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Time-dependent aggrecan gene expression of articular chondrocytes in response to hyperosmotic loading
G D Palmer1, P H Chao Ph, F Raia
1Cellular Engineering Laboratory, Department of Biomedical Engineering, Columbia University, New York, NY, 10027, USA.
Objective:
To investigate the effects of increasing extracellular osmolality on aggrecan gene expression and cell size in cultured chondrocytes.
Design:
Aggrecan promoter activity and mRNA levels were measured in bovine monolayer chondrocytes subjected to hyperosmotic loading for different time periods, using transient transfection assays or RT-PCR. Cell size changes were also determined using an epifluorescence microscopy system.
Results:
Hyperosmotic loading for 24 h suppressed aggrecan promoter activity and mRNA levels approximately two-fold. However no suppression of promoter activity was observed when exon 1 was deleted from the human aggrecan promoter construct. Osmotic regulation of aggrecan gene expression was time-dependent and found to correlate with cell shrinking and swelling. No suppression in promoter activity was observed when the hyperosmotic stimulus was applied in a cyclic manner, or when serum was present in the culture medium.
Conclusion:
Hyperosmotic loading regulates aggrecan gene expression and cell size in isolated chondrocytes. Osmotic regulation of gene expression is also affected by the time-varying nature of loading and the presence of serum.