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1.
Dev Dyn ; 249(6): 711-722, 2020 06.
Article in English | MEDLINE | ID: mdl-32022343

ABSTRACT

BACKGROUND: Cruciate ligament (CL) and patellar tendon (PT) are important elements of the knee joint, uniting femur, patella, and tibia into a single functional unit. So far, knowledge on the developmental mechanism of CL, PT, and patella falls far behind other skeletal tissues. RESULTS: Here, employing various lineage tracing strategies we investigate the cellular sources and dynamics that drive CL, PT, and patella formation during mouse embryonic development. We show that Gdf5 and Gli1 are generally expressed in the same cell population that only contributes to CL, but not PT or patella development. In addition, Col2 is expressed in two independent cell populations before and after joint cavitation, where the former contributes to the CL and the dorsal part of the PT and the latter contributes to the patella. Moreover, Prrx1 is always expressed in CL and PT progenitors, but not patella progenitors where it is switched off after joint cavitation. Finally, we reveal that patella development employs different cellular dynamics before and after joint cavitation. CONCLUSIONS: Our findings delineate the expression changes of several skeletogenesis-related genes before and after joint cavitation, and provide an indication on the cellular dynamics underlying ligament, tendon, and sesamoid bone formation during embryogenesis.


Subject(s)
Patella/cytology , Patella/metabolism , Posterior Cruciate Ligament/cytology , Posterior Cruciate Ligament/metabolism , Animals , Female , Knee Joint/cytology , Knee Joint/metabolism , Mice , Patellar Ligament/cytology , Patellar Ligament/metabolism , Pregnancy , Tendons/cytology , Tendons/metabolism , Transcription Factors/metabolism
2.
Int Orthop ; 39(1): 183-91, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25416119

ABSTRACT

PURPOSE: The adult human posterior cruciate ligament (PCL) has poor functional healing response. The synovial tissue, which surrounds the PCL ligament, might be the major regulator of the microenvironment in the joint cavity after PCL injury, thus affecting the healing process. Here we establish a novel co-culture system for PCL fibroblasts and synovial cells (SC) in vitro to explore the direct influence of paracrine on PCL cells by characterizing the different expressions of the lysyl oxidase family (LOXs) and matrix metalloproteinases (MMP-1, 2, 3), which respectively facilitate extracellular matrix (ECM) repair and degradation. METHODS: Total RNA was harvested, reverse transcribed and assessed by semi-quantitative PCR and real-time PCR for the expression of LOXs and MMP-1, 2, 3 messenger RNAs. MMP-2 activity was assayed from the collected culture media samples by using zymography. RESULTS: We found co-culture could promote gene expressions of the LOXs and MMP-1, 2, 3 in normal PCL fibroblasts. But in injured PCL, we found that matrix crosstalk induced an increase of the MMP-1, 2, 3 expressions and a down-regulation of the LOXs. CONCLUSION: Based on these results, the crosstalk between PCL and SC strongly modified homeostatic balance of ECM and appeared to have a significant impact on PCL wound healing; decreased expression of cross-linking enzymes (LOXs) and increased expression of ECM-degrading proteinases (MMP-1, 2, 3) might be of great contribution to poor healing ability of PCL ligament.


Subject(s)
Fibroblasts/metabolism , Matrix Metalloproteinases/metabolism , Posterior Cruciate Ligament/cytology , Protein-Lysine 6-Oxidase/metabolism , Synovial Membrane/metabolism , Adult , Cells, Cultured , Coculture Techniques , Extracellular Matrix/metabolism , Fibroblasts/cytology , Gene Expression , Humans , Posterior Cruciate Ligament/metabolism , Real-Time Polymerase Chain Reaction , Synovial Membrane/cytology , Wound Healing/physiology
3.
J Arthroplasty ; 26(3): 504.e9-504.e12, 2011 Apr.
Article in English | MEDLINE | ID: mdl-20462737

ABSTRACT

Histologic analysis of the posterior cruciate ligament has been reported in the normal and osteoarthritic knee but not after cruciate-retaining (CR) total knee arthroplasty (TKA). Retention of the posterior cruciate ligament during TKA has been debated as to whether it is beneficial in stability and function. If the presence of mechanoreceptors is shown to be maintained in CR TKA, then there may be an argument for retention. This case report used a retrieval of a well-functioning TKA specimen that had a CR TKA. To prove the presence of mechanoreceptors within the ligament, immunohistochemistry techniques using S100 protein and neurofilament protein were used. This specimen had pacini and lamellar type of mechanoreceptors present on immunohistochemistry analysis. The presence or retention of mechanoreceptors and innervations of the ligament may indicate an advantage when retained during TKA.


