Tullio Meloni

IDDM and Early Infant Feeding: Sardinian case-control study

OBJECTIVE To further investigate the association between the type of feeding in infancy and the development of IDDM. RESEARCH DESIGN AND METHODS We have carried out a case-control study in the area of Sassari (northern Sardinia, Italy), which is characterized by an ethnically homogenous population at high risk of IDDM. The study subjects comprised 100 IDDM patients and 100 control subjects, matched for sex and age and selected from children admitted at the Department of Pediatrics of the University of Sassari. Diabetic children (53 boys, 47 girls) had been diagnosed between 1983 and 1994, and their age at diagnosis ranged between 1 and 15 years. Information on feeding patterns during the 1st year of life was collected through questionnaires administered to the mothers. The questionnaire was designed to evaluate the duration of complete or partial breast-feeding and the age at which dietary products containing cows milk were introduced into the diet. RESULTS A larger proportion of the diabetic children rather than the control children had been breast-fed, and the risk of IDDM among children who had not been breast-fed was below unity (odds ratio [OR] 0.41; 95% CI 0.19–0.91). No clear difference was observed between diabetic and control subjects in the duration of breast-feeding (medians: 3 and 2 months, respectively), even if, overall, the data suggested a slight increase in the risk of IDDM with longer duration of breast-feeding (OR 1.10; 95% CI 0.99–1.22 per month). Although a larger proportion of control children rather than diabetic children had been given cows milk–derived formula and solid food before the age of 3 months, there was no time-risk relationship. CONCLUSIONS Our data do not support the existence of a protective effect of breast-feeding on the risk of IDDM, nor do the data indicate that early exposure to cows milk and dairy products has any influence on the development of IDDM in a high-risk population.

Membrane cross bonding in red cells in favic crisis: a missing link in the mechanism of extravascular haemolysis

Red cells of G6PD (D‐glucose‐6‐phosphate:NADP+ 1‐oxidoreductase; G6PD) deficient (Mediterranean variant) subjects were studied during a fava bean haemolytic crisis. Two representative cases are described. In Case 1, haemolysis was still going on. In more than 50% of the red cells the Hb was confined to one part of the cell, leaving the other part as transparent as a Hb‐free ghost. In this part the membranes appeared tightly bonded because swelling did not peel apart the bonded membrane areas. This feature is defined as membrane cross bonding (MCB). In Case 2. haemolysis had terminated and MCB‐cells were less than 1%. MCB was reproduced in vitro by incubating G6PD‐deficient whole blood with 1 mM divicine for up to 10 h. Subsequent shrinkage of red cells in hypertonic plasma (400 mOsm) resulted in the rapid formation of MCB. Membrane modifications by divicine, contained in fava beans, followed by osmotic shrinkage in the kidney and/or squeezing in the microcirculation are proposed as the cause of MCB during the favic crisis. MCB reduces the effective surface area of red cells. This is a plausible cause for sequestration by the reticulo‐endothelial system. Intravascular haemolysis observed in favic crisis cannot be explained by mechanical forces, but it is possible that the effective surface area is reduced by MCB to such an extent that red cells lyse osmotically.

Favism in a female newborn infant whose mother ingested fava beans before delivery

We describe a case of favism in a female newborn infant with glucose-6-phosphate dehydrogenase (G6PD) deficiency whose mother had ingested fava beans 5 days before delivery. At birth there were clinical and hematologic signs of hemolytic anemia, hemoglobinuria, and no blood group immunization. Study of the G6PD activity and 2-deoxy-glucose-6-phosphate utilization rate revealed that the infant and the mother were heterozygous for G6PD deficiency.

Marked decline of favism after neonatal glucose-6-phosphate dehydrogenase screening and health education: the Northern Sardinian experience

Favism is a potentially fatal manifestation of glucose-6-phosphate dehydrogenase (G6PD) deficiency, and it is therefore a public health problem in areas where this genetic abnormality is common. In the district of Sassari (northern Sardinia), the frequency of G6PD male hemizygotes is approximately 7.5%, and therefore all newborns since 1971 have been screened for G6PD deficiency. We have analyzed the incidence of favism in this community in two 10-year periods: (1) 1961-1970; and (2) 1981-1990. In period 1, there were 508 cases of favism, of which 76% occurred in boys. In period (2) there were 144 cases of favism, of which only 52% in boys. Thus, between the two periods there was an overall decrease in the incidence of favism of 75%, whereas the proportion of girls affected has approximately doubled. These data suggest that neonatal screening and health education programs can produce a substantial decrease in the number of cases of favism, and that the relative increase in favism in girls is possibly due to failure of the screening method used to detect all the heterozygotes for G6PD deficiency.

Mechanisms of perturbation of erythrocyte calcium homeostasis in favism.

