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Type II Collagen (type + ii_collagen)
Kinds of Type II Collagen Terms modified by Type II Collagen Selected AbstractsCis-Acting Intronic Elements That Regulate Cartilage-Specific Alternative Splicing of the Type II Collagen (Col2) Pre-mRNA Lie at or Near Splice Site Junction Sequences Flanking Exon 2 of the Gene,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 9 2003Takayuki Nishiyama Abstract Knowledge of the cis-acting elements is required for identifying trans-acting splicing factors underlying cartilage-specific alternative splicing of Col2 pre-mRNA. By performing desired deletions in the mouse Col2 pre-mRNA, location of the intronic cis-acting elements was narrowed down to be at or near splice-junction sequences flanking exon 2 of the gene. Introduction: Type II collagen (Col2) pre-mRNA undergoes cartilage-specific alternative splicing involving exon 2 during chondrocyte differentiation. Thus, the trans-acting protein factors that regulate the splicing are associated with the differentiation of chondrocytes. Knowledge of the cognate cis-acting elements is necessary to eventually identify the trans-acting factors. Materials and Methods: To localize the cis-acting sequences, we created several deletions within a minigene containing exon 1 to exon 4 of mouse Col 2 gene and evaluated alternative splicing of the resulting pre-mRNAs in ATDC5 cells, a model of insulin-stimulated chondrocyte differentiation. The first deletion reduced intron 1 from 3799 to 259 bp, the second reduced intron 2 from 1108 to 94 bp, the third combined the above two deletions, and the fourth was derived from the third by removing intron 3 and exon 4. ATDC5 cells harboring these constructs were cultured for up to 21 days with or without insulin. Alternative splicing was evaluated by determining the ratio of Col2B (lacks exon 2) to Col2A (has exon 2) RNAs by reverse transcription-polymerase chain reaction. Results: The deletion in intron 1 had no effect on the alternative splicing while other deletions affected splicing (demonstrated by the presence of splicing intermediates) in cells cultured without insulin or with insulin for 1 week. The splicing intermediates were not seen from any construct when cells were cultured longer (14,21 days) with insulin. Conclusion: These results show that the 259-bp intron 1, the 94-bp intron 2, and exon 2 sequences retained in the fourth construct provide cis-acting signal sufficient for insulin-induced cartilage-specific alternative splicing of Col2 pre-mRNA. [source] The Response of Articular Chondrocytes to Type II Collagen,Au NanocompositesARTIFICIAL ORGANS, Issue 12 2007Shan-hui Hsu Abstract:, The nanocomposites (denoted "CII,Au") of porcine type II collagen (CII) with 0.05, 0.1, 0.5, 1, or 2.5% (wt/wt) Au nanoparticles (,5 nm) were fabricated for potential use in cartilage tissue engineering. Au formed clusters on the surface of all nanocomposites and appeared to distribute along the collagen fibrils inside the matrix. The addition of Au at low concentrations (,0.5%) increased the modulus and viscosity, as well as the free radical-scavenging ability. These effects decreased at higher concentrations of Au. The chondrocytes on CII,Au became spindle-like with lamellipodia formation. Cell proliferation on CII,Au 0.1% was promoted. Nitric oxide (NO) in the culture medium was reduced by CII,Au 0.05% and CII,Au 0.1%. Type I collagen, aggrecan, and Sox 9 gene expressions increased with an increased Au content, but slightly decreased at 2.5% Au. There was no significant difference in the CII gene expression. The cellular uptake of Au was observed but less than that which occurred when 10 ppm of Au was added in culture medium. Chondrocytes cultured with ,10 ppm of Au nanoparticles showed neither cytotoxicity nor change in gene expression. Au at an appropriate amount could be well dispersed in CII, and enhanced the material modulus, antioxidant effect, as well as the chondrocyte growth and matrix production. [source] Type II collagen without adjuvant induces eosinophilic arthritisEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 2 2007Robert Bockermann Abstract Eosinophilia is a characteristic feature of many inflammatory diseases including inflammatory bowel disease and asthma. It also occurs in a subtype of rheumatoid arthritis but the role of eosinophils has been unclear and animal models have been lacking. Here, we introduce a new mouse model to study the role of eosinophilia in arthritis. Intraperitoneal injection of type II collagen alone, without any adjuvant, was sufficient to induce chronic arthritis in a mouse with transgenic T cells specific for type II collagen. The arthritis was accompanied by infiltration of eosinophils into the synovial tissue and the disease could be blocked with neutralizing anti-IL-5 antibodies. To our knowledge, this is the first description of an eosinophilic disease form of destructive arthritis. [source] The hip joint: the fibrillar collagens associated with development and ageing in the rabbitJOURNAL OF ANATOMY, Issue 1 2001YVETTE S. BLAND The fibrillar collagens associated with the articular cartilages, joint capsule and ligamentum teres of the rabbit hip joint were characterised from the 17 d fetus to the 2-y-old adult by immunohistochemical methods. Initially the putative articular cartilage contains types I, III and V collagens, but when cavitation is complete in the 25 d fetus, type II collagen appears. In the 17 d fetus, the cells of the chondrogenous layers express type I collagen mRNA, but not that of type II collagen. Types III and V collagens are present throughout life, particularly pericellularly. Type I collagen is lost. In all respects, the articular cartilage of the hip joint is similar to that of the knee. The joint capsule contains types I, III and V collagens. In the fetus the ligamentum teres contains types I and V collagens and the cells express type I collagen mRNA; type III collagen is confined mainly to its surface and insertions. After birth, the same distribution remains, but there is more type III collagen in the ligament, proper. The attachment to the cartilage of the head of the femur is marked only by fibres of type I collagen traversing the cartilage; the attachment cannot be distinguished in preparations localising types III and V collagens. The attachment to the bone at the lip of the acetabulum is via fibres of types I and V collagens and little type III is present. The ligament is covered by a sheath of types III and V collagens. Type II collagen was not located in any part of the ligamentum teres. The distribution of collagens in the ligamentum teres is similar to that in the collateral ligaments of the knee. Its insertions are unusual because no fibrocartilage was detected. [source] Cis-Acting