Mariangela Ronchi

Effects of ursodeoxycholic acid therapy for liver disease associated with cystic fibrosis.

The hydrophilic bile acid ursodeoxycholic acid (UDCA) has recently been shown to improve indexes of liver function in adult patients with various liver diseases. The clinical and biochemical responses to UDCA administration (10 to 15 mg/kg body weight per day) were therefore investigated in nine patients with cystic fibrosis and evidence of liver disease. All patients were receiving pancreatic enzymes and taurine supplementation. Liver function tests were done and serum bile acid concentrations and biliary bile acid composition were determined before and during UDCA therapy; fat balance studies and fecal bile acid excretion were carried out before and 6 months after UDCA treatment. After 2 months of bile acid therapy, biliary bile acid composition was enriched in UDCA from approximately 5% before treatment to 25%, at the expense of cholic and chenodeoxycholic acids, thus making the pool more hydrophilic. This enrichment is lower than that reported for adults with chronic liver diseases. Serum concentrations of UDCA increased significantly but variably. UDCA became the predominant fecal bile acid excreted (12% to 67%), indicating a variable absorption of the administered bile acid. Liver function improved in all patients after 2 to 6 months of therapy, although the degree of improvement (aspartate aminotransferase, -34%; alanine aminotransferase, -41%; gamma-glutamyltranspeptidase, -41% alkaline phosphatase, -19%) was lower than that observed in adults with chronic liver diseases. Mean coefficient of fat absorption and growth rate were, on average, unaffected by UDCA therapy, although an improvement was noted for three patients with greater severity of steatorrhea. The study indicates that UDCA can be used safely in this patient population but that higher doses of UDCA may be of greater benefit in the treatment of the liver disease associated with cystic fibrosis.

Correlation between Fetal and Maternal Serum Bile Acid Concentrations

ABSTRACT: Serum concentrations of different bile acids (BA) were determined by radioimmunoassay in 56 human fetuses and mothers. Serum was obtained immediately after legal abortion, performed between the 14th and the 21st wk of gestation. Conjugated cholic (CCA) and chenodeoxycholic acid (CCDCA) concentrations were determined in 33 cases, conjugated lithocholic (CLCA) and deoxycholic acid (CDCA) in 20, and sulfolithocholyglycine (SLCG) in 15. In fetal blood, mean concentrations of CCA (0.80 ± 0.40 μmol/liter), CCDCA (4.50 ± 2.70 μmol/liter), and CLCA (1.70 ± 1.04 μmol/liter) were significantly higher than those in the mother (CCA 0.34 ± 0.17 μmol/liter; CCDCA 0.79 ± 0.34 μmol/liter; CLCA: 0.70 ± 0.30 μmol/liter; p < 0.001); fetal serum levels of CDCA (0.46 ± 0.32 μmol/liter) and SLCG (0.15 ± 0.09 μmol/liter) were lower than in the mothers (CDCA 1.20 ± 0.80 μmol/liter, p < 0.001; SLCG 0.40 ± 0.30 μmol/liter, p < 0.01). There was no correlation between levels of BA and gestational age. Serum total protein and albumin concentrations were both reduced in 10 fetuses as compared with the mothers. These data support the concept of a state of physiologic cholestasis during development and suggest that placental transfer of primary BA occurs mostly in the fetal to maternal direction. This transfer could be facilitated by the reduced fetal plasma albumin concentration, since BA in free solution diffuse more easily through the placenta. There is evidence of lithocholic acid synthesis in the fetal liver, while deoxycholic acid appears to be mostly of maternal origin. Finally, sulfation of BA is poorly developed at this age of gestation.