Subject(s)
Arthroplasty, Replacement, Knee/instrumentation , Knee Prosthesis , Mechanoreceptors/cytology , Posterior Cruciate Ligament/cytology , Biomechanical Phenomena , Device Removal , Humans , Knee Joint/physiology , Mechanoreceptors/metabolism , Neurofilament Proteins/metabolism , Posterior Cruciate Ligament/metabolism , S100 Proteins/metabolism
4.
Genesis ; 48(11): 635-44, 2010 Nov.
Article in English | MEDLINE | ID: mdl-20806356

ABSTRACT

Sox9 expression defines cell progenitors in a variety of tissues during mouse embryogenesis. To establish a genetic tool for cell-lineage tracing and gene-function analysis, we generated mice in which the CreERT2 gene was targeted to the endogenous mouse Sox9 locus. In Sox9(CreERT2/+) ;R26R embryos, tamoxifen activated Cre recombinase exclusively in Sox9-expressing tissues. To determine the suitability of this mouse line for developmental stage-specific gene recombination, we investigated the cellular origins of the cruciate ligaments of the knee joint and the limb tendons, in which precursor cells have not been defined. The cells in these tissues were labeled after tamoxifen treatment before or at the stage of chondrogenic mesenchymal condensation, indicating that ligament and tendon cells originated from Sox9-expressing cells and that cell fate determination occurred at mesenchymal condensation. This mouse line is a valuable tool for the temporal genetic tracing of the progeny of, and inducible gene modification in Sox9-expressing cells.


Subject(s)
Anterior Cruciate Ligament/embryology , Knee Joint/embryology , Posterior Cruciate Ligament/embryology , SOX9 Transcription Factor/metabolism , Stem Cells/physiology , Tendons/embryology , Animals , Anterior Cruciate Ligament/cytology , Anterior Cruciate Ligament/metabolism , Anterior Cruciate Ligament/physiology , Cell Differentiation/genetics , Cells, Cultured , Embryo, Mammalian , Extremities/embryology , Extremities/physiology , Female , Knee Joint/metabolism , Mice , Mice, Transgenic , Organogenesis/genetics , Organogenesis/physiology , Posterior Cruciate Ligament/cytology , Posterior Cruciate Ligament/metabolism , Posterior Cruciate Ligament/physiology , Pregnancy , SOX9 Transcription Factor/genetics , Stem Cells/metabolism , Tendons/metabolism
5.
J Orthop Res ; 17(5): 748-54, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10569487

ABSTRACT

A method for measuring the expression of integrin subunits on the cell surface of knee ligament fibroblasts was developed with use of flow cytometry and immunofluorescence. The ligament cells exhibited uniform size and density, as shown by forward and side-scatter properties, and showed minimal nonspecific binding of isotype control antibodies compared with unstained cells. All cells expressed the alpha5 integrin subunit; lateral collateral ligament cells stained with antibody to alpha5 showed a mean fluorescence intensity 2-fold higher than that of medial collateral ligament cells, 1.5-fold higher than that of posterior cruciate ligament cells, and 3-fold higher than that of anterior cruciate ligament cells, indicating a greater expression of the alpha5 subunit by lateral collateral ligament cells than by medial collateral, posterior cruciate, and anterior cruciate ligament cells. All cells expressed the beta1 integrin subunit; the expression by posterior cruciate ligament cells was 3-fold higher than that by medial collateral ligament or lateral collateral ligament cells and 5-fold higher than that by anterior cruciate ligament cells. All cells expressed the beta3 integrin subunit; the expression by posterior cruciate ligament cells was 1.5, 3, and 4.5-fold greater than that by lateral collateral, anterior cruciate, and medial collateral ligament cells, respectively. Our data suggest there is a differential expression of integrin subunits in knee ligament fibroblasts, and this in part may explain differences in their attachment and adherence to extracellular matrix molecules.


