摘要:

AbstractBone marrow contains a population of rare progenitor cells capable of differentiating into bone, cartilage, tendon, and other connective tissues. These cells, referred to as mesenchymal stem cells, can be purified and culture‐expanded from animals and humans and have been shown to regenerate functional tissue when delivered to the site of musculoskeletal defects in experimental animals. To test the ability of purified human mesenchymal stem cells to heal a clinically significant bone defect, mesenchymal stem cells isolated from normal human bone marrow were culture‐expanded, loaded onto a ceramic carrier, and implanted into critical‐sized segmental defects in the femurs of adult athymic rats. For comparison, cell‐free ceramics were implanted in the contralateral limb. The animals were elthanized at 4, 8, or 12 weeks, and healing bone defects were compared by high‐resolution radiography, immunohistochemistry, quantitative histomorphometry, and biomechanical testing. In mesenchymal stem cell‐loaded samples, radiographic and histologic evidence of new bone was apparent by 8 weeks and histomorphometry demonstrated increasing bone formation through 12 weeks. Biomechanical evaluation confirmed that femurs implanted with mesenchymal stem cell‐loaded ceramics were significantly stronger than those that received cell‐free ceramics. These studies demonstrate that human mesenchymal stem cells can regenerate bone in a clinically significant osseous defect and may therefore provide an alternative to autogen

ISSN:0736-0266    

DOI:10.1002/jor.1100160202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractThe measured fatigue strength of a material can be affected by specimen size: tests using a large stressed volume may show a low fatigue strength due to the increased probability of finding weak regions. A Weibull analysis revealed an important size effect in bone and predicted this effect with an accuracy of 12%. This approach also explained apparent inconsistencies in the published data and made it possible to separate and quantify the effects of frequency, loading mode, and material source. The effect of frequency is the same for human and bovine bone, and the differences between different types of loading (tension, compression, and bending) are small (maximum: 12%). By extrapolating to the volume of whole bones, it is concluded that large bones will have a fatigue strength much lower, by a factor of 2–3, than that measured by conventional tests. Failure within 105cycles is expected to occur at cyclic stresses of 23–30 MPa in human long bones and of 32–43 MPa in bovine bones. Repair is therefore needed to prevent failure at physiological stress l

ISSN:0736-0266    

DOI:10.1002/jor.1100160203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractThe structural integrity of microfilaments has been shown to be necessary for the signal transduction of mechanical stimuli within osteoblasts. Qualitative and quantitative changes within the cytoskeleton of osteoblasts may therefore be crucial components of the signal transduction processes of these cells in response to mechanical stimulation. Avian osteoblasts were strained with a device that deforms a flexible, cell‐laden membrane at a defined frequency and intensity in a uniform biaxial manner. We examined the effects of mechanical strain on the accumulation of protein and the expression of the major cytoskeletal elements and specific integrin‐binding (arginine‐glycine‐aspartic acid) proteins of these cells. Mechanical strain increased the level of total extracellular matrix‐accumulated fibronectin by approximately 150% and decreased that of osteopontin by approximately 60% but had no quantifiable effect on the accumulation of β1 integrin subunit or collagen type I. An examination of the major elements of the cytoskeleton demonstrated that neither the level of actin nor that of the intermediate filament protein vimentin changed; however, the amount of tubulin decreased by approximately 75% and the amount of vinculin, a major protein of focal adhesion complexes, increased by approximately 250%. An analysis of protein synthesis by two‐dimensional gel electrophoresis of [35S]methionine‐labeled cytoskeletal proteins demonstrated that the changes in the accumulation of vinculin and tubulin resulted from their altered synthesis. Messenger RNA analysis confirmed that the changes in accumulation and protein synthesis observed for vinculin, fibronectin, and osteopontin were controlled at a pretranslational level. Immunofluorescent microscopy demonstrated that mechanical strain led to increased formation and thickening of actin stress fibers, with a commensurate dissociation in microtubules and a clear increase in levels of vinculin at the peripheral edges of the cells. In conclusion, the elevated rate of synthesis and the increased accumulation of vinculin and fibronectin, as well as the increase in the number and size of stress fibers and focal adhesion complexes, suggest that mechanical strain leads to a coordinated change both in the cytoskeleton and in extracellular matrix proteins that will facilitate tighter adhesion of an osteoblast to its extrac

