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A novel co-culture model for the study of osteoarthritis in dogsStreppa, Heather Kirsten. January 2004 (has links)
Thesis (M.S.)--University of Missouri--Columbia, 2004. / Typescript. Includes bibliographical references (leaves 63-70). Also issued on the Internet. Also available on the Internet.
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TP508 maintains chondrocyte cell viability through blocking apoptosis in an NO-dependent mannerZhong, Ming 27 November 2006 (has links)
TP508 is a 23 amino acid peptide derived from human prothrombin. It helps
wound healing in both soft tissues and bones. In our previous study, we have
demonstrated that TP508 retains chondrocyte in a less mature differentiation
state while expanding the cartilage mass, indicating it may partly help bone
healing by expand the cartilage template in the endochondral bone formation
stage. In our current study, we want to demonstrate that TP508 also blocks
chondrocyte apoptosis. We used rat costochondral growth plate chondrocytes
as our model. We first established chelerythrine as an apoptogen in
chondrocytes. TP508 is able to block apoptosis caused by chelerythrine.
Chelerythrine also causes an increase in NO production, which is known to
cause both pathological and physiological apoptosis of chondrocyte, and
blocking NO production can in turn block apoptosis caused by them. TP508 is
also able to block NO production caused by chelerythrine. Therefore, TP508
may partially block chondrocyte apoptosis by blocking NO production. From all above,
we conclude that besides decreasing chondrocyte differentiation, TP508 also
blocks their apoptosis, so as to conserve the cartilage template in
endochondral bone formation
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Caveolae and Caveolin-1 are important for Vitamin D signallingWong, Kevin L. 20 October 2010 (has links)
The most active form of Vitamin D, 1alpha,25(OH)2D3, modulates cells via receptor mediated mechanisms. While studies have elucidated the pathway via the classical nuclear Vitamin D Receptor (VDR), little is known about the membrane-associated Vitamin D Receptor (ERp60). Caveolae and its characteristic protein Caveolin-1 have been involved in many signaling pathways due to its specific structure and physical configuration. Other studies have shown that many components of the Vitamin D pathway have been found in caveolae. This study hypothesizes that caveolae and Caveolin-1 are important for the effects of 1,25 Vitamin D signaling via ERp60. Research up to date have shown that in rat and mouse growth zone chondrocytes, cells deprived of intact caveolae either through disruption through beta-Cyclodextrin or genetic knockout do not exhibit the characteristic responses to Vitamin D through ERp60 when compared to chondrocytes with functional caveolae. Studies using immunofluorescence co-localization and caveolae fractionation have shown that ERp60 is localized in the caveolae domains. Cellular fractionation was also performed to examine the localization of the ERp60 receptor in lipid rafts and caveolae. Histology and transmission electron microscopy were also used to examine the physiological importance of caveolae and Caveolin-1 in growth plate morphology and cellular characteristics.
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Matrix-induced autologous chondrocyte implantation for articular cartilage injury : biology, histology and clinical outcomesWillers, Craig Robert January 2008 (has links)
[Truncated abstract] Articular cartilage has no vascular, neural, or lymphatic supply, and hence no intrinsic capacity to self-repair following injury. These physiological limitations, combined with the inability of local chondrocytes to contribute to the repair process, translate to poor structural and functional outcomes in these troublesome defects, and osteoarthritic deterioration with time. Subsequently, many surgical therapies have been trialed to stimulate cartilage repair, but none have produced reliable outcomes. Hence, cartilage repair research has been broadened, with many investigators now focused on cell-based treatment. Smith began a revolution of autologous cell research when in 1965 she isolated chondrocytes from articular cartilage and transplanted them into fresh cartilage nodules (Smith, 1965). Since, new technologies and improved techniques have seen autologous chondrocyte implantation (ACI) widely accepted for use in clinical orthopaedics (Bentley et al., 2003; Brittberg et al., 1994; Grande et al., 1989; Peterson et al., 2002). At present, matrix-induced autologous chondrocyte implantation (MACI) is the most surgically simple form of ACI, boasting clinical outcomes comparable to any technique on the market, and far less complications compared to the first generation of ACI - periosteal ACI (Bartlett et al., 2005; Behrens et