Gino Morisi

Toxicity of wheat flour proteins and protein-derived peptides for in vitro developing intestine from rat fetus.

Summary: A peptic-tryptic-cotazym (PTC) digest of a crude wheat gliadin preparation was obtained under experimental conditions simulating in vivo protein digestion and then fractionated into 10 peaks by ion-exchange chromatography. PTC-gliadin digest and one of its subfractions (coded as fraction 9 according to its elution pattern) were very active in inhibiting in vitro development and morphogenesis of small intestine from 17− and 18-day-old rat fetuses, whereas they were harmless for the culture of jejunum from 21-day-old fetuses. PTC-digest also induced extensive tissue degeneration and necrosis of in vitro cultured small intestinal mucosa from patients with active celiac disease (gluten-induced entheropathy), but did not cause any detectable effect on histolog-ically normal human small intestinal mucosa. Some wheat albumin and gliadin fractions were also tested on in vitro developing small intestine from 17-day-old rat fetus. Among all the tested protein fractions, only one gliadin fraction (coded as α10-gliadin from its gel electrophoretic mobility) exhibited a toxic effect; morphologic alterations induced by α10-gliadin were similar to those induced by PTC-digest and fraction 9.Speculation: Peptides obtained from wheat gliadins under in vitro conditions simulating protein digestion by humans as well as whole wheat gliadin are very active in damaging in vitro developing jejunum from 17− and 18-day-old rat fetuses, but have no toxic effect on the culture of jejunum from 21-day-old rat fetus. The transient character of such an effect is suggestive of a protective mechanism against the toxic protein components from wheat, associated with the age-dependent maturation of rat intestinal mucosa. It is speculated that also human normal intestine develops with maturation similar specific detoxifying mechanisms and that wheat gliadin and gliadin-derived peptides might exert a toxic action when human intestinal mucosa is not yet fully mature (i.e., during early extrauterine life) or when adult human mucosa regresses towards immature steps of development (i.e., in intestinal diseases with mucosal atrophy).

Free amino acids in fish brain: normal levels and changes upon exposure to high ammonia concentrations in vivo, and upon incubation of brain slices.

1. 1. When fish (Carassius auratus) were kept in water containing controlled concentrations of ammonia, the cerebral concentration of glutamine increased up to ten times, that of several other amino acids increased to a lesser extent. 2. 2. The changes observed were much more pronounced in brain than in other organs. 3. 3. Fish brain slices incubated aerobically in a medium containing glucose or glutamate with or without NH4C1 showed an increase in the level of most amino acids. Glutamine and histidine increased the most, in absolute terms. 4. 4. The amino acid changes of incubated rat brain slices differed, in several aspects, from those observed in fish.

Serum triglycerides in the prediction of coronary artery disease (an Italian experience)

An occupational group living in Rome and composed of 3,007 men aged 46 to 65 years who were free from previous major coronary events was screened for a number of coronary risk factors and then followed up for 10 years. In all, 107 coronary deaths occurred in 10 years. There was a positive relation between coronary death rate and increasing levels of triglycerides, but the difference between the extreme quintile classes was not any more significant after adjustment for cholesterol levels. A cross-classification involving low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol and triglyceride levels showed a higher coronary artery disease (CAD) mortality in subjects with a higher LDL cholesterol/HDL cholesterol ratio and higher triglycerides. However, the excess risk in this subgroup was largely explained by the mean levels of total cholesterol. The multivariate proportional-hazards Cox model with coronary deaths as the end point, and age and triglycerides as predictors, produced a significant coefficient for triglycerides that became nonsignificant when other lipids alone or in combination (total, HDL and LDL cholesterol and some of their ratio) were fed into the model as further covariates. It is concluded that there is no evidence of an independent role of triglycerides in the prediction of coronary deaths.

Determination of ascorbic acid in blood plasma or serum and in seminal plasma using a simplified sample preparation and high-performance liquid chromatography coupled with UV detection

Abstract A simplified method of sample preparation and high-performance liquid chromatography procedure using UV detection is described for the determination of ascorbic acid (AA) in blood plasma or serum and seminal plasma. Within two hours from collection samples are treated with dithioerythritol (DTE) and then stored under Argon at -80°C. Prior to analysis, protein precipitation is initiated with the addition of cold methanol. AA elution is carried out on a C18 reverse phase column using dodecyltrimethylammonium bromide as an ion-pairing agent. the detection is accomplished by measuring ultraviolet absorption at 265 nm. the analysis time for sample is 10.5 min, the retention time of AA being 9.7 min. Within- and between-day coefficients of variation are 2.9% and 4.9% for blood serum, 1.0 % and 2.3 % for seminal plasma. Mean analytical recovery of 102.5 ± 3.7% was found analyzing a serum pool after addition of a standard amount of AA. AA levels are stable for at least 43 days under the described storage...

