A study of three polymorphic sites of the ADA gene in children with type 1 diabetes mellitus.

BACKGROUND: Adenosine deaminase is a polymorphic enzyme that has an important role in immune functions and in the regulation of intracellular and extracellular concentrations of adenosine and adenosine receptor activity. AIM: To search for possible association of type 1 diabetes mellitus (DM1) with three loci haplotypes (ADA1, ADA2, ADA6) of the adenosine deaminase gene. PATIENTS: One hundred and eighty-nine consecutive children with DM1 from Sassari, Sardinia, and a control sample of 239 children from the same area were studied. METHODS: ADA loci genotypes were determined by DNA analysis. RESULTS: Compared to controls, diabetic boys show a decrease of the 2(2)/6(1) haplotype while diabetic girls show an increase of the same haplotype. This association was replicated in an independent sample from Continental Italy. CONCLUSIONS: The 2(2)/6(1) haplotype may exert a protective action in males but may increase susceptibility to DM1 in females: OR = 0.398, 95% CI 0.16-0.96 for males, and OR = 2.31, 95% CI 1.32-4.06 for females.

Feto-maternal ACP1 activity ratio and intrauterine survival.

OBJECTIVE: Genetic differences in the activity of phosphotyrosine phosphatases between mother and embryo could result in a differential activation of signals induced by growth factors in the two sides of placenta. Previous observations suggest that this may have important effects on intrauterine development and survival. The aim of the present study is to confirm previous observations and show new data. STUDY DESIGN: We have studied 573 mother/newborn pairs, 169 wife/husband couples with repeated spontaneous abortion and 34 fertile wife/husband couples RESULTS: In mother/newborn pairs, the analysis of joint mother/infant ACP1 distribution has shown a deficit of pairs with the mother having low ACP1 S isoform concentration and the infant having high S isoform concentration, and an excess of pairs with the mother having high S isoform concentration and the infant having low S isoform concentration. In RSA couples there is an excess of couples in which the wife has low S isoform concentration and the husband has high S isoform concentration and a deficit of couples in which the wife has high S isoform concentration and the husband has low S isoform concentration. In fertile couples the pattern is reversed. CONCLUSION: The data suggest that when the mother to fetus S isoform concentration ratio is in favour of the mother, the probability of survival of the fetus is greater than in the opposite situation.

ACP1 and offspring sex ratio in smoking puerperae: a study at population level.

BACKGROUND: Data from previous study by our group suggest that in smoking women sex ratio of offspring is higher in newborns carrying ACP1C allele than in other ACP1 genotypes, suggesting that differences observed among human population concerning the effect of smoking may depend in part on this genetic factor. OBJECTIVES: In order to further explore this issue we have studied another population and have analysed the relationship between sex ratio and ACP1C gene frequency at population level. METHODS: The analysis includes 719 consecutive births from Central Italy considered in a previous paper and 5510 consecutive births from Sardinia. Data from English and Japanese populations have also been considered in the analysis. RESULTS: Among newborns not carrying ACP1C there is a decrease of SR among the offspring of smoking mothers, while among newborns carrying the ACP1C allele there is an increase of SR among the offspring of smoking mothers relative to non-smoking mothers. Considering Sardinian, Italian, English and Japanese population there is a linear positive relationship between C allele frequency and SR in smoking mothers. CONCLUSIONS: The present observation suggests an interaction between smoking and ACP1 regarding their effects on sex ratio, by which the presence of the ACP1C allele appears to counteract the effect of smoking. This suggests that genetic background may modify the effects of toxic environmental factors on gamete production and functionality and/or on intrauterine survival.

Risk of type 1 diabetes in childhood and maternal age at delivery, interaction with ACP1 and sex.