al., 2006; Gigante et al., 2006; Henderson et al., 2004; Marlovits et al., 2005; Minas, 2001; Willers et al., 2007; Zheng et al., 2007). But whilst MACI has been adopted by the orthopaedic surgeon for articular cartilage repair, many of the molecular, histological, and clinical factors governing patient outcomes are still largely understudied. Firstly we assessed the bioactivity of fibrin sealant (FS - Tisseel®), a critical component of MACI, on the migration and proliferation of human articular chondrocytes in vitro. We also looked to elucidate the associated molecular mechanisms of thrombin, a key active ingredient in FS, by examining the expression and activation of proteaseactivated receptors (PARs), established thrombin receptors. All four PAR isoforms were detected in human chondrocytes, with PAR-1 being the major isoform expressed. '...' This thesis has demonstrated biological, histological, and clinical features of the MACI technique. Our in vitro has supported the use of fibrin sealant and collagen membrane as the major material components of MACI, illustrating improved chondrocyte proliferation, migration, and chondrogenic differentiation. We have evidenced that MACI stimulates successful production of hyaline-like cartilage by 6 months, while also showing that revised and clinically failed repair tissues are predominantly hyaline-like and fibrocartilage with inferior composition. Clinically, we have documented significant improvements in patient repair structure, function, symptoms, quality of life, and satisfaction, whilst concurrently confirming sentiment within the literature regarding the importance of exercise/ rehabilitation for maximising MACI outcome. In summary, the findings presented in this thesis suggest that MACI is a biologically sound and clinically efficacious cell-based treatment option for repairing articular cartilage defects.
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Mathematical modeling of transport and reaction in cellular and tissue engineeringPragyansri, Pathi. Locke, Bruce R. January 2005 (has links)
Thesis (Ph. D.)--Florida State University, 2005. / Advisor: Bruce R. Locke, Florida State University, FAMU-FSU College of Engineering, Dept. of Chemical and Biomedical Engineering. Title and description from dissertation home page (viewed Jan. 12, 2006). Document formatted into pages; contains xix, 250 pages. Includes bibliographical references.
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The role of cultured chondrocytes and mesenchymal stem cells in the repair of acute articular cartilage injuriesSecretan, Charles Coleman. January 2010 (has links)
Thesis (Ph.D.)--University of Alberta, 2010. / A thesis submitted to the Faculty of Graduate Studies and Research in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Experimental Surgery, Department of Surgery. Title from pdf file main screen (viewed on April 24, 2010). Includes bibliographical references.
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Laser scanning confocal arthroscopy in orthopaedics : examination of chondrial and connective tissues, quantification of chondrocyte morphology, investigation of matirx-induced autologous chondrocyte implantation and characterisation of osteoarthritisJones, Christopher Wynne January 2007 (has links)
[Truncated abstract] Articular cartilage (AC) covers the surface of synovial joints providing a nearly frictionless bearing surface and distributing mechanical load. Joint trauma can damage the articular surface causing pain, loss of mobility and deformation. Currently there is no uniform treatment protocol for managing focal cartilage defects, with most treatment options targeted towards symptomatic relief but not limiting the progression into osteoarthritis (OA). Autologous chondrocyte implantation (ACI) and more recently matrix-induced autologous chondrocyte implantation (MACI), have emerged as promising methods for producing hyaline or hyaline-like repair tissue, however there remains some controversy regarding the exact histological nature of the tissue formed. Histological characterisation of AC repairs requires destructive tissue biopsy potentially inducing further joint pathology thereby negating the treatment effect. OA is recognised as a major cause of pain, loss of function and disability in Western populations, however the exact aetiology is yet to be elucidated. The assessment of both OA and cartilage repair has been limited to macroscopic observation, radiography, magnetic resonance imaging (MRI) or destructive biopsy. The development of non-destructive AC assessment modalities will facilitate further development of AC repair techniques and enable early monitoring of OA changes in both experimental animal models and clinical subjects. Confocal laser scanning microscopy (CLSM) is a type of fluorescence microscopy that generates high-resolution three-dimensional images