Intronic Elements That Regulate Cartilage-Specific Alternative Splicing of the Type II Collagen (Col2) Pre-mRNA Lie at or Near Splice Site Junction Sequences Flanking Exon 2 of the Gene,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 9 2003Takayuki Nishiyama Abstract Knowledge of the cis-acting elements is required for identifying trans-acting splicing factors underlying cartilage-specific alternative splicing of Col2 pre-mRNA. By performing desired deletions in the mouse Col2 pre-mRNA, location of the intronic cis-acting elements was narrowed down to be at or near splice-junction sequences flanking exon 2 of the gene. Introduction: Type II collagen (Col2) pre-mRNA undergoes cartilage-specific alternative splicing involving exon 2 during chondrocyte differentiation. Thus, the trans-acting protein factors that regulate the splicing are associated with the differentiation of chondrocytes. Knowledge of the cognate cis-acting elements is necessary to eventually identify the trans-acting factors. Materials and Methods: To localize the cis-acting sequences, we created several deletions within a minigene containing exon 1 to exon 4 of mouse Col 2 gene and evaluated alternative splicing of the resulting pre-mRNAs in ATDC5 cells, a model of insulin-stimulated chondrocyte differentiation. The first deletion reduced intron 1 from 3799 to 259 bp, the second reduced intron 2 from 1108 to 94 bp, the third combined the above two deletions, and the fourth was derived from the third by removing intron 3 and exon 4. ATDC5 cells harboring these constructs were cultured for up to 21 days with or without insulin. Alternative splicing was evaluated by determining the ratio of Col2B (lacks exon 2) to Col2A (has exon 2) RNAs by reverse transcription-polymerase chain reaction. Results: The deletion in intron 1 had no effect on the alternative splicing while other deletions affected splicing (demonstrated by the presence of splicing intermediates) in cells cultured without insulin or with insulin for 1 week. The splicing intermediates were not seen from any construct when cells were cultured longer (14,21 days) with insulin. Conclusion: These results show that the 259-bp intron 1, the 94-bp intron 2, and exon 2 sequences retained in the fourth construct provide cis-acting signal sufficient for insulin-induced cartilage-specific alternative splicing of Col2 pre-mRNA. [source] Type II collagen modulates the composition of extracellular matrix synthesized by articular chondrocytesJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 2 2003Wen-Ning Qi Abstract The articular cartilage extracellular matrix (ECM) interfaces with chondrocytes and influences many biological processes important to cartilage homeostasis and repair. The alginate bead culture system can be viewed as a model of cartilage repair in which the chondrocyte attempts to recreate the pericellular matrix while maintaining a differentiated phenotype. The purpose of this study was to evaluate the alteration in epitopes of proteoglycan and tenascin synthesized by chondrocytes in the presence of exogenous extracellular type II collagen. We evaluated the effects on four biomarkers associated with the creation of the denovo matrix using ELISA and immunohistochemistry: keratan sulfate epitope (5D4), 3B3(,) neoepitope of chondroitin-6- sulfate, 3B3(+) chondroitinase-generatedepitope of chondroitin-6-sulfate, and tenascin-C expression. TGF-,1 stimulated the production of 3B3(+), 5D4, and tenascin-C in a dose-dependent manner and decreased 3B3(,) levels. Following the addition of exogenous type II collagen, 3B3(,) increased and tenascin-C decreased but did not change the direction of TGF-,1 effects. In contrast, 5D4 expression decreased in the presence of collagen II as TGF-,1 increased to 10 ng/ml. Interestingly, the amount of 3B3(+) epitope was not affected by the incorporation of type II collagen. Immunohistochemistry found there was no significant difference in distribution of these biomarkers in the presence and absence of extracellular type II collagen incorporation. These results elucidate the subtle biochemical differences in ECM synthesized by chondrocytes in the presence of type II collagen and further characterize the role played by ECM in the TGF-,1 regulation of the articular cartilage physiology. © 2002 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved. [source] Induction of a neoarthrosis by precisely controlled motion in an experimental mid-femoral defectJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 3 2002Dennis M. Cullinane Bone regeneration during fracture healing has been demonstrated repeatedly, yet the regeneration of articular cartilage and joints has not yet been achieved. It has been recognized however that the mechanical environment during fracture healing can be correlated to the contributions of either the endochondral or intramembranous processes of bone formation, and to resultant tissue architecture. Using this information, the goal of this study was to test the hypothesis that induced motion can directly regulate osteogenic and chondrogenic tissue formation in a rat mid-femoral bone defect and thereby influence the anatomical result. Sixteen male Sprague Dawley rats (400 ± 20 g) underwent production of a mid-diaphyseal, non-critical sized 3.0 mm segmental femoral defect with rigid external fixation using a custom designed four pin fixator. One group of eight animals represented the controls and underwent surgery and constant rigid fixation. In the treatment group the custom external fixator was used to introduce daily interfragmentary bending strain in the eight treatment animals (12°s angular excursion), with a hypothetical symmetrical bending load centered within the gap. The eight animals in the treatment group received motion at 1.0 Hz, for 10 min a day, with a 3 days on, one day off loading protocol for the first two weeks, and 2 days on, one day off for the remaining three weeks. Data collection included histological and immunohistological identification of tissue types, and mean collagen fiber angles and angular conformity between individual fibers in superficial, intermediate, and deep zones within the cartilage. These parameters were compared between the treatment group, rat knee articular cartilage, and the control group as a structural outcome assessment. After 35 days the control animals demonstrated varying degrees of osseous union of the defect with some animals showing partial union. In every individual within the mechanical treatment group the defect completely failed to unite. Bony arcades developed in the experimental group, capping the termini of the bone segments on both sides of the defect in four out of six animals completing the study. These new structures were typically covered with cartilage, as identified by specific histological