BACKGROUND: We have investigated the possible role of ACP1 (also known as cLMWPTP: cytosolic low molecular weight phosphotyrosine phosphatase), a highly polymorphic enzyme involved in signal transduction of T-cell receptor, insulin receptor and other growth factors in the relationship between maternal age at delivery and risk of type 1 diabetes in the offspring. METHODS: One hundred and eighty-nine consecutive children with type 1 diabetes (TIDM) diagnosed at the Department of Pediatrics of the University of Sassari (Sardinia) were studied. A control sample of 5460 consecutive newborns from the same population was also studied. RESULTS: Maternal age at birth of children with type 1 diabetes has shifted towards high values. There is also an effect of birth order on the susceptibility to type 1 diabetes, which is independent of that due to maternal age. The proportion of low activity ACPl genotypes is much higher among children born from older mothers than among diabetic children born from relatively young mothers. There is a significant effect of sex, maternal age, sex-ACPl two-way interaction and sex-ACP1-maternal age three-way interaction on the age at diagnosis of diabetes. CONCLUSIONS: The present data confirm the strong association between maternal age at delivery and risk of type 1 diabetes in the child. In addition, our analysis suggests a complex interaction among maternal age, sex of infant and ACP1 concerning age at diagnosis of diabetes. Thus, risk and clinical course of type 1 diabetes seem to be dependent on both maternal environment during intrauterine development and foetal genetic factors.

Seroprevalence of Helicobacter pylori in children with type 1 diabetes mellitus in Sardinia.

A few studies have reported an increased prevalence of Helicobacterpylori (HP) infection in diabetic subjects, which may be one of the causes of gastrointestinal symptoms and chronic atrophic gastritis frequently seen in diabetes of long duration. We determined the prevalence of HP infection in children and adolescents with Type 1 diabetes mellitus (T1DM) in the area of Sassari (northern Sardinia, Italy), which is characterized by an ethnically homogenous population at high risk of T1DM. HP IgG and IgA titres were measured in 138 patients with T1DM and 138 age-matched healthy controls. The percentage of infected subjects did not differ between T1DM patients (29.7%) and controls (32.6%). Globally, infected subjects were more than 1 yr older (13.0 +/- 2.7 yr) than non-infected ones (11.8 +/- 2.9 yr), independently of the presence of T1DM; in most HP-positive subjects infection was asymptomatic, and only 2 subjects in each group reported clinically relevant symptoms. HP-positive and HP-negative diabetic patients had the same duration of the disease (5.6 +/- 3.5 vs 5.5 +/- 3.6 yr) and received very similar doses of insulin (0.94 +/- 0.27 vs 0.96 +/- 0.4 IU/kg/d), whereas mean HbA1c was significantly lower in HP-positive patients (7.8 +/- 1.6% vs 8.6 +/- 1.7%,p=0.02). We conclude that the prevalence of HP infection is not higher in Sardinian children with T1DM as compared to controls of similar age, and the overall clinical impact of HP infection in terms of gastrointestinal symptoms and diabetic control seems to be low.

Genotypes of cytosolic low-molecular-weight protein-tyrosine-phosphatase correlate with age at onset of type 1 diabetes in a sex-specific manner.

We investigated the possible role of cytosolic low-molecular-weight protein-tyrosine-phosphatase (cLMWPTP or acid phosphatase locus 1 [ACP1]) in the mediation of age at onset of type 1 diabetes. ACP1 is an enzyme involved in signal transduction of T-cell receptors, insulin, and other growth factor receptors. We studied acid phosphatase polymorphism in 189 consecutive children with type 1 diabetes admitted to the Pediatric Clinic of Sassari University (Sardinia) and in 86 adolescent patients with recently diagnosed type 1 diabetes from continental Italy. In both populations, females with medium-high activity acid phosphatase genotypes had onset of disease significantly earlier than males. The data suggest that acid phosphatase genotype affects the age of onset and probably also the sex ratio in type 1 diabetes. Sex hormones might modulate the susceptibility to autoimmune diseases, including type 1 diabetes, through the influence of signal transduction pathways involved in immune functions. Elucidation of the molecular basis for gender differences in the course and severity of type 1 diabetes could have important implications for treatment as well, because there might be gender-specific effects in the response to immunotherapy.

Maternal-fetal interaction in the ABO system: a comparative analysis of healthy mothers and couples with recurrent spontaneous abortion suggests a protective effect of B incompatibility.

We investigated the possible differential effects of A and B blood group materno-fetal incompatibility on human fertility through a comparative analysis of couples with recurrent spontaneous abortion (RSA) and healthy mothers. ABO phenotype was determined in 5180 healthy mothers and their newborn babies from the population of Sassari (Sardinia) and in 1359 healthy puerperae (women who have just given birth) from the population of Rome. Mother-newborn joint ABO distribution in healthy mothers was compared with wife-husband joint ABO distribution in RSA couples. Distortions from expected distribution were evaluated by symmetry analysis. In both RSA couples and healthy mothers significant deviation from expected symmetry patterns were observed. Deviations in RSA are in the opposite direction to those observed in healthy puerperae. The most important difference observed concerned the symmetric joint phenotypes mother (women) A/infant (husband) B (B incompatible) and mother (women) B/infant (husband) A (A incompatible). A low number of B incompatible in RSA couples and a high number of B incompatible in healthy mothers was observed. The phenomenon is much more evident in women aged 24-28 years, a period of maximum fecundity. It is possible that the presence of anti-B immunoglobulin in the mother might have a protective effect against fetal loss in some cases of mother-infant ABO incompatibility.

