New High-performance Liquid Chromatography-DAD Method for Analytical Determination of Arbutin and Hydroquinone in Rat Plasma.

Natural substances present in herbal preparations should be carefully used because they can give toxic or therapeutic effects despite of their amount or the way of administration. The safety of products of vegetable origin must be assessed before commercialisation by monitoring the active ingredients and their metabolites. This study was therefore designed to identify and quantify arbutin and its metabolite hydroquinone, naturally present in Arctostaphylos uva-ursi (L.) Spreng plant in rat plasma, after an acute and subacute administration of aqueous arbutin solution in Wistar rats. For this purpose a reversed-phase high-performance liquid chromatography coupled with photodiode array detection was developed to assess the pharmacokinetic of arbutin and hydroquinone in plasma of female rats treated with aqueous arbutin solutions. The detection (arbutin: 0.0617 µg/ml and hydroquinone 0.0120 µg/ml) and quantification (arbutin: 0.2060 µg/ml and hydroquinone: 0.0400 µg/ml) limits were determined. At the arbutin concentration level of 10.7 µg/ml repeatability was 13.33% and its recovery 93.4±6.93%, while at the hydroquinone concentration level of 10.6 µg/ml repeatability was 11.66% and its recovery 92.9±7.75%. Furthermore the method was fully validated and the obtained data indicate that the new method provides good performances.

Diffusing capacity for carbon monoxide in children with type 1 diabetes.

AIMS/HYPOTHESIS: Few data are available on lung dysfunction in children with diabetes. We studied the association of pulmonary function variables (flows, volumes and alveolar capillary diffusion) with disease-related variables in children with type 1 diabetes mellitus. METHODS: We studied 39 children with type 1 diabetes (mean age 10.9+/-2.6 years, disease duration 3.6+/-2.4 years, insulin.kg(-1).day(-1) 0.77+/-0.31) and 30 healthy control children (mean age 10.4+/-3.0 years). Pulmonary function tests included spirometry, N(2) wash-out and the single-breath diffusing capacity for carbon monoxide (DL(CO)) corrected for the alveolar volume (DL(CO)/V(A)). Glycaemic control was assessed on the basis of HbA(1)c, with HbA(1)c values of 8% or less considered to indicate good glycaemic control, and HbA(1)c values of 8% or more considered to indicate poor control. RESULTS: Children with poor glycaemic control had comparable percentage values for predicted flows and volumes but lower DL(CO)/V(A) values than children with good glycaemic control and healthy control children (86.7+/-12.6 vs 99.8+/-18.4 and 102.0+/-15.7; p<0.05). The predicted DL(CO)/V(A) percentages correlated with HbA(1)c levels (r=-0.39, p=0.013). A multiple regression analysis (stepwise model) controlling for HbA(1)c levels and other disease-related variables (age of disease onset, disease duration, daily insulin dose/kg, sex) identified HbA(1)c levels as the sole predictor of DL(CO)/V(A) in percent. CONCLUSIONS/INTERPRETATION: In children with type 1 diabetes, the diffusing capacity diminishes early in childhood and is associated with poor metabolic control. Although low DL(CO)/V(A) levels in these children probably reflect pulmonary microangiopathy induced by type 1 diabetes, other factors presumably influencing CO diffusion capacity measurements (e.g. a left shift in HbA(1)c resulting in high O(2) binding and low CO binding) could explain the apparent capillary and alveolar basal membrane dysfunction.

Genetic prediction of type 1 diabetes in a population with low frequency of HLA risk genotypes and low incidence of the disease (the DIABFIN study).