staining for Type II collagen and proteoglycans. The distribution of collagen within analogous superficial, intermediate, and deep zones of the newly formed cartilage tissue demonstrated preferred fiber angles consistent with those seen in articular cartilage. Although not resulting in complete joint development, these neoarthroses show that the induced motion selectively controlled the formation of cartilage and bone during fracture repair, and that it can be specifically directed. They further demonstrate that the spatial organization of molecular components within the newly formed tissue, at both microanatomical and gross levels, are influenced by their local mechanical environment, confirming previous theoretical models. © 2002 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved. [source] Type II collagen levels correlate with mineralization by articular cartilage vesiclesARTHRITIS & RHEUMATISM, Issue 9 2009Brian Jubeck Objective Pathologic mineralization is common in osteoarthritic (OA) cartilage and may be mediated by extracellular organelles known as articular cartilage vesicles (ACVs). Paradoxically, ACVs isolated from OA human cartilage mineralize poorly in vitro compared with those isolated from normal porcine cartilage. We recently showed that collagens regulate ACV mineralization. We sought to determine differences between collagens and collagen receptors on human and porcine ACVs as a potential explanation of their different mineralization behaviors. Methods ACVs were enzymatically released from old and young human and porcine hyaline articular cartilage. Western blotting was used to determine the presence of types I, II, VI, and X collagen and various collagen receptors on ACVs. Type II collagen was quantified by enzyme-linked immunosorbent assay. Biomineralization was assessed by measuring the uptake of 45Ca by isolated ACVs in agarose gels and by ACVs in situ in freeze-thawed cartilage. Results As previously shown, isolated human ACVs mineralized poorly in response to ATP compared with porcine ACVs, but human and porcine ACVs mineralized similarly in situ in freeze-thawed cartilage. Type II collagen levels were 100-fold higher in isolated human ACVs than in porcine ACVs. Type II collagen in human ACVs was of high molecular weight. Transglutaminase-crosslinked type II collagen showed increased resistance to collagenase, suggesting a possible explanation for residual collagen on human ACVs. Expression of other collagens and collagen receptors was similar on human and porcine ACVs. Conclusion Higher levels of type II collagen in human ACV preparations, perhaps mediated by increased transglutaminase crosslinking, may contribute to the decreased mineralization observed in isolated human ACVs in vitro. [source] Type II collagen without adjuvant induces eosinophilic arthritisEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 2 2007Robert Bockermann Abstract Eosinophilia is a characteristic feature of many inflammatory diseases including inflammatory bowel disease and asthma. It also occurs in a subtype of rheumatoid arthritis but the role of eosinophils has been unclear and animal models have been lacking. Here, we introduce a new mouse model to study the role of eosinophilia in arthritis. Intraperitoneal injection of type II collagen alone, without any adjuvant, was sufficient to induce chronic arthritis in a mouse with transgenic T cells specific for type II collagen. The arthritis was accompanied by infiltration of eosinophils into the synovial tissue and the disease could be blocked with neutralizing anti-IL-5 antibodies. To our knowledge, this is the first description of an eosinophilic disease form of destructive arthritis. [source] Heterozygous SOX9 Mutations Allowing for Residual DNA-binding and Transcriptional Activation Lead to the Acampomelic Variant of Campomelic Dysplasia,HUMAN MUTATION, Issue 6 2010Alex Staffler Abstract Campomelic dysplasia is a malformation syndrome with multiple symptoms including characteristic shortness and bowing of the long bones (campomelia). CD, often lethal due to airway malformations, is caused by heterozygous mutations in SOX9, an SRY-related gene regulating testis and chondrocyte development including expression of many cartilage genes such as type II collagen. Male to female sex reversal occurs in the majority of affected individuals with an XY karyotype. A mild form without campomelia exists, in which sex-reversal may be also absent. We report here two novel SOX9 missense mutations in a male (c.495C>G; p.His165Gln) and a female (c.337A>G; p.Met113Val) within the DNA-binding domain leading to non-lethal acampomelic CD. Functional analyses of mutant proteins demonstrate residual DNA-binding and transactivation of SOX9-regulated genes. Combining our data and reports from the literature we postulate a genotype-phenotype correlation: SOX9 mutations allowing for residual function lead to a mild form of CD in which campomelia and sex reversal may be absent. © 2010 Wiley-Liss, Inc. [source] Antibodies to citrullinated proteins: molecular interactions and arthritogenicityIMMUNOLOGICAL REVIEWS, Issue 1 2010Hüseyin Uysal Summary:, The discovery of antibodies specific for citrullinated protein epitopes [anti-citrullinated protein antibodies (ACPAs)] is a hallmark for the diagnosis and prognosis of rheumatoid arthritis (RA) and will also be a useful tool for understanding the fundamental pathologic processes. There are several essential questions pertaining to ACPA that remain to be explored, such as understanding the early specificity of the underlying T-cell recognition, whether the production of ACPA is a primary or secondary process, and in the event of such antibodies being arthritogenic, whether they could possibly regulate the disease development. To answer these questions, animal models are needed, but unfortunately ACPA is not a prominent feature of any of the classical animal models of RA. However, we showed recently that ACPA can be isolated from animals susceptible to collagen-induced arthritis that are specific for citrullinated type II collagen (CII). The citrulline specificity could be visualized, and the specificity is determined primarily by a direct interaction with citrulline. We also demonstrated that these antibodies are specific for the citrullinated epitopes and are pathogenic in vivo. A new hypothesis to explain how inflammation in RA can be directed to cartilaginous joints and be self-perpetuating is suggested, which involves recognition of post-translational modifications (glycosylation and citrullination) on CII by T and B cells that can have both arthritogenic and regulatory consequences. [source] Sex preference of ankylosis in collagen-induced arthritis in B10.RIII miceINTERNATIONAL JOURNAL OF RHEUMATIC DISEASES, Issue 1 2006Jun ITO Abstract Aim:, The sex