Search PubMed⌕ Search

Biomedical subjects

A Ratcliffe

Publications and source records attributed to A Ratcliffe.

At least 19 recordsLinked to original sources
PubMed2001

Scaffold-based three-dimensional human fibroblast culture provides a structural matrix that supports angiogenesis in infarcted heart tissue.

BACKGROUND: We have developed techniques to implant angiogenic patches onto the epicardium over regions of infarcted cardiac tissue to stimulate revascularization of the damaged tissue. These experiments used a scaffold-based 3D human dermal fibroblast culture (3DFC) as an epicardial patch. The 3DFC contains viable cells that secrete angiogenic growth factors and has previously been shown to stimulate angiogenic activity. The hypothesis tested was that a viable 3DFC cardiac patch would stimulate an angiogenic response within an area of infarcted cardiac tissue. METHODS AND RESULTS: A coronary occlusion of a branch of the left anterior descending coronary artery was performed by thermal ligation in severe combined immunodeficient mice. 3DFCs with or without viable cells were sized to the damaged area, implanted in replicate mice onto the epicardium at the site of tissue injury, and compared with animals that received infarct surgery but no implant. Fourteen and 30 days after surgery, hearts were exposed and photographed, and tissue samples were prepared for histology and cytochemistry. Fourteen and 30 days after surgery, the damaged myocardium receiving viable 3DFC exhibited a significantly greater angiogenic response (including arterioles, venules, and capillaries) than nonviable and untreated control groups. CONCLUSIONS: In this animal model, viable 3DFC stimulates angiogenesis within a region of cardiac infarction and can augment a repair response in damaged tissue. Therefore, a potential use for 3DFC is the repair of myocardial tissue damaged by infarction.

Animals↗
PubMed2001

Effects of flow patterns on endothelial cell migration into a zone of mechanical denudation.

Vascular endothelial cells (ECs) in vivo are subject to different flow conditions due to the variation in vessel geometry. The aim of this study is to elucidate the effects of different flow conditions on EC monolayer migration into a mechanically denuded zone and their underlying mechanisms. Both laminar and disturbed flows significantly enhanced EC migration. EC migration speed was the fastest under laminar flow, which preferentially promoted directional EC migration from the upstream side of the wounded monolayer. C3 exoenzyme (a Rho inhibitor) inhibited EC migration under static and flow conditions, and markedly reduced the effects of flow on EC migration. These results indicate that flow promotes EC migration through the Rho signaling pathway. Genistein (a tyrosine kinase inhibitor) selectively retarded EC migration under disturbed flow, suggesting that tyrosine phosphorylation may play a role in EC migration under disturbed flow. This study has demonstrated that different flow patterns differentially affect EC monolayer migration into the denuded zone involving multiple mechanisms.

ADP Ribose Transferases↗
PubMed2001

Insulin-like growth factor-I gene expression patterns during spontaneous repair of acute articular cartilage injury.

This study evaluated the constitutive insulin-like growth factor-I (IGF-I) gene expression pattern in spontaneously healing cartilage defects over the course of 16 weeks, and correlated the tissue morphology and matrix gene expression with IGF-I mRNA levels. Full-thickness 15 mm cartilage defects were debrided in the femoral trochlea of both femoropatellar joints of 8 horses and the healing defects examined 2, 4, 8, or 16 weeks after surgery. Samples were harvested for histologic assessment of tissue healing using H&E staining, toluidine blue histochemical reaction for proteoglycan deposition, and in situ hybridization and immunohistochemistry procedures to demonstrate collagen type II mRNA and protein expression. Total RNA was isolated for Northern analysis to measure cartilage matrix molecule expression, and for semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) to determine IGF-I gene expression patterns in healing cartilage defects. Full-thickness cartilage defects in horses were slow to heal compared to smaller lesions in similar locations in other animals. However, a progressive decline in tissue cellularity and vascularity, and increased tissue organization were observed on H&E stained specimens over the 16-week experiment. Evidence of early chondrogenic repair was detected through collagen type II in situ hybridization and immunohistochemistry. However, levels of collagen type II and aggrecan mRNA in lesions were not abundant on Northern analysis indicating incomplete chondrogenesis. IGF-I message expression followed a cyclic pattern with low levels at 2 weeks, followed by an increase at 4 and 8 weeks, and a subsequent decline at 16 weeks. There was no direct correlation between the stage of healing and cartilage matrix message expression, and the abundance of IGF-I mRNA in the healing lesions. In conclusion, this study demonstrated that the spontaneous healing of articular defects was accompanied by a temporal fluctuation in IGF-I gene expression which was discoordinate to the steady rise in expression of cartilage matrix molecules such as procollagen type II.

