Effect of spironolactone on the expression of rat hepatic UDP-glucuronosyltransferase.
Spironolactone (SL) increases the glucuronidation rate of several compounds. We analyzed the molecular basis of changes occurring in major rat liver UDP-glucuronosyltransferase (UGT) family 1 isoforms and in UGT2B1, a relevant isoform of family 2, in response to SL. UGT activity toward bilirubin, ethynylestradiol and p-nitrophenol was assayed in native and activated microsomes. Protein and mRNA levels were determined by Western and Northern blotting. The lipid composition and physicochemical properties of the microsomal membrane were also analyzed. Glucuronidation rates of bilirubin and ethynylestradiol (at both 3-OH and 17 beta-OH positions), determined in UDP-N-acetylglucosamine-activated membranes, were increased in SL group. Western blot analysis revealed increased levels of UGT1A1 and 1A5 (bilirubin and 3-OH ethynylestradiol conjugation), and 2B1 (17 beta-OH ethynylestradiol conjugation). Northern blot studies suggested transcriptional regulation by the steroid. Analysis of UGT activity in native vs. alamethicin-activated microsomes indicated increased latency, which was not associated to changes in physicochemical properties of the microsomal membrane. p-Nitrophenol glucuronidation rate and mRNA and protein levels of UGT1A6, the main isoform conjugating planar phenols, were not affected by the inducer. The data suggest transcriptional regulation of specific isoforms of hepatic UGT by SL, thus explaining previously reported increases in UGT activity toward selective substrates.
Catania
Viviana A
VA
Instituto de Fisiología Experimental, CONICET-Universidad Nacional de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Suipacha 570, Rosario 2000, Argentina. vcatania@fbioyf.unr.edu.ar
Luquita
Marcelo G
MG
Sánchez Pozzi
Enrique J
EJ
Ikushiro
Shin-ichi
S
Emi
Yoshikazu
Y
Iyanagi
Takashi
T
Mottino
Aldo D
AD
D000818:Animals; D001663:Bilirubin / Q000494:pharmacology; D015152:Blotting, Northern; D004232:Diuretics / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects*; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D007425:Intracellular Membranes / Q000737:chemistry; D007527:Isoenzymes / Q000378:metabolism*; D008099:Liver / Q000187:drug effects* / Q000201:enzymology; D008297:Male; D008560:Membrane Fluidity / Q000187:drug effects; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism; D009596:Nitrophenols / Q000494:pharmacology; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013148:Spironolactone / Q000494:pharmacology*
D004232:Diuretics; D007527:Isoenzymes; D009596:Nitrophenols; D012333:RNA, Messenger; D013148:Spironolactone; D014453:Glucuronosyltransferase; D001663:Bilirubin