preference of ankylosis in collagen-induced arthritis is evaluated. Method:, Mice were immunized with bovine type II collagen emulsified with complete Freund's adjuvant H37Ra. The incidence of arthritis, arthritic score, number of paws with whole rear-paw swelling (WRPS), and number of paws with ankle ankylosis (Ankank) were monitored at various intervals after immunization. Results:, Arthritic score and number of paws with WRPS in each mouse were significantly higher in male than in female mice at 3, 5, and 8 weeks. On the other hand, there was no difference between male and female mice in arthritic score, number of paws with WRPS in each mouse, and number of paws with Ankank among ankles with WRPS at 10 and 15 weeks. Conclusion:, There was a suppressive effect on the development of arthritis in female mice. On the other hand, there was no sex difference in the incidence of ankylosis in arthritic ankles after arthritis was established. [source] An immunohistochemical study of the triangular fibrocartilage complex of the wrist: regional variations in cartilage phenotypeJOURNAL OF ANATOMY, Issue 1 2007S. Milz Abstract The triangular fibrocartilage complex (TFCC) transmits load from the wrist to the ulna and stabilizes the distal radioulnar joint. Damage to it is a major cause of wrist pain. Although its basic structure is well established, little is known of its molecular composition. We have analysed the immunohistochemical labelling pattern of the extracellular matrix of the articular disc and the meniscal homologue of the TFCC in nine elderly individuals (age range 69,96 years), using a panel of monoclonal antibodies directed against collagens, glycosaminoglycans, proteoglycans and cartilage oligomeric matrix protein (COMP). Although many of the molecules (types I, III and VI collagen, chondroitin 4 sulphate, dermatan sulphate and keratan sulphate, the oversulphated epitope of chondroitin 6 sulphate, versican and COMP) were found in all parts of the TFCC, aggrecan, link protein and type II collagen were restricted to the articular disc and to entheses. They were thus not a feature of the meniscal homologue. The shift in tissue phenotype within the TFCC, from a fibrocartilaginous articular disc to a more fibrous meniscal homologue, correlates with biomechanical data suggesting that the radial region is stiff and subject to considerable stress concentration. The presence of aggrecan, link protein and type II collagen in the articular disc could explain why the TFCC is destroyed in rheumatoid arthritis, given that it has been suggested that autoimmunity to these antigens results in the destruction of articular cartilage. The differential distribution of aggrecan within the TFCC is likely to be reflected by regional differences in water content and mobility on the radial and ulnar side. This needs to be taken into account in the design of improved MRI protocols for visualizing this ulnocarpal complex of the wrist. [source] Expression of extracellular matrix molecules typical of articular cartilage in the human scapholunate interosseous ligamentJOURNAL OF ANATOMY, Issue 6 2006S. Milz Abstract The scapholunate interosseous ligament (SLIL) connects the scaphoid and lunate bones and plays a crucial role in carpal kinematics. Its rupture leads to carpal instability and impairment of radiocarpal joint function. As the ligament is one of the first structures affected in rheumatoid arthritis, we conducted an immunohistochemical study of cadaveric tissue to determine whether it contains known autoantigens for rheumatoid arthritis. We immunolabelled the ligament from one hand in 12 cadavers with monoclonal antibodies directed against a wide range of extracellular matrix (ECM) molecules associated with both fibrous and cartilaginous tissues. The labelling profile has also enabled us to comment on how the molecular composition of the ligament relates to its mechanical function. All regions of the ligament labelled for types I, III and VI collagens, chondroitin 4 and 6 sulphates, keratan sulphate, dermatan sulphate, versican, tenascin and cartilage oligomeric matrix protein (COMP). However, both entheses labelled strongly for type II collagen, aggrecan and link protein and were distinctly fibrocartilaginous. In some regions, the ligament attached to bone via a region of hyaline cartilage that was continuous with articular cartilage. Labelling for cartilage molecules in the midsubstance was most evident dorsally. We conclude that the SLIL has an ECM which is typical of other highly fibrocartilaginous ligaments that experience both tensile load and shear. The presence of aggrecan, link protein, COMP and type II collagen could explain why the ligament may be a target for autoantigenic destruction in some forms of rheumatoid arthritis. [source] Fibrocartilage at the entheses of the suprascapular (superior transverse scapular) ligament of man,a ligament spanning two regions of a single boneJOURNAL OF ANATOMY, Issue 5 2001B. MORIGGL The suprascapular ligament converts the suprascapular notch into a foramen separating the vessels and nerve of the same name. It connects 2 regions of the same bone and does not cross any joint, and no mechanical function has yet been attributed to it. Nevertheless, variations in its thickness and length, and its tendency to ossify, suggest that the ligament responds to changes in mechanical load. This should be reflected in the composition of the extracellular matrix. The primary purpose of the present study is to demonstrate that the suprascapular ligament has fibrocartilaginous entheses (i.e. insertion sites), even though there is no obvious change in insertional angle that directly results from joint movement. Such a change is more typical of tendons or ligaments that cross highly mobile joints. The complete ligament (including both entheses) was removed from 7 cadavers shortly after death and fixed in 90% methanol. Cryosections were immunolabelled with a panel of monoclonal antibodies against collagens (types I, II, III, VI), glycosaminoglycans (chondroitin 4 sulphate, chondroitin 6 sulphate, dermatan sulphate and keratan sulphates), proteoglycans (aggrecan and versican) and link protein. Both entheses were strongly fibrocartilaginous, and a moderately fibrocartilaginous matrix was also detected throughout the remainder of the ligament. The extracellular matrix of both entheses labelled strongly for type II collagen, aggrecan and link protein. The fibrocartilaginous character of the entheses suggests that the insertion sites of the ligament are subject to both compressive and tensile loading and are regions of stress concentration. This in turn probably reflects the complex shape of the scapula and the presence of a conspicuous indentation (the suprascapular notch) near the ligament. The loading patterns may reflect either