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Browsing by Author "Frisbie, David D., advisor"

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    Cartilage repair using trypsin enzymatic pretreatment combined with growth-factor functionalized self-assembling peptide hydrogel
    (Colorado State University. Libraries, 2019) Zanotto, Gustavo Miranda, author; Frisbie, David D., advisor; Grodzinsky, Alan, committee member; McIlwraith, C. Wayne, committee member; Barrett, Myra F., committee member; Puttlitz, Christian, committee member
    Treatment of cartilage defects remains challenging in the orthopedic field. Several techniques are currently available to treat cartilage defects, with subchondral bone microfracture being the most commonly used marrow stimulation technique. However, despite satisfactory results in the short-term, clinical and functional outcomes of microfracture treated patients tend to decline over time. Improving microfracture technique using tissue engineering principles may be a more attractive way to treat cartilage defects compared to other more complex and expensive alternatives. Self-assembling peptide hydrogel has been extensively studied as a scaffold for cartilage repair. This hydrogel is biocompatible within the joint environment and has been shown to increase cartilage healing and improve clinical and functional outcomes in both rabbit and equine models of cartilage repair. Recently, a clinically applicable technique was described using trypsin enzymatic pretreatment of the surrounding cartilage combined with local delivery of heparin binding insulin growth factor-1 (HB-IGF-1). The results of this study demonstrated improved cartilage integration in vitro when this technique is utilized. Thus, in the present study we evaluated the combination of trypsin enzymatic pretreatment with a self-assembling peptide hydrogel functionalized with growth factors to improve cartilage repair. First, the effect of trypsin enzymatic pretreatment alone or combined with self-assembling peptide hydrogel functionalized with HB-IGF-1 and/or platelet-derived growth factor- BB (PDGF-BB) was tested using a rabbit model (48 rabbits). Subsequently, trypsin enzymatic pretreatment combined with self-assembling peptide hydrogel functionalized with HB-IGF-1 and PDGF-BB was used to augment microfracture augmentation in an equine model of cartilage defects (8 horses). In the small animal model, trypsin enzymatic pre-treatment resulted in an overall increase in defect filling, as well as improvements in subchondral bone reconstitution, surface regularity, cartilage firmness, reparative tissue color, cell morphology and chondrocyte clustering. The presence of PDGF-BB alone improved subchondral bone reconstitution and basal integration, while the combination of HB-IGF-1 and PDGF-BB resulted in an overall improvement in tissue and cell morphology. In the equine model, microfracture augmentation using trypsin enzymatic pretreatment combine with self-assembling peptide hydrogel functionalized with growth factors (HB-IGF-1 and PDGF-BB) resulted in better functional outcomes, better defect healing on second look arthroscopy at 12 months, as well as improved reparative tissue histology and increased biomechanical proprieties of the adjacent cartilage compared to defects treated with microfracture only. In conclusion, trypsin enzymatic pretreatment combined with self-assembling peptide hydrogel functionalized with growth factors (HB-IGF-1 and PDGF-BB) resulted in successful microfracture augmentation. These therapeutic approaches can result in a more cost effective way to improve cartilage healing in patients undergoing subchondral bone microfracture.
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    Evaluation of allogeneic freeze-dried platelet lysate and conditioned serum in joint tissues under inflammatory condition in vitro
    (Colorado State University. Libraries, 2017) Camargo Garbin, Livia, author; Frisbie, David D., advisor; McIlwraith, C. Wayne, committee member; Kisiday, John D., committee member; Olver, Christine S., committee member
    To view the abstract, please see the full text of the document.
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    Gene expression in phenotypically homogeneous chondrocytes from different articular cartilage layers of equine osteoarthritic and control joints: method validation and gene array analysis
    (Colorado State University. Libraries, 2007) Düsterdieck, Katja Friederike, author; Frisbie, David D., advisor
    Osteoarthritis remains a common and debilitating disease in horses, despite advances in diagnosis and treatment. Cartilage is commonly considered to play a central role in the pathophysiology of osteoarthritis. The investigation of differences in gene expression in cells from osteoarthritic and control cartilage is expected to yield genes possibly playing a role in the pathophysiology of osteoarthritis, representing new targets for treatment of the disease. The goals of this investigation were to develop a methodology to isolate RNA from phenotypically homogeneous cells of various cartilage layers for gene array analysis and to determine differentially expressed genes in these cells in osteoarthritic and control cartilage. A methodology to isolate phenotypically homogeneous chondrocytes from frozen sections of adult equine articular cartilage was developed using laser capture microdissection, RNA isolation, amplification and qrt-PCR. Expression levels of candidate genes were compared to those in conventionally isolated RNA from paired cartilage samples. The methodology was adequate to produce sufficient amounts of RNA for gene array analysis. Gene expression was found to be altered, but in a consistent fashion. The validated methodology, followed by gene array analysis was utilized to compare expression patterns in chondrocytes from tangential and radial layers of experimentally induced osteoarthritic and control cartilage. 154 genes were differentially expressed between tangential and radial cartilage layers and 17 genes were differentially expressed between osteoarthritic and control cartilage. The gene expression pattern of the tangential layer reflected support of cell proliferation, suppression of apoptosis and several genes involved in cell-matrix interactions or inflammatory processes. In contrast, the gene expression pattern of the radial layer was dominated by genes supporting the synthesis of proteins and proteoglycans. The gene expression pattern from osteoarthritic cartilage suggested an active response to oxidative stress, activation of the NF-κB pathway, decreased anti-apoptotic ability and downregulation of proteoglycan synthesis and glycolysis. This study was the first to determine gene expression patterns between two different layers of articular cartilage, improving our understanding of cartilage homeostasis in health and disease.