BACKGROUND: To develop a sensitive, specific screening strategy for predicting genetic risk for type 1 diabetes mellitus (T1DM) in the low-incidence continental Italian population, and to define with this tool, a cohort of high-to-moderate risk infants for an immunological follow-up study aimed at identifying environmental risk factors for T1DM. METHODS: 4855 newborns in three regions of continental Italy were screened for T1DM HLA-DRB1-DQB1 risk genotypes using a reverse line blot typing method. Risk classification was based on odds ratios (OR) found in a preliminary case-control study (356 T1DM patients, 412 controls). Screening efficiency was optimized by allele subtyping. RESULTS: Screening for well-known T1DM susceptibility genotypes [DRB1*03/*04-DQB1*0302; DRB1*03/*03; DRB1*04/*04-DQB1*0302; DRB1*04-DQB1*0302/X where X is not equal to DRB1*03, DRB1*04-DQB1*0302, DQB1*0602 or DQB1*0603] was associated with <60% sensitivity due to their low frequencies in the general Italian population. Inclusion of an additional genotype from which protective DRB1 and DQB1 alleles had been excluded [DRB1*03/X degrees where DQB1 is not equal to *0301, *0503, *0602, or *0603 and X degrees not equal DRB1*03, DRB1*04-DQB1*0302 or DRB1*07] increased screening sensitivity to 75% (specificity: 85%). Among 4855 newborns, we have found the high-risk genotype [DRB1*03/*04-DQB1*0302; estimated absolute risk (AR) 1/23] to be present in only 0.9%. The moderate-risk genotypes were found in 13.8% of newborns (estimated AR 1/177). CONCLUSIONS: Risk classification must be tailored to the characteristics of the individual population, in particular, the allelic frequencies in the background population and T1DM prevalence. We have developed a screening strategy with good levels of sensitivity that should prove effective for use throughout the Italian peninsula.

Involvement of oxygen radicals in cytarabine-induced apoptosis in human polymorphonuclear cells.

We investigated apoptosis in polymorphonuclear neutrophils (PMNs) induced by cytarabine (Ara-C). This drug increased apoptosis by 100% with respect to the controls after 3 hr of incubation. This increase was inhibited by N-acetyl-L-cysteine (NAC) or diphenyleneiodonium chloride (DPI). Ara-C alone caused an early increase (after a 30-min incubation) in intracellular oxidant generation (inhibitable by rotenone, fumonisin b1, and DPI) and in protein tyrosine phosphorylations (inhibitable by NAC). The drug also affected the observed reduction of dimethylthiazol diphenyltetrazolium bromide (MTT). No extracellular release of reactive oxygen species (ROS) was elicited by the addition of Ara-C, while the drug increased the release of ROS by N-formyl-leucyl-phenylalanine-(f-MLP) but not phorbol 12-myristate 13-acetate-stimulated PMNs. This phenomenon was abolished by the addition of genistein, whereas such an effect was not observed following the addition of 1-(5-isoquinolynilsulfonyl)-2-methylpiperazine (H7). Ara-C induced ROS release from PMNs in the presence of subthreshold concentrations of f-MLP (priming effect). These results indicate that intracellular ROS production from mitochondria promotes Ara-C-induced apoptosis. Ara-C primes plasma membranes by a mechanism involving protein tyrosine phosphorylations and may also contribute to ROS generation from the granules.

Peritoneal dialysis in an infant with type 1 diabetes and hyperosmolar coma.

Hyperosmolar coma which is characterized by severe hyperglycemia in absence of chetosis is very rare in pediatric age with only 11 cases reported in the literature. The outcome of the condition is usually poor with mental retardation being the most common event. Here a case of hyperosmolar coma is described in a female of three months of age who was treated with peritoneal dialysis 11 hours after admittance to hospital. This female patient has been receiving insulin from three months of age and today at the age of 10 years she leads a normal life despite being on insulin therapy. A very low level of C-peptide (<0.3 ng/ml) clearly confirms she is affected by Type 1 diabetes. To our knowledge this is the first case report of hyperosmolar coma in a neonate with Type 1 diabetes who survived this condition without late neurological consequences.

No effect of oral insulin on residual beta-cell function in recent-onset type I diabetes (the IMDIAB VII). IMDIAB Group.