the attachment of muscles and/or the forces transmitted to the suprascapular ligament from the neighbouring coracoclavicular ligament. [source] The hip joint: the fibrillar collagens associated with development and ageing in the rabbitJOURNAL OF ANATOMY, Issue 1 2001YVETTE S. BLAND The fibrillar collagens associated with the articular cartilages, joint capsule and ligamentum teres of the rabbit hip joint were characterised from the 17 d fetus to the 2-y-old adult by immunohistochemical methods. Initially the putative articular cartilage contains types I, III and V collagens, but when cavitation is complete in the 25 d fetus, type II collagen appears. In the 17 d fetus, the cells of the chondrogenous layers express type I collagen mRNA, but not that of type II collagen. Types III and V collagens are present throughout life, particularly pericellularly. Type I collagen is lost. In all respects, the articular cartilage of the hip joint is similar to that of the knee. The joint capsule contains types I, III and V collagens. In the fetus the ligamentum teres contains types I and V collagens and the cells express type I collagen mRNA; type III collagen is confined mainly to its surface and insertions. After birth, the same distribution remains, but there is more type III collagen in the ligament, proper. The attachment to the cartilage of the head of the femur is marked only by fibres of type I collagen traversing the cartilage; the attachment cannot be distinguished in preparations localising types III and V collagens. The attachment to the bone at the lip of the acetabulum is via fibres of types I and V collagens and little type III is present. The ligament is covered by a sheath of types III and V collagens. Type II collagen was not located in any part of the ligamentum teres. The distribution of collagens in the ligamentum teres is similar to that in the collateral ligaments of the knee. Its insertions are unusual because no fibrocartilage was detected. [source] Structure of the human tibialis anterior tendonJOURNAL OF ANATOMY, Issue 4 2000WOLF PETERSEN The structure and vascular pattern of the human tibialis anterior tendon was investigated using injection techniques, light and transmission electron microscopy and immunohistochemistry. From the well vascularised peritenon, blood vessels penetrate the tendon tissue and anastomose with a longitudinally oriented intratendinous network. The distribution of blood vessels within the tibialis anterior tendon was not homogenous. The posterior part of the tendon had a complete vascular network that extends from the musculotendinous junction to the insertion at the first metatarsal and medial cuneiform bones. In the anterior half, the tissue was avascular in a zone with a length of 45,67 mm. This zone was covered by a single layer (,30 ,m) of oval shaped cells. Transmission electron microscopy showed that these cells have the characteristics of chondroid cells. This region was stained by Alcian blue at pH 1 which indicates a high concentration of acid glycosaminoglycans and immunohistochemical staining for chondroitin-4-sulphate, chondroitin-6-sulphate and aggrecan was positive. However, immunostaining for the typical cartilage specific type II collagen within this zone was negative. The location of the avascular zone corresponds to the region where the tibialis anterior tendon wraps around the superior and inferior retinacula which serve as fibrous pulleys. This is the region where most spontaneous ruptures of the tibialis anterior tendon occur. The presence of fibrocartilage within gliding tendons is a functional adaptation to compressive and shearing forces. In contrast to reports from the literature about the structure of gliding tendons wrapping around a bony pulley, the gliding zone of the tibialis anterior tendon has only a narrow layer of chondroid cells and proof of type II collagen is lacking. [source] Safety and therapeutic efficacy of undenatured type-ii collagen (UC-II) in comparison to glucosamine and chondroitin in arthritic horsesJOURNAL OF ANIMAL PHYSIOLOGY AND NUTRITION, Issue 2 2009R. C. Gupta Osteoarthritis (OA) is the most common form of arthritis, which causes severe inflammation and loss of cartilage. It is a debilitating disease that commonly affects thousands of horses each year. Recently, we assessed the anti-arthritic and anti-inflammatory potential of undenatured type II collagen (UC-II) in horses. This comparative investigation evaluated arthritic pain in horses receiving daily placebo, UC-II 320 mg/day (providing 80 mg active UC-II), 480 mg/day (providing 120 mg active UC-II), or 640 mg/day (providing 160 mg active UC-II), and glucosamine and chondroitin (5.4 g/day and 1.8 g/day, respectively, bid for the first month, and thereafter once daily) for 150 days. Pain in each leg was evaluated using the flexion test and the lameness-grading system after the initial two strides. Average pain of all four legs represented the pain for each horse. Horses receiving placebo showed no change in arthritic condition, while those receiving 320, 480, or 640 mg UC-II exhibited significant reduction in arthritic pain (P < 0.05). UC-II at 480 mg dose provided optimal effects. With this dose, reduction in overall pain was from 5.7 ± 0.0.42 (100%) to 0.7 ± 0.42 (12%); and in pain upon limb manipulation from 2.35 ± 0.37 (100%) to 0.52 ± 0.18 (22%). In regards to glucosamine and chondroitin treated group, although reduction in pain was significant compared to pretreated values, the efficacy was significantly less compared with that observed with UC-II. UC-II was found to be twice as effective as glucosamine and chondroitin in arthritic horses. Clinically, physical condition, and liver (ALP, GGT, and bilirubin), kidney (BUN and creatinine), and heart (CK) functions remained unchanged, suggesting that these supplements were well tolerated. Overall, these results demonstrate that UC-II was significantly more efficacious than glucosamine and chondroitin in arthritic horses. [source] The Assembly and Remodeling of the Extracellular Matrix in the Growth Plate in Relationship to Mineral Deposition and Cellular Hypertrophy: An In Situ Study of Collagens II and IX and Proteoglycan,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 2 2002Fackson Mwale Abstract The recent development of new specific immunoassays has provided an opportunity to study the assembly and resorption of type II and IX collagens of the extracellular matrix in relationship to endochondral calcification in situ. Here, we describe how in the bovine fetal physis prehypertrophic chondrocytes deposit an extensive extracellular matrix that, initially, is rich in both type II and type IX collagens and proteoglycan (PG; principally, aggrecan). The majority of the ,1(IX)-chains lack the NC4 domain consistent with our previous