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Objective: Adult articular cartilage is a physiologically hypoxic tissue with a proposed gradient of oxygen tension ranging from <10% oxygen at the cartilage surface to <1% in the deepest layers. This gradient may be disturbed during diseases of the joint, for example in rheumatoid arthritis when synovial fluid pO(2)falls. We investigated whether changes in oxygen tension modulate gene expression in articular chondrocytes., Design: Bovine articular chondrocytes were cultured in alginate beads in medium maintained at <0.1, 5, 10 or 20% oxygen. A modified RNA arbitrarily primed polymerase chain reaction (RAP-PCR) technique was used to identify several genes whose mRNA abundance in articular chondrocytes was dependent upon oxygen tension. Northern hybridization slot blots were used to quantify changes in mRNA level relative to a housekeeping gene, beta-actin., Results: Genes found by RAP-PCR to undergo up-regulation in hypoxia included TIMP-1 and integrin-linked kinase. Collagen V mRNA levels were down-regulated in hypoxic chondrocytes. This led us to examine mRNA levels for various cytokines, matrix structural molecules and beta1 integrin. Interleukin 1beta, transforming growth factor beta and connective tissue growth factor were all up-regulated by low oxygen tensions, as was beta1 integrin. Collagen II (COL2A1) was down-regulated by hypoxia but aggrecan mRNA levels remained unchanged. The mRNA levels for GAPDH, the archetypal hypoxia responsive gene, were not modulated in articular chondrocytes by changes in oxygen tension., Conclusions: Oxygen tension modulates the abundance of mRNAs encoding structural molecules, several cytokines, beta1 integrin and integrin-linked kinase in articular chondrocytes. This may be important during disease progression. Chondrocytes are unusual in their response to hypoxia, presumably because they exist physiologically in a low oxygen environment., (Copyright 2001 OsteoArthritis Research Society International.)

Macrophages accumulate in areas of inflammation and necrosis that are likely to be hypoxic. Chemotaxis of monocytes and macrophages towards chemokines is rapidly (within 60-90 min) inhibited by hypoxia. Exposure to the inflammatory cytokine TNF-alpha has a similar effect on monocyte migration. We report here that neither changes in mitochondrial respiration nor intracellular pH are involved in migration arrest. However, hypoxic inhibition of migration was mimicked using chemical activators of hypoxia-inducible factor-1 and reversed by transcriptional inhibition. We used RNA arbitrarily primed PCR, a differential display technique, to investigate which genes were up-regulated within 90-min exposure to hypoxia. Of several thousand mRNA screened, only one was consistently up-regulated by hypoxia and this was identified as MAPK phosphatase 1 (MKP-1), which modulates MAPK activity. Levels of MKP-1 mRNA and protein were rapidly elevated in monocytic cells and primary macrophages after hypoxia or TNF-alpha treatment. The functional significance of MKP-1 was illustrated by hypoxia-induced decreases in phosphorylated MAPK in these cells and arrest of chemotaxis by MAPK inhibitors. We suggest that one of the important events in an 'emergency stop' response in monocytic cells and macrophages may be inhibition of the chemoattractant signaling cascade.

Articular cartilage is a physiologically hypoxic tissue with a proposed gradient of oxygen tension ranging from about 10% oxygen at the cartilage surface to less than 1% in the deepest layers. The position of the chondrocyte within this gradient may modulate the cell's behavior and phenotype. Moreover, the oxygen gradient is likely to be disturbed during joint diseases in which the pO(2)of the synovial fluid declines which may cause changes in chondrocyte behavior and gene expression. Thus, there is a need to understand the chondrocyte's response to different oxygen tensions. We compared the behavior of bovine articular chondrocytes cultured in alginate beads for 7 days in medium maintained at <0.1, 5, 10 or 20% oxygen. The chondrocytes' survival, differentiation, cell division, viability and matrix production were assessed at each oxygen tension and rRNA and mRNA abundance was measured. Chondrocytes were able to survive under all oxygen tensions for at least 7 days but cells cultured under anoxic conditions were metabolically less active than cells maintained in higher oxygen tensions; this was associated with a decrease in matrix production. In <0.1% oxygen there was a marked decrease in rRNA and mRNA abundance in the cells. There were no differences in cell division or differentiation between any oxygen tensions. These findings indicate that articular chondrocytes can be cultured successfully in the pO(2)range in which they are thought to exist in vivo (5-10% pO(2)) and are fully active under these conditions. Under anoxic conditions (<0.1% pO(2)) function is severely compromised., (Copyright 2000 OsteoArthritis Research Society International.)