Effect Of Dexamethasone And Ciprofloxacin On Canine Chondrocytes

Objective: Glucocorticoids, with powerful anti-inflammatory effects, may inhibit the toxic effect of tumor necrosis factor on cells, which are widely used to treat various diseases, such as nonbacterial osteoarthritis and so on. Dexamethason(DEX) may relieve articular inflammation, but DEX used abundantly for a long-term should lead to some complications such as osteoporosis or femoral head necrosis. DEX combined with ciprofloxacin(CIP) or other quinolones are usually used to cure infected arthritis,but there were many reports about quinolones leading cartilage damage. Therefore, studying thoroughly on the effects of DEX and CIP, commonly used drugs of glucocorticoids or quinolones in clinic, on the biological characteristics of chondrocytes is of great significance to guiding clinical medication.Methods: The methods of culturing tissue mass were used to isolate immature canine articular chondrocytes in the experiment in vitro, and then pure cultivated chondrocytes in the third generation were used for experimental studies, the serum-free culture medium containing 10-4, 10-6, or 10-8g/L DEX, respectively, or containing 0.15, 0.1, 0.05g/L CIP, respectively.The cell proliferation and protein synthesis in chondrocytes were regularly observed, and type-â…¡collagen gene expression was detected by fluorescent quantitative PCR in order to analysis the effects of DEX and CIP on the activity of chondrocytes. Apoptosis rate was detected by the flow cytometer, the mRNA expression of the Fas gene and Bcl-2 gene were detected by the fluorescent quantitative PCR, the ultrastructure of chondrocyte were observed under the scanning electron microscope and transmission electron microscope, which were significant in analyzing whether DEX and CIP had injured chondrocytes.Results: when DEX was added into the cell culture fluid, the cells proliferated rapidly in 24h, the 10-6g/L DEX with obvious effects, and then the cell proliferation stepped down, the 10-4g/L DEX with evident inhibition. Total protein in chondrocytes increased slightly, the 10-6g/L DEX with significant effects. But 24 hours after adding CIP into the cell culture fluid, it increased significantly the cell proliferation, and the inhibition enhanced along with the time extended, and the larger the dose, the stronger the inhibition. CIP might also reduce the total protein content in chondrocytes, and the 0.15, 0.1g/L CIP did obviously (P<0.01).The detection results by fluorescent quantitation PCR indicated that both DEX and CIP could make the mRNA of type-â…¡collagen gene show down- regulated expression in chondrocytes (P<0.01), the Fas gene up-regulated (P<0.05), and Bcl-2 gene down-regulated (P<0.05), and the effects of CIP was stronger than that of DEX.Under the electron microscope, in DEX groups the chondrocyte surface became rough, with obvious nucleoli and more abnormal nuclei, there were many vesicular degenerative mitochondria in cytoplasm, and mitochondria became medullary degeneration. In CIP groups the microvillus on the chondrocyte surface decreased, with nucleoli lysis or shrinkage, and the chromatin formed en plaque and dispersed in the nucleus, and the cell size increased, organelle decreased obviously and the vacuoles increased in the cytoplasm, and endoplasmic reticulum became large cystic vacuoles, some membrane of organelles broke and desquamated in the capsular space to become flocc, a part of the mitochondria appeared medullary degeneration.Conclusion: A primary culture method for canine articular chondrocytes had been established through the experiments. Both of the DEX and CIP could inhibit the cell proliferation, synthetic of the protein and type-â…¡collagen in the chondrocytes, and promote chondrocyte apoptosis, which showed Fas gene up-regulated expression, Bcl-2 gene down-regulated expression, and the cell ultrastructure destroyed.