Subject(s)
Integrins/biosynthesis , Ligaments, Articular/cytology , Animals , Anterior Cruciate Ligament/cytology , Anterior Cruciate Ligament/metabolism , Antigens, CD/analysis , Antigens, CD/biosynthesis , Dogs , Fibroblasts/chemistry , Fibroblasts/metabolism , Flow Cytometry , Fluorescent Antibody Technique , Integrin alpha5 , Integrin alphaV , Integrin beta1/analysis , Integrin beta1/biosynthesis , Integrins/analysis , Knee , Ligaments, Articular/metabolism , Male , Medial Collateral Ligament, Knee/cytology , Medial Collateral Ligament, Knee/metabolism , Posterior Cruciate Ligament/cytology , Posterior Cruciate Ligament/metabolism
6.
Anat Embryol (Berl) ; 200(3): 325-34, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10463347

ABSTRACT

The structure and vascularization of the human anterior and posterior cruciate ligament were investigated by light microscopy, transmission electron microscopy,, injection techniques and by immunohistochemistry. The major part of the anterior and posterior cruciate ligament is composed of bundles of type I collagen. Type III collagen-positive fibrils separate the bundles. The major cell type is the elongated fibroblast, lying solitarily between the parallel collagen fibrils. The histologic structure of the cruciate ligaments is not homogeneous. In both ligaments there is a zone where the tissue resembles fibrocartilage. In the anterior cruciate ligament the fibrocartilaginous zone is located 5-10 mm proximal of the tibial ligament insertion in the anterior portion of the ligament. In the posterior cruciate ligament the fibrocartilage is located in the central part of the middle third. Within those zones the cells are arranged in columns and the cell shape is round to ovoid. Transmission electron microscopy reveals typical features of chondrocytes. The chondrocytes are surrounded by a felt-like pericellular matrix, a high content of cellular organelles and short processes on the cell surface. The pericellular collagen is positive for type II collagen. The major blood supply of the cruciate ligaments arises from the middle geniculate artery. The distal part of both cruciate ligaments is vascularized by branches of the lateral and medial inferior geniculate artery. Both ligaments are surrounded by a synovial fold where the terminal branches of the middle and inferior arteries form a periligamentous network. From the synovial sheath blood vessels penetrate the ligament in a horizontal direction and anastomose with a longitudinally orientated intraligamentous vascular network. The density of blood vessels within the ligaments is not homogeneous. In the anterior cruciate ligament an avascular zone is located within the fibrocartilage of the anterior part where the ligament faces the anterior rim of the intercondylar fossa. The fibrocartilaginous zone of the middle third of the posterior cruciate ligament is also avascular. According to Pauwel's theory of the "causal histogenesis" (1960) the stimulus for the development of fibrocartilage within dense connective tissue is shearing and compressive stress. In the anterior cruciate ligament this biomechanical situation may occur when the ligament impinges on the anterior rim of the intercondylar fossa when the knee is fully extended. Compressive and shearing stress in the center of the middle third of the posterior cruciate ligament may result from twisting of the fiber bundles.


Subject(s)
Anterior Cruciate Ligament/anatomy & histology , Posterior Cruciate Ligament/anatomy & histology , Adult , Aged , Anterior Cruciate Ligament/blood supply , Anterior Cruciate Ligament/cytology , Blood Vessels/cytology , Cartilage/cytology , Chondrocytes/diagnostic imaging , Collagen/analysis , Female , Humans , Immunohistochemistry , Knee Joint/anatomy & histology , Knee Joint/cytology , Laminin/analysis , Male , Microscopy, Electron , Middle Aged , Posterior Cruciate Ligament/blood supply , Posterior Cruciate Ligament/cytology , Ultrasonography
7.
Folia Morphol (Warsz) ; 59(4): 285-90, 2000.
Article in English | MEDLINE | ID: mdl-11107700

ABSTRACT

Investigations were carried out on 43 serially sectioned human embryos of developmental stages 18 to 23. The homogeneous interzone of the future knee joint is observed in embryos at stage 18. During stage 19 this interzone is differentiated into dense, intensively stained, peripheral parts, which are the primordia of menisci and the medial portion, in which the cruciate ligaments are formed. All structures of the interior of the knee joint are more clearly delineated during stage 20, and they are well developed during the last embryonic week (stages 21-23).