The steatocrit: a simple method for monitoring fat malabsorption in patients with cystic fibrosis

Steatocrit was determined through microcentrifugation of fecal homogenate from 110 pediatric controls and 107 patients with cystic fibrosis (CF). For 74 CF patients, steatocrit was determined in the same fecal material collected to determine a fat balance. In controls, steatocrit value was 0.7 +/- 1.0%, which was significantly lower than values found in CF patients with a coefficient of fat excretion less than 10% of intake (1.7 +/- 1.2%). Significantly increased values were found in CF patients with a coefficient of fat excretion ranging between 10 and 25% of intake (4.7 +/- 1.7%) and in those whose coefficient of fat excretion was greater than 25% of intake (11.3 +/- 4.3%). In the 74 CF patients, steatocrit was directly correlated to the coefficient of fat excretion (r = 0.93; P less than 0.001). We performed steatocrit several times in the course of the 1st year of life in 33 infants with CF diagnosed by means of CF screening. Values obtained at the time of diagnosis, before starting enzymatic therapy, were relatively high; they showed a progressive decrease when, using steatocrit as a guide, the dose of pancreatic enzymes had been increased. The normalization of steatocrit values was accompanied by a better growth rate in the majority of these infants, confirming the importance of an optimal early correction of pancreatic insufficiency. We propose that this simple semiquantitative test can be usefully performed for the frequent monitoring of fat absorption and for checking the response to enzymatic therapy in patients with CF.

Effect of Taurine Supplementation on Fat and Bile Acid Absorption in Patients with Cystic Fibrosis

Eleven children with cystic fibrosis (CF) and pancreatic insufficiency were given supplementation with taurine (30-40 mg/kg/day) for 2 months, while taking their usual dosage of enzymatic therapy. One patient dropped out of the study because she developed severe constipation. In the other 10 patients, urinary taurine excretion (88 +/- 30.1 mg/m2s.a./24 h) was similar to that of controls (86.2 +/- 6 mg/m2s.a./24 h) before taurine and increased markedly after supplementation (618.2 +/- 79.97 mg/m2s.a./24 h), indicating efficient intestinal absorption. Their coefficient of fat absorption was 81.2 +/- 2.3% and increased significantly after taurine (91.3 +/- 1.13%; p less than 0.01); the area under the curve of plasma triglyceride postprandial levels (1 +/- 0.1 mg X min/ml) also increased significantly after taurine (1.4 +/- 0.3 mg X min/ml; p less than 0.05), showing values very similar to those of controls. Conversely, no change was observed in the serum postprandial levels of glycocholic acid: the maximum postprandial peak before (1.2 +/- 0.3 mumol/l) and after taurine (1 +/- 0.1 mumol/l) remained significantly lower than in controls (2.4 +/- 0.3 mumol/l); p less than 0.01 and p less than 0.001, respectively. Mean total fecal bile acid (BA) excretion was 10.24 +/- 2.15 mg/kg/day before taurine and 12.8 +/- 4.27 mg/kg/day after taurine (normal pediatric values, 2.91 +/- 1.1 mg/kg/day); however, in the individual patients we found a variable trend, four of them showing a net increase in fecal BA excretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Value of a commercial kit for detecting anti-C1q autoantibodies and correlation with immunological and clinical activity of lupus nephritis

Abstract Background: The association of anti-C1q antibodies (anti-C1q) with the renal activity of lupus nephritis (LN) and the methods for their determination is still a matter of debate. Methods: In 116 serum samples of 66 patients with biopsy proven LN, we aimed: 1) to compare the results of the determination of anti-C1q obtained by a commercial kit with a clinically validated in-house ELISA; 2) to evaluate the correlation of anti-C1q with the most important immunological and clinical parameters employed in LN, i.e., antibodies to dsDNA (anti-dsDNA), C3 and C4 complement component, haemoglobin and haematuria. Results: Good correlation and agreement between the two methods (r=0.81, p<0.0001; contingency coefficient=0.70, p<0.0001, respectively) were demonstrated. No differences were observed between the two assays by ROC curves comparison. Anti-C1q levels were significantly higher in patients with active LN [44 arbitrary units (AUs)] in comparison to those with inactive LN (23 AUs, p=0.047) and significantly correlated with anti-dsDNA (r=0.44, p<0.0001), complement fractions (C3: r=−0.33, p=0.001; C4: r=−0.29, p=0.003), haemoglobin levels (r=−0.34, p=0.0004) and the number of urinary red blood cells (r=0.26, p=0.01). Conclusions: Our results suggest the validity of this commercial assay in detecting anti-C1q and confirm the association of anti-C1q with renal involvement of LN and the importance of introducing this parameter in the analytical panel for the evaluation of LN activity.

