Brunetta Porcelli

Overcoming a "Probable" diagnosis in antimitochondrial antibody negative primary biliary cirrhosis: Study of 100 sera and review of the literature

Serum anti-mitochondrial antibodies (AMA) are the serological hallmark of primary biliary cirrhosis (PBC), yet up to 15% of PBC sera are AMA negative at routine indirect immunofluorescence (IIF) while being referred to as “probable” cases. The diagnostic role of PBC-specific antinuclear antibodies (ANA) remains to be determined. We will report herein data on the accuracy of new laboratory tools for AMA and PBC-specific ANA in a large series of PBC sera that were AMA-negative at IIF. We will also provide a discussion of the history and current status of AMA detection methods. We included IIF AMA-negative PBC sera (n = 100) and sera from patients with other chronic liver diseases (n = 104) that had been independently tested for IIF AMA and ANA; sera were blindly tested with an ELISA PBC screening test including two ANA (gp210, sp100) and a triple (pMIT3) AMA recombinant antigens. Among IIF AMA-negative sera, 43/100 (43%) manifested reactivity using the PBC screening test. The same test was positive for 6/104 (5.8%) control sera. IIF AMA-negative/PBC screen-positive sera reacted against pMIT3 (11/43), gp210 (8/43), Sp100 (17/43), both pMIT3 and gp210 (1/43), or both pMIT3 and Sp100 (6/43). Concordance rates between the ANA pattern on HEp-2 cells and specific Sp100 and gp210 ELISA results in AMA-negative subjects were 92% for nuclear dots and Sp100 and 99% for nuclear rim and gp210. Our data confirm the hypothesis that a substantial part of IIF AMA-negative (formerly coined “probable”) PBC cases manifest disease-specific autoantibodies when tested using newly available tools and thus overcome the previously suggested diagnostic classification. As suggested by the recent literature, we are convinced that the proportion of AMA-negative PBC cases will be significantly minimized by the use of new laboratory methods and recombinant antigens.

Comparative determination of purine compounds in carotid plaque by capillary zone electrophoresis and high-performance liquid chromatography

Allantoin, uric acid (UA), hypoxanthine (Hx) and xanthine (X) were determined on carotid plaque by capillary zone electrophoresis (CZE) and high-performance liquid chromatography (HPLC). Comparison of the results showed that capillary zone electrophoresis may have similar or even superior analytical performance to HPLC, especially for the determination of allantoin in biological samples.

Effects of testosterone on cholesterol levels and fatty acid composition in the rat.

The effects of testosterone treatment on cholesterol levels and its fatty acid components were studied in adult rats. Cholesterol levels increased both in the liver and in the serum of castrated rats. Androgen administration restored the normal values only in the serum. A general decrease in unsaturated and essential fatty acids in cholesterol esters was evident after testosterone administration. In the liver, only the C16:O/C16:1 ratio clearly increased after testosterone administration, which inhibited the delta 9 unsaturation of palmitic acid, but not of stearic acid. In the serum the C16:O/C16:1, C18:O/C18:1, and C18:2/C2O:4 ratios decreased after castration and were restored by testosterone. The results indicate a clear inhibition of delta 9 unsaturation of palmitic and stearic acids, of delta 5 unsaturation and elongation in organs other than the liver. No effect was evident on delta 6 unsaturation. This suggests that fatty acid unsaturations are regulated differently by testosterone in different tissues. For delta 9 unsaturation in the liver, the effect also seems to be substrate-dependent.

Fast-atom bombardment mass spectrometry for mapping of endogenous methylated purine bases in urine extracts.

Fast-atom bombardment (FAB) mass spectrometry, linked with tandem mass spectrometry (MS/MS), was employed for the identification of methylated purine bases in four urinary extracts of healthy subjects and fourteen urinary extracts of patients bearing colorectal tumors. In order to obtain an easy structural identification of the species present in urinary extracts, the MS/MS spectra of MH+ species of twenty nine diagnostically relevant purine bases were studied. Even if definitive quantitative data cannot be obtained by this approach, FAB mass spectra of urine extracts lead to a readily reproducible mapping of endogenous purine bases, allowing a distinction between healthy and sick subjects. Bases such as 9-ethyladenine, N6-2-isopentenyladenine and N6-benzyladenine were detected only in urine samples of colorectal tumor bearing patients. The detection in urine of compounds such as 7-methylguanine and 1-methylguanine, and their increase in the urine of colorectal tumor bearing patients, has been justified either by a more rapid turnover of nucleic acids in tumor tissue or by an increase in the extent of their methylation. The obtained results indicate that the method can be employed for diagnostic purposes.

Association between stressful life events and autoimmune diseases: A systematic review and meta-analysis of retrospective case–control studies

BACKGROUND Evidence of a relationship between stressful life events and the onset of autoimmune diseases is not univocal and there are no meta-analyses in the literature on the question. AIM To look for differences in the number and type of stressful life events in the premorbid period between patients with autoimmune diseases and healthy subjects. METHOD Review of the literature in PubMed and Scopus (January 1963-May 2015). INCLUSION CRITERIA We included retrospective case-control studies that compared patients diagnosed with autoimmune disorders and controls regarding the incidence of stressful events occurring before diagnosis, and investigated said events with validated questionnaires. EFFECT-SIZE INDEXES By random effect meta-analysis, two independent researchers calculated effect-size indexes as the difference between the means of the clinical groups and the control group in relation to the combined standard deviation. RESULTS The database searches produced 2490 articles, 14 of which were selected (3201 patients). Analysis showed a moderate but significant mean effect-size index [d=0.63, p<0.01], suggesting that autoimmune disorders are effectively associated with major stressful events in the premorbid period. The relationship between stressful events and autoimmune disease was weaker in studies with a high proportion of female subjects [β=-0.004, p<0.01] and stronger in studies that considered a longer interval between stressors and onset of disease [β=0.16, p<0.01]. CONCLUSIONS The results of this meta-analysis suggest that stressors may play an important role in the etiopathogenesis of autoimmune disorders. Only prospective studies can provide more certain inference about the causality of this relationship.