Acute Disease↗
PubMed2000

Mitogen-activated protein kinase signaling in bovine articular chondrocytes in response to fluid flow does not require calcium mobilization.

In the present study, the role of mitogen-activated protein kinases (MAPKs) in chondrocyte mechanotransduction was investigated. We hypothesized that MAPKs participate in fluid flow-induced chondrocyte mechanotransduction. To test our hypothesis, we studied cultured chondrocytes subjected to a well-defined mechanical stimulus generated with a laminar flow chamber. The extracellular signal-regulated kinases 1 and 2 (ERK1/2) were activated 1.6-3-fold after 5-15 min of fluid flow exposure corresponding to a chamber wall shear stress of 1.6 Pa. Activation of ERK1/2 was observed in the presence of both 10% FBS and 0.1% BSA, suggesting that the flow effects do not require serum agonists. Treatment with thapsigargin or EGTA had no significant effect on the ERK1/2 activation response to flow, suggesting that Ca2+ mobilization is not required for this response. To assess downstream effects of the activated MAPKs on transcription, flow studies were performed using chondrocytes transfected with a chimeric luciferase construct containing 2.4 kb of the promoter region along with exon 1 of the human aggrecan gene. Two-hour exposure of transfected chondrocytes to fluid flow significantly decreased aggrecan promoter activity by 40%. This response was blocked by treatment of chondrocytes with the MEK-1 inhibitor PD98059. These findings demonstrate that, under the conditions of the present study, fluid flow-induced signals activate the MEK-1/ERK signaling pathway in articular chondrocytes, leading to down-regulation of expression of the aggrecan gene.

Aggrecans↗
PubMed2000

Tissue engineering of vascular grafts.

The challenge of tissue engineering blood vessels with the mechanical properties of native vessels, and with the anti-thrombotic properties required is immense. Recent advances, however, indicate that the goal of providing a tissue-engineered vascular graft that will remain patent in vivo for substantial periods of time, is achievable. For instance, collagen gels have been used to fabricate a tissue in vitro that is representative of a native vessel: an acellular collagen tubular structure, when implanted as a vascular graft, was able to function, and to become populated with host cells. A completely cellular approach culturing cells into tissue sheets and wrapping these around a mandel was able to form a layered tubular structure with impressive strength. Culture of cells onto a biodegradable scaffold within a dynamic bioreactor, generated a tissue-engineered vascular graft with substantial stiffness and, when lined with endothelial cells, was able to remain patent for up to 4 weeks in vivo. In our experiments, use of a non-degradable polyurethane scaffold and culture with smooth muscle cells generated a construct with mechanical properties similar to native vessels. This composite tissue engineered vascular graft with an endothelial layer formed using fluid shear stress to align the endothelial cells, was able to remain patent with an neointima for up to 4 weeks. These results show that tissue engineering of vascular grafts has true potential for application in the clinical situation.

Biocompatible Materials↗
PubMed1999

A method for tissue engineering of cartilage by cell seeding on bioresorbable scaffolds.

This chapter describes a method for the in vitro generation of a 3-dimensional cartilage matrix from articular chondrocytes seeded onto a bioresorbable polymeric scaffold. This particular growth system was chosen for the subject of this chapter owing to the relative simplicity of the methods required and the ease with which the necessary materials can be obtained. The tissue produced using this protocol is a cellular, metabolically active hyaline-like matrix, containing the major cartilage constituents: sulfated proteoglycan, collagen type II, and water. It serves as a useful in vitro tool for studying the influence of various mechanical and chemical factors on cartilage metabolism, as well as providing an implantable material for in vivo cartilage repair studies.

