O. Mäentausta

Complete amino acid sequence of human placental 17β‐hydroxysteroid dehydrogenase deduced from cDNA

CDNA clones for 17β‐hydroxysteroid dehydrogenase (17‐HSD; EC 1.1.1.62) were isolated from a placental λgt11 expression library using polyclonal antibodies against placental 17‐HSD. The largest cDNA contained 1325 nucleotides, consisting of a short 5′‐noncoding segment, a coding segment of 987 nucleotides terminated by a TAA codon, and a 329 nucleotide long 3′‐noncoding segment. The open reading frame encoded a polypeptide of 327 amino acid residues with a predicted M r of 34853. The amino acid sequence of 23 N‐terminal amino acids determined from purified 17‐HSD agreed with the sequence deduced from cDNA. The deduced amino acid sequence also contained two peptides previously characterized from the proposed catalytic area of placental 17‐HSD.

CHANGES IN SERUM BILE ACID CONCENTRATIONS DURING NORMAL PREGNANCY, IN PATIENTS WITH INTRAHEPATIC CHOLESTASIS OF PREGNANCY AND IN PREGNANT WOMEN WITH ITCHING

Two primary bile acids, cholic acid (CA) and chenodeoxycholic acid (CDCA), and one secondary bile acid, deoxycholic acid (DCA), were measured by radioimmuno‐assay in pregnancy serum from 30 healthy women, 49 women with itching and 45 with intrahepatic cholestasis of pregnancy. All subjects were studied serially from between 16 and 20 weeks gestation until 35–60 days post partum. In healthy pregnant women, serum CA and DCA levels did not change significantly at any time. The mean CDCA level rose significantly towards term. In women with intrahepatic cholestasis, serum levels of CA and CDCA were increased ten‐ and five‐fold, respectively, at the time of appearance of clinical Symptoms and the CA/CDCA ratio rose from l/l to 2/1; there was also a moderate increase in the serum concentration of DCA. In 4 of 8 women studied prospectively an increase in serum bile acid levels preceded the appearance of Symptoms or other laboratory evidence of intrahepatic cholestasis. Nine of the women with itching with normal routine liver function test results had increases in serum CA and CDCA concen‐trations suggesting mild cholestasis.

Serum bile acid levels in intrahepatic cholestasis of pregnancy during treatment with phenobarbital or cholestyramine

Nineteen patients suffering from the intrahepatic cholestasis (IHC) of pregnancy were studied. Twelve of them were treated with phenobarbital (100 mg/day) and seven with cholestyramine (18 g/day). The overnight fasting levels of serum cholic acid (CA), chenodeoxycholic acid (CDCA) and deoxycholic acid (DCA) were measured by radioimmunoassay. The activities of serum transaminases, gamma-glutamyltranspeptidase, alkaline phosphatase and total and conjugated bilirubins were also analyzed. It was found that there was no correlation between the itching symptom and the serum bile acid levels. During phenobarbital treatment serum bile acid concentrations did not change. Also, the other measured parameters as well as the CA/CDCA ratio did not change significantly. Transaminases had, however, a slight tendency to decrease. The therapy successfully relieved itching in half of the cases. There was no relationship between the relief of the itching and the change in the bile acid concentrations. Cholestyramine treatment did not decrease the CA level significantly, but that of the CDCA decreased (P less than 0.05) and the ratio of CA/CDCA increased (P less than 0.05). In the other analyzed liver function test results, an increase (P less than 0.05) occurred only in the concentrations of conjugated bilirubin. The itching was relieved in five of the seven cases during the first week of treatment, but after that the symptom tended to reappear. There was a slight correlation between the decrease in the CDCA level and in the relief of the itching. The two drugs did not cause any particular side effects.

Immunological measurement of human 17β-hydroxysteroid dehydrogenase

Abstract Human placental 17β-hydroxysteroid dehydrogenase (17-HSD) was purified to apparent homogeneity using ammonium sulfate precipitation and chromatography on Red-Agarose and DEAE-Sepharose columns. Electrophoresis on polyacrylamide gels under denaturing conditions and using silver staining showed a single protein with an apparent molecular weight of 37,800. Antibodies to the purified protein were raised in rabbits and were found by immunoblotting to be specific to 17-HSD. A sensitive radioimmunoassay was established using 125I-labeled 17-HSD as a tracer, an appropriate dilution of the antibody, and a kaolin-coupled double antibody for separating the antibody-bound and free fractions. The detection limit of the assay was approximately 150 pg/tube (1.5 μg/l). The cytosol fraction (105,000 g) of term placental tissue contained approximately 0.7 mg of 17-HSD per gram of protein, and the concentrations of 17-HSD measured by immunoassay and enzymatic activity proved to be strictly parallel in different partly purified placental preparations. The supernatants from centrifugations of human endometrial homogenates at 800 g and 105,000 g (after detergent treatment) displayed cross-reactivity with the antibody. The mean concentration of the cross-reacting substance in the radioimmunoassay was 14.1 μg/g protein (range 2–62.3) in specimens taken on different days in the cycle. These concentrations showed a significant correlation with the 17-HSD activities measured in the endometrial specimens (r = 0.722, P

Steroid Biosynthetic Enzymes: 17 β Hydroxysteroid Dehydrogenase

Polyclonal antibodies produced against human placental 17 β-hydroxysteroid dehydrogenase (17HSD), purified to homogeneity, and the corresponding cDNA for the enzyme were used to study the expression of 17HSD in a number of human tissues using various immunological methods together with RNA hybridization techniques. In addition, two 17HSD genes and their putative regulatory elements were sequenced. Immunoblotting analysis showed that the placental-type enzyme is expressed in granulosa-luteal cells, breast cancer tissue and breast cancer cell lines. An immunologically identical antigen was also detected in normal and carcinomatous human endometrium. The same antiserum, following affinity purification, was used for immunohistochemical studies of the endometrium and breast tissue, whereupon staining of the cytoplasm of the epithelial cells alone was observed. Immunostalning was also present in cultured human granulosa cells and in about half of the endometrial and breast carcinoma specimens investigated. Prog...

