|
| ||
|
|
||
Chondrocyte Cultures (chondrocyte + culture)
Selected AbstractsChemInform Abstract: Synthesis and in vitro Evaluation of 5-Arylidene-3-hydroxyalkyl-2-phenylimino-4-thiazolidinones (V) with Antidegenerative Activity on Human Chondrocyte Cultures.CHEMINFORM, Issue 11 2008Rosaria Ottana Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source] Intracellular Na+ and Ca2+ modulation increases the tensile properties of developing engineered articular cartilageARTHRITIS & RHEUMATISM, Issue 4 2010Roman 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] Smad3-Deficient Chondrocytes Have Enhanced BMP Signaling and Accelerated Differentiation,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 1 2006Tian-Fang Li Abstract Smad3 deficiency accelerates chondrocyte maturation and leads to osteoarthritis. Primary chondrocytes without Smad3 lack compensatory increases of TGF-, signaling factors, but BMP-related gene expression is increased. Smad2 or Smad3 overexpression and BMP blockade abrogate accelerated maturation in Smad3,/, chondrocytes. BMP signaling is increased in TGF-, deficiency and is required for accelerated chondrocyte maturation. Introduction: Disruption of TGF-, signaling results in accelerated chondrocyte maturation and leads to postnatal dwarfism and premature osteoarthritis. The mechanisms involved in this process were studied using in vitro murine chondrocyte cultures. Materials and Methods: Primary chondrocytes were isolated from the sterna of neonatal wildtype and Smad3,/, mice. Expressions of maturational markers, as well as genes involved in TGF-, and BMP signaling were examined. Chondrocytes were treated with TGF-, and BMP-2, and effects on maturation-related genes and BMP/TGF-, responsive reporters were examined. Recombinant noggin or retroviral vectors expressing Smad2 or Smad3 were added to the cultures. Results: Expression of colX and other maturational markers was markedly increased in Smad3,/, chondrocytes. Smad3,/, chondrocytes lacked compensatory increases in Smad2, Smad4, TGFRII, Sno, or Smurf2 and had reduced expression of TGF - ,1 and TGFRI. In contrast, Smad1, Smad5, BMP2, and BMP6 expression was increased, suggesting a shift from TGF-, toward BMP signaling. In Smad3,/, chondrocytes, alternative TGF-, signaling pathways remained responsive, as shown by luciferase assays. These non-Smad3-dependent TGF-, pathways reduced colX expression and alkaline phosphatase activity in TGF-,-treated Smad3,/, cultures, but only partially. In contrast, Smad3,/, chondrocytes were more responsive to BMP-2 treatment and had increased colX expression, phosphoSmads 1, 5, and 8 levels, and luciferase reporter activity. Overexpression of both Smad2 and Smad3 blocked spontaneous maturation in Smad3-deficient chondrocytes. Maturation was also abrogated by the addition of noggin, an extracellular BMP inhibitor. Conclusions: These findings show a key role for BMP signaling during the chondrocyte maturation, occurring with loss of TGF-, signaling with important implications for osteoarthritis and cartilage diseases. [source] Functional Differences Between Growth Plate Apoptotic Bodies and Matrix Vesicles,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 10 2003Thorsten Kirsch Abstract Mineralization often occurs in areas of apoptotic changes. Our findings indicate that physiological mineralization is mediated by matrix vesicles. These matrix vesicles use mechanisms to induce mineralization that are different from the mechanisms used by apoptotic bodies released from apoptotic cells. Therefore, different therapeutic approaches must be chosen to inhibit pathological mineralization depending on the mechanism of mineralization (matrix vesicles versus apoptotic bodies). Introduction: Physiological mineralization in growth plate cartilage is restricted to regions of terminally differentiated and apoptotic chondrocytes. Pathological mineralization of tissues also often occurs in areas of apoptosis. We addressed the question of whether apoptotic changes control mineralization events or whether both events are regulated independently. Methods: To induce mineralization, we treated growth plate chondrocytes with retinoic acid (RA); apoptosis in these cells was induced by treatment with staurosporine, anti-Fas, or TNF,. The degrees of mineralization and apoptosis were determined, and the structure and function of matrix vesicles and apoptotic bodies were compared. Results: Release of matrix vesicles and mineralization in vivo in the growth plate occurs earlier than do apoptotic changes. To determine the functional relationship between apoptotic bodies and matrix vesicles, growth plate chondrocytes were treated with RA to induce matrix vesicle release and with staurosporine to induce release of apoptotic bodies. After 3 days, approximately 90% of staurosporine-treated chondrocytes were apoptotic, whereas only 2,4 % of RA-treated cells showed apoptotic changes. RA- and staurosporine-treated chondrocyte cultures were mineralized after 3 days. Matrix vesicles isolated from RA-treated cultures and apoptotic