Favism is an acute hemolytic anemia triggered by ingestion of fava beans in genetically susceptible subjects with severe deficiency of glucose-6-phosphate dehydrogenase (G6PD) activity. Erythrocytes from 10 favic patients had constantly and markedly increased calcium levels, as compared with values detected in 4 asymptomatic G6PD-deficient controls. Correspondingly, the calcium permeability of erythrocytes, estimated as the fraction of intracellular calcium exchangeable with externally added 45Ca2+, was invariably enhanced in favism and returned to normal patterns after several months from the acute hemolytic crisis. In favic patients, the levels of erythrocyte calcium ATPase activities showed wide variability, ranging from 2.0-12.9 mumol Pi/ml RBC/h, while control values in asymptomatic G6PD-deficient subjects were 10.62 +/- 2.03 mumol Pi/ml RBC/h. Analysis of the calcium ATPase in situ in erythrocyte membranes from favic patients showed the same molecular mass of 134 kD as observed in the control subjects. Exposure of G6PD-deficient erythrocytes in vitro to autoxidizing divicine, a pyrimidine aglycone strongly implicated in the pathogenesis of favism which leads to late accumulation of intracellular calcium, caused: (i) a marked inactivation of calcium ATPase, without changes in the molecular mass of 134 kD; and (ii) the concomitant loss of spectrin, band 3 and band 4.1, all known substrates of the calcium activated procalpain-calpain proteolytic system. Thus, the increased intraerythrocytic calcium apparently results in the degradation of calcium ATPase observed in some favic patients. It is proposed that both enhanced calcium permeability and a calcium-stimulated degradation of the calcium pump are the mechanisms responsible for the perturbation of erythrocyte calcium homeostasis in favism.

Phenobarbital for prevention of hyperbilirubinemia in glucose-6-phosphate dehydrogenase-deficient newborn infants.

Summary Prophylactic treatment with barbituric acid has proved effective in preventing severe hyperbilirubinemia in G-6-PD-deficient newborn infants. The changes in hemoglobin and hematocrit during the first days of life showed only moderate erythrocyte destruction in the patients studied.

Favism and Hemolytic Anemia in Glucose-6-Phosphate Dehydrogenase-Deficient Subjects in North Sardinia

The present paper reports the incidence from 1965 to 1979 of acute hemolytic anemia for a total of 948 cases in G-6-PD-deficient subjects due to the ingestion of fresh or dried fava beans or certain drugs and to viral infections. The highest percentage of hemolytic crises was due to fresh fava beans (94.4%). No cases of favism were observed in breast-fed babies whose mothers had eaten fava beans or from pollen inhalation. The male sex proved to be the hardest hit. Hemoglobin values were lower than or equal to 7 g/dl in about 75% of males and 50% of females. Total bilirubin values were lower than 103 mumol/l (6 mg/dl) in about 75% of males and 85% of females. Both the hemoglobin and bilirubin values were statistically significant. Mean transaminase values (SGPT) were significantly higher than those of normal controls. No correlation between favism and blood groups was found.

Favism: Erythrocyte Metabolism during Haemolysis and Reticulocytosis

Summary. The reduced activity of glucose‐6‐phosphate dehydrogenase (d‐glucose‐6‐phosphate; NADP+ i‐oxidoreductase; G6PD) in Mediterranean erythrocytes explains the precarious equilibrium of the hexose monophosphate pathway (HMP) and the susceptibility of these cells to haemolytic agents. G6PD‐deficient erythrocytes, in steady‐state conditions, have a low NADPH/NADP+ ratio, thus allowing the HMP to operate at its maximal intracellular rate and to compensate the intrinsic erythrocyte enzyme deficiency. Studies started soon after accidental intake of fava beans by sensitive G6PD‐deficient subjects demonstrate a decrease of both NADPH/NADP+ ratio and reduced glutathione. The metabolic effects induced by fava beans may be attributed to oxidative stress probably associated with an inhibitor effect of some unknown metabolite on the HMP. The availability of erythrocytes from subjects recovering from haemolysis with high reticulocyte counts and increased G6PD activity, provides new information on the rate of synthesis as well as on the in vivo decay of the mutant enzyme. Correlation of G6PD activity to reticulocyte count and extrapolation to an ideally homogenous population of reticulocytes reveal that the mutant enzyme is synthesized at a nearly normal rate. Furthermore, the present correlation allows an estimate of the in vivo half‐life of Mediterranean G6PD. The rate of decline of about 8 d observed in this study well correlates to the intracellular metabolic aspects of G6PD Mediterranean erythrocytes.

Neonatal hyperbilirubinaemia in heterozygous giucose‐6‐phosphate dehydrogenase deficient females

Summary. Glucose‐6‐phosphate dehydrogenase (G6PD, D‐glucose 6‐phosphate: NADP oxidoreductase, E.C. 1.1.1.49) activity and the percentage of G6PD deficient erythrocytes was determined in 50 girls heterozygous for G6PD deficiency, 25 of whom had had hyperbilirubinaemia at birth and 25 who had normal bilirubin levels. The enzymatic activity was 2 32 ±0·87 l.U./g Hb in the first group and 3·31 ±0·92 l.U./g Hb in the second group. The percentage of G6PD deficient erythrocytes was 54.1 ±15.3 and 65.3 ±0.87, respectively. The level of enzymatic activity exceeded 4 I.U./g Hb and the percentage of G6PD deficient cells fell below 40% in only one of the subjects who had developed hyperbilirubinaemia. Levels of enzymatic activity below 4 I.U./g Hb, or percentages of G6PD deficient erythrocytes higher than 40% can therefore be considered to be associated with a high risk of developing neonatal hyperbilirubinaemia. In our opinion, these babies should receive prophylactic treatment with phenobarbital, as do G6PD deficient Mediterranean males.

Glucose 6-phosphate dehydrogenase deficiency and cataract of patients in Northern Sardinia

We determined the activity level of glucose 6-phosphate dehydrogenase in 467 patients with cataract from northern Sardinia. Of 226 men, 18 (8%) had glucose 6-phosphate dehydrogenase deficiency. Of 241 women, 30 (12%) were heterozygous and two (1%) were homozygous for glucose 6-phosphate dehydrogenase deficiency. These prevalences were not significantly different from those expected in the general population. We concluded that patients with glucose 6-phosphate dehydrogenase deficiency do not have a higher risk of developing cataract.