Serum levels of immunoreactive trypsin during development: Comparison with levels of lipase and amylase

In order to obtain additional information on serum pancreatic enzyme levels during development, we have measured immunoreactive trypsin (IRT), immunoreactive lipase (IRL), and total amylase in paired fetal and maternal sera. Samples were obtained during early gestation (14–21 week of gestation) and at the time of normal delivery. IRT levels were lower in maternal sera as compared to paired fetal and neonatal (p < 0.005); conversely, IRL and amylase, although present in measurable concentrations, were significantly lower in fetal and neonatal sera than in the maternal (p < 0.001). We also serially monitored serum pancreatic enzyme levels in a group of premature infants during the first 10 days of life. Concentrations of IRT showed a significant increase over time (p < 0.05) and those of IRL remained stable while amylase levels decreased sharply, suggesting possible maternal origin of this enzyme. Serum concentrations of the three pancreatic enzymes in newborns at term (second day of life) were higher than in infants aged 0.5–6 months; however, only IRT levels were above the normal range for adults. Beyond the neonatal period, IRT levels were stable and comparable to adults, whereas amylase and IRL levels were very low in infants younger than 6 months and increased significantly with age (p < 0.001). These data seem to indicate that “physiologic hypertrypsinemia” occurs early during development and may be accentuated by postnatal events. They provide an indirect indication of both early fetal production of trypsinogen and possible placental transfer of pancreatic enzymes from the maternal circulation. The overall results would suggest that only beyond the neonatal period can determination of serum pancreatic enzyme levels be used as an indirect index of their asynchronous development.

92 SERUM GLYCINE (G) AND TAURINE (T) CONUGATED BILE ACIDS (BA) IN PREMATURE FORMULA-FED INFANTS BEFORE AND AFTER TAURINE SUPPLEMENTATION

Tauro-conjugated BA, which are predominant during development, are more polar and more resistant to passive intestinal absorption than G-conjugated. To further elucidate the possible impact of postnatal BA conjugation pattern on the dynamic of enterohepatic circulation, we have studied 16 premature formula-fed infants (29-36 gw): at the age of 2 weeks, they were randomly assigned to receive an adapted formula(F)with or without T (25μmol/dl). Before and after 4 weeks of this diet, fasting serum levels of cholic (CA) and chenodeoxycholic (CDCA) acids were determined by competitive solid phase enzyme immunoassay, after TLC separation of the G and T conjugated forms.The predominance of G-conjugates (mainly of CA)in serum of prematures, contrasts with the reported excess of T-conjugated BA in their duodenal fluid and may be due to later appearance of the active ileal transport system, for which T-conjugates are almost exclusively dependent to be absorbed. T-feeding produced a slight change in the relative proportion of G and T conjugates in serum, which seems limited to CA, in agreement with the “in vitro” observation of higher synthesis of conjugates for CA than CDCA. The sensible decrease of serum CA levels after T supplementation may be related to the inability of prematures to absorb increased amounts of tauro-CA, while extensive passive absorption would permit conservation of the more hydrophobic CDCA.

The diagnostic accuracy of biomarkers for diagnosis of primary biliary cholangitis (PBC) in anti-mitochondrial antibody (AMA)-negative PBC patients: A review of literature

Abstract Primary biliary cholangitis (PBC), also known as primary biliary cirrhosis, is an autoimmune disease of the liver characterized by anti-mitochondrial antibodies (AMA) in 90%–95% of patients. The aim of this study was to evaluate the diagnostic value of several serum biomarkers in patients with PBC but negative for AMA. Some antinuclear antibodies (ANA) pattern, detected by indirect immunofluorescence (IIF), such as multiple nuclear dot (MND) and rim-like patterns are well-known to be specific for PBC. The corresponding nuclear antigens are the components of the nuclear pore complex (Gp210 for rim-like pattern) and Sp100, PML proteins (for MND pattern) detectable by immunoblotting and ELISA methods. More recently, new biomarkers have been evaluated in order to improve the diagnostic sensitivity, such as kelch-like 12 (KLHL12) and hexokinase-1. Considering these different serum biomarkers, studies evaluating their diagnostic role in AMA-negative PBC patients compared to AMA-positive ones and controls were included in this review. Pooled sensitivity and specificity were 37% and 85%, respectively. The overall PPV and NPV mean values were 45% and 83%. Even if all biomarkers are very specific for PBC, the overall sensitivity was poor and therefore is necessary to identify a marker with a greater sensitivity for PBC in AMA-negative patients.

