The mechanism of aggrecan release from cartilage differs with tissue origin and the agent used to stimulate catabolism

2002 ◽  
Vol 362 (2) ◽  
pp. 465-472 ◽  
Author(s):  
Robert SZTROLOVICS ◽  
Robert J. WHITE ◽  
Peter J. ROUGHLEY ◽  
John S. MORT
Keyword(s):  
Retinoic Acid ◽  
Matrix Metalloproteinases ◽  
Culture Media ◽  
Explant Culture ◽  
Epiphyseal Cartilage ◽  
Human Cartilage ◽  
Nasal Cartilage ◽  
Tissue Extracts ◽  
Significant Release ◽  
Stimulatory Agent

The mechanisms of aggrecan degradation in adult human articular, adult bovine nasal and fetal bovine epiphyseal cartilage in response to either interleukin-1β (IL-1β) or retinoic acid were compared using an explant culture system. Bovine nasal cartilage cultured with either IL-1β or retinoic acid exhibited significant release of glycosaminoglycan (GAG). For both factors, aggrecan proteolysis occurred predominantly at the ‘aggrecanase’ site, with no evidence for the action of matrix metalloproteinases, and resulted in the appearance of the corresponding G1 fragment in tissue extracts and in culture media. In human cartilage, little effect of IL-1β was seen, but abundant release of GAG occurred in the presence of retinoic acid, with evidence of aggrecanase action. Treatment of fetal epiphyseal cartilage with retinoic acid resulted in significant GAG release, whereas treatment with IL-1β did not. In the retinoic acid-treated tissue, however, no evidence for the cleavage of aggrecan in the interglobular region was apparent. Thus, in the fetal system, agents in addition to aggrecanase and matrix metalloproteinases appear to be active. Taken together, these data demonstrate that the pathways utilized for aggrecan catabolism may vary between different cartilages for a given stimulatory agent, and that, for a given tissue, different factors may elicit aggrecan release via different pathways.

scholarly journals Triggering cultured human osteoblast-like cells’ maturation by an extremely low magnitude alternating electromagnetic field

2020 ◽  
Vol 3 (1) ◽  
pp. 12-17
Author(s):  
Nahum Rosenberg ◽  
Keyword(s):  
Electromagnetic Field ◽  
Cellular Response ◽  
Specific Activity ◽  
Culture Media ◽  
Colorimetric Method ◽  
Explant Culture ◽  
Cell Counting ◽  
Human Osteoblast ◽  
Background Magnetic Field ◽  
Pulsed Electromagnetic

Introduction: Alternating and pulsed electromagnetic magnetic fields (AEMF and PEMF) of different amplitudes and frequencies can induce metabolic and proliferative effects in osteoblasts, but there is no clearly directed tendency of these effects. I hypothesize that there are extremely low triggering parameters of alternating electromagnetic field (EMF) intensity, i.e., above the background magnetic field on earth but below the lowest AEMF and PEMF that have been investigated to date (above 0.07 mT and below 0.4 mT) that induce cellular response. Methods: Accordingly, human monolayer explant culture replica were exposed four times in 24-hour intervals to two minutes of 10 kHz AEMF or PEMF (10 Hz pulses at a basic 5 kHz frequency) with a maximal EMF intensity of 0.2 mT for both. Cell proliferation was estimated from microscopic cell counting and cell death by lactate dehydrogenase (LDH) specific activity in culture media (measured using a colorimetric method). The early marker of osteoblast maturation, cellular alkaline phosphatase (AP) specific activity, was measured using a colorimetric method (n=6 for all experiment conditions). Results: No difference was found in cell numbers in the culture samples exposed either to AEMF or PEMF and in the LDH’s specific activity in culture media in comparison to the unexposed controls (p>0.05, for both). The cellular AP’s specific activity increased significantly only in cell cultures exposed to the 10 kHz AEMF (p=0.011). Conclusions: The triggering for human osteoblast activation for maturation by an extremely low AEMF (10 kHz) is at least 0.2 mT, which is distinct and below the previously found triggering range of a PEMF for proliferation induction. Therefore, application of these EMF parameters in a clinical setup by a separate finetuning of osteoblast proliferation and maturation might have a therapeutic value in enhancing damaged bone regeneration.