studies with cultured chondrocytes. During assembly, the molar ratio of type II/COL2 domain of the ,1(IX)-chain varied from 8:1 to 25:1. An increase in the content of Ca2+ and inorganic phosphate (Pi) was initiated in the prehypertrophic zone when the NC4 domain was removed selectively from the ,1(IX)-chain. This was followed by the progressive loss of the ,1(IX) COL2 domain and type II collagen. In the hypertrophic zone, the Ca2+/Pi molar ratio ranged from 1.56 to a maximum of 1.74, closely corresponding to that of mature hydroxyapatite (1.67). The prehypertrophic zone had an average ratio Ca2+/Pi ranging from 0.25 to 1, suggesting a phase transformation. At hypertrophy, when mineral content was maximal, type II collagen was reduced maximally in content coincident with a peak of cleavage of this molecule by collagenase when matrix metalloproteinase 13 (MMP-13) expression was maximal. In contrast, PG (principally aggrecan) was retained when hydroxyapatite was formed consistent with the view that this PG does not inhibit and might promote calcification in vivo. Taken together with earlier studies, these findings show that matrix remodeling after assembly is linked closely to initial changes in Ca2+ and Pi to subsequent cellular hypertrophy and mineralization. These changes involve a progressive and selective removal of types II and IX collagens with the retention of the PG aggrecan. [source] Differential gene expression analysis using paraffin-embedded tissues after laser microdissectionJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2003Joung-Ok Kim Abstract Recent advances in laser microdissection allow for precise removal of pure cell populations from morphologically preserved tissue sections. However, RNA from paraffin-embedded samples is usually degraded during microdissection. The purpose of this study is to determine the optimal fixative for RNA extractions from laser microdissected paraffin-embedded samples. The integrity of RNA was evaluated with the intactness of 18S and 28S ribosomal RNA by electrophoresis and by the length of individual gene transcripts using RT-PCR. The various fixatives were methacarn (a combination of methanol, chloroform, and acetic acid) and several concentrations of ethanol and isopropanol. Methacarn was the optimal fixative for RNA preservation in paraffin-embedded tissues, which included liver, lung, kidney, muscle, and limb. Based on RT-PCR analysis, methacarn fixed samples exhibited the expected RNA sizes for individual genes such as glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) and bone-related genes (e.g., alkaline phosphatase and osteonectin). The laser microdissection technique with methacarn fixation was then applied to analyze the differential gene expression between hypertrophic and proliferative chondrocytes in the growth plate of long bone. The expression of type X collagen (ColX,1), a specific gene for hypertrophic chondrocytes, was only observed in hypertrophic chondrocytes, while type II collagen (Col2,1) was observed more broadly in the growth plate as anticipated. Thus, combining laser microdissection with methacarn fixation facilitates the examination of differentially expressed genes from various tissues. © 2003 Wiley-Liss, Inc. [source] Immortalized cell lines from mouse xiphisternum preserve chondrocyte phenotypeJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2006Manas K. Majumdar Chondrocytes are unique to cartilage and the study of these cells in vitro is important for advancing our understanding of the role of these cells in normal homeostasis and disease including osteoarthritis (OA). As there are limitations to the culture of primary chondrocytes, cell lines have been developed to overcome some of these obstacles. In this study, we developed a procedure to immortalize and characterize chondrocyte cell lines from mouse xiphisternum. The cells displayed a polygonal to fibroblastic morphology in monolayer culture. Gene expression studies using quantitative PCR showed that the cell lines responded to bone morphogenetic protein 2 (BMP-2) by increased expression of matrix molecules, aggrecan, and type II collagen together with transcriptional factor, Sox9. Stimulation by IL-1 results in the increased expression of catabolic effectors including MMP-13, nitric oxide synthase, ADAMTS4, and ADAMTS5. Cells cultured in alginate responded to BMP-2 by increased synthesis of proteoglycan (PG), a major matrix molecule of cartilage. IL-1 treatment of cells in alginate results in increased release of PG into the conditioned media. Further analysis of the media showed the presence of Aggrecanase-cleaved aggrecan fragments, a signature of matrix degradation. These results show that the xiphisternum chondrocyte cell lines preserve their chondrocyte phenotype cultured in either monolayer or 3-dimensional alginate bead culture systems. In summary, this study describes the establishment of chondrocyte cell lines from the mouse xiphisternum that may be useful as a surrogate model system to understand chondrocyte biology and to shed light on the underlying mechanism of pathogenesis in OA. J. Cell. Physiol. 209: 551,559, 2006. © 2006 Wiley-Liss, Inc. [source] Bone morphogenetic protein-mediated type II collagen expression in pilomatricoma and cutaneous mixed tumorJOURNAL OF CUTANEOUS PATHOLOGY, Issue 3 2005Hideki Mieno Background:, We have previously reported that type II collagen deposition in overlying dermo,epidermal junction (DEJ) of pilomatricoma is mediated by bone morphogenetic protein 2/4 (BMP 2/4) expressed by shadow cells (SCs) of pilomatricoma. Objective:, This time, we studied the expression of type II collagen and BMP in a large number of cases of pilomatricoma and extended our study to cutaneous mixed tumor (CMT). Results:, We found type II collagen deposition in the overlying DEJ (16 of 50 cases) and in the SCs (19/50) of pilomatricoma. The number of case of type II collagen deposition in DEJ (DEJ+) and in SCs (SC+) of pilomatricoma correlated to the chronological stage of pilomatricoma. We also found type II collagen deposition in overlying DEJ (two of 11) and in the stromal chondroid tissue (four of 11) of CMT. BMP 2 was expressed in most cases of pilomatricoma (37/50) and CMT (seven of 11). Conclusions:, The expression of type II collagen in pilomatricoma is dependent upon the chronological stage of pilomatricoma. Type II collagen expression in the overlying DEJ and chondroid matrix in CMT may be induced by BMP via the same mechanism as in pilomatricoma. [source] Nanostructure of collagen fibrils in human nucleus pulposus and its correlation with macroscale tissue mechanicsJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 4 2010Darwesh M.K. Aladin Abstract Collagen fibrils are the main structural components of the nucleus pulposus tissue in the intervertebral