Comparative effects of lovastatin and chenodeoxycholic acid on plasma cholestanol levels and abnormal bile acid metabolism in cerebrotendinous xanthomatosis.

We investigated the effect of the hepatic hydroxymethyl glutaryl coenzyme A (HMG-CoA) reductase inhibitor lovastatin and the primary bile acid chenodeoxycholic acid (CDCA) on plasma sterol and bile alcohol concentrations and the excretion of bile alcohols in urine in a 38-year-old homozygote with cerebrotendinous xanthomatosis (CTX). Untreated, plasma cholesterol concentrations were less than normal (171 +/- 5 v 185 +/- 3 mg/dL, P < .05) while plasma cholestanol levels were more than 20 times higher than the control mean (2.26 +/- 0.17 v 0.1 +/- 0.1 mg/dL, P < .0001). Plasma and urinary bile alcohol concentrations were markedly increased (12.6 +/- 0.6 and 154 micrograms/mL, respectively, v trace amounts in controls), with the ratio of 5 beta-cholestane-3 alpha,7 alpha,12 alpha, 25-tetrol to 5 beta-cholestane, 3 alpha,7 alpha,12 alpha,23 (22 and 24),25-pentols being 1.6 in plasma and reversed to 0.15 in urine. Treatment with lovastatin (40 mg/d) reduced plasma cholesterol concentrations 13%, but failed to decrease plasma cholestanol or bile alcohol levels. Abundant amounts of bile alcohols continued to be excreted in urine. In contrast, CDCA (750 mg/d) inhibited abnormal bile acid synthesis, as evidence by a 17-fold decrease in total bile alcohol levels in plasma and a 29-fold decrease in urine and the virtual elimination of cholic acid and deoxycholic acid from the bile. Plasma cholestanol concentrations also decreased 85%, but cholesterol levels increased 14%. The combination of CDCA with lovastatin did not improve plasma cholestanol or bile alcohol concentrations compared with CDCA treatment alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Determination of the glucurono-conjugated position in bile alcohol glucuronides excreted in urine of a patient with cerebrotendinous xanthomatosis by a nuclear magnetic resonance study.

This paper describes the determination of the glucurono-conjugated position in two bile alcohol glucuronides excreted in urine of a patient with cerebrotendinous xanthomatosis by a nuclear magnetic resonance study. The urine sample was extracted with reversed-phase resin, and chromatographed on a reversed-phase partition column and a silica gel column to isolate glucurono-conjugates of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha, 25-tetrol and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha, 23,25- pentol. Proton and carbon-13 nuclear magnetic resonance spectra of the natural tetrol glucuronide were identical with those of the chemically synthesized tetrol glucuronide, 7 alpha, 12 alpha, 25-trihydroxy-5 beta-cholestane-3 alpha-O-beta-D- glucopyranosyluronic acid. Hence, the glucurono-conjugated position of the natural tetrol glucuronide was determined to be the C-3 position. By comparison of the 13C chemical shift data with that of the unconjugated pentol, 5 beta-cholestane-3 alpha, 7 alpha, 23,25-pentol, the glucurono-conjugated position of the natural pentol glucuronide was determined to be C-23. Thus the natural pentol glucuronide can be formulated as 3 alpha, 7 alpha, 12 alpha, 25- tetrahydroxy-5 beta-cholestane-23-O-beta-D-glucopyranosyluronic acid. The difference in the glucurono-conjugated position between the 25-tetrol glucuronide and the 23,25-pentol glucuronide indicates that the former is not the biosynthetic precursor of the latter.

Capillary gas-liquid chromatographic separation of bile alcohols.

Gas-liquid chromatographic separation of C23, C24, C25, C26, and C27 bile alcohols with either 3 alpha, 7 alpha-dihydroxylated or 3 alpha, 7 alpha, 12 alpha-trihydroxylated ring system on two capillary columns, CP-Sil-19 CB and CP-Sil-5 CB, is described. Bile alcohols with two ring hydroxyl groups at 3 alpha- and 7 alpha-positions consistently showed larger retention times on CP-Sil-19 CB columns and smaller retention times on CP-Sil-5 CB columns than the corresponding bile alcohols with three ring hydroxyl groups at 3 alpha-, 7 alpha-, and 12 alpha-positions. Resolutions of all bile alcohols were better on CP-Sil-19 CB columns; however, we hope that the gas-liquid chromatographic characteristics on the two columns will be useful for better identification of bile alcohols in biological fluids.