from relatively thick sections of tissue. ... Biomechanical analysis suggested that the mechanical properties of MACI tissue remain inferior for at least three months. This study showed the potential of a multi-site sheep model of articular cartilage defect repair and validated its assessment via LSCA. Finally, the LSCA was used to arthroscopically image the cartilage of an intact fresh frozen cadaveric knee from a patient with clinically diagnosed OA. Images were correlated to ICRS (Outerbridge) Grades I-IV and histology. The LSCA gave excellent visualization of cell morphology and cell density to a depth of up to 200'm. Classical OA changes including clustering chondrocytes, surface fibrillation and fissure formation were imaged. Fair to moderate agreement was demonstrated with statistically significant correlations between all modalities. This study confirmed the viability of the LSCA for non-destructive imaging of the microstructure of the OA cartilage. In conclusion, the LSCA identified histological features of orthopaedic tissues, accurately quantified chondrocyte morphology and demonstrated classical OA changes. While the development and investigation of an ovine model of cartilage repair showed the treatment benefit of MACI, some biomechanical issues remain. Ultimately, the LSCA has been demonstrated as a reliable nondestructive imaging modality capable of providing optical histology without the need for mechanical biopsy. Medical Subject Headings (MESH): articular cartilage; autologous chondrocyte implantation; matrix-induced autologous chondrocyte implantation; biomechanics; cartilage; confocal microscopy; diagnosis; histology; image analysis; immunohistochemistry; magnetic resonance imaging; microscopy; osteoarthritis
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Effect of Shear Stress on RhoA Activities and Cytoskeletal Organization in ChondrocytesWan, Qiaoqiao 05 September 2013 (has links)
Indiana University-Purdue University Indianapolis (IUPUI) / Mechanical force environment is a major factor that influences cellular homeostasis and remodeling. The prevailing wisdom in this field demonstrated that a threshold of mechanical forces or deformation was required to affect cell signaling. However, by using a fluorescence resonance energy transfer (FRET)-based approach, we found that C28/I2 chondrocytes exhibited an increase in RhoA activities in response to high shear stress (10 or 20 dyn/cm2), while they showed a decrease in their RhoA activities to intermediate shear stress at 5 dyn/cm2. No changes were observed under low shear stress (2 dyn/ cm2). The observed two-level switch of RhoA activities was closely linked to the shear stress-induced alterations in actin cytoskeleton and traction forces. In the presence of constitutively active RhoA (RhoA-V14), intermediate shear stress suppressed RhoA activities, while high shear stress failed to activate them. Collectively, these results herein suggest that intensities of shear stress are critical in differential activation and inhibition of RhoA activities in chondrocytes.
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Expression and Function of the PRL Family of Protein Tyrosine PhosphataseDumaual, Carmen Michelle 06 March 2013 (has links)
Indiana University-Purdue University Indianapolis (IUPUI) / The PRL family of enzymes constitutes a unique class of protein tyrosine phosphatase, consisting of three highly homologous members (PRL-1, PRL-2, and PRL-3). Family member PRL-3 is highly expressed in a number of tumor types and has recently gained much interest as a potential prognostic indicator of increased disease aggressiveness and poor clinical outcome for multiple human cancers. PRL-1 and PRL-2 are also known to promote a malignant phenotype in vitro, however, prior to the present study, little was known about their expression in human normal or tumor tissues. In addition, the biological function of all three PRL enzymes remains elusive and the underlying mechanisms by which they exert their effects are poorly understood. The current project was undertaken to expand our knowledge surrounding the normal cellular function of the PRL enzymes, the signaling pathways in which they operate, and the roles they play in the progression of human disease. We first characterized the tissue distribution and cell-type specific localization of PRL-1 and PRL-2 transcripts in a variety of normal and diseased human tissues using in situ hybridization. In normal, adult human tissues we found that PRL-1 and PRL-2 messages were
