Influence of oxygen tension on nitric oxide and prostaglandin E2 synthesis by bovine chondrocytes.
OBJECTIVE
To determine the in vitro effects of oxygen tension on interleukin (IL)-1beta induced nitric oxide (*NO) and prostaglandin E(2) (PGE(2)) production by bovine chondrocytes.


METHODS
Enzymatically isolated bovine chondrocytes were cultured for different periods in suspension in 21 (atmospheric), 5 or 1% (low) oxygen tension and in the absence or in the presence of increased amounts (0.01 to 1nM) of IL-1beta. Nitrite and nitrate concentrations in the culture supernatants were determined by a spectrophotometric method based upon the Griess reaction. PGE(2) production was quantified by a specific radioimmunoassay (RIA). Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) mRNA steady state levels were also quantified by real-time polymerase chain reaction (PCR).


RESULTS
In the absence of IL-1beta, ()NO production remained stable whatever the oxygen tension used. IL-1beta dose-dependently increased *NO production in both atmospheric and low oxygen conditions but the effect was more pronounced in low (1 and 5%) than in atmospheric (21%) oxygen tension (P<0.001). Under low and atmospheric oxygen tension, iNOS gene expression was increased by IL-1beta, but to a lesser extent in 21% than in 1 or 5% oxygen (P<0.01). In the basal condition, bovine chondrocytes spontaneously produced PGE(2) whatever the oxygen tension used. At 21% oxygen, IL-1beta dose-dependently increased PGE(2) production while no significant effect was observed at 1 or 5% oxygen. COX-2 gene expression was significantly upregulated by IL-1beta in both low and atmospheric oxygen tension. No significant difference between oxygen tension conditions was observed.


CONCLUSIONS
This study demonstrates that a hypoxic environment fully blocks COX-2 activity but favours iNOS gene expression in chondrocytes culture. These findings indicate that O(2) tension modulates cellular behaviour in culture and supports the concept of chondrocyte culture in low oxygen tension to reproduce in vitro the life conditions of chondrocytes.
Mathy-Hartert
M
M
Bone and Cartilage Metabolism Research, Department of Physical Medicine, Institute of Pathology, CHU, University of Liège, Belgium.
Burton
S
S
Deby-Dupont
G
G
Devel
P
P
Reginster
J-Y
JY
Henrotin
Y
Y
D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D002417:Cattle; D015687:Cell Hypoxia / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism*; D051546:Cyclooxygenase 2; D004247:DNA / Q000032:analysis; D015232:Dinoprostone / Q000096:biosynthesis*; D005786:Gene Expression Regulation / Q000187:drug effects; D007375:Interleukin-1 / Q000494:pharmacology; D008297:Male; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D010100:Oxygen / Q000494:pharmacology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics
D007375:Interleukin-1; D012333:RNA, Messenger; D009569:Nitric Oxide; D004247:DNA; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone; D010100:Oxygen