Association of selected social, environmental and constitutional factors to blood lead levels in men aged 55–75 years

Blood lead (B-Pb) levels were determined in 1802 out of 1856 non-occupationally exposed men aged 55-75 years living in the Rome area who participated, between 1989 and 1990, in an epidemiological survey for coronary heart disease (New Risk Factors Project). The median B-Pb level was 113 micrograms/l (10th-90th centiles: 74-180 micrograms/l) and only 0.7 per cent (n = 14) of the subjects had B-Pb values higher than 300 micrograms/l. B-Pb levels were significantly and positively associated to alcohol consumption. Moderate and heavy drinkers had median B-Pb level of 143 micrograms/l (10th-90th centiles: 92-233) and 165 micrograms/l (10th-90th centiles: 102-285) respectively, whereas non-drinkers had a median B-Pb level of 96 micrograms/l (10th-90th centiles: 66-143). The influence of smoking habits was less relevant. Subjects who never smoked and subjects smoking more than 20 cigarettes daily had median B-Pb levels of 103 and 133 micrograms/l, respectively. Individuals classified as habitual car-drivers had slightly higher Pb levels than non-drivers. Subjects classified as manual workers had higher B-Pb levels in comparison with non-manual workers and retired subjects. B-Pb levels were directly related to HDL-cholesterol (HDL-C, r = 0.2252) and gamma-glutamyltransferase (gamma-GT, r = 0.2207) serum levels. The alleged alcohol consumption was more related to B-Pb level (r = 0.3848) than to serum level of HDL-C (r = 0.2474) or gamma-GT (r = 0.2469). A significant correlation (r = 0.2409) also existed between B-Pb and blood cadmium levels (B-Cd). Subjects with a low Gaensler ratio, an index of respiratory function, had higher B-Pb levels. In multiple regression analyses alcohol intake was the most important predictor of B-Pb level, explaining more (14.27%) of the total variance than did B-Cd (4.98%), HDL-C (1.89%), driving habits (1.46%), gamma-GT (1.09%), skinfold thickness (0.96%), and Gaensler index (0.38%). The risk ratio of having B-Pb level higher than 180 micrograms/l (90th centile of B-Pb distribution in our subjects) was 5.3 (95% CI: 2.7-10.4) for drinkers versus non-drinkers and 1.9 (95% CI: 1.2-3.1) for current smokers versus subjects who had never smoked. B-Pb was, at least in our subjects, a more specific and sensitive objective index of alcohol consumption than gamma-GT and HDL-C.(ABSTRACT TRUNCATED AT 400 WORDS)

Gamma-aminobutyric acid (GABA) uptake by the developing mouse brain in vivo

Abstract (1) Cerebral uptake of GABA was studied following the subcutaneous administration of different doses of the amino acid to mice at various prenatal and postnatal stages. (2) The cerebral level of GABA could be significantly increased at all ages studied, but the doses injected in order to produce a measurable net uptake needed to be higher in more mature animals. High doses of GABA were toxic for immature mice. (3) In pregnant females injected with doses lethal for newborn mice, GABA was taken up by the placenta, reached the embryonic blood and was taken up by the embryonic brain to a greater extent than at any postnatal stage studied. GABA concentration decreased more rapidly in maternal plasma than in the placenta or fetal plasma. (4) The rate of excretion and metabolism of GABA was greater in mature mice, where plasma levels decreased much more rapidly than in immature animals. (5) Large doses of γ-amino-β-hydroxybutyric acid inhibited the metabolic degradation and the rate of excretion of exogenous GABA. Moreover, γ-amino-β-hydroxybutyric acid caused a statistically significant inhibition of GABA uptake by brain. (6) The concept of the blood-brain barrier to GABA and of its evolution during development is discussed. It is concluded that decreased permeability of the blood-brain interface to GABA is at best one of the factors contributing to the reduced elevation of cerebral GABA levels observed in mature animals after parenteral administration. Other important factors are increased rates of metabolic breakdown, of excretion, and possibly of active transport from the brain.

