24 h did not affect NO production. Addition of IL-1 provoked a large increase in NO synthesis which was abrogated in the presence of LMA. Application of stretch decreased the IL-1 induced NO synthesis, but did not modify the effect of LMA (being a competitive inhibitor of the inducible NO synthase) inhibiting IL-1 induced NO production. Glucosaminoglycan production was noted as proteoglycan synthesis showing almost no effect of cyclic stretch alone in comparison to the control condition, which correlates with the missing NO production in control and stretch conditions. Addition of IL-1 strongly inhibited proteoglycan synthesis, which was partly restored in the presence of LMA. However, cyclic stretch acted as a stronger restorer of proteoglycan synthesis in IL-1 treated conditions in the absence, and even more in the presence, of LMA. It was concluded that motion in the form of cyclic tensile stretch is a remarkable anti-inflammatory stimulus reversing the IL-1 induced suppression of proteoglycan synthesis in chondrocytes. These findings have therapeutic implications for the treatment of temporomandibular joint disorders, supporting early onset of postoperative and post-traumatic continuous passive motion therapy.

Animals↗
PubMed1999

Modulation of the biologic activity of the rabbit intervertebral disc by gene therapy: an in vivo study of adenovirus-mediated transfer of the human transforming growth factor beta 1 encoding gene.

STUDY DESIGN: In vivo studies using a rabbit model to determine the biologic effects of direct, adenovirus-mediated transfer of a therapeutic gene to the intervertebral disc. OBJECTIVES: 1) To deliver an exogenous therapeutic gene to rabbit lumbar intervertebral discs in vivo, 2) to quantify the resulting amount of gene expression, and 3) to determine the effect on the biologic activity of the discs. SUMMARY OF BACKGROUND DATA: Although growth factors such as transforming growth factor beta 1 appear to have promising therapeutic properties, there currently is no practical method for sustained delivery of exogenous growth factors to the disc for the management of certain chronic types of disease (e.g., disc degeneration). A possible solution is to modify the disc cells genetically through gene transfer such that the cells manufacture the desired growth factors endogenously on a continuous basis. METHODS: Saline, with or without virus, was injected directly into lumbar discs of 22 skeletally mature female New Zealand white rabbits. Group 1 (n = 11) received the adenovirus construct Ad/CMV-hTGF beta 1 containing the therapeutic human transforming growth factor beta 1-encoding gene. Group 2 (n = 6) received adenovirus containing the luciferase marker gene. Group 3 (n = 5) received saline only. The rabbits were killed 1 week after injection. Immunohistochemical staining for human transforming growth factor beta 1 was performed on the disc tissues of one rabbit from Group 1. Nucleus pulposus tissues from the remaining rabbits were cultured in serumless medium. Bioassays were performed to determine human transforming growth factor beta 1 production and proteoglycan synthesis. RESULTS: Discs injected with Ad/CMV-hTGF beta 1 exhibited extensive and intense positive immunostaining for transforming growth factor beta 1. The nucleus pulposus tissues from the discs injected with Ad/CMV-hTGF beta 1 exhibited a 30-fold increase in active transforming growth factor beta 1 production, and a 5-fold increase in total (active + latent) transforming growth factor beta 1 production over that from intact control discs (P < 0.05). Furthermore, these tissues exhibited a 100% increase in proteoglycan synthesis compared with intact control tissue, which was statistically significant (P < 0.05). CONCLUSIONS: The results of this study suggest that the intervertebral disc is an appropriate site for adenovirus-mediated transfer of exogenous genes and subsequent production of therapeutic growth factors. Gene therapy therefore may have useful applications for study of the basic science of the intervertebral disc and for clinical management of degenerative disc disease.

Adenoviridae↗
PubMed1999

Cyclic tensile stress exerts antiinflammatory actions on chondrocytes by inhibiting inducible nitric oxide synthase.

Continuous passive motion manifests therapeutic effects on inflamed articular joints by an as-yet-unknown mechanism. Here, we show that application of cyclic tensile stress (CTS) in vitro abrogates the catabolic effects of IL-1beta on chondrocytes. The effects of CTS are mediated by down-regulation of IL-1beta-dependent inducible NO production, and are directly attributed to the inhibition of inducible NO synthase (iNOS) mRNA expression and protein synthesis. The inhibition of iNOS induction by CTS is paralleled by abrogation of IL-1beta-induced down-regulation of proteoglycan synthesis. Furthermore, CTS inhibits iNOS expression and up-regulates proteoglycan synthesis at concentrations of IL-1beta frequently observed in inflamed arthritic joints, suggesting that the actions of CTS may be clinically relevant in suppressing the sustained effects of pathological levels of IL-1beta in vivo. These results are the first to demonstrate that mechanisms of the intracellular actions of CTS in IL-1beta-activated chondrocytes are mediated through inhibition of a key molecule in the signal transduction pathway that leads to iNOS expression.