Prevalence of silent celiac disease in patients with autoimmune thyroiditis from Northern Sardinia.

Celiac disease (CD) is frequently associated with other autoimmune diseases such as Type 1 diabetes mellitus, autoimmune thyroiditis (AT), and Addison's disease. The frequency of these associations varies with the populations studied. We conducted this study to ascertain the prevalence of CD in patients with AT from Sardinia, an area with a very high prevalence of CD. To this aim, 297 consecutive patients with AT (as defined by elevated antithyroid antibody levels and a positive ultrasound scan) were studied. Immunoglobulin A and G-class antigliadin antibodies were assayed in serum; if either or both were positive, antiendomysium antibodies were determined. If two markers were positive, serum ferritin, folate, and vitamin B12 levels were measured and jejunal biopsy was suggested. Thirteen out of the 14 patients who showed at least two positive markers consented to jejunal biopsy and all of them showed histological features of CD. The prevalence of CD in AT patients was 4-fold greater than that observed in the general population (4.37 vs 1.06%, p<0.0001). Ferritin was low in 6 and vitamin B12 in 2 out of 13 patients; serum folates were normal in all patients. Molecular typing of HLA class II alleles showed an increased frequency of the extended haplotype DRB1*0301/DQA1*0501/DQB1*0201. None of our patients had a history of gastrointestinal symptoms. We confirm the increased prevalence of silent CD in patients with AT. Patients with AT ought to be regarded as a high-risk group for CD and should be screened routinely for it; if negative, screening tests should be repeated at regular intervals.

High prevalence of lactose absorbers in Northern Sardinian patients with type 1 and type 2 diabetes mellitus.

BACKGROUND: Increased intestinal lactase activity has been shown to occur in alloxan and streptozotocin diabetic rats. OBJECTIVE: The objective of this study was to determine whether increased intestinal lactase activity is present in humans with diabetes mellitus. DESIGN: We assessed the capacity to digest lactose by measuring breath-hydrogen production after oral administration of lactose in 50 patients with type 1 diabetes, 50 patients with type 2 diabetes, and 50 healthy control subjects from Sassari, Sardinia, Italy, a population characterized by a low prevalence of lactase persistence (lactose absorbers). RESULTS: Fourteen percent of control subjects were lactose absorbers, compared with 48% of patients with type 1 diabetes and 52% of patients with type 2 diabetes (P < 0.005). The odds ratio of lactase persistence in patients with type 1 diabetes was 5.3 (95% CI: 2.0, 14.0) and in patients with type 2 diabetes was 5.5 (95% CI: 2.1, 14.5). CONCLUSIONS: Diabetes is associated with increased intestinal lactase activity in humans. Consequently, there is a greater exposure to glucose and galactose in diabetic patients with high lactose consumption. This may explain the association between diabetes and the risk of cataract.

Rh system and intrauterine growth. Interaction with season of birth.

Based on the hypothesis that maternal-fetal genetic differences in membrane transport and signal transduction may influence intrauterine development, the recent acquisition on transport function of Rh protein prompted us to study the relationship between joint maternal-fetal Rh phenotype and birth weight. Considering that metabolic effect of maternal-fetal competition could be amplified by environmental conditions, we have investigated possible seasonal effects on such relationship. We have studied 5291 infants born in Sardinia in the period January 1993--December 1996 and 984 infants born in Rome during 1996. In Rh(-) mothers there is a significant association between season of birth and birth weight that shows the highest mean value in infants born in autumn (i.e. conceived in winter). The association is much more evident in male than in female infants. In male infants from Rh(-) mothers, the association between birth weight and season is significant in Rh(+) male newborns only. Recent observations by our group in NIDDM suggest that glucose transport in RBC may be related to D protein, thus we propose an interpretation of the present observation in terms of transport function. When the density of D protein in the infant is greater than in the mother, the balance is in favour of the infant who may attain a significant developmental advantage when conceived in the cold season.