AIMS/HYPOTHESIS: Induction of tolerance to insulin is achievable in animal models of Type I (insulin-dependent) Diabetes mellitus by oral treatment with this hormone, which can lead to prevention of the disease. In the Diabetes Prevention Trial of Type I diabetes (DPT-1), oral insulin is given with the aim of preventing disease insurgence. We investigated whether if given at diagnosis of Type I diabetes in humans, oral insulin can still act as a tolerogen and therefore preserve residual beta-cell function, which is known to be substantial at diagnosis. METHODS: A double-blind trial was carried out in patients (mean age +/- SD: 14 +/- 8 years) with recent-onset Type I diabetes to whom oral insulin (5 mg daily) or placebo was given for 12 months in addition to intensive subcutaneous insulin therapy. A total of 82 patients with clinical Type I diabetes ( < 4 weeks duration) were studied. Basal C peptide and glycated haemoglobin were measured and the insulin requirement monitored every 3 months up to 1 year. Insulin antibodies were also measured in 27 patients treated with oral insulin and in 18 patients receiving placebo at the beginning of the trial and after 3, 6 and 12 months of treatment. RESULTS: The trial was completed by 80 patients. Overall and without distinction between age at diagnosis, at 3, 6, 9 and 12 months baseline mean C-peptide secretion in patients treated with oral insulin did not differ from that of those patients treated with placebo. In patients younger than 15 years a tendency for lower C-peptide values at 9 and 12 months was observed in the oral insulin group. Insulin requirement at 1 year was similar between the two groups as well as the percentage of glycated haemoglobin. Finally, IgG insulin antibodies were similar in the two groups at each time point. CONCLUSION/INTERPRETATION: The results of this study indicate that the addition of 5 mg of oral insulin does not modify the course of the disease in the first year after diagnosis and probably does not statistically affect the humoral immune response against insulin.

Sleep apnoea in children with diabetes mellitus: effect of glycaemic control.

AIMS/HYPOTHESIS: Patients with diabetes mellitus commonly have cardiovascular autonomic dysfunction and an abnormal ventilatory pattern during sleep. Few data are available on these changes in childhood diabetes. We investigated whether young diabetic children with or without diabetic neuropathy have ventilatory dysfunction during sleep and if so, whether these autonomic changes are related to the duration of diabetes and glycaemic control. METHODS: We studied 25 children with insulin-dependent diabetes mellitus (19 boys, mean age 7.72 +/- 1.99 years). All patients were insulin-dependent at diagnosis; blood samples for HbA1c assay were collected on the morning before testing and at 3-month intervals during the preceding year. Patients and control subjects (20 age-matched healthy children, 15 boys) underwent overnight polysomnography. RESULTS: More diabetic patients than control subjects had sleep apnoeas (p = 0.006); apnoeas in patients also lasted longer (p = 0.07). Patients with poorly controlled diabetes had more apnoeas than patients with well-controlled diabetes and than healthy control subjects (p < 0.0001). Respiratory events during sleep correlated significantly with glycaemic control (r = 0.360; p = 0.09) and with the duration of diabetes (r = 0.430; p = 0.04). CONCLUSION/INTERPRETATION: We conclude that respiratory control is compromised very early in children with diabetes. These anomalies are closely related to the duration of diabetes and to glycaemic control. In young children with diabetes, screening of ventilatory control using recording techniques that are simpler than polysomnography could provide an early indication that an adverse cardiopulmonary reaction has begun.

Autoantibody negative new onset type 1 diabetic patients lacking high risk HLA alleles in a caucasian population: are these type 1b diabetes cases?

