Coexpression of GTP cyclohydrolase I and inducible nitric oxide synthase mRNAs in mouse osteoblastic cells activated by proinflammatory cytokines

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Abstract
Proinflammatory cytokines, a combination of IL‐1β, TNF‐α, and IFN‐γ, caused mRNA expression of GTP cyclohydrolase I (GTP‐CH), the rate‐limiting enzyme in tetrahydrobiopterin (BH4) biosynthesis, and of inducible nitric oxide synthase (iNOS) in a well‐characterized osteoblastic clone MC3T3‐E1 cell line. We found the expression of the GTP‐CH gene in osteoblasts for the first time. The expression of GTP‐CH and iNOS mRNAs was found to be maximal at 3 and 9 h, respectively. The expression of both genes elicited increases in BH4 and NO levels. Pharmacological studies using 2,4‐diamino‐6‐hydroxypyrimidine, an inhibitor of GTP‐CH activity, showed that BH4 is involved in the activity of iNOS, but not in the induction of iNOS mRNA. The results using an inhibitor of nuclear factor (NF)‐κB and activating protein‐1 (AP‐1) activation suggested that coinduction of the two genes in response to cytokines occurred via activation of NF‐κB and AP‐1. In MC3T3‐E1 cells BH4 and sepiapterin, producing BH4, could protect against apoptosis, i.e. the degradation of nuclear DNA in the cells, induced by NO derived from S‐nitroso‐N‐acetyl‐d,l‐penicillamine. These results suggest that the induction of BH4 together with NO by proinflammatory cytokines could protect against NO‐induced apoptosis in MC3T3‐E1 cells.

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