Animals↗
PubMed1999

Human articular chondrocyte adhesion and proliferation on synthetic biodegradable polymer films.

The effect of polymer chemistry on adhesion, proliferation, and morphology of human articular cartilage (HAC) chondrocytes was evaluated on synthetic degradable polymer films and tissue culture polystyrene (TCPS) as a control. Two-dimensional surfaces of poly(glycolide) (PGA), poly(L-lactide) (L-PLA), poly(D,L-lactide) (D,L-PLA), 85:15 poly(D,L-lactide-co-glycolide) (D,L-PLGA), poly(epsilon-caprolactone) (PCL), 90:10 (D,L-lactide-co-caprolactone) (D,L-PLCL), 9:91 D,L-PLCL, 40:60 L-PLCL, 67:33 poly(glycolide-co-trimethylene carbonate) (PGTMC), and poly(dioxanone) (PDO) were made by spin-casting into uniform thin films. Adhesion kinetics were studied using TCPS and PCL films and revealed that the rate of chondrocyte adhesion began to level off after 6 h. Degree of HAC chondrocyte adhesion was studied on all the substrates after 8 h, and ranged from 47 to 145% of the attachment found on TCPS. The greatest number of chondrocytes attached to PGA and 67:33 PGTMC polymer films, and attachment to PCL and L-PLA films was statistically lower than that found on PGA (p < 0.05). There was no correlation between amount of chondrocyte attachment to the substrates and the substrates' water contact angle. Chondrocytes proliferated equally well on all the substrates resulting in equivalent cell numbers on all the substrates at both day 4 and day 7 of the culture. However, these total cell numbers were reached as a result of a 88- and 42-fold expansion on PDO and PLA, respectively, which was significantly higher than the 11-fold expansion found on TCPS (p < 0.05). The greater fold expansion of the cells on PDO and L-PLA films may be attributed to the availability of space for cells to grow, since their numbers at the start of culture were fewer following the 8 h attachment period. This suggests that regardless of initial seeding density on these degradable polymer substrates (i.e., if some minimum number of cells are able to attach), they will eventually populate the surfaces of all these polymers given sufficient space and time.

Cartilage, Articular↗
PubMed1999

The relationship between parafunctional masticatory activity and arthroscopically diagnosed temporomandibular joint pathology.

PURPOSE: The purpose of this investigation was to assess the relationship between parafunctional masticatory activity and arthroscopically visualized changes in patients with severe, unremitting symptoms caused by intra-articular temporomandibular joint pathology. The working hypothesis was that the presence of parafunctional activity leads to increased arthroscopically diagnosed pathology. MATERIALS AND METHODS: Temporomandibular joint arthroscopy was performed on 124 joints in 83 patients (female:male, 5.4:1; mean age, 35 years; mean duration of symptoms, 49 months) with severe symptoms unresponsive to nonsurgical management. Preoperatively, the presence of parafunctional habits (bruxism, clenching) was assessed, and joints were classified as either with or without parafunctional influences. Joints were diagnosed arthroscopically and assessed for the presence or absence of osteoarthritis, synovitis, and adhesions. Analyses were performed to determine significant relationships between parafunctional activity and the presence of osteoarthritis, synovitis, and adhesions. RESULTS: Parafunctional influences were present in 82 of 124 joints (66%). Clinically diagnosed osteoarthritis was present in 59 of 124 joints (48%) and arthroscopically diagnosed osteoarthritis was seen in 82 of 124 joints (66%). Arthroscopically, synovitis was diagnosed in 123 of 124 joints (99%) and adhesions in 93 of 124 joints (75%). Statistical analyses showed a significant relationship between parafunction and clinically diagnosed osteoarthritis, and suggested a close relationship between parafunction and arthroscopically diagnosed osteoarthritis. A significant association between clinically and arthroscopically diagnosed osteoarthritis and adhesions was also demonstrated. There also was no significant relationship detected between parafunction and the presence of synovitis or adhesions seen arthroscopically. CONCLUSIONS: It was concluded that parafunctional masticatory activity and its influence on joint loading contribute to osteoarthritis of the temporomandibular joint. Such osteoarthritis is associated with adhesions of the joint. Arthroscopically diagnosed synovitis is not specifically associated with parafunction, and it appears that numerous other causative factors may contribute to its development in the TMJ. Because abnormal joint loading is a major causative factor in cartilage degradation, biochemical and biomechanical abnormalities, and intraarticular temporomandibular pathology, clinicians must identify and address parafunctional masticatory activity during nonsurgical, surgical, and postsurgical treatment regimens.