BACKGROUND: In Caucasians, a small number of Type 1 diabetic patients do not show evidence of humoral islet autoimmunity at disease onset, at least with common screening procedures. In African- and Hispanic-American diabetic children at time of diagnosis, many show no evidence of autoimmunity but have an atypical clinical form of the disease. According to the recent American Diabetes Association classification, this subgroup of autoantibody negative patients is referred to as Type 1b diabetic subjects. In the present study, a homogeneous Caucasian Type 1 diabetic clinic-based cohort has been evaluated at diagnosis using a large panel of diabetes-related antibodies and then characterized for various genetic features in order to identify newly diagnosed Type 1 diabetics who are potentially autoantibody negative, i.e. possibly referrable to as idiopathic Type 1b diabetes. METHODS: Newly diagnosed Type 1 diabetic patients of Italian origin (n=141, mean age 12.0+/-7.6 years) were tested for anti-islet cell, anti-insulin, anti-65 kDa isoform of glutamic acid decarboxylase and anti-amino acid residues 256-979 of the tyrosine-phosphatase IA-2 molecule autoantibodies (Step 1). Only those patients found to be autoantibody negative were tested for anti-disialo-ganglioside GD3, anti-thyroid peroxidase, anti-thyroglobulin, anti-21-OH hydroxylase, anti-gastric parietal cell and anti-transglutaminase antibodies (Step 2). Sera negative for the presence of these six autoantibodies as well were characterized in terms of HLA DRB1, DQB1 and CTLA-4. RESULTS: Six out of 141 subjects (3.5%) were autoantibody negative in the first step of the study and five out of six in the second. These five autoantibody negative patients underwent genetic analysis. Three of them had at least one Type 1 diabetes-related high risk HLA haplotype (3/141, 2.1%) while the remaining two cases showed neutral (DR5-DQB1*0301/DR5-DQB1*0301) or strongly protective (DR2-DQB1*0602/DR2-DQB1*0602) HLA genotypes, respectively (2/141, 1. 4%). CONCLUSIONS: Clinically defined Type 1 diabetic patients with no sign of autoimmunity do exist in a Caucasian population. These patients (2 out of 141) that cannot be classified as Type 1a diabetic patients lack clinical characteristics of Type 1b diabetes and have to be reconsidered for a more appropriate ADA classification. These data suggest the need of further large population-based studies to understand if Type 1b diabetes really occurs in a Caucasian population. The patient with a strongly protective HLA genotype is particularly interesting considering that among Caucasians only a few sporadic cases with Type 1 diabetes and DQB1*0602, have been reported, none of whom was homozygous at DQB1 locus.

T-cell mediated autoimmunity to the insulinoma-associated protein 2 islet tyrosine phosphatase in type 1 diabetes mellitus.

The target molecules of the T-cell response in type 1 diabetes, despite their pathogenic importance, remain largely uncharacterized, especially in humans. Interestingly, molecules such as insulin and glutamic acid decarboxylase (GAD) have been shown to be a target not only of autoantibodies, but also of autoreactive T-lymphocytes both in man and in the non-obese diabetic (NOD) mouse. In the present study we aimed to determine the existence of a specific T-cell response towards the insulinoma-associated protein 2 (IA-2) islet tyrosine phosphatase, a recently identified autoantigen which is the target of autoantibodies strongly associated with diabetes development. Human recombinant IA-2 produced in Escherichia coli, was tested for its reactivity with peripheral blood lymphocytes obtained from 16 newly diagnosed type 1 diabetic patients and from 25 normal controls, 15 of whom were HLA-DR-matched. A T-cell proliferation assay was performed in triplicate employing freshly isolated cells in the absence or in the presence of the antigen to be tested (at two different concentrations: 2 microg/ml and 10 microg/ml). A specific T-cell proliferation (defined as a stimulation index (S.I.) >/=3) was observed against IA-2 used at a concentration of 10 microg/ml (but not of 2 microg/ml) in 8/16 diabetic patients, in 1/15 HLA-DR-matched control subjects (P<0.01 by Fisher exact test) and in 0/10 of the remaining normal individuals. A statistically significant difference (P<0.003 by Mann-Whitney U test) was also observed in S.I. values between patients (3.1+/-1.4) and HLA-DR-matched controls (1.7+/-0.54) employing IA-2 at a concentration of 10 microg/ml. However, when IA-2 was used at a concentration of 2 microg/ml, the difference in S. I. between patients (1.65+/-0.8) and controls (1.0+/-0.3) did not reach statistical significance. In conclusion, these data show the presence of a specific, dose-dependent T-lymphocyte response against the IA-2 islet tyrosine phosphatase at the onset of type 1 diabetes. Consequently, this molecule appears to be a target not only at the B-lymphocyte but also at the T-lymphocyte level, reinforcing the potential pathogenic role of this autoantigen in the islet destructive process.

