Cartilage Development (cartilage + development)

Distribution by Scientific Domains

Selected Abstracts

Expanding field of purinergic signaling

Geoffrey Burnstock
Abstract This article attempts to paint a broad picture of the extraordinary explosive recent developments in the purinergic signaling field. After a brief historical review and update of purinoceptor subtypes, the focus is on the physiological roles of purines and pyrimidines. These are considered both in terms of short-term signaling in neurotransmission, secretion, and vasodilatation and in long-term (trophic) signaling in development, regeneration, proliferation, and cell death. Examples of trophic signaling include cartilage development in limb buds, glial cell proliferation, development of skeletal muscle, changes in receptor expression in smooth-muscle phenotypes, maturation of testicular spermatids, and bone remodeling. Plasticity of purinoceptor expression in pathological conditions is described, including the increase in the purinergic component of parasympathetic nervous control of the human bladder in interstitial cystitis and outflow obstruction and in sympathetic cotransmitter control of blood vessels in hypertensive rats, the appearance of P2X7 receptors in the glomeruli of the kidney from diabetic and transgenic hypertensive animal models, and up-regulation of P2X1 and P2Y2 receptor mRNA in hearts of rats with congestive heart failure. The role of P2X3 receptors in nociception is considered, and a new hypothesis about purinergic mechanosensory transduction in the gut is explored. A personal view of some of the areas ripe for future development concludes this article, including a discussion of different strategies that could lead to the development of purinergic therapeutic agents. Drug Dev. Res. 52:1,10, 2001. © 2001 Wiley-Liss, Inc. [source]

The distribution of Notch receptors and their ligands during articular cartilage development

A. J. Hayes
Abstract We examined the distribution of Notch family members and their ligands during the development of articular cartilage and the growth plate. Notch 1 was expressed by the chondrocytes of the developing articular surface but became increasingly restricted to the deeper layers after birth whilst expression of this family member was restricted to hypertrophic chondrocytes in the growth plate. Notch 2 and 4, Delta and Jagged 2 showed a broadly similar distribution, being present throughout the articular cartilage during development and becoming increasingly restricted to deeper layers with age. Hypertrophic chondrocytes within the growth plate also expressed Notch 2 and 4, Delta and Jagged 2 (which was also expressed in prehypertrophs). Notch 3 and Jagged 1 were absent from developing articular cartilage but were present in deeper layers at later time points (> 1 month) and both receptor and ligand were expressed in hypertrophic chondrocytes at all ages examined. These results highlight the complex Notch signalling interactions that result in the formation of the heterogeneous articular cartilage and allow for the co-ordinated ossification and elongation of the growth plate. Mechanisms by which these processes are controlled are discussed in light of recent advances in the understanding of Notch signalling pathways. [source]

Therapeutic Effects of Anti-FGF23 Antibodies in Hypophosphatemic Rickets/Osteomalacia,,

Yukiko Aono
Abstract X-linked hypophosphatemia (XLH), characterized by renal phosphate wasting, is the most common cause of vitamin D-resistant rickets. It has been postulated that some phosphaturic factor plays a causative role in XLH and its murine homolog, the Hyp mouse. Fibroblast growth factor 23 (FGF23) is a physiological phosphaturic factor; its circulatory level is known to be high in most patients with XLH and Hyp mice, suggesting its pathophysiological role in this disease. To test this hypothesis, we treated Hyp mice with anti-FGF23 antibodies to inhibit endogenous FGF23 action. A single injection of the antibodies corrected the hypophosphatemia and inappropriately normal serum 1,25-dihydroxyvitamin D. These effects were accompanied by increased expressions of type IIa sodium-phosphate cotransporter and 25-hydroxyvitamin-D-1,-hydroxylase and a suppressed expression of 24-hydroxylase in the kidney. Repeated injections during the growth period ameliorated the rachitic bone phenotypes typically observed in Hyp mice, such as impaired longitudinal elongation, defective mineralization, and abnormal cartilage development. Thus, these results indicate that excess actions of FGF23 underlie hypophosphatemic rickets in Hyp mice and suggest a novel therapeutic potential of the FGF23 antibodies for XLH. [source]

Inactivation of Pten in Osteo-Chondroprogenitor Cells Leads to Epiphyseal Growth Plate Abnormalities and Skeletal Overgrowth,