ISSN:0736-0266    

DOI:10.1002/jor.1100160204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractBone marrow stromal cells, progenitor cells involved in repair of bone and cartilage, can potentially provide a source for autologous skeletal tissue engineering. We investigated which factors were required to inducein vitrodifferentiation of avian bone marrow stromal cells into three‐dimensional cartilaginous and bone‐like tissues. Bone marrow stromal cells from embryonic chicks were expanded in monolayers, seeded onto biodegradable polyglycolic acid scaffolds, and cultured for 4 weeks in orbitally mixed Petri dishes. Cell‐polymer constructs developed an organized extracellular matrix containing glycosaminoglycans and collagen, whereas control bone marrow stromal cell pellet cultures were smaller and consisted predominantly of fibrous tissue. Bone marrow stromal cells expanded with fibroblast growth factor‐2 and seeded onto polymer scaffolds formed highly homogeneous three‐dimensional tissues that contained cartilage‐specific molecular markers and had biochemical compositions comparable with avian epiphyseal cartilage. When cell‐polymer constructs were cultured in the presence of beta‐glycerophosphate and dexamethasone, the extracellular matrix mineralized and bone‐specific proteins were expressed. Our work shows that cell expansion in the presence of fibroblast growth factor‐2 and cultivation on a three‐dimensional polymer scaffold allows differentiation of chick bone marrow stromal cells into three‐dimensional cartilaginous tissues. In thein vitrosystem studied, the same population could be selectively induced to regenerate either cartilagi

ISSN:0736-0266    

DOI:10.1002/jor.1100160205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractPrevious studies have suggested that moderate cooling increases the responsiveness of vascular α2‐adrenoceptors. However, limited information is available documenting the influence of temperature changes on adrenoceptor responses in the microvasculature of thermoregulatory organs (e.g., the human digit and the rabbit ear) subjected to a wide range of temperatures. In the present study, the effect of local cooling (24°C) on cutaneous microvascular adrenoceptors in the ear was observedin vivoin male New Zealand White rabbits (total: 66 ears). The rabbit ear was studied in a temperature‐controlled tissue bath; the ear preparation was pretreated with terazosin (an α1‐adrenoceptor antagonist) (10−5M) or a combination of terazosin (10−5M) and propranolol (a β‐adrenoceptor antagonist) (10−6M). The microvascular diameter responses of the ear to norepinephrine (10−l1‐10−4M) then were determined at 24 or 34°C, respectively, to determine the influences of low temperature on adrenoceptor responses to norepinephrine stimulation. The results demonstrated that low concentrations of norepinephrine induced vasodilation in arterioles and arteriovenous anastomoses. This vasodilation was followed by vasoconstriction with an increased concentration of norepinephrine in animals with α1‐adrenergic blockade at 34°C. Moderate tissue cooling increased the microvascular maximal response of the rabbit ear to norepinephrine and abolished the vasodilatation induced by a low concentration of norepinephrine. There was no significant difference in the microvascular response to norepinephrine between the two temperature conditions after simultaneous blockade of α1‐adrenoceptors and β‐adrenoceptors. Data from the present study indicate that moderate cooling does not enhance the responsiveness of α2‐adrenoceptors to norepinephrine. In contrast, cooling reduced the β‐adrenergic activity of arterioles and arteriovenous anas

ISSN:0736-0266    

DOI:10.1002/jor.1100160206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractA standardized and reliable model for muscle laceration injuries was developed. The biomechanical and morphological changes during the process of muscle repair after injury were analysed, and the reproducibility of the methods was evaluated. The soleus muscles of Sprague‐Dawley rats were completely transected and were allowed to heal for 5, 7, 10, 14, 21, 28, or 56 days, when the muscles either were pulled to failure to measure different parameters of tensile strength or were removed for morphological analysis. During the repair process, the regenerating myofibers penetrated into the connective‐tissue scar and formed new myotendinous junctions, thus restoring the functional continuity across the muscle stumps. The muscle atrophied significantly during the recovery period. Mechanical failure occurred in the scar until day 10, and thereafter it occurred within myofibers. Until day 10, the failure load, strain, and specific energy increased to as much as 46.59. and 36% of the control level, respectively: thereafter, there were only minor changes. Stress (tensile strength per cross‐sectional area) reached 86% of the control level by day 21 and further increased to as much as 96% of the control level until day 56. These results indicate that the scar becomes stronger than muscle within 14 days: thereafter, the weakest point is the atrophic muscle. The fact that the stress value was most rapidly normalized suggests that, qualitatively, the regenerated muscle had virtually regained its pretrauma strength by day 56 and that the low values of failure load, strain, and specific energy were mainly due to atrophy of the muscle. Thus, further increase in the tensile strength of the regenerated muscle‐tendon unit may require active exercise to reverse muscle

ISSN:0736-0266    

DOI:10.1002/jor.1100160207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractChondrocytes that were isolated from adult human articular cartilage changed phenotype during monolayer tissue culture, as characterized by a fibroblastic morphology and cellular proliferation. Increased proliferation was accompanied by downregulation of the cartilage‐specific extracellular matrix proteoglycan, aggrecan, by cessation of type‐II collagen expression, and by upregulation of type‐I collagen and versican. This phenomenon observed in monolayer was reversible after the transfer of cells to a suspension culture system. The transfer of chondrocytes to suspension culture in alginate beads resulted in the rapid upregulation of aggrecan and type‐II collagen and the downregulation of expression of versican and type‐I collagen. Type‐X collagen and osteopontin, markers of chondrocyte hypertrophy and commitment to endochondral ossification, were not expressed by adult articular chondrocytes cultured in alginate, even after 5 months. In contrast, type‐X collagen was expressed within 2 weeks in a population of cells derived from a fetal growth plate. The inability of adult articular chondrocytes to express markers of chondrocyte hypertrophy has underscored the fundamental distinction between the differentiation pathways that lead to articular cartilage or to bone. Adult articular chondrocytes expressed only hyaline articular cartilage markers without evidence o