Adult↗
PubMed1999

Repair of osteochondral defects with allogeneic tissue engineered cartilage implants.

The objective of this study was to evaluate the effect of allogeneic tissue engineered cartilage implants on healing of osteochondral defects. Rabbit chondrocytes were cultured in monolayer, then seeded onto biodegradable, three-dimensional polyglycolic acid meshes. Cartilage constructs were cultured hydrodynamically to yield tissue with relatively more (mature) or less (immature) hyalinelike cartilage, as compared with adult rabbit articular cartilage. Osteochondral defects in the patellar grooves of both stifle joints either were left untreated or implanted with allogeneic tissue engineered cartilage. Histologic samples from in and around the defect sites were examined 3, 6, 9, and 12, and 24 months after surgery. By 9 months after surgery, defects sites treated with cartilage implants contained significantly greater amounts of hyalinelike cartilage with high levels of proteoglycan, and had a smooth, nonfibrillated articular surface as compared to untreated defects. In contrast, the repair tissue formed in untreated defects had fibrillated articular surfaces, significant amounts of fibrocartilage, and negligible proteoglycan. These differences between treated and untreated defects persisted through 24 months after surgery. The results of this study suggest that the treatment of osteochondral lesions with allogenic tissue engineered cartilage implants may lead to superior repair tissue than that found in untreated osteochondral lesions.

Animals↗
PubMed1998

Load-controlled compression of articular cartilage induces a transient stimulation of aggrecan gene expression.

The effects of short- and long-term load-controlled compression on the levels of aggrecan mRNA have been determined. Results show that a compressive stress of 0.1 MPa on bovine articular cartilage explants for 1, 4, 12, and 24 h produces a transient up-regulation of aggrecan mRNA synthesis. At 1 h, aggrecan mRNA levels in loaded explants were increased 3.2-fold compared to control explants. At longer times (>/=4 h), the levels of aggrecan mRNA returned to baseline values or stayed slightly higher. There is a dose dependence in the response of the explant to increasing levels of compressive stress (0-0.5 MPa) for 1 h. Aggrecan mRNA levels increased 2- to 3-fold at 0-0.25 MPa. At 0.5 MPa, the level of aggrecan mRNA was lower than those at 0.1 and 0.25 MPa. This dose-dependent effect suggests a reversal of the stimulatory effects of compression on aggrecan gene expression at higher loads. After 24 h of compression, the levels of aggrecan mRNA in explants subjected to any of the stress levels were not significantly different from those in control explants. The stimulatory effect of 0.1 MPa compressive stress on aggrecan mRNA levels was blocked by Rp-cAMP and U-73122, indicating the involvement of the classical signal transduction pathways in the mechanical modulation of aggrecan gene expression. The responses of link protein mRNA to compression paralleled those of aggrecan, while there was no significant change in expression of the gene for the housekeeping protein elongation factor-1 alpha. The results indicate that articular cartilage chondrocytes can respond to short-term compressive loads by transiently up-regulating expression of the aggrecan gene. The fact that long-term compression did not significantly alter aggrecan mRNA levels suggests that previously observed inhibitory effects of prolonged static compression on proteoglycan synthesis in articular cartilage may be, for the most part, mediated through mechanisms other than suppression of aggrecan mRNA levels.

Aggrecans↗
PubMed1998

Regulatory activities of the 5'- and 3'-untranslated regions and promoter of the human aggrecan gene.

