Effects of S-petasin on cyclic AMP production and enzyme activity of P450scc in rat zona fasciculata-reticularis cells.
Petasites hybridus is used in Chinese herbal medicine. S-petasin is a bioactive compound isolated from leaves or roots of P. hybridus, which has been used to relieve gastrointestinal pain, lung disease, and spasms of urogenital tract. We have demonstrated that S-petasin inhibited corticosterone release from rat zona fasiculata-reticularis cells. However, the mechanism and molecular effects of S-petasin on zona fasiculata-reticularis cells are still unclear. This study explored the effects of S-petasin on cellular adenosine 3':5'-cyclic monophosphate (cAMP) production, the functions of steroidogenic enzymes including cytochrome P450 side-chain cleavage enzyme (P450scc), 11beta-hydroxylase, and the expression levels of steroidogenic acute regulatory protein or P450scc. In this experiment, zona fasciculata-reticularis cells were incubated with S-petasin in the presence or absence of adrenocorticotropin (ACTH), 8-bromo-adenosine 3':5'-cyclic monophosphate (8-Br-cAMP), forskolin, 25-OH-cholesterol, deoxycorticosterone at 37 degrees C for 0.5, 1 or 3 h. The media were used to measure the concentration of corticosterone or pregnenolone by radioimmunoassay. The cells were used to measure the content of cAMP by radioimmunoassay and extracted protein for Western blot or messenger RNA (mRNA) for reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Our data demonstrated that (1) S-petasin inhibits ACTH- or forskolin-stimulated cellular cAMP production, (2) S-petasin increased the Michaelis constants of P450scc and 11beta-hydroxylase and (3) S-petasin decreased the expression levels and mRNA of steroidogenic acute regulatory protein. In summary, the actions of S-petasin mediate the inhibition of cAMP formation, decrease the activities of key enzymes P450scc and 11beta-hydroxylase, and reduce mRNA of steroidogenic acute regulatory protein and expression of steroidogenic acute regulatory protein.
Chang
Ling-Ling
LL
Department of Chemical Engineering, Chinese Culture University, Shih-Lin, Taipei 111, Taiwan, ROC. llchiang@staff.pccu.edu.tw
Wun
Wan-Song Alfred
WS
Lin
Yun-Lian
YL
Wang
Paulus S
PS
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D003345:Corticosterone / Q000096:biosynthesis; D000242:Cyclic AMP / Q000096:biosynthesis*; D007700:Kinetics; D008297:Male; D010750:Phosphoproteins / Q000096:biosynthesis; D011284:Pregnenolone / Q000096:biosynthesis / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D011863:Radioimmunoassay; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012717:Sesquiterpenes / Q000494:pharmacology*; D013252:Steroid 11-beta-Hydroxylase / Q000378:metabolism; D015383:Zona Fasciculata / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D015385:Zona Reticularis / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D000098682:Steroidogenic Acute Regulatory Protein
D010750:Phosphoproteins; D012333:RNA, Messenger; D012717:Sesquiterpenes; D000098682:Steroidogenic Acute Regulatory Protein; D005576:Colforsin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D011284:Pregnenolone; D000324:Adrenocorticotropic Hormone; C418727:petasin; D000242:Cyclic AMP; D013252:Steroid 11-beta-Hydroxylase; D002786:Cholesterol Side-Chain Cleavage Enzyme; D003345:Corticosterone