Steroids and bile acids in intrahepatic cholestasis of pregnancy : effects of treatment with ursodeoxycholic acid

Abstract

A method permitting simultaneous analysis of individual steroid hormone metabolites and bile acids in different forms of conjugation in serum and urine was established. Total bile acids and neutral steroids were extracted from human serum or urine with octadecylsilane-bonded silica, separated into different groups of conjugates by anion-exchange chromatography and analyzed by fast atom bombardment mass spectrometry.

After sequential hydrolysis/solvolysis, derivatized bile acids and steroids were semi-quantified by gas-liquid chromatography and characterized by gas chromatography-mass spectrometry. A temperature program was used that permitted detection of compounds in a range from C18 steroids to sugar conjugates of C21 steroids. Novel metabolites of progesterone were identified in urine from pregnant women. These were double conjugates with N-acetylglucosamine and either glucuronic or sulfuric acids. The positions of conjugation were determined by collision-induced dissociation of the anions produced by fast atom bombardment ionization. 5A-Pregnane-3a,20a-diol 3-glucuronide, 20 N-acetylglucosaminide and 3-sulfate, 20-N-acetylglucosaminide were found to be the major double conjugates, the latter being the major steroid monosulfate. Thus, conjugation with N acetylglucosamine constitutes a quantitatively important pathway of progesterone metabolism in human pregnancy (mean urinary excretion 32 umol/g creatinine in five healthy women in late pregnancy).

Patients with intrahepatic cholestasis of pregnancy (ICP) usually had elevated levels of aminoacyl amidated bile acids in serum and increased urinary excretion of these and sulfated bile acids. 7a,12a-Dihydroxy-3-oxo-4-cholenoic acid was identified as a major bile acid in urine of both healthy pregnant women and patients with ICP. The formation of this bile acid might reflect a rate limitation in the activity of 3-oxo-~4-steroid 5B-reductase in pregnancy. Sulfated progesterone metabolites, particularly isomers with a 3a-hydroxy configuration and their conjugates with N-acetylglucosamine, were significantly increased in ICP, both in serum and urine, whereas glucuronidated metabolites were unchanged or decreased.

The changes of the steroid sulfates in some cases preceded the increase of serum bile acids, supporting the suggestion of a selective defect in the biliary secretion of steroid metabolites in ICP. An abnormality of the reductive metabolism of progesterone cannot be excluded. oral administration of UDCA effectively reduced the elevated levels of both bile acids and sulfated steroids in patients with ICP and changed the compositions of bile acids and sulfated steroids towards normal. The changes were accompanied by improvements of pruritus and liver function tests. Low levels of 7a-hydroxy-4-cholesten-3-one in serum of patients with ICP indicated a reduced formation of bile acids, which was not changed by the oral administration of UDCA for up to four weeks. UDCA was hydroxylated in the 1B-, 5B, 6a/B- and 22-positions in pregnancy A 4-hydroxy UDCA was tentatively identified, which occurred predominantly as a double conjugate with amino acids and glucuronic acid as did other 4-hydroxylated bile acids of probable fetal origin. Possible mechanisms behind the beneficial effects of UDCA on ICP are discussed.