Alice Fiona Ford-Hutchinson
Abstract To study the role of the Pten tumor suppressor in skeletogenesis, we generated mice lacking this key phosphatidylinositol 3,-kinase pathway regulator in their osteo-chondroprogenitors. A phenotype of growth plate dysfunction and skeletal overgrowth was observed. Introduction: Skeletogenesis is a complex process relying on a variety of ligands that activate a range of intracellular signal transduction pathways. Although many of these stimuli are known to activate phosphatidylinositol 3,-kinase (PI3K), the function of this pathway during cartilage development remains nebulous. To study the role of PI3K during skeletogenesis, we used mice deficient in a negative regulator of PI3K signaling, the tumor suppressor, Pten. Materials and Methods:Pten gene deletion in osteo-chondrodroprogenitors was obtained by interbreeding mice with loxP-flanked Pten exons with mice expressing the Cre recombinase under the control of the type II collagen gene promoter (Ptenflox/flox:Col2a1Cre mice). Phenotypic analyses included microcomputed tomography and immunohistochemistry techniques. Results: ,CT revealed that Ptenflox/flox:Col2a1Cre mice exhibited both increased skeletal size, particularly of vertebrae, and massive trabeculation accompanied by increased cortical thickness. Primary spongiosa development and perichondrial bone collar formation were prominent in Ptenflox/flox:Col2a1Cre mice, and long bone growth plates were disorganized and showed both matrix overproduction and evidence of accelerated hypertrophic differentiation (indicated by an altered pattern of type X collagen and alkaline phosphatase expression). Consistent with increased PI3K signaling, Pten-deficient chondrocytes showed increased phospho-PKB/Akt and phospho-S6 immunostaining, reflective of increased mTOR and PDK1 activity. Interestingly, no significant change in growth plate proliferation was seen in Pten-deficient mice, and growth plate fusion was found at 6 months. Conclusions: By virtue of its ability to modulate a key signal transduction pathway responsible for integrating multiple stimuli, Pten represents an important regulator of both skeletal size and bone architecture. [source]

Microenvironment regulation of PRG4 phenotype of chondrocytes

Megan E. Blewis
Abstract Articular cartilage is a heterogeneous tissue with superficial (S), middle (M), and deep (D) zones. Chondrocytes in the S zone secrete the lubricating PRG4 protein, while chondrocytes from the M and D zones are more specialized in producing large amounts of the glycosaminoglycan (GAG) component of the extracellular matrix. Soluble and insoluble chemicals and mechanical stimuli regulate cartilage development, growth, and homeostasis; however, the mechanisms of regulation responsible for the distinct PRG4-positive and negative phenotypes of chondrocytes are unknown. The objective of this study was to determine if interaction between S and M chondrocytes regulates chondrocyte phenotype, as determined by coculture in monolayer at different ratios of S:M (100:0, 75:25, 50:50, 25:75, 0:100) and at different densities (240,000, 120,000, 60,000, and 30,000 cells/cm2), and by measurement of PRG4 secretion and expression, and GAG accumulation. Coculture of S and M cells resulted in significant up-regulation in PRG4 secretion and the percentage of cells expressing PRG4, with simultaneous down-regulation of GAG accumulation. Tracking M cells with PKH67 dye in coculture revealed that they maintained a PRG4-negative phenotype, and proliferated less than S cells. Taken together, these results indicate that the up-regulated PRG4 expression in coculture is a result of preferential proliferation of PRG4-expressing S cells. This finding may have practical implications for generating a large number of phenotypically normal S cells, which can be limited in source, for tissue engineering applications. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 25:685,695, 2007 [source]

PerioGlas® Regulates Osteoblast RNA Interfering

Annalisa Palmieri PhD
Abstract Purpose: PerioGlas® (PG) is an alloplastic material that has been used for grafting periodontal osseous defects since the 1990s. In animal models, it has been proven that PG achieves histologically good repairs of surgically created defects. In clinical trials, PG is effective as an adjunct to conventional surgery in the treatment of intrabony defects; however, how PG alters osteoblast activity to promote bone formation is poorly understood. We therefore attempted to address this question by using microRNA (miRNA) microarray techniques to investigate the translation process in osteoblasts exposed to PG. Materials and Methods: By using miRNA microarrays containing 329 probes designed from human miRNA sequences, we identified several miRNA whose expression was significantly modified in osteoblast-like cell lines (MG-63) cultured with PG. Results: There were ten up-regulated miRNA (mir-337, mir-377, mir-9, mir-516, mir-515-3p, mir-496, mir-200b, mir-489, mir-25, mir-423) and two down-regulated miRNA (mir-26a, mir-30d). Conclusion: PG acts on miRNAs, which in turn regulate several messengers. Among them there are mRNAs related to bone formation and skeletal and cartilage development. The vast majority of detected genes are down-regulated, and some are homeobox genes like NOG, EN1, and CHRD. Other down-regulated genes are receptors (like GHRHR) and extracellular matrix proteins (like COMP). Although the exact mechanism of PG action on osteoblasts is still incompletely understood, these data demonstrate that PG has not only an osteoconductive effect, but also regulates bone formation. [source]

Extraskeletal myxoid chondrosarcoma: Updated clinicopathological and molecular genetic characteristics

