MEASUREMENT OF INTERLEUKIN-1 STIMULATED CONSTITUTIVE PROSTAGLANDIN G H SYNTHASE (CYCLOOXYGENASE) MESSENGER-RNA LEVELS IN OSTEOBLASTIC MC3T3-E1 CELLS USING COMPETITIVE REVERSE-TRANSCRIPTASE POLYMERASE CHAIN-REACTION/

To assess regulation of constitutive prostaglandin G/H synthase (PGHS- 1) by interleukin-1 (IL-1) in osteoblastic MC3T3-E1 cells, we compared analysis by competitive reverse transcriptase-polymerase chain reacti on (RT-PCR) with Northern blot analysis. Using RT-PCR, IL-1 increased PGHS-1 mRNA levels by 1.84 +/- 0.10 or 2.07 +/- 0.17; depending oh the method of calculation. Using Northern blot analysis, the effect of IL -1 on PGHS-1 mRNA levels was more variable, and the variability was in creased by normalization of PGHS-1 mRNA levels to the housekeeping gen es, beta-actin and glyceraldehyde phosphate dehydrogenase (GAPDH), bec ause their mRNA level's were also regulated by IL-1. We conclude that competitive RT-PCR is a reproducible and accurate method for studying small changes in mRNA levels.