Animals↗
PubMed1999

Direct adenoviral gene transfer of viral IL-10 to rabbit knees with experimental arthritis ameliorates disease in both injected and contralateral control knees.

IL-10, a cytokine produced primarily by macrophages, B lymphocytes, and Th2 cells, has both immunostimulatory and immunosuppressive properties. A homologue of IL-10 encoded by EBV, known as viral IL-10 (vIL-10), is also able to suppress the immune response, but may lack some of the immunostimulatory properties of IL-10. To evaluate the potential of vIL-10 to block the progression of rheumatoid arthritis, we have utilized a replication-defective adenovirus vector to deliver the gene encoding vIL-10 to the knee joints of rabbits with Ag-induced arthritis. Intraarticular expression of vIL-10 significantly reduced leukocytosis, cartilage matrix degradation, and levels of endogenous rabbit TNF-alpha, as well as the degree of synovitis, while maintaining high levels of cartilage matrix synthesis. Interestingly, an antiarthritic effect was also observed in opposing contralateral control knee joints that received only a marker gene. An adenoviral vector carrying the enhanced green fluorescent protein marker gene was used to demonstrate that a morphologically similar subset of cells infected in the injected knee joint are able to traffic to the uninjected contralateral knee joint. Our results suggest that direct, local intraarticular delivery of the vIL-10 gene may have polyarticular therapeutic effects.

Adenoviruses, Human↗
PubMed1999

Lessons learned from gene transfer approaches.

Recent technological advances allow the transfer of genes to the synovial lining of joints. As well as opening novel opportunities for therapy, these techniques provide valuable new tools for the study of synovitis and other aspects of the biology of joints in health and disease. This article reviews briefly the results of experiments in which selected genes have been transferred to the knee joints of healthy rabbits and rabbits with antigen-induced arthritis.

Animals↗
PubMed1999

Adenoviral transfer of the viral IL-10 gene periarticularly to mouse paws suppresses development of collagen-induced arthritis in both injected and uninjected paws.

Gene therapy is a promising new approach in the treatment of rheumatoid arthritis. Gene delivery to diseased joints offers the prospect of achieving high, local concentrations of a therapeutic gene product in a sustained manner, while minimizing exposure of nontarget organs. We report that a single administration of a modified adenovirus encoding the Epstein-Barr-derived homologue of IL-10 can suppress the development of disease for extended periods of time when injected locally within the periarticular tissue surrounding the ankle joints of mice with collagen type II-induced arthritis. Furthermore, we show that injection of an adenoviral vector carrying the IL-10 gene into a single paw can suppress development of arthritis in other, noninjected paws of the same individual. The systemic protection resulting from local gene therapy occurred in the absence of detectable levels of viral IL-10 in the serum. Circulating Ab levels to heterologous collagen were unaffected; however, treatment with viral IL-10 significantly suppressed the development of Abs to autologous mouse type II collagen. Thus, the treatment of a single joint by local delivery of the vIL-10 gene may protect multiple joints of the same individual while avoiding deleterious side effects often associated with systemic therapy.

Adenoviridae↗
PubMed1999

Inhibition of transforming growth factor beta production by nitric oxide-treated chondrocytes: implications for matrix synthesis.