Masanori Hisaoka
Extraskeletal myxoid chondrosarcoma (EMC) is a rare soft-tissue sarcoma characterized by distinctive morphological and cytogenetical features. As its name implies, EMC was believed to represent a variant of soft-tissue chondrosarcoma owing to its histological resemblance to chondroblastic tissue in the early stages of cartilage development or chondroid tumors such as skeletal chondrosarcoma. However, the chondroid nature has been a subject of controversy, and its line of differentiation remains to be determined. Consequently, the tumor is provisionally classified into a group of tumors of uncertain differentiation in the revised World Health Organization classification of tumors of soft tissue and bone. Moreover, immunohistochemical and ultrastructural features of neural or neuroendocrine differentiation have been recently reported in a subset of EMC, providing a new insight into their histogenetic nature. Chromosomal rearrangements involving 9q22, such as t(9;22)(q22;q12), and resultant NR4A3 fusion genes are tumor-type specific or pathognomotic for this entity and are assumed to play an important role in the development of EMC. Although the biological mechanisms and functions are largely unknown, the NR4A3-related pathway is considered a potential molecular target for future therapeutic intervention. Because of its protracted but resilient nature, a tenacious and long-term follow up is necessary for any patient. [source]

Endothelial nitric oxide synthase deficiency in mice results in reduced chondrocyte proliferation and endochondral bone growth

Qian Yan
Objective Nitric oxide (NO) and aberrant chondrocyte differentiation have both been implicated in the pathogenesis of osteoarthritis, but whether these processes are connected is unknown, and the role of specific NO synthase (NOS) enzymes in chondrocyte physiology is unclear. This study was undertaken to examine the effects of inactivation of endothelial cell NOS (eNOS) on cartilage development in mice. Methods Skeletal growth and development of mice carrying a null mutation in the eNOS gene was compared with that of their control littermates. In situ analyses were complemented by experiments with primary chondrocytes and tibial explants from these mice. Results Mice that were deficient in eNOS showed increased fatality and reduced bone growth, with hypocellular growth plates and a marked reduction in the number of proliferating chondrocytes. In vitro studies demonstrated lower chondrocyte numbers and reduced endochondral bone growth in mutant mice, suggesting that the role of eNOS signaling in chondrocyte proliferation is cell autonomous. Reduced chondrocyte numbers appear to be caused by decreased cyclin D1 and increased p57 expression in mutant mice, resulting in slower cell cycle progression and earlier cell cycle exit. In addition, expression of early chondrocyte markers such as SOX9 was reduced, and prehypertrophic markers were expressed prematurely in mutant mice. Conclusion Our findings identify a novel and important role of eNOS in chondrocyte proliferation and endochondral bone growth and demonstrate that loss of eNOS results in premature cell cycle exit and prehypertrophic chondrocyte differentiation during cartilage development. [source]

Intracellular Na+ and Ca2+ modulation increases the tensile properties of developing engineered articular cartilage

Roman M. Natoli
Objective Significant collagen content and tensile properties are difficult to achieve in tissue-engineered articular cartilage. The aim of this study was to investigate whether treating developing tissue-engineered cartilage constructs with modulators of intracellular Na+ or Ca2+ could increase collagen concentration and construct tensile properties. Methods Inhibitors of Na+ ion transporters and stimulators of intracellular Ca2+ were investigated for their ability to affect articular cartilage development in a scaffoldless, 3-dimensional chondrocyte culture. Using a systematic approach, we applied ouabain (Na+/K+ -ATPase inhibitor), bumetanide (Na+/K+/2Cl, tritransporter inhibitor), histamine (cAMP activator), and ionomycin (a Ca2+ ionophore) to tissue-engineered constructs for 1 hour daily on days 10,14 of culture and examined the constructs at 2 weeks or 4 weeks. The gross morphology, biochemical content, and compressive and tensile mechanical properties of the constructs were assayed. Results The results of these experiments showed that 20 ,M ouabain, 0.3 ,M ionomycin, or their combination increased the tensile modulus by 40,95% compared with untreated controls and resulted in an increased amount of collagen normalized to construct wet weight. In constructs exposed to ouabain, the increased percentage of collagen per construct wet weight was secondary to decreased glycosaminoglycan production on a per-cell basis. Treatment with 20 ,M ouabain also increased the ultimate tensile strength of neo-tissue by 56,86% at 4 weeks. Other construct properties, such as construct growth and type I collagen production, were affected differently by Na+ modulation with ouabain versus Ca2+ modulation with ionomycin. Conclusion These data are the first to show that treatments known to alter intracellular ion concentrations are a viable method for increasing the mechanical properties of engineered articular cartilage and identifying potentially important relationships to hydrostatic pressure mechanotransduction. Ouabain and ionomycin may be useful pharmacologic agents for increasing tensile integrity and directing construct maturation. [source]

Transcriptional profiling and biochemical analysis of mechanically induced cartilaginous tissues in a rat model