The prevalence of coeliac disease in infertility.

An increased incidence of reproductive problems, including infertility, miscarriage, low birth weight newborns, and shorter duration of breast-feeding, are known to exist in women with coeliac disease; some of these conditions are improved by a gluten-free diet. We have tried to ascertain the prevalence of coeliac disease in 99 couples who were being evaluated for infertility, compared with the known prevalence of silent disease in the population of Northern Sardinia, in which it is endemic. Of all women, four tested positive for at least two out of three markers: immunoglobulin A (IgA) antigliadin, immunoglobulin (IgG) antigliadin, and anti-endomysium antibodies, and underwent a jejunal biopsy; three had histological evidence of coeliac disease. One male partner was positive for two markers, and had a diagnostic jejunal biopsy. The prevalence of coeliac disease in infertile women seems higher (three out of 99, 3. 03%) in the study group than in the general population (17 out of 1607, 1.06%), and particularly in the subgroup with unexplained infertility (two out of 25, 8%, P < 0.03). Screening for coeliac disease should be part of the diagnostic work-up of infertile women, particularly when no apparent cause can be ascertained after standard evaluation.

Lactose absorption in patients with ovarian cancer.

To determine whether lactase persistence might be related to ovarian cancer risk, in 1994-1995 the authors assessed the capacity to digest lactose by measuring breath hydrogen production after oral administration of lactose in 50 women with ovarian cancer and 100 healthy controls. All of the women came from Sassari (Sardinia), Italy, an area where the population has a high frequency of lactose malabsorption. Thirty percent of cases were lactose absorbers, as compared with 15% of controls. The odds ratio for ovarian cancer among lactose absorbers was 2.51 (95% confidence interval 1.10-5.68). These results provide some support for a role of lactose ingestion and galactose cytotoxicity in the pathogenesis of ovarian cancer.

Bilirubin levels in the acute hemolytic crisis of G6PD deficiency are related to Gilbert's syndrome.

In this study we analyzed the effect of the (TA)7 polymorphism of the UGT1A gene associated with Gilbert's syndrome in G6PD-deficient subjects during an acute hemolytic crisis (fabic crisis). DNA from 44 subjects originating from the same geographic area in Sardinia was analyzed for the UGT1A promoter polymorphism. The increase of unconjugated bilirubin level during fabic crisis and its relationship with UGT1A polymorphism was evaluated. The UGT1A (TA)7 TATA box variant was found in 9/44 (21%) of the G6PD deficient subjects examined. The median value for unit of increase of bilirubin (mg/dl)/unit of decrease of hemoglobin (g/dl) was higher in variant homozygous than in heterozygous and normal subjects. These findings imply a contribution of the UGT1A polymorphism associated to Gilbert's syndrome to development of the hyperbilirubinemia in G6PD deficient subjects during acute hemolytic anemia.

Gilbert's syndrome and jaundice in glucose-6-phosphate dehydrogenase deficient neonates.

BACKGROUND AND OBJECTIVE: The pathogenesis of the hyperbilirubinemia present in approximately 30% of neonates affected by glucose-6-phosphate dehydrogenase deficiency is an unsolved problem. We evaluated the effect of Gilbert's syndrome, the most common defect of bilirubin conjugation, on the hyperbilirubinemia of these neonates. DESIGN AND METHODS: One hundred and two neonates affected by glucose-6-phosphate dehydrogenase deficiency were enrolled in this study: 56 had hyperbilirubinemia and 46 had normal bilirubin levels. The analysis of the A(TA)nTAA motif in the promoter region of the UGT1A gene was performed by means of PCR, followed by separation on 6% denaturing polycrylamide gel. RESULTS: The frequency of the three different genotypes of the A(TA)nTAA motif was similar in the study and control groups. Our results demonstrated no difference in the percentage of homozygotes for the UGT1A (TA)7 variant associated with Gilbert's syndrome. INTERPRETATION AND CONCLUSIONS: These findings indicate that Gilbert's syndrome does not account for the hyperbilirubinemia occurring in some neonates with glucose-6-phosphate dehydrogenase deficiency. Furthermore our results suggest that hemolysis is not the major event in the pathogenesis of hyperbilirubinemia in these patients.

Primary lactase deficiency and past malarial endemicity in Sardinia.