A multi-centre randomized trial of two different doses of nicotinamide in patients with recent-onset type 1 diabetes (the IMDIAB VI).

BACKGROUND: Intensive insulin therapy is the gold standard by which Type 1 diabetes is treated. In addition to this therapy, administration of nicotinamide (NA) can be beneficial. This concept is reinforced by the results of a recent meta-analysis of the use of NA in patients with recent-onset Type 1 diabetes. METHODS: In this study we compared two different doses of NA in 74 patients with duration of Type 1 diabetes <4 weeks (mean age 13 years). Patients were randomly allocated in blind to two treatment groups: 38 patients received a dose of 25 mg/kg (b.w.) of NA and 36 patients received a dose of 50 mg/kg (b.w.) of NA. Intensive insulin therapy was carried out in order to optimize metabolic control as soon as possible after diagnosis and to maintain blood glucose level as near to normal as possible. Response to therapy was monitored throughout the study by investigating the occurrence of clinical (complete) remission defined, according to the recommendations of the International Diabetes Immunotherapy Group, as restoration of normal fasting and post-prandial blood glucose without any insulin administration for more than 2 weeks. Moreover, the integrated measures of metabolic control (C-peptide, HbA(1c) and insulin dose) were analysed at 3- month intervals up to 1 year after diagnosis. RESULTS: There were no significant differences in the integrated measures of metabolic control between the two NA treated groups either at onset of the disease or at each 3-month interval up to 1 year after diagnosis, although there was a tendency toward higher insulin dosages in the 50 mg NA group. No significant differences were observed in the rate of clinical remission between the two groups. CONCLUSION: We conclude that patients with recent-onset Type 1 diabetes treated with two different doses of NA, in addition to intensive insulin therapy, show similar residual beta-cell function 1 year later. Since both doses of NA are likely to be effective in reducing beta-cell dysfunction, the smaller dose of 25 mg/kg NA would be sufficient as a higher dose may induce insulin resistance.

[Nutritional status, obesity and metabolic control in children with type 1 diabetes mellitus]. / Stato nutrizionale, obesità ed equilibrio metabolico in soggetti in età evolutiva affetti da diabete mellito di tipo 1.

BACKGROUND AND AIMS: The aim of this study was to evaluate the nutritional status of children with type 1 diabetes and to search for possible influences of changes in body composition on aspects of diabetes. METHODS: A group of 96 diabetic subjects (41 males and 55 females) were studied, aged between 3 and 19 years old. The following parameters were examined: weight, stature, 5 skin folds, 7 circumferences, bioelectric impedance, arterial pressure, cholesterolemia, triglyceridemia, insulin dose, HbA1c and duration of disease. RESULTS: Obesity and overweight were present in 34.5% of the sample, but obesity was only observed in females (25.5%). There was also a high percentage of underweight subjects (11.5% of the entire sample). The mean values of weight BMI, 5 skin folds, 4 circumferences, FM (calculated using fold measurement and BIA) and AFA were higher in females, whereas mean values of waist/hip ratio and waist/thigh ratio and FFM (in % of body weight) were higher in males. A close correlation was also found between the 4 weight classes (underweight, normal weight, overweight, obese) and the majority of marker parameters for adiposity (5 folds, 4 circumferences, BIA, FM calculated using BIA, fold measurement and AFA). Of the other parameters examined (mean duration of disease, HbA1c assay, daily insulin dose, total cholesterolemia, triglycerididemia, arterial pressure), only the daily insulin dose showed higher values in females in 3 weight classes (underweight, normal weight and obese). Following a comparison with the control population (2469 subjects), higher mean values were found in the latter compared to diabetic subjects, but only in relation to 3 skin folds (tricipital, subscapular and suprailiac) and one circumference (forearm). CONCLUSIONS: The study shows a high frequency of overweight and obesity in children with type 1 diabetes, comparable to that in the healthy population. The finding of a higher frequency of obesity in diabetic females might be explained by their advanced puberal status, given that almost all the obese diabetic females were aged between 10 and 19 years old. The study confirms the validity of a number of anthropometric measurements (BMI, folds, circumference) and BIA in the evaluation of nutritional status in terms of body composition.

