BETAMETHASONE MODULATION OF SPHINGOMYELIN HYDROLYSIS UP-REGULATES CTP- CHOLINEPHOSPHATE CYTIDYLYLTRANSFERASE ACTIVITY IN ADULT-RAT LUNG

Glucocorticoids appear to play an integral role in stimulating surfact ant synthesis by activating the rate-regulatory enzyme for phosphatidy lcholine synthesis, CTP: cholinephosphate cytidylyltransferase (CT). T he activity of liver CT, in vitro, has been shown to be inhibited by t he sphingomyelin hydrolysis product, sphingosine. In order to investig ate the mechanisms by which glucocorticoids alter CT activity, in vivo , we administered beta-methasone (1 mg/kg intraperitoneally) sequentia lly to adult male rats for 5 days. Betamethasone increased CT activity 2-fold relative to control in whole lung. The hormone also increased membrane-bound activity, but did not affect cytosolic enzyme activity. Betamethasone modestly increased CT mRNA as determined by the reverse -transcription PCR and Southern analysis of PCR products, but did not alter the levels of immunoreactive enzyme in lung membranes as demonst rated by Western blotting. The hormone did, however, produce a nearly 3-fold increase in membrane-associated sphingomyelin, and co-ordinatel y a substantial decrease in the levels of sphingosine in lung membrane s. Sphingosine, but not sphinganine, was a competitive, reversible inh ibitor of lung CT with respect to the enzyme activator, phosphatidylgl ycerol. Betamethasone decreased the activities of the sphingomyelin hy drolases: acid sphingomyelinase by 33%, and of alkaline ceramidase by 21%. The hormone also inhibited the generation of sphingosine from lys osphingomyelin in lung membranes. There was no significant effect of t he hormone on serine palmitoyltransferase activity, the first committe d enzyme for sphingolipid biosynthesis. Further, administration of L-c ycloserine, an inhibitor of sphingosine formation, was shown to stimul ate CT activity by 74%, and increase disaturated phosphatidylcholine i n alveolar lavage by 52% relative to control. These observations sugge st that glucocorticoids upregulate surfactant synthesis at the level o f a key regulatory enzyme by significantly altering the availability o f inhibitory metabolites resulting from sphingomyelin hydrolysis.