Identification of (23S)-5 alpha-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol in urine of patients with cerebrotendinous xanthomatosis.

This paper describes the identification of a new bile alcohol possessing the 5 alpha-cholestane structure that was found in the urine of patients with cerebrotendinous xanthomatosis. The urine samples were extracted with reversed-phase resin, treated with beta-glucuronidase, and separated on silica gel and reversed-phase column chromatography. The new bile alcohol isolated was the second component of the urinary bile alcohols and was identified as (23S)-5 alpha-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol by means of gas-liquid chromatography/mass spectrometry and nuclear magnetic resonance spectroscopic studies.

Identification of short side chain bile acids in urine of patients with cerebrotendinous xanthomatosis.

Urine from patients with cerebrotendinous xanthomatosis (CTX) was found to contain a number of minor bile acids along with three major bile acids, 7-epicholic acid, norcholic acid, and cholic acid. The following minor bile acids were identified by combined gas-liquid chromatography-mass spectrometry: 7-ketobisnordeoxycholic acid; 12-ketobisnorchenodeoxycholic acid; 7-ketonordeoxycholic acid; 12-ketochenodeoxycholic acid; 7-ketodeoxycholic acid; 12-ketochendeoxycholic acid; bisnorcholic acid; allonorcholic acid; allocholic acid; 1 beta-hydroxybisnorcholic acid; 1 beta-hydroxynorcholic acid; 1 beta-hydroxycholic acid; 2 beta-hydroxybisnorcholic acid; 2 beta-hydroxy-norcholic acid; 2 beta-hydroxycholic acid. The presence of C22 and C23 bile acids in urine of the CTX patients suggests that bile alcohols having a hydroxyl group at C22 or C23 in the side chain may be further degraded to these bile acids.

Cerebrotendinous xanthomatosis.

A case report on a 23-year-old female patient with cerebrotendinous xanthomatosis (CTX) is presented. From 8 years of age, the patient clinically showed multiple xanthoma masses on both knees, both heels, and the nasal bridge, juvenile cataracts, multiple abnormal neurologic dysfunctions, and dementia. The level of cholestanol in urine, serum, and xanthoma mass tissues was increased, as determined by capillary gas chromatography.

Abnormal urinary bile acids in a patient suffering from cerebrotendinous xanthomatosis during oral administration of ursodeoxycholic acid.

The urinary bile acid profile, obtained by capillary gas chromatography, of a patient suffering from cerebrotendinous xanthomatosis and treated with ursodeoxycholic acid demonstrated, besides the occurrence of 23-norcholic acid and (23R)-hydroxycholic acid (as a consequence of this disease), six additional unknown bile acids and three known bile acids, viz. ursodeoxycholic acid, hyocholic acid and omega-muricholic acid. The structure of two of the unknown bile acids were elucidated and proven by organic syntheses. These were 23-norursodeoxycholic acid and 3 beta-ursodeoxycholic acid. The structures of three bile acids were tentatively elucidated as being 1 beta-hydroxyursodeoxycholic acid, 21-hydroxyursodeoxycholic acid and 22-hydroxyursodeoxycholic acid, and the possibility that the structure of the remaining bile acid is that of 5-hydroxyursodeoxycholic acid is discussed. Two of these bile acids (1 beta-hydroxyursodeoxycholic acid and 5-hydroxyursodeoxycholic acid) also occurred in urine of a healthy individual during oral ursodeoxycholic acid treatment, whereas 23-norcholic acid, 23-norursodeoxycholic acid, (23R)-hydroxycholic acid, 21-hydroxyursodeoxycholic acid and 22-hydroxyursodeoxycholic acid were only present in urine of the patient suffering from cerebrotendinous xanthomatosis. The metabolism of ursodeoxycholic acid, both in the normal state and in the cerebrotendinous xanthomatosis, is discussed.

Screening for cerebrotendinous xanthomatosis by using an enzymatic assay for 7 alpha-hydroxylated steroids in urine.

We used a commercial enzymatic kit for measuring 7 alpha-hydroxylated bile acids to screen urines from normal subjects, liver-transplant recipients, and patients with various liver diseases, cerebro-hepato-renal syndrome, or cerebrotendinous xanthomatosis (CTX). Because of their high concentrations of 7 alpha-hydroxylated compounds excreted, the CTX patients were clearly distinguished from all other groups except for a slight overlap with the patients with cerebro-hepato-renal syndrome and liver-transplant recipients. Gas chromatography for bile alcohols completed the differential diagnosis.