[Nutritional status, obesity, and metabolic balance in pediatric patients with type I diabetes mellitus]. / Stato nutrizionale, obesità ed equilibrio metabolico in soggetti in età evolutiva affetti da diabete mellito di tipo 1.

BACKGROUND AND AIMS: The aim of this study was to evaluate the nutritional status of children with type 1 diabetes and to search for possible influences of changes in body composition on aspects of diabetes. METHODS: A group of 96 diabetic subjects (41 males and 55 females) were studied, aged between 3 and 19 years old. The following parameters were examined: weight, stature, 5 skin folds, 7 circumferences, bioelectric impedance, arterial pressure, cholesterolemia, triglyceridemia, insulin dose, HbA1c and duration of disease. RESULTS: Obesity and overweight were present in 34.5% of the sample, but obesity was only observed in females (25.5%). There was also a high percentage of underweight subjects (11.5% of the entire sample). The mean values of weight BMI, 5 skin folds, 4 circumferences, FM (calculated using fold measurement and BIA) and AFA were higher in females, whereas mean values of waist/hip ratio and waist/thigh ratio and FFM (in % of body weight) were higher in males. A close correlation was also found between the 4 weight classes (underweight, normal weight, overweight, obese) and the majority of marker parameters for adiposity (5 folds, 4 circumferences, BIA, FM calculated using BIA, fold measurement and AFA). Of the other parameters examined (mean duration of disease, HbA1c assay, daily insulin dose, total cholesterolemia, triglycerididemia, arterial pressure), only the daily insulin dose showed higher values in females in 3 weight classes (underweight, normal weight and obese). Following a comparison with the control population (2469 subjects), higher mean values were found in the latter compared to diabetic subjects, but only in relation to 3 skin folds (tricipital, subscapular and suprailiac) and one circumference (forearm). CONCLUSIONS: The study shows a high frequency of overweight and obesity in children with type 1 diabetes, comparable to that in the healthy population. The finding of a higher frequency of obesity in diabetic females might be explained by their advanced puberal status, given that almost all the obese diabetic females were aged between 10 and 19 years old. The study confirms the validity of a number of anthropometric measurements (BMI, folds, circumference) and BIA in the evaluation of nutritional status in terms of body composition.

Three novel missense mutations in the glucokinase gene (G80S; E221K; G227C) in Italian subjects with maturity-onset diabetes of the young (MODY). Mutations in brief no. 162. Online.

The maturity-onset diabetes of the young (MODY), an autosomal dominant form of non-insulin dependent diabetes mellitus (NIDDM), is caused by mutations in the glucokinase (GK, MODY 2) and in the hepatocyte nuclear factor 1a (MODY 3) and 4a (MODY 1) genes. We have screened the glucokinase gene by the polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) in fifteen subjects with clinical characteristics of MODY and one parent with NIDDM, impaired glucose tolerance or gestational diabetes. PCR products with abnormal mobility in DGGE were directly sequenced. We have identified four mutant alleles, three of them (G80S, E221K, G227C) are new missense mutations located in or near the region of the active site cleft of the enzyme. The mutations co-segregate with hyperglycemia in the families of the three probands, whose biochemical and clinical phenotype is similar to other individuals with MODY 2 mutations.

Association between insulin dependent diabetes mellitus and coeliac disease. A study on 175 diabetes patients.