Cholic acid and chenodeoxycholic acid concentrations in serum during infancy and childhood.

Abstract. Heikura, S., Similä, S., Finni, K., Mäentausta, O. and Jänne, O. (Departments of Clinical Chemistry, Biochemistry and Paediatrics, University of Oulu, Oulu, Finland). Cholic acid and chenodeoxycholic acid concentrations in serum during infancy and childhood. Acta Paediatr Scand, 69: 659, 1980.—Concentrations of two primary bile acids (cholic and chenodeoxycholic acids) were determined by radioimmunoassay in the serum of infants and children at ages ranging from 1 hour to 15 years. The same bile acids were also measured in umbilical cord serum. Concentrations of the primary bile acids were significantly higher in the serum of 1‐hour old infants than those in the umbilical cord serum or the peripheral vein serum of adults. The levels of cholic and chenodeoxycholic acid remained high until the age of 6 months, being about 5‐fold higher than those in the sera of adults. Primary bile acid concentrations reached the adult level by the age of 1–2 years. These results indicate that developmental changes occur in the metabolism and excretion of bile acids in man. The relatively high concentrations of the primary bile acids in serum during the first 6 months of life suggest that up to this age, the mature ability of the liver to excrete the bile salts into the bile and/or to clear them from the circulation has not yet been reached.

Gall bladder volume and serum bile acids in cholestasis of pregnancy.

Summary. Gall bladder volume was measured by ultrasound in eight patients with cholestasis of pregnancy, in 21 normal pregnant women and in 14 non‐pregnant women, and at the same time serum cholic and nodeoxycholic acid levels were estimated. The gall bladder volume was 60% larger in cholestasis than that in normal pregnancy and more than two times larger in normal pregnancy than in non‐pregnant women. The serum cholic and chenodeoxycholic acid concentrations were significantly higher in cholestasis of pregnancy than those in normal pregnancy.

Gallbladder function and maternal bile acids in intrahepatic cholestasis of pregnancy

Ultrasonic measurement of the gallbladder volume was taken in 8 nonpregnant, healthy women, in 7 women with normal pregnancies and in 7 women whose pregnancies were complicated by intrahepatic cholestasis of pregnancy, in the fasting state and 30, 60, 120 and 180 min after a test meal. At the same time the serum concentrations of cholic acid and chenodeoxycholic acid were measured. The fasting and ejection volumes of the gallbladder in cholestasis of pregnancy were greater than in normal pregnancy. The fasting volume of the gallbladder was greater, but the ejection volume smaller in normal pregnancy than in nonpregnant women. No difference in the ultrasonic appearance of the intra- and extrahepatic bile ducts was found between the groups. Serum bile acids were increased in cholestasis of pregnancy and did not display any decreasing tendency after the postprandial rise during the following 3 h. The results indicate that in cholestasis of pregnancy the gallbladder function and the enterohepatic circulation of bile acids are different from normal pregnancy. This may be associated with the great tendency to gallstones in these women. The large size of the gallbladder in cholestasis of pregnancy has differential diagnostic importance in the ultrasonic evaluation of a pregnant woman with liver disease.

Serum Cholic Acid and Chenodeoxycholic Acid Concentrations in Neonatal Hyperbilirubinemia

Primary bile acid concentrations were measured in serum of 332 newborns with neonatal hyperbilirubinemia (serum total bilirubin level greater than 200 mumol/l) and compared with those of 95 nonhyperbilirubinemic neonates (serum total bilirubin level less than 200 mumol/l). The serum concentrations (mumol/l; mean +/- SEM) for cholic acid (8.78 +/- 0.44) and chenodeoxycholic acid (10.5 +/- 0.68) were significantly higher (p less than 0.001) in the hyperbilirubinemic group than in the controls (7.16 +/- 0.48 and 6.67 +/- 0.48, respectively). 80 (24%) of the hyperbilirubinemic newborns had true cholestasis (serum levels of cholic and/or chenodeoxycholic acid higher than mean +/- 2 SD in the reference group). The ratio of cholic to chenodeoxycholic acid was significantly higher (p less than 0.05) in the cholestatic group than in the hyperbilirubinemic newborns without cholestasis. There was no significant differences in the serum concentrations of alkaline phosphatase or lactate dehydrogenase between the cholestatic and noncholestatic groups. In the hyperbilirubinemic newborns, the primary bile acids were indiscriminately raised. Only 8 infants from the 332 newborns had jaundice at the age of 1 month. Of these 8 infants only 2 had neonatal cholestatic hyperbilirubinemia. It thus appears that measurement of serum primary bile acid concentrations has only limited diagnostic value in assessing the severity or prognosis of neonatal hyperbilirubinemia.

Serum bile acids and lipids during treatment of climacteric symptoms with natural oestrogen--progestin combinations.

Climacteric symptoms of 21 women were treated for 6 mth with sequential combination preparations containing natural oestrogen (oestradiol and oestriol) and norethisterone acetate as progestin. There were no significant changes during the treatment period in the serum alanine aminotransferase activity or concentrations of cholesterol, high-density lipoprotein (HDL)-cholesterol, triglycerides, cholic acid and deoxycholic acid. The concentration of chenodeoxycholic acid was, however, significantly decreased after 6 mth treatment. It thus appears that the above natural oestrogen--progestin combinations do not have adverse effects on hepatic function and lipid metabolism.