Identification and characterization of the regulatory elements of the human aggrecan gene are necessary first steps in addressing the molecular mechanisms through which the gene is regulated. Using luciferase reporter constructs driven by the human aggrecan promoter or the cytomegalovirus promoter, the 5'- and 3'-untranslated regions of the human aggrecan gene were found to regulate gene expression transcriptionally in a promoter- and/or cell type-specific manner. Independent of cell type, the 5'-untranslated region was inhibitory with respect to the cytomegalovirus promoter, but it was stimulatory to the human aggrecan promoter. The 5'-untranslated region inhibited the cytomegalovirus promoter by approximately 60% in both chondrocytes and NIH 3T3 cells, but it stimulated the activity of the human aggrecan promoter about 8-fold in chondrocytes and 40-fold in NIH 3T3 cells. In contrast, the 3'-untranslated region inhibited the activities of the human aggrecan promoter by 40-70% in both cell types, but it stimulated the cytomegalovirus promoter activities by 50-60% in NIH 3T3 cells and inhibited its activity by 70% in chondrocytes. The differential effects of the untranslated regions on the two types of promoters may be a reflection of differences in regulation of TATA-less promoters, such as the human aggrecan promoter, and TATA-containing promoters, such as the cytomegalovirus promoter.

Aggrecans↗
PubMed1998

Osteoarthritis and synovitis as major pathoses of the temporomandibular joint: comparison of clinical diagnosis with arthroscopic morphology.

PURPOSE: The purposes of this investigation were to determine how common osteoarthritis and synovitis are in patients with severe, recalcitrant temporomandibular joint (TMJ) symptoms using clinical diagnostic criteria as well as arthroscopic examination, and to compare the accuracy of the clinical and arthroscopic diagnoses with respect to specificity and sensitivity. PATIENTS AND METHODS: Clinical and arthroscopic diagnoses were established in 126 joints of 84 patients with severe TMJ symptoms recalcitrant to conservative therapy. All joints were classified as having osteoarthritis (OA) or no osteoarthritis (non-OA) and synovitis (syn) or no synovitis (non-syn) using clinical and arthroscopic criteria. Chi-squared analysis was used to determine whether there was a relationship between the clinical and arthroscopic diagnoses. Preoperative clinical diagnoses were compared with arthroscopic morphologic diagnoses to determine the specificity and sensitivity of the clinical diagnostic criteria for synovitis and osteoarthritis. RESULTS: A preoperative clinical diagnosis of OA was established in 59 of 126 joints (47%) compared with an arthroscopic diagnoses of OA in 82 of 126 joints (65%). Chi-squared analysis showed a significant relationship between the clinical and arthroscopic diagnosis of OA. A clinical diagnosis of OA was associated with a high specificity (.977); however, there were 23 of 82 (.293) false-negative findings and a sensitivity of only .707. A preoperative clinical diagnosis of synovitis was established in 114 of 126 joints (90%), compared with an arthroscopic diagnosis of synovitis in 112 of 126 (89%). Chi-squared analysis did not show a significant relationship between the clinical and arthroscopic diagnosis of synovitis. A clinical diagnosis of synovitis was associated with a high sensitivity (.920); however, there were 11 of 14 false-positive findings (.786) associated with a low specificity (.214). CONCLUSIONS: Although there was high specificity for the clinical diagnosis of OA, the sensitivity was very low. (Comparison of clinical and arthroscopic diagnoses showed that osteoarthritis frequently escapes clinical detection. The clinical diagnosis of synovitis showed that low specificity and symptoms may be caused by other pathoses.

Adolescent↗
PubMed1998

Proteoglycans in the synovial fluid of the temporomandibular joint as an indicator of changes in cartilage metabolism during primary and secondary osteoarthritis.