bodies isolated from staurosporine-treated cultures were associated with calcium and phosphate. However, matrix vesicles were bigger than apoptotic bodies. Furthermore, matrix vesicles but not apoptotic bodies contained alkaline phosphatase and Ca2+ channel-forming annexins II, V, and VI. Consequently, matrix vesicles but not apoptotic bodies were able to take up Ca2+ and form the first mineral phase inside their lumen. Mineralization of RA-treated cultures was inhibited by antibodies specific for annexin V but not mineralization of staurosporine-treated cultures. Conclusion: Physiological mineralization of growth plate chondrocytes is initiated by specialized matrix vesicles and requires alkaline phosphatase and annexins. In contrast, mineral formation mediated by apoptotic bodies occurs by a default mechanism and does not require alkaline phosphatase and annexins. [source] Combined role of type IX collagen and cartilage oligomeric matrix protein in cartilage matrix assembly: Cartilage oligomeric matrix protein counteracts type IX collagen,induced limitation of cartilage collagen fibril growth in mouse chondrocyte culturesARTHRITIS & RHEUMATISM, Issue 12 2009K. Blumbach Objective Defects in the assembly and composition of cartilage extracellular matrix are likely to result in impaired matrix integrity and increased susceptibility to cartilage degeneration. The aim of this study was to determine the functional interaction of the collagen fibril,associated proteins type IX collagen and cartilage oligomeric matrix protein (COMP) during cartilage matrix formation. Methods Primary chondrocytes from mice deficient in type IX collagen and COMP (double-deficient) were cultured in monolayer or alginate beads. Anchorage of matrix proteins, proteoglycan and collagen content, collagen crosslinks, matrix metalloproteinase activity, and mechanical properties of the matrix were measured. Electron microscopy was used to study the formation of fibrillar structures. Results In cartilage lacking both type IX collagen and COMP, matrilin 3 showed decreased matrix anchorage. Less matrilin 3 was deposited in the matrix of double-deficient chondrocytes, while larger amounts were secreted into the medium. Proteoglycans were less well retained in the matrix formed in alginate cultures, while collagen deposition was not significantly affected. Electron microscopy revealed similar cartilage collagen fibril diameters in the cultures of double-deficient and wild-type chondrocytes. In contrast, a larger fibril diameter was observed in the matrix of chondrocytes deficient in only type IX collagen. Conclusion Our results show that type IX collagen and COMP are involved in matrix assembly by mediating the anchorage and regulating the distribution of other matrix macromolecules such as proteoglycans and matrilins and have counteracting effects on collagen fibril growth. Loss of type IX collagen and COMP leads to matrix aberrations that may make cartilage more susceptible to degeneration. [source] Extent of cell differentiation and capacity for cartilage synthesis in human adult adipose-derived stem cells: Comparison with fetal chondrocytesBIOTECHNOLOGY & BIOENGINEERING, Issue 2 2010Nastaran Mahmoudifar Abstract This study evaluated the extent of differentiation and cartilage biosynthetic capacity of human adult adipose-derived stem cells relative to human fetal chondrocytes. Both types of cell were seeded into nonwoven-mesh polyglycolic acid (PGA) scaffolds and cultured under dynamic conditions with and without addition of TGF-,1 and insulin. Gene expression for aggrecan and collagen type II was upregulated in the stem cells in the presence of growth factors, and key components of articular cartilage such as glycosaminoglycan (GAG) and collagen type II were synthesized in cultured tissue constructs. However, on a per cell basis and in the presence of growth factors, accumulation of GAG and collagen type II were, respectively, 3.4- and 6.1-fold lower in the stem cell cultures than in the chondrocyte cultures. Although the stem cells synthesized significantly higher levels of total collagen than the chondrocytes, only about 2.4% of this collagen was collagen type II. Relative to cultures without added growth factors, treatment of the stem cells with TGF-,1 and insulin resulted in a 59% increase in GAG synthesis, but there was no significant change in collagen production even though collagen type II gene expression was upregulated 530-fold. In contrast, in the chondrocyte cultures, synthesis of collagen type II and levels of collagen type II as a percentage of total collagen more than doubled after growth factors were applied. Although considerable progress has been achieved to develop differentiation strategies and scaffold-based culture techniques for adult mesenchymal stem cells, the extent of differentiation of human adipose-derived stem cells in this study and their capacity for cartilage synthesis fell considerably short of those of fetal chondrocytes. Biotechnol. Bioeng. 2010;107: 393,401. © 2010 Wiley Periodicals, Inc. [source] | ||||||||||