EFFECTS OF TAURAIN (T) AND URSODEOXYCHOLIC ACID (UDCA) ON LIVER FUNCTION TESTS IN PATIENTS WITH CYSTIC FIBROSIS

Patients with Cystic Fibrosis (CF) often develop liver disease and although the mechanism leading to the pathognomonic lesion (focal biliary cirrhosis) is still unknown, quantitative/qualitative alterations in bile secretion may of of major importance. Correction of the complex abnormalities of bile acid (BA) metabolism described in these patients, (BA malabsorption with predominance of relatively more hydrophobic glycine-conjugated BA), may thus slow the progress of liver involvement associated to CF. Recently, a shift of the BA pool composition towards more hydrophilic-less hepatotoxic components, induced by UDCA and, in one report, by T administration, has been shown to be beneficial in adults with Primary Biliary Cirrhosis and Chronic Hepatitis. In 6 CF patients (aged 8-12 years) with clinical and biochemical signs of liver involvement but no portal hypertension, T (30 mg/Kg/die) was administered one month before and during the successive treatment with UDCA (10-15 mg/Kg/die). Liver functin tests were determined before and during each period of treatment, fecal fat and BA (GLC) exretion and biliary BA composition (HPLC) before and after long term administration of UDCA and T. Before treatment, both mean coefficient of fat absorption n enzymatic therapy (83.7 ± 10%) and fecal BA excretion (12.3± 5.9 mg/Kg/die) were abnormal. Chenodeoxycholic acid was predominant among biliary BA (44.1 ± 9.1%) and the glycine to taurine conjugate ratio was 2.7 ± 2.0. T administration produced only inconsistent changes of lvier function tests from basal abnormal values, whereas in patients treated fo two months with T + UDCA a substantial improvement was observed: AST -28%, ALT -40%. GT -33%, Alkaline Phosphatase -19%. The effects of longer periods of treatment are currently investigated, with purpose of establishing their relationship with changes in the BA pool composition.

13. BILE ACID (BA) PATTERN AM) CONJUGATION IN THE HUMAN FETUS

The characterization of BA metabolism during development is essential to understand the clinical problems related to the immaturity of the entero-hepatic circulation of BA in early life. We have previously examined primary BA ccncentrations in fetal bile during early gestation: these were low prior week 16, but after this time showed a surge in the range of 10-30 fold. We report here the results of qualitative BA analysis of 32 samples of fetal bile (14th - 21st gw)by means of GLC-MS and HPLC. In all the samples, the predominant BA were chenodeoxycholic and cholic acids, while the main secondary BA were present only in small amounts. Several “atypical BA” were found, with hydroxyl groups in unusual positions in the BA steroid nucleus and they constituted around 40% of total BA in each sample; hyocholic acid and a trihydroxy-BA of unknown structure were quantitatively the most important and together these frequently exceeded the amounts of cholic acid. The presence of 3β-Hydroxy-5-cholenoic ac. and 3-oxo-7α-hydroxy-5β-cholanoic acid seems to indicate fetal synthetic pathways in which side chain oxidation preceeds nuclear changes in structure, HPLC analysis of fetal bile revealed the major proportion of BA (around 80 %) to be taurine conjugated, while a much lower proportion was in the glycine conjugated fraction; less than 5 % of biliary BA were found to be sulphated.These unique characteristics of BA metabolism may in part explain the cholestatic tendency during development; these data may also be of help to elucidate the etiology of certain forms of neonatal cholestasis due to abnormalities in BA metabolism.