PURPOSE: The specific aim of this investigation was to assess differences between primary and secondary osteoarthritis (OA) of the temporomandibular joint (TMJ) using clinical evaluation and synovial fluid analysis for proteoglycans. MATERIALS AND METHODS: Arthroscopic surgery was performed on 101 TMJs from patients with significant pain or dysfunction and who had failed to respond to treatment. Joints were assessed for primary and secondary osteoarthritis. Synovial fluid aspirates were obtained and analyzed to determine the levels of keratan sulfate (KS) epitope and a novel 3B3(-) epitope by enzyme-linked immunosorbent assay (ELISA). RESULTS: Fifty-four patients and 67 joints had OA diagnosed by both clinical examination and arthroscopy. Primary OA was diagnosed in 14 joints (20%), and the remaining 53 joints were regarded as having secondary OA. No differences were detected in the levels of KS in the synovial fluid from the primary and secondary OA joints. Furthermore, the 3B3(-) epitope was not detectable in the synovial fluid aspirates of any TMJ. CONCLUSION: Secondary OA is a common disorder of the TMJ. However, there is no apparent difference in the metabolism of the joints with primary and secondary OA as assessed by proteoglycans in the synovial fluid. The apparent absence of the 3b3(-) epitope, in contrast to its presence in OA of other major synovial joints, suggests that there are some differences between the cartilage metabolism of the TMJ and these other joints during OA.

Adult↗
PubMed1997

Correlation between arthroscopic diagnosis of osteoarthritis and synovitis of the human temporomandibular joint and keratan sulfate levels in the synovial fluid.

PURPOSE: The specific aims of this investigation were to determine if there is a relationship between an arthroscopic diagnosis of synovitis and osteoarthritis, and if the presence of synovitis influences the level of cartilage degradation, as evidenced by keratan sulfate levels in the synovial fluid. PATIENTS AND METHODS: Arthroscopic surgery was performed on 114 temporomandibular joints in 88 patients who had significant pain or dysfunction and whose condition had failed to improve with conservative treatment. Synovial fluid aspirates were obtained immediately before arthroscopy and used for the determination of keratan sulfate levels. Arthroscopic examination included assessment of the presence or absence of osteoarthritis and synovitis. RESULTS: Synovitis was present in 90% of joints, and osteoarthritis was present in 62% of joints examined arthroscopically. Both osteoarthritis and synovitis existed in 57% of the joints. Joints with an arthroscopic diagnosis of synovitis had significantly lower levels of keratan sulfate in the synovial fluid aspirates than joints with osteoarthritis. Synovial fluid aspirates from temporomandibular joints with osteoarthritis had significantly higher levels of keratan sulfate than synovial fluids from joints without osteoarthritis. CONCLUSIONS: Osteoarthritis and synovitis are common diagnoses and are often present concurrently in patients with symptomatic temporomandibular joints. Osteoarthritis is associated with elevated keratan sulfate levels; however, the elevation of keratan sulfate is less in patients with concomitant synovitis.

Analysis of Variance↗
PubMed1997

Differences in patellofemoral joint cartilage material properties and their significance to the etiology of cartilage surface fibrillation.

OBJECTIVE: To determine if differences in biomechanical properties and biochemical composition exist between human patellar articular cartilage and the opposing femoral articular cartilage. DESIGN: The biomechanical properties and biochemical composition of the articular cartilage of 17 knees from 13 donors were determined for four sites on the patella and three sites on the femur representing regions of contact at 30 degrees and 90 degrees of flexion. The material properties were determined by biphasic indentation testing, yielding the compressive aggregate modulus, HA, permeability, k, and Poisson's ratio, vs. The thickness of the cartilage at the indentation site, h, was also measured using a needle probe. Full-thickness samples of cartilage adjacent to each indentation site were used for wet weight, sulfated glycosaminoglycan content and hydroxyproline content determinations. RESULTS: The patellar cartilage was found to have a lower compressive aggregate modulus by 30% (P < 0.001), higher permeability to fluid flow by 66% (P < 0.001) and greater thickness by 23% (P = 0.017) than that of the opposing femoral cartilage. The Poisson's ratios for both surfaces were found to be nearly zero. The water content of the patella was higher by 5% (P = 0.031) and the proteoglycan content lower by 19% (P = 0.030) than that of the femur. However, no differences were found between the collagen contents of the cartilages. CONCLUSIONS: Significant differences were found between the intrinsic material properties of the patellar cartilage and those of the femoral-trochlear cartilage. This variability of cartilage material properties with the patellofemoral joint may help explain why patellar cartilage has been frequently observed clinically to exhibit earlier and more severe fibrillation changes than the opposing femoral cartilage.

Adolescent↗