Ultrastructural, tomographic and confocal imaging of the chondrocyte primary cilium in situ

Hyaline cartilage chondrocytes express one primary cilium per cell, but its function remains unknown. We examined the ultrastructure of chick embryo sternal chondrocyte cilia and their interaction with extracellular matrix molecules by transmission electron microscopy (TEM) and, for the first time, double‐tilt electron tomography. Ciliary bending was also examined by confocal immunohistochemistry. Tomography and TEM showed the ciliary axoneme to interdigitate amongst collagen fibres and condensed proteoglycans. TEM also revealed the presence of electron‐opaque particles in the proximal axoneme which may represent intraciliary‐transport (ICT) particles. We observed a wide range of ciliary bending patterns. Some conformed to a heavy elastica model associated with shear stress. Others were acutely deformed, suggesting ciliary deflection by collagen fibres and proteoglycans with which the cilia make contact. We conclude that mechanical forces transmitted through these matrix macromolecules bend the primary cilium, identifying it as a potential mechanosensor involved in skeletal patterning and growth.

[1]  R. Loeser,et al.  CD44 and integrin matrix receptors participate in cartilage homeostasis , 2002, Cellular and Molecular Life Sciences CMLS.

[2]  S. Jimenez,et al.  Ultrastructural study of chondrocytes from fibrillated and non-fibrillated human osteoarthritic cartilage. , 1996, Osteoarthritis and cartilage.

[3]  W. Dentler,et al.  Microtubule-membrane interactions in cilia and flagella. , 1981, International review of cytology.

[4]  D. Wheatley,et al.  CONFOCAL ANALYSIS OF PRIMARY CILIA STRUCTURE AND COLOCALIZATION WITH THE GOLGI APPARATUS IN CHONDROCYTES AND AORTIC SMOOTH MUSCLE CELLS , 1997, Cell biology international.

[5]  M. Knight,et al.  Mechanical compression influences intracellular Ca2+ signaling in chondrocytes seeded in agarose constructs. , 2001, Journal of applied physiology.

[6]  A. Vogel,et al.  Single cilia in the articular cartilage of the cat. , 1979, Experimental cell biology.

[7]  M Marko,et al.  The Emergence of Electron Tomography as an Important Tool for Investigating Cellular Ultrastructure , 2001, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[8]  Lesilee S. Rose,et al.  Role of a Class Dhc1b Dynein in Retrograde Transport of Ift Motors and Ift Raft Particles along Cilia, but Not Dendrites, in Chemosensory Neurons of Living Caenorhabditis elegans , 1999, The Journal of cell biology.

[9]  M. A. R. Freeman,et al.  Adult Articular Cartilage , 1973 .

[10]  J. Rosenbaum,et al.  Localization of intraflagellar transport protein IFT52 identifies basal body transitional fibers as the docking site for IFT particles , 2001, Current Biology.

[11]  K. Spring,et al.  Removal of the MDCK Cell Primary Cilium Abolishes Flow Sensing , 2003, The Journal of Membrane Biology.

[12]  P. Beech,et al.  Chlamydomonas Kinesin-II–dependent Intraflagellar Transport (IFT): IFT Particles Contain Proteins Required for Ciliary Assembly in Caenorhabditis elegans Sensory Neurons , 1998, The Journal of cell biology.

[13]  E. A. Schwartz,et al.  Analysis and modeling of the primary cilium bending response to fluid shear. , 1997, The American journal of physiology.

[14]  A. Bajer,et al.  Effects of dehydration on the microtubules of the mitotic spindle. Studies in vitro and with the electron microscope. , 1969, Journal of ultrastructure research.

[15]  C. Jacobs,et al.  Effects of fluid flow on intracellular calcium in bovine articular chondrocytes. , 1997, The American journal of physiology.

[16]  J. Frank,et al.  Double-tilt electron tomography. , 1995, Ultramicroscopy.

[17]  K. Kozminski,et al.  A motility in the eukaryotic flagellum unrelated to flagellar beating. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[18]  N. Wilsman.  Cilia of adult canine articular chondrocytes. , 1978, Journal of ultrastructure research.

[19]  C A Poole,et al.  Analysis of the morphology and function of primary cilia in connective tissues: a cellular cybernetic probe? , 1985, Cell motility.

[20]  Helen Muir,et al.  The chondrocyte, architect of cartilage. Biomechanics, structure, function and molecular biology of cartilage matrix macromolecules , 1995, BioEssays : news and reviews in molecular, cellular and developmental biology.

[21]  C. Farnum,et al.  Incidence and morphology of equine and murine chondrocytic cilia , 1980, The Anatomical record.

[22]  C. Rieder,et al.  Flexible-substratum technique for viewing cells from the side: some in vivo properties of primary (9+0) cilia in cultured kidney epithelia. , 1988, Journal of cell science.

[23]  A Khodjakov,et al.  The disassembly and reassembly of functional centrosomes in vitro. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[24]  K. R. Spring,et al.  Bending the MDCK Cell Primary Cilium Increases Intracellular Calcium , 2001, The Journal of Membrane Biology.

[25]  N. Wilsman,et al.  Cilia of neonatal articular chondrocytes incidence and morphology , 1978, The Anatomical record.

[26]  V C Mow,et al.  The mechanical properties of articular cartilage. , 1983, Bulletin of the Hospital for Joint Diseases Orthopaedic Institute.

[27]  Buu P. Tu,et al.  Mechanisms of chondrocyte adhesion to cartilage: role of β1‐integrins, CD44, and annexin V , 2001, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[28]  J. Rosenbaum,et al.  Intraflagellar transport , 2002, Nature Reviews Molecular Cell Biology.

[29]  Mechanically induced calcium waves in articular chondrocytes are inhibited by gadolinium and amiloride , 1999, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[30]  D. Wheatley,et al.  Primary cilia in cultured mammalian cells: detection with an antibody against detyrosinated alpha-tubulin (ID5) and by electron microscopy. , 1994, Journal of submicroscopic cytology and pathology.

[31]  D. Cole.  The Intraflagellar Transport Machinery of Chlamydomonas reinhardtii , 2003, Traffic.

[32]  C. A. Poole,et al.  The differential distribution of acetylated and detyrosinated alpha‐tubulin in the microtubular cytoskeleton and primary cilia of hyaline cartilage chondrocytes , 2001, Journal of anatomy.

[33]  Susan K Dutcher,et al.  Three-dimensional organization of basal bodies from wild-type and delta-tubulin deletion strains of Chlamydomonas reinhardtii. , 2003, Molecular biology of the cell.