Subject(s)
Anterior Cruciate Ligament/embryology , Embryo, Mammalian/physiology , Embryonic and Fetal Development , Ligaments, Articular/embryology , Posterior Cruciate Ligament/embryology , Anterior Cruciate Ligament/cytology , Embryo, Mammalian/cytology , Gestational Age , Humans , Knee Joint/cytology , Knee Joint/embryology , Ligaments, Articular/cytology , Posterior Cruciate Ligament/cytology
8.
Acta Orthop Belg ; 60 Suppl 1: 57-61, 1994.
Article in English | MEDLINE | ID: mdl-8053342

ABSTRACT

Surgical reconstruction of the PCL has not yet gained the acceptance that ACL reconstruction has achieved. However, in selecting an autograft to restore PCL function in symptomatic posterior knee instability, the free patellar tendon autograft is commonly used at present. Knowledge of the basics in graft healing and of factors regulating this healing process are still limited. It is of interest to determine the biologic response and final morphology of a patellar tendon autograft after PCL replacement. Based on morphological studies in PCL replacement in a sheep model the patellar tendon autograft under-goes necrosis and degeneration followed by a gradual healing process comprising revitalization (i.e. revascularization and cellular proliferation), formation of extracellular matrix components and remodeling. The autograft bone pegs become osseointegrated by 6 weeks. After 2 years, the autograft tissue differs structurally from a ligament, suggesting that the autograft may never approach normal ligament characteristics. Degenerative alterations in the core region of the autograft, the widespread presence of type III collagen and fibronectin, as well as the predominance of thin collagen fibrils do not favor a ligamentization process. The understanding of the autograft healing process remains the prerequisite for a realistic assessment of the biologic PCL replacement and will be a baseline of studies with the goal of influencing the healing process and thus improving the clinical results.


Subject(s)
Bone Transplantation/methods , Patellar Ligament/transplantation , Posterior Cruciate Ligament/surgery , Animals , Cell Division , Collagen/analysis , Extracellular Matrix/chemistry , Fibronectins/analysis , Osseointegration , Posterior Cruciate Ligament/cytology , Sheep , Transplantation, Autologous
9.
Cell Prolif ; 42(4): 448-60, 2009 Aug.
Article in English | MEDLINE | ID: mdl-19489981

ABSTRACT

OBJECTIVES: Mesenchymal stem cells have great potential for tissue regeneration, and these cells can be harvested from a variety of tissues; however, up to now it has not been clear whether stem cells could be isolated from cruciate ligaments of the knee joint. The aim of our study was to isolate and characterize stem cells from both anterior and posterior cruciate ligaments (ACL and PCL) of humans. MATERIALS AND METHODS: Cruciate ligaments were obtained from patients receiving total knee arthroplasty for advanced osteoarthritis and plastic-adherent cells were serially passaged. In vitro chondrogenic, osteogenic and adipogenic abilities of the cells were evaluated by reverse transcriptase-polymerase chain reaction and histological study. Karyotyping and surface immunophenotyping of the cells were performed. RESULTS: It was found that a population of ligament-derived cells could be expanded and subcultured extensively. These cells were able to differentiate into osteoblasts, chondrocytes and adipocytes under appropriate inductions. Their phenotypic characteristics were similar to those of bone marrow mesenchymal stem cells. Karyotyping was normal after serial passage. CONCLUSIONS: In summary, our study demonstrates that human multipotent stem cells can be isolated and expanded from human ACL and PCL, which are easily obtained from patients following total knee or cruciate ligament reconstructive surgery. Self-renewal and mesodermal differentiation potential of these cells make them a viable alternative source for use in regenerative medicine.


Subject(s)
Anterior Cruciate Ligament/cytology , Cell Culture Techniques , Knee Joint/cytology , Mesenchymal Stem Cells/cytology , Multipotent Stem Cells/cytology , Posterior Cruciate Ligament/cytology , Adipogenesis , Aged , Cell Differentiation , Cell Proliferation , Cell Separation , Chondrogenesis , Female , Gene Expression , Humans , Karyotyping , Male , Mesenchymal Stem Cells/immunology , Middle Aged , Multipotent Stem Cells/immunology , Osteogenesis , Phenotype , Synovial Membrane/cytology
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