The performance of Italian laboratories in the determination of trace elements in blood

Abstract The analytical performance of a group of Italian laboratories with regard to the determination of blood lead (BPb), blood cadmium (BCd), serum aluminum (SAl), serum copper (SCu), and serum zinc (SZn) was estimated from the results of quality assurance programs carried out in Italy between 1983 and 1990. Mean imprecision, mean inaccuracy, and percentage of acceptable results provided, according to preset limits, were adopted as indexes of performance. Positive trends were observed, over the course of the years, toward smaller bias and lower percentage of laboratories with poor performance. At present, taking into account results from all participants, mean absolute inaccuracy can be evaluated as 40 μg/liter (mean concentration: 440 μg/liter) for BPb, 0.55 μg/liter (mean concentration: 3.6 μg/liter) for BCd, and 13.4 μg/liter (mean concentration: 77 μg/liter) for SAl. At mean concentrations of 0.94 mg/liter of SCu and 1.08 mg/liter of SZn, mean absolute inaccuracies were 0.130 mg/liter and 0.162 mg/liter, respectively. The percentage of laboratories providing results within established limits for at least 80% of the examined samples is rather high for BCd (61.9%), SCu (43.8%), and BPb (41.2%). On the contrary, only 15.7% of laboratories achieved good performance in SZn analysis and none in SAl determination.

Organization of interlaboratory quality assurance programs for the analysis of trace elements in blood: Evaluation of procedures and analysis of data☆

Abstract Some of the problems arising from the organization of quality assurance programs for the analysis of trace elements in blood are discussed. These include the preparation of suitable control materials by the organizing center and the elaboration of valuable strategies of sample distribution, treatment of data, and evaluation of results, which can be applied even to a small number of participants. The procedures used in the Italian quality assurance scheme for trace elements in blood are reported. Their reliability was partly confirmed by the results of some experiments carried out within the framework of the program. They dealt with the commutability of control and fresh samples in blood lead analysis, the analytical behavior of liquid compared to that of lyophilized control materials for serum aluminum determination, and the evaluation of laboratory performance in the analysis of “open” or “blind” duplicates.

Lead levels in whole blood of an adult population group from Rome

Pb-blood levels of 801 adult non-occupationally exposed subjects from Rome are reported. The investigation was carried out according to EEC Directive No. 77/312 with acceptable quality control of analytical data. A mean Pb-blood level of 173 micrograms/l (198 micrograms/l for males and 150 micrograms/l for females) was found; good correlations were found between Pb-blood levels and age, sex and smoking habits but no correlation was found in relation to drinking habits, residence and other variables which were examined. In the present survey all the three EEC reference levels were observed.

Blood lead and cadmium determination: Results of the Italian external quality assessment scheme

The procedures adopted in the Italian external quality assessment scheme (EQAS) for blood Pb (B-Pb) and Cd (B-Cd) determination, including the preparation of control materials, are described. Each scheme involves the use of internal quality control materials and the participation in periodical external quality assessment trials. All control materials are prepared at the Laboratory of Clinical Biochemistry, Italian National Institute of Health (Istituto Superiore di Sanità, ISS), as the reference centre. Computerized procedures have been adopted whenever possible, i.e. sample randomization, data transfer and treatment. The analytical performance of the participating laboratories, evaluated from the results obtained in ten years of activity, is reported. A total of eight phases has been carried out between 1983 and 1993. Over the course of these ten years, a positive trend was observed for B-Pb towards a smaller bias and a lower percentage of laboratories with poor performance. In the eighth phase, taking into account the results from all participants, the mean absolute inaccuracy for B-Pb was 32 μg/l (mean Pb concentration in control samples, 165 μg/l). The percentage of laboratories providing results within established acceptability limits for at least 80% of the examined samples (good performers) increased with time from 28% in the first phase to 48% in the last phase. The laboratories providing less than 50% of acceptable results (poor performers) decreased from 35% in the first phase to 22% in the last phase. As regards B-Cd, mean absolute inaccuracy decreased from 0.96 μg/l to 0.55 μg/l between the first phase and the sixth phase. The percentage of good performers increased from 45% to 62% and that of poor performers decreased from 31% to 5%. From the seventh phase a large group of new laboratories agreed to participate in the EQAS for B-Cd and the inaccuracy rose to 0.75 μg/l (mean concentration: 2.7 μg/l) and the percentage of poor performers rose to 24%, whereas the percentage of good performers remained almost unchanged.