BACKGROUND: The association between diabetes mellitus and coeliac disease has been known for many years. In a random group of 175 insulin dependent diabetes mellitus patients of varying ages the following tests have been carried out: serum antigliadin antibodies (AGA) of IgA and IgG class, antireticulin antibodies (ARA) and antiendomisyum antibodies (AEA), both of IgA class. MATERIALS AND METHODS: The patients, 85 males and 90 females, had ages ranging from 1 yr to 30 yrs (102 in paediatric age--mainly between 6 and 14 years--and 73 adults). Patients with pathological values for AEA and/or ARA underwent an intestinal biopsy. RESULTS: Out of 175 patients studied, 21 had pathological values for AEA with or without pathological values for ARA and AGA, and 2 patients had only pathological values for ARA. 23 patients (21 with pathological values for AEA with or without ARA and AGA, 2 only for ARA ) underwent intestinal biopsy, all patients with pathological values for AEA had villous atrophy. The prevalence of coeliac disease among IDDM patients was 8.8% (95% CI 3.3 to 14.3) for the children and 16.4% (95% CI 7.9 to 24.9) for the adults. In patients with mucous atrophy, ARA, AGA IgA and IgG were pathological in 85%, 71% and 61% respectively. Symptoms and insulin requirements in all patients affected by coeliac disease before and after one year on a gluten free diet were also evaluated. The patients had clinical features with prevalently one or only few atypical symptoms which disappeared on a gluten free diet. Insulin requirements after one year on a gluten free diet appeared unchanged in coeliac patients. CONCLUSIONS: The need to screen all diabetic patients for coeliac disease is underlined.

Vitamin E and nicotinamide have similar effects in maintaining residual beta cell function in recent onset insulin-dependent diabetes (the IMDIAB IV study)

OBJECTIVE: Protection of residual beta cell function at the time of diagnosis of insulin-dependent diabetes mellitus (IDDM) by intensive insulin therapy and the addition of nicotinamide (NA) has been established. The objective of this study was to evaluate the effect of a free oxygen radical scavenger such as vitamin E (Vit E) on residual beta cell function and parameters of metabolic control in patients with recent onset IDDM undergoing intensive insulin therapy. DESIGN: The effect of Vit E was compared with that of NA (control group) in a randomized multicentre trial. METHODS: Eighty-four IDDM patients between 5 and 35 years of age (mean age 15.8 +/- 8.4 (s.d.) years) entered a one year prospective study. One group of patients (n = 42) was treated with Vit E (15 mg/kg body weight/day) for one year; the other group (n = 42) received NA for one year (25 mg/kg body weight/day). All patients were under intensive insulin therapy with three to four injections a day. Basal and stimulated (1 mg i.v. glucagon) C-peptide secretion, glycosylated haemoglobin and insulin dose were evaluated at diagnosis and at three-monthly intervals up to one year. RESULTS: Preservation and slight increase of C-peptide levels at one year compared with diagnosis were obtained in the two treated patient groups. No statistically significant differences were observed in basal or stimulated C-peptide levels between the two groups of patients for up to one year after diagnosis. Glycosylated haemoglobin and insulin dose were also similar between the two groups; however patients receiving Vit E under the age of 15 years required significantly more insulin than NA-treated patients one year after diagnosis (P < 0.04). CONCLUSIONS: Our data indicate that Vit E and NA possess similar effects in protecting residual beta cell function in patients with recent onset IDDM. Since their putative mechanism of protection on beta cell cytotoxicity is different, combination of these two vitamins may be envisaged for future trials of intervention at IDDM onset.

Incidence of insulin-dependent diabetes mellitus among Sardinian-heritage children born in Lazio region, Italy.