Celiac and non-celiac gluten sensitivity: a review on the association with schizophrenia and mood disorders

An association between many psychiatric and gluten-related disorders has been known for some time. In the case of schizophrenia and mood disorders, the major psychiatric disorders, there is much evidence, not without contradictions, of a possible association between schizophrenia and celiac disease. The association between mood disorders and gluten-related disorders, especially celiac disease, has only been studied for depression, often coupled with anxiety, and very recently for bipolar disorder. Since non-celiac gluten sensitivity is now known to be different from celiac disease, many studies have shown that gluten sensitivity is also associated with major psychiatric disorders. Here we review the literature on the association between schizophrenia/mood disorders and celiac disease/gluten sensitivity, pointing out the differences between these associations.

Assessment of a Test for the Screening and Diagnosis of Celiac Disease

Celiac disease (CD) is an immune‐mediated intolerance to dietary gluten, affecting genetically predisposed individuals. ELISA‐based serological tests help to decide if further duodenal biopsy is necessary, for this the diagnostic kits have to be accurate, specific, and sensible. In this study, we investigate the performance of an ELISA that uses the purified cross‐linked complex of tissue transglutaminase and gliadin, referred as the “neoepitope” (AESKULISA® tTG New Generation), as antigen.

Serum erythropoietin and blood lead concentrations

The aim of the study was to test the hypothesis that high blood lead levels are associated with depressed serum erythropoietin concentrations in workers occupationally exposed to lead. The results in exposed workers and in a control group of unexposed subjects were compared. Blood lead values were ≤20 μg/dl in unexposed subjects and ≥30 μg/dl in exposed subjects. The two groups of exposed workers and the control population were matched for sex and age. Hemoglobin levels were not affected by blood lead values and did not differ significantly between the three groups. The two-tailed, nonparametric Mann-Whitney U-test was used to compare unpaired groups. The Spearman rank correlation test was used to evaluate the dose-effect relationship between Pb and EPO. The analysis of the data indicate that erythropoietin values are significantly lower in exposed subjects than in controls. However no correlation was demonstrated between blood lead concentrations and erythropoietin in any group.

Pattern of Vitamin B12 and Folic Acid during Pregnancy

The aim of this study was to evaluate folate and vitamin B12 serum concentrations and their reciprocal relationships during pregnancy in relation to gestational age and levels of hemoglobin. Serum levels of vitamin B12 (B12) and folic acid (FA) were assessed by RIA in 213 women between the 6th and 43rd week of pregnancy. For 195 of these subjects, hemoglobin and hematocrit values were available. The logarithm (log) of B12 levels was found to be inversely correlated with weeks of pregnancy (R = -0.261; p < 0.001). A decrease in B12 levels occurred before the 27th week and was significant only in the subgroup of patients having hemoglobin levels above 11 g/dl. The log FA concentrations did not show any significant correlation with weeks of pregnancy irrespective of Hb levels. There was also a highly significant correlation between log B12 and log FA after 27 weeks and in the subgroups divided according to Hb levels (< or = and > 11 g/dl). It is therefore concluded that: (1) the demand for vitamin B12 is high in the first 27 weeks of pregnancy due to increasing maternal and embryo-fetal erythropoiesis and in order to sustain normal maternal Hb levels in the last weeks of pregnancy; (2) vitamin B12 might favor the absorption and utilization of FA after 27 weeks of pregnancy.

Autoantibody profiling of patients with primary biliary cirrhosis using a multiplexed line-blot assay.

OBJECTIVE To evaluate the autoantibody profile in patients with primary biliary cirrhosis (PBC) using a new multiplexed line-blot assay specifically designed for the diagnosis of autoimmune liver diseases. METHODS Sera of 58 consecutive PBC patients and 191 disease controls (144 with autoimmune liver diseases other than PBC, and 67 with non-autoimmune chronic liver diseases) were tested by both the multiplexed line-blot Autoimmune Liver Disease Profile 2 (ALD2) and by IIF on HEp-2 cells and on rat kidney/liver/stomach tissues. ALD2 contains the following PBC-associated antigens: AMA-M2, natively purified from bovine heart; M2-E3, a recombinant fusion protein including the E2 subunits of PDC, BCOADC and OGDC; sp100, PML and gp210 recombinant proteins. RESULTS With the ALD2 assay, a positive reaction to AMA-M2, M2-E3, sp100, PML and gp210 in PBC patients was observed in 77.6%, 84.5%, 34.5%, 15.1% and 18.9%, respectively, of the PBC sera. The overall sensitivity and specificity for PBC were 98.3% and 93.7%. Using IIF, positivity rates to AMA, and to antinuclear autoantibodies with membranous/rim-like and multiple nuclear dot patterns were 86.2%, 8.6% and 22.4%, respectively. The overall sensitivity and specificity for PBC of the IIF method were 86.2% and 97.9%, respectively. CONCLUSIONS The ALD2 line-blot showed a good diagnostic accuracy for PBC and a higher sensitivity than the IIF method to detect sp100 and gp210 autoantibodies.