discs. The structure,property relationship of the nucleus pulposus (NP) tissues is still unclear. We investigated the structure of individual collagen fibrils of the NP and evaluated its correlation with the bulk mechanical properties of the tissue. Collagen fibrils were extracted from the NP of discs retrieved from adolescents during scoliosis correction surgery, and the extracts were confirmed by SDS-PAGE. The diameters of the individual collagen fibrils were measured through atomic force microscopy, and the compressive mechanical properties of the tissues were evaluated by confined compression. The correlations between the nanoscale morphology of the collagen fibrils and the macroscale mechanical properties of the tissues were evaluated by linear regression. The SDS-PAGE results showed that the fibril extracts were largely composed of type II collagen. The mean diameter of the collagen fibrils was 92.1,±,26.54 nm; the mean swelling pressure and compressive modulus of the tissues were 6.15,±,4.3 kPa and 1.23,±,0.7 MPa, respectively. The mean fibril diameter had no linear correlation (R2,=,0.30) with the swelling pressure of the tissues. However, it had a mild linear correlation with the compressive modulus (p,=,0.023, R2,=,0.68). This is the first study, to our knowledge, to evaluate the nanostructure of the individual collagen fibrils of the nucleus pulposus and its relationship with macroscale mechanical properties of the NP tissues. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:497,502, 2010 [source] Low-intensity pulsed ultrasound (LIPUS) increases the articular cartilage type II collagen in a rat osteoarthritis modelJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 3 2010Kiyohito Naito Abstract In this study, the effect of low-intensity pulsed ultrasound (LIPUS) on cartilage was evaluated in a rat osteoarthritis (OA) model using serum biomarkers such as CTX-II (type II collagen degradation) and CPII (type II collagen synthesis) as well as histological criteria (Mankin score and immunohistochemical type II collagen staining). OA was surgically induced in the knee joint of rats by anterior cruciate/medial collateral ligament transection and medial meniscus resection (ACLT,+,MMx). Animals were divided into three groups: sham-operated group (Sham), ACLT,+,MMx group without LIPUS (,LIPUS), and ACLT,+,MMx group with LIPUS (+LIPUS; 30 mW/cm2, 20 min/day for 28 days). CTX-II levels were elevated in both ,LIPUS and +LIPUS groups compared to that in the Sham group after the operation, but there was no significant difference between +LIPUS and ,LIPUS groups, suggesting that LIPUS does not affect the degradation of type II collagen in this model. In contrast, CPII was significantly increased in +LIPUS group compared to ,LIPUS and Sham. Moreover, histological damage on the cartilage (Mankin score) was ameliorated by LIPUS, and type II collagen was immunohistochemically increased by LIPUS in the cartilage of an OA model. Of interest, mRNA expression of type II collagen was enhanced by LIPUS in chondrocytes. Together these observations suggest that LIPUS is likely to increase the type II collagen synthesis in articular cartilage, possibly via the activation of chondrocytes and induction of type II collagen mRNA expression, thereby exhibiting chondroprotective action in a rat OA model. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:361,369, 2010 [source] Formation of cartilage repair tissue in articular cartilage defects pretreated with microfracture and covered with cell-free polymer-based implants,JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 10 2009Christoph Erggelet Abstract The aim of our study was to evaluate the mid-term outcome of a cell-free polymer-based cartilage repair approach in a sheep cartilage defect model in comparison to microfracture treatment. Cell-free, freeze-dried implants (chondrotissue®) made of a poly-glycolic acid (PGA) scaffold and hyaluronan were immersed in autologous serum and used for covering microfractured full-thickness articular cartilage defects of the sheep (n,=,4). Defects treated with microfracture only served as controls (n,=,4). Six months after implantation, cartilage implants and controls were analyzed by immunohistochemical staining of type II collagen, histological staining of proteoglycans, and histological scoring. Histological analysis showed the formation of a cartilaginous repair tissue rich in proteoglycans. Histological scoring documented significant improvement of repair tissue formation when the defects were covered with the cell-free implant, compared to controls treated with microfracture. Immunohistochemistry showed that the cell-free implant induced cartilaginous repair tissue and type II collagen. Controls treated with microfracture showed marginal formation of a mixed-type repair tissue consisting of cartilaginous tissue and fibro-cartilage. Covering of microfractured defects with the cell-free polymer-based cartilage implant is suggested to be a promising treatment option for cartilage defects and improves the regeneration of articular cartilage. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1353,1360, 2009 [source] Paracrine effect of transplanted rib chondrocyte spheroids supports formation of secondary cartilage repair tissue,JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 9 2009Kolja Gelse Abstract The study's objective was to investigate if transplanted chondrocyte or periosteal cell spheroids have influence on ingrowing bone marrow-derived cells in a novel cartilage repair approach in miniature pigs. Autologous rib chondrocytes or periosteal cells were cultured as spheroids and press-fitted into cavities that were milled into large, superficial chondral lesions of the patellar joint surface. Within the milled cavities, the subchondral bone plate was either penetrated or left intact (full-thickness or partial-thickness cavities). The transplantation of chondrocyte spheroids into full-thickness cavities induced the formation of additional secondary repair cartilage that exceeded the original volume of the transplanted spheroids. The resulting continuous tissue was rich in proteoglycans and stained positive for type II collagen. Cell labeling revealed that secondarily invading repair cells did not originate from transplanted spheroids, but rather from arroded bone marrow. However, secondary invasion of repair cells was less pronounced following transplantation of periosteal cells and absent in partial-thickness cavities. According to in vitro analyses, these observations could be ascribed to the ability of chondrocyte spheroids to secrete relevant amounts of bone morphogenetic protein-2, which was not detected for periosteal cells. Transplanted chondrocyte spheroids exert a dual function: they provide cells for the repair tissue and have a stimulatory paracrine activity, which promotes ingrowth and chondrogenesis of bone marrow-derived cells. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res [source] Comparison of articular and auricular cartilage as a cell source for the autologous chondrocyte implantationJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 7 2009Elvira Mali Abstract Articular (medial femoral condyle) and auricular cartilage (anithelix) was compared as a cell source for the autologous joint repair. Cells isolated from five human cadaveric donors were cultured parallel in the monolayer cultures and in the 3D alginate hydrogel constructs for 1 week. Cell morphology was controlled by the fluorescent microscopy and gene expressions of type I collagen (COL1), type II collagen (COL2), aggrecan (AGR), versican (VER), and elastin (ELS) were analyzed by the real-time polymerase chain reaction. COL1 and ELS, predominant in the phenotype of auricular biopsy, were statistically lower in the articular biopsies. Even though COL2 and AGR decreased in monolayers of both cell sources, the dedifferentiation process affected auricular cells intensely. Cells embedded in the alginate hydrogel directly after the isolation did not exhibit the dedifferentiated phenotype. Additionally, COL1, COL2, AGR, and VER were comparable between the two sources. ELS however, remained higher in the auricular cells regardless of the culture type. The study indicates that auricular chondrocytes cultured in a 3D environment immediately after the isolation have a neo-cartilage potential for the articular surface reconstruction. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 943,948, 2009 [source] Prostaglandin E2 inhibits BMP signaling and delays chondrocyte maturationJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 6 2009Christine A. Clark Abstract While cyclooxygenases are important in endochondral bone formation during fracture healing, mechanisms involved in prostaglandin E2 (PGE2) regulation of chondrocyte maturation are incompletely understood. The present study was undertaken to determine if PGE2 effects on chondrocyte differentiation are related to modulation of the bone morphogenetic protein (BMP) signaling pathway. In primary murine sternal chondrocytes, PGE2 differentially regulated genes involved in differentiation. PGE2 induced type II collagen and MMP-13, had minimal effects on alkaline phosphatase, and inhibited the expression of the maturational marker, type X collagen. In BMP-2,treated cultures, PGE2 blocked the induction of type X collagen. All four EP receptors were expressed in chondrocytes and tended to be inhibited by BMP-2 treatment. RCJ3.1C5.18 chondrocytes transfected with the protein kinase A (PKA) responsive reporter, CRE-luciferase, showed luciferase induction following exposure to PGE2, consistent with activation of PKA signaling and the presence of the EP2 and EP4 receptors. Both PGE2 and the PKA agonist, dibutyryl cAMP, blocked the induction of the BMP-responsive reporter, 12XSBE, by BMP-2 in RCJ3.1C5.18 chondrocytes. In contrast, PGE2 increased the ability of TGF-, to activate the TGF-,-responsive reporter, 4XSBE. Finally, PGE2 down-regulated BMP-mediated phosphorylation of Smads 1, 5, and 8 in RCJ3.1C5.18 cells and in primary murine sternal chondrocytes. Altogether, the findings show that PGE2 regulates chondrocyte maturation in part by targeting BMP/Smad signaling and suggest an important role for PGE2 in endochondral bone formation. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 785,792, 2009 [source] Sustained hypoxia enhances chondrocyte matrix synthesisJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 6 2009Christian H. Coyle Abstract Articular cartilage is an avascular tissue with chondrocytes in the deeper zones existing under conditions of sustained hypoxia. Using a hypoxic chamber to provide controlled hypoxia, this study was performed to determine whether sustained hypoxia enhances the production of cartilage matrix proteins. Freshly isolated primary bovine articular chondrocytes were encapsulated in three-dimensional alginate beads and maintained at 2% oxygen with media changes using media pre-equilibrated to 2% oxygen. Immunolocalization of HIF-1, was performed to verify hypoxic conditions. Sustained hypoxia resulted in an increase in proteoglycan synthesis after only 1 day, as measured by 35S-sulfate incorporation. This increase was maintained for the duration of the 17 day study. After 17 days of hypoxic culture, increases in total type II collagen and COL2A1 gene expression were probed by indirect immunofluorescence, type II collagen ELISA, and real-time qPCR; in addition, increased glycosaminoglycan deposition was observed as determined by chemical analysis. These studies show that sustained hypoxia enhances articular chondrocyte matrix synthesis and viability in three-dimensional alginate culture. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 793,799, 2009 [source] Differential effects of static and dynamic compression on meniscal cell gene expressionJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 6 2003Maureen L. Upton Abstract Cells of the meniscus are exposed to a wide range of time-varying mechanical stimuli that may regulate their metabolic activity in vivo. In this study, the biological response of the meniscus to compressive stimuli was evaluated in vitro, using a well-controlled explant culture system. Gene expression for relevant extracellular matrix proteins was quantified using real-time RT-PCR following a 24 h period of applied static (0.1 MPa compressive stress) or dynamic compression (0.08,0.16 MPa). Static and dynamic compression were found to differentially regulate mRNA levels for specific proteins of the extracellular matrix. Decreased mRNA levels were observed for decorin (,2.1 fold-difference) and type II collagen (,4.0 fold-difference) following 24 h of dynamic compression. Decorin mRNA levels also decreased following static compression (,4.5 fold-difference), as did mRNA levels for both types I (,3.3 fold-difference) and II collagen (,4.0 fold-difference). Following either static or dynamic compression, mRNA levels for aggrecan, biglycan and cytoskeletal proteins were unchanged. It is noteworthy that static compression was associated with a 2.6 fold-increase in mRNA levels for collagenase, or MMP-1, suggesting that the homeostatic balance between collagen biosynthesis and catabolism was altered by the mechanical stimuli. These findings demonstrate that the biosynthetic response of the meniscus to compression is regulated, in part, at the transcriptional level and that transcription of types I and II collagen as well as decorin may be regulated by common mechanical stimuli. © 2003 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved. [source] | ||||||||||||||||||||||||||||