Retinoic acid increases the sensitivity of the rat embryo fibroblast transformation assay

1988 ◽  
Vol 8 (4) ◽  
pp. 1845-1848
Author(s):  
T D Halazonetis ◽  
C Daugherty ◽  
P Leder
Keyword(s):  
Tissue Culture ◽  
Retinoic Acid ◽  
Culture Media ◽  
Rat Embryo ◽  
Transformation Assay ◽  
Tissue Culture Media ◽  
Focus Assay ◽  
Fibroblast Transformation

The rat embryo fibroblast focus assay is used to evaluate the transforming potential of several oncogenes. The sensitivity of this assay increased fivefold when retinoic acid was added to tissue culture media. Retinoic acid probably acts by selectively inhibiting the proliferation of nontransformed cells.

scholarly journals Role of Matrix Metalloproteinases 7 in the Pathogenesis of Laryngopharyngeal Reflux: Decreased E-cadherin in Acid exposed Primary Human Pharyngeal Epithelial Cells

2019 ◽  
Vol 20 (21) ◽  
pp. 5276 ◽  
Author(s):  
Nu-Ri Im ◽  
Doh Young Lee ◽  
Byoungjae Kim ◽  
Jian Kim ◽  
Kwang-Yoon Jung ◽  
...  
Keyword(s):  
Matrix Metalloproteinases ◽  
Culture Media ◽  
Laryngopharyngeal Reflux ◽  
Acid Exposure ◽  
Immunocytochemical Staining ◽  
Mmp Inhibitor ◽  
Transepithelial Permeability ◽  
Laryngopharyngeal Reflux Disease ◽  
E Cadherin

Cleavage of E-cadherin and the resultant weakness in the cell-cell links in the laryngeal epithelium lining is induced by exposure to acidic contents of the refluxate. Herein, we aimed to evaluate the role of matrix metalloproteinases (MMPs) in inducing E-cadherin level changes following acid exposure to the human pharyngeal mucosal cells. E-cadherin levels were inversely correlated with the duration of acid exposure. Treatment with actinonin, a broad MMP inhibitor, inhibited this change. Immunocytochemical staining and transepithelial permeability test revealed that the cell surface staining of E-cadherin decreased and transepithelial permeability increased after acid exposure, which was significantly inhibited by the MMP inhibitor. Among the various MMPs analyzed, the mRNA for MMP-7 in the cellular component was upregulated, and the secretion and enzymatic activity of MMP-7 in the culture media increased with the acid treatment. Consequently, MMP-7 plays a significant role in the degradation of E-cadherin after exposure to a relatively weak acidic condition that would be similar to the physiologic condition that occurs in Laryngopharyngeal reflux disease patients.

scholarly journals Immunochemical analysis of cartilage proteoglycans. Cross-reactivity of molecules isolated from different species

1981 ◽  
Vol 199 (1) ◽  
pp. 81-87 ◽  
Author(s):  
J Wieslander ◽  
D Heinegård
Keyword(s):  
Hyaluronic Acid ◽  
Articular Cartilage ◽  
Chondroitin Sulphate ◽  
Limb Bud ◽  
Binding Region ◽  
Human Cartilage ◽  
Nasal Cartilage ◽  
Cartilage Proteoglycans ◽  
Rabbit Articular Cartilage ◽  
Hyaluronic Acid Binding

Antibodies directed against whole bovine nasal-cartilage proteoglycan and against the hyaluronic acid-binding region and chondroitin sulphate peptides from the same molecule were used in immunodiffusion and immunoelectromigration experiments. Proteoglycans from bovine nasal and tracheal cartilage showed immunological identity, with all three antisera. Proteoglycans from pig hip articular cartilage, dog hip articular cartilage, human tarsal articular cartilage and rat chondrosarcoma reacted with all the antisera and showed immunological identity with the corresponding structures isolated from bovine nasal-cartilage proteoglycans. In contrast, proteoglycans from rabbit articular cartilage, rabbit nasal cartilage and cultured chick limb buds did not react with the antibodies directed against the hyaluronic acid-binding region, though reacting with antibodies raised against whole proteoglycan monomer and against chondroitin sulphate peptides. All the proteoglycans gave two precipitation lines with the anti-(chondroitin sulphate peptide) antibodies. Similarly, the proteoglycans reacting with the anti-(hyaluronic acid-binding region) antibodies gave two precipitation lines. The results indicate the presence of at least two populations of aggregating proteoglycan monomers in cartilage. The relative affinity of the antibodies for cartilage proteoglycans and proteoglycan substructures from various species was determined by radioimmunoassay. The affinity of the anti-(hyaluronic acid-binding region) antibodies for the proteoglycans decreased in the order bovine, dog, human and pig cartilage. Rat sternal-cartilage and rabbit articular-cartilage proteoglycans reacted weakly, whereas chick limb-bud and chick sternal-cartilage proteoglycans did not react. In contrast, the affinity of antibodies to chondroitin sulphate peptides for proteoglycans increased in the order bovine cartilage, chick limb bud and chick sternal cartilage, dog cartilage, rat chondrosarcoma, human cartilage, pig cartilage, rat sternal cartilage and rabbit cartilage.

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