BACKGROUND: It has recently been suggested that primary lactase deficiency might have been selected for by malaria, as occurred for beta-thalassaemia and glucose 6-phosphate dehydrogenase deficiency. However, recently we have found that the prevalence of primary lactase deficiency in the area of Sassari (Northern Sardinia), where, in the past, there was intermediate malarial endemicity, is comparable to that observed in the adult population from other areas of Southern Italy where malaria was less endemic. AIMS: To address the problem further, we have determined the prevalence of primary lactase deficiency, glucose 6-phosphate dehydrogenase deficiency deficiency and beta-thalassaemia trait in the populations of three Sardinian villages which differ in altitude above sea-level, socioeconomic features, history of endemic malaria and prevalence of b-thalassaemia and glucose 6-phosphate dehydrogenase deficiency. SUBJECTS: We tested 138 adult males: 53 were from Fonni (a non-malarial mountain village, with a strong pastoral tradition), 38 from Lodé (a village with a similar pastoral tradition, but high malarial endemicity in the past) and 47 from Terralba (a lowland fishing village with an agricultural tradition and heavy malarial morbidity and mortality). METHODS: A blood sample was obtained in all subjects for determination of HbA2 and glucose 6-phosphate dehydrogenase activity. Lactase deficiency was assessed by measuring breath hydrogen production after oral administration of lactose (50 g), by gas-chromatography. RESULTS: The frequencies of glucose 6-phosphate dehydrogenase deficiency and of beta-thalassaemia trait in the non-malarial village of Fonni were strikingly low, compared to frequencies found in the two villages (Terralba and Lodé) with a very high past malarial morbidity. In contrast, there was no significant difference in the prevalence of lactase deficiency in the three groups of subjects from the three villages. CONCLUSIONS: These data obtained in Northern Sardinia do not support the hypothesis of a selection of primary lactase deficiency by malaria. For definitive conclusions, however, the malaria hypothesis should be tested in other parts of the world.

Lactose absorption in patients with glucose 6-phosphate dehydrogenase deficiency with and without favism.

BACKGROUND: It has recently been suggested that primary lactase deficiency might have been selected for by malaria, as has been previously shown to occur for thalasaemia and glucose 6-phosphate dehydrogenase (G6PD) deficiency. AIMS: To test this hypothesis, the prevalence of primary lactase deficiency in G6PD deficient subjects and in controls from the area of Sassari (Northern Sardinia) was determined, which in the past was characterised by an intermediate malarial endemicity. SUBJECTS: 70 adult subjects with G6PD deficiency, 34 of whom had a past history of favism, and 50 age matched control subjects. METHODS: The capacity to absorb lactose was assessed by measuring breath hydrogen production after oral administration of lactose (50 g) by a gas chromatographic method. RESULTS: Twenty per cent of G6PD deficient subjects with a positive history of favism and 22% of G6PD deficient subjects without a positive history of favism were lactose absorbers compared with 14% lactose absorbers in the control group. The differences were not statistically significant. CONCLUSIONS: These data show that the prevalence of primary lactase deficiency in the area of Sassari is relatively high, but comparable to that seen in the adult population from another area of southern Italy (Naples) where malaria was less endemic.

Active involvement of catalase during hemolytic crises of favism.

The endemic occurrence of favism in certain Mediterranean regions provided an investigative opportunity for testing in vivo the validity of claims as to the role of catalase in protecting human erythrocytes against peroxidative injury. Reduced activity of catalase was found in the erythrocytes of six boys who were deficient in erythrocytic glucose-6-phosphate dehydrogenase (G6PD) and who were studied while suffering hemolysis after ingesting fava beans. Activity of catalase was further reduced when their red blood cells were incubated with aminotriazole. In contrast, minimal reduction of catalase activity was found, both with and without incubation with aminotriazole, in erythrocytes of a G6PD-deficient boy who had ingested fava beans 7 days earlier and in erythrocytes of seven G6PD-deficient men with a past history of favism. These results confirmed earlier studies in vitro indicating that catalase is a major disposer of hydrogen peroxide in human erythrocytes and, like the glutathione peroxidase/reductase pathway, is dependent on the availability of reduced nicotinamide adenine dinucleotide phosphate (NADPH). The effect of divicine on purified catalase and on the catalase of intact G6PD-deficient erythrocytes was similar to the previously demonstrated effect on catalase of a known system for generating hydrogen peroxide. This effect of divicine strengthens earlier arguments that divicine is the toxic peroxidative component of fava beans.

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.