almost ubiquitously expressed. Only highly specialized cell types, such as fibrocartilage cells, the taste buds of the tongue, and select neural cells displayed little to no expression of either transcript. In almost every other tissue and cell type examined, PRL-2 was expressed strongly while PRL-1 expression levels were variable. Each transcript was widely expressed in both proliferating and quiescent cells indicating that different tissues or cell types may display a unique physiological response to these genes. In support of this idea, we found alterations of PRL-1 and PRL-2 transcript levels in tumor samples to be highly tissue-type specific. PRL-1 expression was significantly increased in 100% of hepatocellular and gastric carcinomas, but significantly decreased in 100% of ovarian, 80% of breast, and 75% of lung tumors as compared to matched normal tissues from the same subjects. Likewise, PRL-2 expression was significantly higher in 100% of hepatocellular carcinomas, yet significantly lower in 54% of kidney carcinomas compared to matched normal specimens. PRL-1 expression was found to be associated with tumor grade in the prostate, ovary, and uterus, with patient gender in the bladder, and with patient age in the brain and skeletal muscle. These results suggest an important, but pleiotropic role for PRL-1 and PRL-2 in both normal tissue function and in the neoplastic process. These molecules may have a tumor promoting effect in some tissue types, but inhibit tumor formation or growth in others. To further elucidate the signaling pathways in which the PRLs operate, we focused on PRL-1 and used microarray and microRNA gene expression profiling to examine the global molecular changes that occur in response to stable PRL-1 overexpression in HEK293 cells. This analysis led to identification of several molecules not previously associated with PRL signaling, but whose expression was significantly altered by exogenous PRL-1 expression. In particular, Filamin A, RhoGDIalpha, and SPARC are attractive targets for novel mediators of PRL-1 function. We also found that PRL-1 has the capacity to indirectly influence the expression of target genes through regulation of microRNA levels and we provide evidence supporting previous observations suggesting that PRL-1 promotes cell proliferation, survival, migration, invasion, and metastasis by influencing multi-functional molecules, such as the Rho GTPases, that have essential roles in regulation of the cell cycle, cytoskeletal reorganization, and transcription factor function. The combined results of these studies have expanded our current understanding of the expression and function of the PRL family of enzymes as well as of the role these important signaling molecules play in the progression of human disease.
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Exploring Chondrocyte Integrin Regulation of Growth Factor IGF-I Expression from a Transient pAAV VectorRatley, Samantha Kay 20 August 2013 (has links)
Indiana University-Purdue University Indianapolis (IUPUI) / Insulin-like Growth Factor I (IGF-I) is a growth factor that stimulates both mitogenic and anabolic responses in articular chondrocytes. While it has been shown that exogenous IGF-I can regulate chondrocyte integrins, little is known regarding regulatory effects of IGF-I produced from a transiently expressed plasmid based adeno-associated virus (pAAV) vector. Because chondrocytes are using cellular machinery
to overexpress IGF-I, it is of interest to see whether or not pAAV IGF-I will significantly upregulate or downregulate chondrocyte integrins. Additionally, it is of interest to know whether chondrocyte adhesion through integrins will have any regulatory effects on the production of IGF-I from the transgene. Therefore, this study will ascertain if pAAV IGF-I will have similar effects that exogenous IGF-I has on integrin regulation and if integrin silencing mechanisms will affect the production of
IGF-I from the transgene.
To test these hypotheses, adult articular chondrocytes were doubly transfected with the pAAV vector for IGF-I and short interference ribonucleic acid (siRNA) for integrins beta 1 and alpha V. Gene products were monitored at the transcriptional levels using
quantitative real time polymerase chain reactions (qPCR) and IGF-I protein production was monitored at the translational level using enzyme linked immunoabsorbant assays (ELISAs). Adult articular chondrocytes doubly transfected were encapsulated in a three dimensional hydrogel system to simulate an in vivo environment. Samples were collected for analysis at days 2, 4, and 6 post encapsulation. Results show that IGF-I treatment with the pAAV vector does not cause significant changes in the
transcriptional regulation of the beta 1 integrin in a three dimensional hydrogel system. The pAAV IGF-I vector did not cause significant regulatory changes on integrin alpha V at any time point during the experiment. Additionally, by knocking down the expression levels of integrins by using siRNA, it was shown that integrin knockdown does not have a significant regulatory effect on transcriptional or translational expression levels of IGF-I from the pAAV vector.
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