Configuration at C-25 in 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol excreted by patients with cerebrotendinous xanthomatosis: circular dichroism and 13C-NMR studies.

The configuration at C-25 in 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha,25,26-pentol isolated from the bile and feces of patients with cerebrotendinous xanthomtosis (CTX) was determined from the lanthanide-induced circular dichroism (CD) Cotton effects and 13C-NMR measurements. Under anhydrous conditions, CD spectra of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha,25,26-pentol in the presence of Eu(fod)3 exhibited a large induced negative Cotton effect at 320 nm. On the basis of the empirical rule (primary-tertiary-alpha-diols) in which R compounds have positive Cotton effects and S compounds have negative Cotton effects at 320 nm, it was concluded that 25,26-pentol has the 1,2,glycol structure with C-25 having the S-configuration. This assignment was based upon comparison with model compounds, 25(R and S),26-dihydroxy cholesterols and 25(R and S),26-dihydroxy cholecalciferols whose single-crystal X-ray structure and 13C-NMR studies have been performed. It is suggested that these data may be helpful to clarify the stereospecificity of the hydroxylation of the terminal methyl group of the cholesterol side chain in CTX.

Capillary gas chromatography of urine samples in diagnosing cerebrotendinous xanthomatosis.

A patient is described with many clinical features of cerebrotendinous xanthomatosis (CTX), but with only slightly elevated cholestanol/cholesterol concentration ratios in serum and xanthomatous tissue. However, with capillary gas chromatographic determinations of urinary bile acids and bile alcohols we demonstrated the typical biochemical abnormalities as seen in CTX patients. This article emphasizes the value of urinary capillary gas chromatography as a specific test to establish the diagnosis of CTX and to monitor the biochemical effectivity of the different treatment regimens.

Increased (23R)-hydroxylase activity in patients suffering from cerebrotendinous xanthomatosis, resulting in (23R)-hydroxylation of bile acids.

Patients suffering from cerebrotendinous xanthomatosis, an inborn error of metabolism in bile acid synthesis, excrete excessive amounts of 23-hydroxylated bile alcohols, 23-norcholic acid and 23-hydroxycholic acid into urine. In this study the configuration of this excreted 23-hydroxycholic acid was established as (23R)-hydroxycholic acid. Urine samples of two treated patients, receiving chenodeoxycholic acid, were investigated to see whether this administered bile acid was partly converted into 23-hydroxychenodeoxycholic acid. One patient was treated with ursodeoxycholic acid for 1 month and subsequently with chenodeoxycholic acid, and the urinary excretion of both (23R)-hydroxychenodeoxycholic acid and (23R)-hydroxyursodeoxycholic acid were followed. Indeed, all three patients excreted (23R)-hydroxylated chenodeoxycholic acid during oral treatment with chenodeoxycholic acid, and the patient treated with ursodeoxycholic acid excreted (23R)-hydroxylated ursodeoxycholic acid. During treatment with chenodeoxycholic acid the excretion of (23R)-hydroxychenodeoxycholic acid increases at first and later on decreases markedly. These findings suggest increased (23R)-hydroxylase activity in patients suffering from cerebrotendinous xanthomatosis, acting both on endogenously synthesized bile alcohols and on exogenously administered bile acids; during continuation of chenodeoxycholic acid treatment in an effective dose (750 mg/day) this enzyme activity gradually disappears.

Detection of carriers of cerebrotendinous xanthomatosis.

Patients suffering from cerebrotendinous xanthomatosis (an autosomal recessive inborn error of metabolism) can easily be distinguished from patients not suffering from this disease, as the first excrete large amounts of the bile alcohol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol, in urine, whereas the second do not. In order to find out, whether carriers of cerebrotendinous xanthomatosis can be detected in a biochemical way, we compared known carriers with controls. The urinary excretions of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol of both groups were practically absent and no selection of carriers with cerebrotendinous xanthomatosis could be made on that basis. When, however, carriers and non-carriers were subjected to cholestyramine treatment, by which endogenous bile acid synthesis was stimulated, the urinary excretion of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol in the carrier rose considerably, whereas this excretion remained essentially the same in the non-carriers. This test can be of value in the genetic counseling of carriers with cerebrotendinous xanthomatosis and helpful in the detection of newborn patients with cerebrotendinous xanthomatosis.

Evaluation of urinary cells in acid cholesteryl ester hydrolase deficiency.