OBJECTIVE: Nitric oxide (NO) is generated copiously by articular chondrocytes activated by interleukin-1beta (IL-1beta). If NO production is blocked, much of the IL-1beta inhibition of proteoglycan synthesis is prevented. We tested the hypothesis that this inhibitory effect of NO on proteoglycan synthesis is secondary to changes in chondrocyte transforming growth factor beta (TGFbeta). METHODS: Monolayer, primary cultures of lapine articular chondrocytes and cartilage slices were studied. NO production was determined as nitrite accumulation in the medium. TGFbeta bioactivity in chondrocyte- and cartilage-conditioned medium (CM) was measured with the mink lung epithelial cell bioassay. Proteoglycan synthesis was measured as the incorporation of 35S-sodium sulfate into macromolecules separated from unincorporated label by gel filtration on PD-10 columns. RESULTS: IL-1beta increased active TGFbeta in chondrocyte CM by 12 hours; by 24 hours, significant increases in both active and latent TGFbeta were detectable. NG-monomethyl-L-arginine (L-NMA) potentiated the increase in total TGFbeta without affecting the early TGFbeta activation. IL-1beta stimulated a NO-independent, transient increase in TGFbeta3 at 24 hours; however, TGFbeta1 was not changed. When NO synthesis was inhibited with L-NMA, IL-1beta increased CM concentrations of TGFbeta1 from 24-72 hours of culture. L-arginine (10 mM) reversed the inhibitory effect of L-NMA on NO production and blocked the increases in TGFbeta1. Anti-TGFbeta1 antibody prevented the restoration of proteoglycan synthesis by chondrocytes exposed to IL-1beta + L-NMA, confirming that NO inhibition of TGFbeta1 in IL-1beta-treated chondrocytes effected, in part, the decreased proteoglycan synthesis. Furthermore, the increase in TGFbeta and proteoglycan synthesis seen with L-NMA was reversed by the NO donor S-nitroso-N-acetylpenicillamide. Similar results were seen with cartilage slices in organ culture. The autocrine increase in CM TGFbeta1 levels following prior exposure to TGFbeta1 was also blocked by NO. CONCLUSION: NO can modulate proteoglycan synthesis indirectly by decreasing the production of TGFbeta1 by chondrocytes exposed to IL-1beta. It prevents autocrine-stimulated increases in TGFbeta1, thus potentially diminishing the anabolic effects of this cytokine in chondrocytes.

Animals↗
PubMed1999

Gene transfer of cytokine inhibitors into human synovial fibroblasts in the SCID mouse model.

OBJECTIVE: To investigate the effects of retrovirus-based gene delivery of inhibitory cytokines and cytokine inhibitors into human synovial fibroblasts in the SCID mouse model of rheumatoid arthritis (RA). METHODS: The MFG vector was used for gene delivery of tumor necrosis factor alpha receptor (TNFalphaR) p55, viral interleukin-10 (IL-10), and murine IL-10 into RA synovial fibroblasts. The effect on invasion of these cells into human articular cartilage and on perichondrocytic cartilage degradation was examined after 60 days of coimplantation into the SCID mouse. RESULTS: TNFalphaR p55 gene transfer showed only a limited effect on inhibition of RA synovial fibroblast invasiveness and cartilage degradation. In contrast, invasion of the RA synovial fibroblasts into the coimplanted cartilage was strongly inhibited by both viral and murine IL-10. Perichondrocytic cartilage degradation was not affected by either form of IL-10. CONCLUSION: The data show that cytokines can be successfully inserted into the genome of human RA synovial fibroblasts using a retroviral vector delivery system, and that the SCID mouse model of human RA is a valuable tool for examining the effects of gene transfer. In addition, inhibition of more than one cytokine pathway may be required to inhibit both synovial- and chondrocyte-mediated cartilage destruction in RA.

Animals↗
PubMed1999

Early expression of marker genes in the rabbit medial collateral and anterior cruciate ligaments: the use of different viral vectors and the effects of injury.

Gene therapy is a technique that may offer advantages over current methods of cytokine delivery to ligaments. To determine if implanted genes could be expressed in normal and injured knee ligaments, the medial collateral ligament and anterior cruciate ligament were studied in 18 rabbits. A retroviral ex vivo technique using allograft medial collateral ligament and anterior cruciate ligament fibroblasts and an adenoviral in vivo technique were compared as methods for delivering the LacZ marker gene to knee ligaments. Bilateral knee surgeries were performed, and the rabbits were equally divided into three groups. Group 1 received the retrovirus and the medial collateral ligament was ruptured, Group 2 received the adenovirus and the medial collateral ligament was ruptured, and Group 3 received the adenovirus and the medial collateral ligament was not injured. The anterior cruciate ligament was not injured in any group. The medial collateral and anterior cruciate ligaments of the right knees received 10(6) allografted, transduced ligament fibroblasts or 10(9) adenovirus particles, whereas the ligaments of the left knee received a similar volume of saline solution only. Equal numbers of rabbits were killed at 10 days, 3 weeks, and 6 weeks following the procedure. Ligament samples were stained with X-gal to detect the expression of the LacZ gene product, beta-galactosidase. LacZ gene expression was evident in ruptured and uninjured medial collateral ligaments as well as in the anterior cruciate ligament. The expression lasted between 10 days and 3 weeks in the medial collateral and anterior cruciate ligaments with use of the retrovirus and between 3 and 6 weeks in the medial collateral ligament and at least 6 weeks in the anterior cruciate ligament with the adenovirus. The length of gene expression in the ruptured and uninjured medial collateral ligaments did not differ. These preliminary studies indicate that gene transfer to normal and injured knee ligaments is possible.

Adenoviridae↗