BACKGROUND: The relative importance of genetic and environmental factors in causing insulin-dependent diabetes mellitus (IDDM) is unknown. We studied this question by assessing the incidence of the disease in children, born in a region with a low incidence of IDDM (Lazio), but whose parents came from a region with high incidence (Sardinia). METHODS: We identified all IDDM cases that occurred between 1989 and 1994. We used as the denominator the number of children aged 0-14 born in Lazio of Sardinian parents to calculate incidence. We compared this rate with the incidences of IDDM in the populations of Lazio and Sardinia. FINDINGS: The age-adjusted incidence of IDDM in Sardinian-heritage children born and living in Lazio was 33.8 per 100,000 per year (95% CI 7.0-99.0) for those with two Sardinian parents, and 15.9 (8.7-26.6) for those with only one parent from Sardinia. The former incidence was not different from that recorded in Sardinia (34.4, 31.3-37.9), but was fourtold that of Lazio-heritage children (7.9, 7.1-8.8). INTERPRETATION: Our results show that two different ethnic groups living in the same region have a fourfold difference in incidence of IDDM. Children of Sardinian-heritage born in Lazio have the same incidence as the population of origin, which is genetically prone to the disease. Moreover, children with one Sardinian parent had a rate half that of Sardinians and double that of the indigenous population. We conclude that in a given population genetic susceptibility determines the frequency of IDDM in response to the environmental challenge.

In vivo measurement of immunoglobulin accumulation in the pancreas of recent onset type 1 diabetic patients.

OBJECTIVE: The possibility to quantify in vivo the severity of the inflammatory process in the pancreas of patients with recent onset insulin dependent diabetes mellitus (IDDM) could be of great relevance for follow-up studies involving immunotherapy. Scintigraphy with radiolabelled human polyclonal immunoglobulins (99mTc-HIG) is currently used for the diagnosis and follow-up of several acute and chronic inflammatory diseases. In this longitudinal study we have investigated to what extent 99mTc-HIG accumulate in the pancreas of patients with recent onset IDDM and in subjects at risk to develop IDDM. METHODS: Combined computerised tomography and gamma camera imaging were used to measure the radioactivity in the pancreatic region, as the pancreas/bone radioactivity ratio (P/B). Patients with IDDM (n = 15) were investigated at the time of diagnosis and after 1 year. Five pre-diabetic ICA+ve subjects and 8 age and sex matched normal subjects were also investigated. RESULTS: Eight out of 15 newly diagnosed IDDM patients and 2/5 ICA+ve subjects showed a significant accumulation of radiolabelled HIG in the pancreas (P/B higher than the upper 1st centile of normal subjects). One year after the diagnosis a significant accumulation of immunoglobulins was still detectable in the pancreas of IDDM patients positive who were positive at diagnosis. CONCLUSIONS: These results suggest that immunoglobulins home and bind to the pancreas of patients with recent onset IDDM and also in some ICA+ve individuals. This may reflect an increased vascular permeability of pancreatic capillaries as a consequence of the inflammatory process involving the islets. Thus, this technique may be useful for monitoring the efficacy of immune intervention at diagnosis.

Anti-ganglioside antibodies in new onset type 1 diabetic patients and high risk subjects.

Insulin dependent (type 1) diabetes mellitus appears to be a genetically determined autoimmune disease. Gangliosides have been implicated in type 1 diabetes as antigenic determinants recognized by islet cell antibodies (ICA) and shown to be able to modulate autoimmune phenomena in experimental diabetes. In order to explore in type 1 diabetes the humoral immune reactivity against gangliosides, taking into account their pancreatic localization and molecular characteristics, antibodies to gangliosides GM3, GM2, GM1, GD3, GD1a, GD1b, and GT1b have been investigated in sera from new onset type 1 diabetics and relatives of type 1 diabetic patients with or without insulin (CIAA) and/or islet cell autoantibodies. Using a purposefully designed sensitive ELISA method we found that presence of antibodies directed against the pacreatic disialo-ganglioside GD3 in a significant percentage of newly diagnosed type 1 diabetics (p < 0.001 vs normal controls) but not in CIAA and/or ICA positive relatives of type 1 diabetics. These findings confirm the involvement of gangliosides in autoimmune phenomena related to type 1 diabetes and suggest disialo-ganglioside GD3 as target of a humoral immune response associated with the onset of insulin-dependent diabetes.