Deficiency in the lysosomal enzyme responsible for cholesteryl ester hydrolysis, acid cholesteryl ester hydrolase (E.C. 3.1.1.13), leads to two clinically recognized diseases: Wolman disease and cholesteryl ester storage including leukocytes, fibroblasts and liver. Analysis of urinary sediment from well characterized cases of Wolman disease and CESD also revealed the shedding of lipid enriched renal tubular cells. Morphologic, enzymic and lipid compositional studies of these cells indicate that the enzyme deficiency observed in fibroblasts and leukocytes from these individuals are reflected in these cells shed in the urine. These findings in renal tubular cells confirm and extend those made in other cell types. These studies indicate that analysis of urinary sediment in suspected cases of acid cholesteryl ester deficiency may provide a meaningful approach for monitoring therapeutic attempts involving enzyme infusion and gene therapy.

Synthesis and structure of 26 (or 27)-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24S,25 xi-pentol isolated from the urine and feces of a patient with sitosterolemia and xanthomatosis.

The urine and feces of a patient with the rare inherited lipid storage disease, sitosterolemia and xanthomatosis, were analyzed. Substantial quantities of C26-bile alcohol, 26 (or 27)-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24S,25 xi-pentol along with 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24-tetrol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24R,25-pentol, and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol were found. The structure of the C26-bile alcohol was confirmed by direct comparison (gas-liquid chromatography-mass spectrometry and thin-layer chromatography) with a standard sample synthesized from cholic acid. The configurational assignment at C-24 was determined by lanthanide-induced circular dichroism Cotton effect measurements. The increased excretion of these C26- and C27-bile alcohols suggests an abnormality of bile acid biosynthesis in this disease.

Capillary gas chromatographic determinations of urinary bile acids and bile alcohols in CTX patients proving the ineffectivity of ursodeoxycholic acid treatment.

Urine samples and serum samples of a patient with cerebrotendinous xanthomatosis (CTX) were investigated by means of capillary gas chromatography, both before and during oral treatment with ursodeoxycholic acid (UDCA), and the results compared with those obtained during chenodeoxycholic acid (CDCA) therapy. The predominantly excreted bile alcohol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol and two abnormal bile acids, i.e. 23-norcholic acid and 23-hydroxycholic acid were determined. In addition, the serum cholestanol/cholesterol ratio was determined. Whereas previous experiments demonstrated that the urinary excretion of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol and the abnormal bile acids decreased within a few weeks during CDCA therapy, the present study shows that their urinary excretions remain essentially the same during UDCA treatment. In contrast to the decrease in the serum cholestanol/cholesterol ratio during CDCA therapy, this ratio remains essentially the same during UDCA therapy. It is therefore concluded that, in contrast to CDCA therapy, UDCA treatment is not effective in the treatment of CTX.

Diagnosis of cerebrotendinous xanthomatosis (CTX) and effect of chenodeoxycholic acid therapy by analysis of urine using capillary gas chromatography.

By means of capillary gas chromatography urine samples of patients with cerebrotendinous xanthomatosis (CTX) were investigated before and during treatment by oral administration of chenodeoxycholic acid. The occurrence of various conjugated bile alcohols, presumably glucuronides, was demonstrated, the major compound being 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha, 23 xi, 25-pentol. In the bile acid fraction norcholic acid and hydroxycholic acid were shown to be present in considerable amounts. In this way the presence of CTX can be demonstrated conclusively. After chenodeoxycholic acid therapy the excretion of both abnormal bile acids as well as of bile alcohols rapidly decreased within a few weeks, showing the effectiveness of the treatment. By early discovery and subsequent therapy it may be possible to prevent the onset of the detrimental symptoms such as mental deficiency, caused by the accumulation of cholestanol and cholesterol in CTX patients.

Urinary cytology in the diagnosis of renal xanthogranulomatosis.

We present our 4-year experience on the preoperative diagnosis of renal xanthogranulomatosis by sequential urinary cytology. The usefulness of this method is evident due to the absence of any other possible means of preoperative diagnosis of this disease. The possibility of obtaining foam cells by ureteral catheter washouts and translumbar aspiration in the patients, where spontaneous urine cytologies were negative, is also pointed out.

Preoperative diagnosis of renal xanthogranulomatosis by serial urinary cytology: preliminary report.

The search for foam cells in the urinary sediment of patients with possible renal xanthogranulomatosis can constitute a harmless and useful diagnostic method for this frequently occurring renal disease, which previously has been diagnosed with certainty only by histological examination or biopsy. Of 5 cases studied the search was positive in 80 per cent.