Kristy T. Salisbury Palomares
Objective To characterize patterns of molecular expression that lead to cartilage formation in vivo in a postnatal setting, by profiling messenger RNA expression across the time course of mechanically induced chondrogenesis. Methods Retired breeder Sprague-Dawley rats underwent a noncritical-sized transverse femoral osteotomy. Experimental animals (n = 45) were subjected to bending stimulation (60° cyclic motion in the sagittal plane for 15 minutes/day) of the osteotomy gap beginning on day 10 after the operation. Control animals (n = 32) experienced continuous rigid fixation. Messenger RNA isolated on days 10, 17, 24, and 38 after surgery was analyzed using a microarray containing 608 genes involved in skeletal development, tissue differentiation, fracture healing, and mechanotransduction. The glycosaminoglycan (GAG) content in the stimulated tissues was compared with that in native articular cartilage as a means of assessing the progression of chondrogenic development of the tissues. Results The majority of the 100 genes that were differentially expressed were up-regulated in response to mechanical stimulation. Many of these genes are associated with articular cartilage development and maintenance, diarthrodial joint development, cell adhesion, extracellular matrix synthesis, signal transduction, and skeletal development. Quantitative real-time polymerase chain reaction results were consistent with the microarray findings. The GAG content of the stimulated tissues increased over time and was no different from that of articular cartilage on day 38 after surgery. Conclusion Our findings indicate that mechanical stimulation causes up-regulation of genes that are principally involved in joint cavity morphogenesis and critical to articular cartilage function. Further study of this type of stimulation may identify key signaling events required for postnatal hyaline cartilage formation. [source]

Profiling microRNA expression in bovine articular cartilage and implications for mechanotransduction

Walter Dunn
Objective Articular cartilage is an avascular tissue with precise polarity and organization comprising 3 distinct functional zones: the surface, middle, and deep zones. Each zone has a different gene expression pattern that plays a specific role in articular cartilage development and maintenance. MicroRNA (miRNA) are small noncoding gene products that play an important regulatory role in determining cell differentiation and function. The purpose of this study was to test our hypothesis that miRNA expression profiles in the different articular cartilage zones as well as between regions subjected to different levels of weight-bearing stresses are unique. Methods Using an miRNA microarray approach in conjunction with quantitative reverse transcription,polymerase chain reaction, we identified miRNA in bovine articular cartilage that were differentially expressed in the different functional zones and in the anterior weight-bearing and posterior non,weight-bearing regions of the medial femoral condyle (M1 and M4, respectively). Results We identified miRNA-221 and miR-222 as part of a subset of differentially expressed miRNA that were up-regulated in articular cartilage in the anterior, M1, greater weight-bearing location. Additionally, miR-126, miR-145, and miR-335 were down-regulated in monolayers of tissue-cultured chondrocytes as compared with levels determined directly from intact native cartilage. Conclusion In conclusion, miR-222 expression patterns in articular cartilage are higher in the weight-bearing anterior medial condyle as compared with the posterior non,weight-bearing medial condyle. Thus, miR-222 might be a potential regulator of an articular cartilage mechanotransduction pathway. These data implicate miRNA in the maintenance of articular cartilage homeostasis and are therefore targets for articular cartilage tissue engineering and regenerative medicine. [source]

Real-time Monitoring of Force Response Measured in Mechanically Stimulated Tissue-Engineered Cartilage

Orahn Preiss-Bloom
Abstract:, Mechanical stimulation improves tissue-engineered cartilage development both in terms of biochemical composition and structural properties. However, the link between the compositional changes attributed to mechanical stimulation and the changing structural properties of the engineered cartilage is poorly understood. We hypothesize that transient events associated with construct stiffening can be documented and used to understand milestones in construct development. To do this, we designed and built a mechanical stimulation bioreactor that can continuously record the force response of the engineered construct in real time. This study documents the transient changes of the stiffness of tissue-engineered cartilage constructs over the first 14 days of their development under cyclic loading. Compressive strain stimulation (15%, 1 Hz) was applied to poly(ethylene glycol) (PEG) hydrogels seeded with primary articular chondrocytes. The average compressive modulus of strain-stimulated constructs was 12.7 ± 1.45 kPa after 2 weeks, significantly greater (P < 0.01) than the average compressive moduli of both unstimulated constructs (10.7 ± 0.94 kPa) and nonviable stimulated constructs (11.2 ± 0.91 kPa). The system was able to document that nearly all of the stiffness increase occurred over the last 2 days of the experiment, where live-cell constructs demonstrated a rapid 20% increase in force response. The system's ability to track significant increases in stiffness over time was also confirmed by Instron testing. These results present a novel view of the early mechanical development of tissue-engineering cartilage constructs and suggest that the real-time monitoring of force response may be used to noninvasively track the development of engineered tissue. [source]