Abstract
Regulation of mRNA levels for the constitutive and inducible prostaglandin endoperoxide synthases, PGHS-1 and PGHS-2, was examined in murine osteoblastic MC3T3-E1 cells. Serum induction of PGHS-2 mRNA levels was rapid, transient, increased by cycloheximide, and inhibited 72% by cortisol. The cortisol inhibition was blocked by cycloheximide. Serum stimulation of PGHS-1 mRNA was slower, decreased by cycloheximide, and inhibited 28% by cortisol. Increased prostaglandin E2 (PGE2) production and induction of PGHS-2 immunoreactive protein paralleled changes in PGHS-2 mRNA. PGHS-2 mRNA was induced at 2 h in serum-free cells by transforming growth factor-beta (TGF-beta), phorbol 12-myristate 13-acetate, and, to a lesser extent, by forskolin. The combination of phorbol 12-myristate 13-acetate and forskolin was synergistic. TGF-beta induction was prolonged compared with serum, inhibited 67% by cortisol, and the inhibition was not blocked by cycloheximide. TGF-alpha had little effect on PGHS-2 mRNA at 2 h, but the combination of TGF-beta and TGF-alpha was synergistic for PGHS-1 and PGHS-2. PGE2 itself induced PGHS-2 mRNA, and inhibition of PGE2 production decreased the serum induction by 55%, suggesting an important role for autoamplification. The rapidity and amplitude of changes in PGHS-2 suggest that it may be involved in bone responses to acute stresses, such as mechanical strain, inflammation, and injury.
Highlights
This study has shown that mRNA for the newly identified PG endoperoxide synthase (PGHS)-2 gene is expressed in osteoblastic MC3T3-El cells and that the mRNA levels for PGHS-2 and PGHS-1 are differentially regulated in these cells by serum, cortisol, and growth factors
Changes in medium PGE2 concentrationduring serum stimulation paralleled changes in PGHS-2mRNA levels, and therewas a large induction of PGHS-2 immunoreactive proteinwithserumstimulation, suggesting that new synthesis of PGHS-2 protein plays the primary role in serumstimulated PGE, production in these cells
The rapid, dose-dependent induction of PGHS-2 mRNA accumulation with serum stimulation was blocked by actinomycin D and superinduced with cycloheximide
Summary
0 1993 by The American Society for Biochemistry and Molecular Biology, Inc. Vol 268, No., Issue of December 5,PP,2P56ri4n3te-2d5i6n49U,.1S9.9A3. Regulation of mRNA levels for the constitutive and inducible prostaglandin endoperoxide synthases, PGHS-1 and PGHS-2, was examined in murine osteoblastic MC3T3-El cells. Serum induction of PGHS-2 mRNA levels wasrapid, transient, increased by cycloheximide, and inhibited 72%by cortisol. Serum stimulation of PGHS-1 mRNA was slower, decreased by cycloheximide, and inhibited 28% by cortisol. PGHS-2 mRNA was induced at 2 h in serum-free cells by transforming growth factor-@(TGF-@),phorbol 12-myristate 13acetate, and, to a lesser extent,by forskolin. PGEz itself induced PGHS-2 mRNA, and inhibition of PGEz production decreased the serum induction by 55%, suggesting an important role for autoamplification. Theyare complex regulators of bone metabolism, being potent stimulators of bone resorption in organ culture [1]and inhibitors of resorption by isolated osteoclasts [2] They can both stimulate and inhibit bone formation [3]. Production of PGs in bone is regulated by systemic hormones, such as parathyroid hormone and glucocorticoid,and by localfactors, such as cytokines and growth factors
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