ISSN:0736-0266    

DOI:10.1002/jor.1100160208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractSmall calcium‐mobilizing inflammatory mediators have been implicated in joint pathology. Here we demonstrate that bradykinin, adenosine 5′‐triphosphate, uridine 5′‐triphosphate, and lysophosphatidic acid raise the intracellular calcium concentration ([Ca2+]i) in human articular chondrocytes. Heterologous cross‐desensitization experiments showed that the uridine 5′‐triphosphate response was abolished by prior treatment with adenosine 5′‐triphosphate and conversely, that the adenosine 5′‐triphosphate response was abolished by prior treatment with uridine 5′‐triphosphate: this indicated competition for the same receptor site, whereas bradykinin and lysophosphatidic acid did not compete with other ligands. Pretreatment with thapsigargin abolished ligand‐mediated Ca2+responses but not vice versa: this confirmed that Ca2+release occurred from intracellular stores. Single‐cell analysis of Fura‐2 acetoxymethyl ester loaded chondrocytes showed mediator‐dependent patterns of oscillatory Ca2+changes in a subset of cells when challenged with submaximal concentrations of bradykinin, adenosine 5′‐triphosphate, or uridine 5′‐triphosphate in the presence of extracellular Ca2+. However, no oscillatory responses were seen after a challenge with lysophosphatidic acid. Therefore, although a number of different Ca2+‐mobilizing ligands activate chondrocytes, the differences that occur in the temporal patterning of Ca2+responses may result in unique me

ISSN:0736-0266    

DOI:10.1002/jor.1100160209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractA mechanically testable tissue was grownin vitrofrom rabbit chondrocytes that were initially plated at high density (approximately 80.000 cells/cm2). The DNA, collagen, and proteoglycan content, as well as the tissue thickness, tensile stiffness, and synthesis rates, were measured at 4, 6, and 8 weeks. The biochemical properties were similar to those for immature cartilage, with predominantly type‐II collagen produced; this indicated that the cells retained their chondrocytic phenotype. The tissue formed a coherent mechanical layer with testable tensile stiffness as early as 4 weeks. The tensile elastic modulus reached 1.3 MPa at 8 weeks, which is in the range of values for native cartilage from the midzone. Collagen density was approximately 24 mg/ml at 8 weeks, which is about one‐half the value for native cartilage, and the collagen fibril diameters were smaller. Chondrocytes in culture responded to culture conditions and were stimulated by cytokine interleukin‐1β. When culture conditions were varied to RPMI nutrient medium with lower fetal bovine serum and higher ascorbic acid concentrations, the thickness decreased and the modulus increased significantly. Interleukin‐1β. added to the 8‐week culture for 2 weeks. caused a decrease of 60% in thickness, a decrease of 81% in proteoglycan content, and a decrease of 31% in collagen content; this is similar to the response of cartilage explants to interleukin‐1β. This cartilage analog may be useful as a model system to study structure‐function relationships in cartilage or as cartilage‐

ISSN:0736-0266    

DOI:10.1002/jor.1100160210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractThe addition of fibronectin fragments to cultured cartilage causes an initial suppression of proteoglycan synthesis, induction of matrix metalloproteinases, and resultant decrease in proteoglycan content by about 50% during the first few days in culture. Because the proteoglycan loss appears to be limited, we investigated whether the fibronectin fragments induce anabolic responses that might counter the damage. The effects of various lengths of exposure of cultured cartilage to the fibronectin fragment on proteoglycan content, proteoglycan synthesis rates, stromelysin‐1 release, and tumor necrosis factor‐α, interleukin‐1α, and interleukin‐6 release were investigated. The results showed that about 7 days of exposure of cultured cartilage to the fibronectin fragment was required for maximal cytokine release, proteoglycan depletion, and stromelysin‐1 release. However, nearly maximal suppression of proteoglycan synthesis occurred within 1 day of the addition of the fibronectin fragment and, after its removal, the rates increased to supernormal levels. Decreasing exposure to 3 days caused only a small decrease in cartilage proteoglycan content, although stromelysin‐1 release still occurred. Decreasing exposure to 1 day caused an immediate increase in proteoglycan synthesis and an increase to supernormal proteoglycan contents. The effect of first treating cartilage with the fibronectin fragment for various periods and then allowing a recovery was to make the cartilage more resistant to secondary exposures. This study shows that cartilage damage can be caused by short exposures to the fibronectin fragment and that exposures either optimal or suboptimal for damage additionally amplify anabolic processes to make the cartilage resistant to further damage and, thus, condition it against pending amplificat

ISSN:0736-0266    

DOI:10.1002/jor.1100160211

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY