PubMed Health⌕ Search

Biomedical subjects

E Bonifacio

Publications and source records attributed to E Bonifacio.

At least 19 recordsLinked to original sources
PubMed1997

Lack of association of DMB polymorphism with insulin-dependent diabetes.

Considerable evidence exists that the genes coding for the HLA class II DQ molecules in the MHC region are major contributors to genetic susceptibility in insulin-dependent diabetes. Located centromeric to the DQ loci are the genes encoding DMA and DMB, two class II-like molecules which play an essential role in the pathway leading to antigen presentation by HLA class II. In this study we have examined the distribution of the DMB allele and studied HLA DQA1-DQB1-TAP2-DMB haplotypes in 52 IDDM families and 65 un-related controls. DMB allele frequencies in IDDM and control subjects were not significantly different. DMB*0101 was present in 85% of patients vs. 76% of controls, DMB*0102 in 12 vs. 17%, DMB*0103 in 3 vs. 5%, DMB*0104 in 0 vs. 2%. The IDDM-susceptible MHC DQA1-DQB1 haplotypes found by analysis of IDDM families were not associated with specific DMB alleles. We conclude that the described DMB polymorphisms are not associated with IDDM susceptibility and DMB genotyping is unlikely to improve the assessment of genetic risk for IDDM.

Alleles↗
PubMed1997

A novel micro-assay for insulin autoantibodies.

Insulin autoantibodies (IAA) are established markers of Type 1 diabetes and are widely used for the prediction of this disease. Standard assays require relatively large serum volumes for reliable measurement of IAA, limiting their use in young children. We have developed a novel small volume assay which is suitable for screening large numbers of samples. For reasons of economy we have adopted a two-stage strategy in which all samples are screened for insulin binding and those with raised levels are quantified in an assay using competitive displacement. Using this assay 126 out of 241 (52%) newly diagnosed IDDM patients (median age 10.2, range 1.3-20.7 years) had IAA levels above the 99th centile of 2860 schoolchildren (median age 11.3, range 9.0-13.8 years), including 81 out of 117 (69%) patients below the age of 10 years. The assay compared well overall when measuring IAA in direct comparison with a conventional assay. We conclude that reliable measurement of IAA is possible on less than 50 microl of serum using this novel assay and that this should facilitate large scale screening, particularly in young children.

Adolescent↗
PubMed1997

Evaluation of islet cell antigen (ICA) 512/IA-2 autoantibody radioassays using overlapping ICA512/IA-2 constructs.

Islet cell antigen (ICA) 512 also termed IA-2 is a novel autoantigen of type 1 diabetes, which has a tyrosine phosphatase-like domain. We have assessed autoantibody RIAs using a series of ICA512/IA-2 constructs to produce in vitro synthesized 35S-methionine-labeled proteins. Levels of ICA512/IA-2 (256-979, truncated aminoterminus) autoantibodies were strongly correlated with those of the full-length ICA512/IA-2 (1-979) autoantibodies (r = 0.96, P < 0.0001) and ICA512/IA-2 (687-979) autoantibodies (r = 0.98, P < 0.0001). RIAs using these 3 constructs had increased sensitivity relative to our initially reported ICA512 autoantibody RIA (amino acids 389-948, truncated carboxy- and aminoterminus). Only 2 of 38 sera examined in this study reacted with an aminoterminus ICA512/IA-2 (1-577) construct. The mean SD score (SD score = (index of unknown sample-mean index of controls)/SD of controls) using the ICA512/IA-2 (256-979) construct was significantly higher than the SD score obtained with other ICA512/IA-2 constructs (P < 0.001). Amongst patients with new-onset diabetes and prediabetic relatives, using RIAs for autoantibodies reacting with ICA512/IA-2 (256-979), insulin, and glutamic acid decarboxylase 65, 98% expressed one or more of these autoantibodies and 78% expressed two or more, whereas no control (n = 208) expressed more than a single autoantibody. A combined ICA512/IA-2 (256-979), glutamic acid decarboxylase 65 autoantibody RIA with differential autoantigen labeling (35S-methionine, 3H-leucine) has been developed that uses 96-well plate membrane filtration and Top Counter beta counting. Concordance between results of dual and single RIAs was greater than 90%. This simple combined autoantibody assay should facilitate large-scale autoantibody screening.

Adolescent↗
PubMed1997

Prediction of IDDM in the general population: strategies based on combinations of autoantibody markers.

Strategies for assessing risk of progression to IDDM, based on single and combined autoantibody measurement, were evaluated in 2,855 schoolchildren (median age 11.4 years) and 256 children with newly diagnosed IDDM (median age 10.2 years), recruited to a population-based study in the Oxford region. In 256 children with IDDM, levels of antibodies > or =97.5th centile of the schoolchild population were found in 225 (88%) for islet cell antibodies (ICAs), in 190 (74%) for antibodies to GAD, in 193 (75%) for antibodies to protein tyrosine phosphatase IA-2 (IA-2), and in 177 (69%) for autoantibodies to insulin (IAAs). Estimates of risk of progression to IDDM within 10 years, derived by comparing the distribution of antibody markers in the two populations (schoolchildren and children with IDDM), were 6.7% (ICAs), 6.6% (GAD antibodies), 5.6% (IA-2 antibodies), and 4.8% (IAAs) for schoolchildren with levels above the 97.5th centile, increasing to 20, 23, 24, and 11%, respectively, for antibody levels >99.5th centile. Most children with IDDM had multiple antibody markers, and 89% of those diagnosed over age 10 years had > or =2 antibodies above the 97.5th centile, as compared against 0.7% of schoolchildren, in whom this combination gave a 27% 10-year estimated risk of IDDM. Risk increased but sensitivity fell as combined antibody thresholds were raised, or the number of antibodies above the threshold was increased. Strategies based on detection of > or =2 antibodies with primary testing for GAD and IA-2 antibodies and second line testing for ICAs and/or IAAs were evaluated. Detection of at least two markers selected from GAD antibodies > or =97.5th centile and/or IA-2 antibodies > or =99.5th centile and/or ICAs > or =97.5th centile identified 0.25% of schoolchildren and 83% of children with newly diagnosed IDDM, with an estimated risk of 71% (95% CI 57-91). Although confirmation from prospective studies is still needed, this analysis suggests that antibody combinations can predict diabetes in the general population.

Adolescent↗
PubMed1997

Low prevalence of islet autoantibodies in patients with gestational diabetes mellitus.

OBJECTIVE: To determine the proportion of patients with gestational diabetes mellitus (GDM) who have serological characteristics typical of IDDM. RESEARCH DESIGN AND METHODS: Islet cell antibodies (ICAs), insulin autoantibodies (IAAs), GAD65, and IA-2 antibodies were measured in 145 pregnant women with GDM, 33 with impaired glucose tolerance (IGT), and in 73 with normal glucose tolerance (NGT). ICAs were measured by indirect immunofluorescence; GAD65 and IA-2 antibodies, by a radio-ligand immunoassay incorporating 35S-labeled recombinant antigen; and IAAs, by a liquid-phase radiobinding assay. RESULTS: The prevalences of islet autoantibodies were low and not significantly different between groups. ICAs were detected at levels ranging from 5 to 45 Juvenile Diabetes Foundation U in 14 (10%) women with GDM, 2 (6%) women with (GT, and in 4 (5%) women with NGT. IAAs were detected at levels between 3 and 4 SD above the mean in 4 (3%) women with GDM, 0 women with IGT, and in 1 (1%) woman with NGT. None had both ICAs and IAAs. Neither GAD65 nor IA-2 antibodies, which have been detected in the majority of pre-IDDM and IDDM patients, were found in NGT, IGT, or GDM patients. CONCLUSIONS: Low-titer ICAs and IAAs are not infrequent in pregnant women, but multiple islet autoantibodies and antibodies to GAD65 or IA-2 were not found in GDM. These findings suggest that the serological characteristics of IDDM are rare in GDM.

Adult↗
PubMed1997

Islet cell antigens in the prediction and prevention of insulin-dependent diabetes mellitus.

Insulin-dependent diabetes mellitus (IDDM) is associated with both antibody and T-cell autoimmunity to pancreatic islet cell components. In recent years, considerable progress has been made in the identification of the molecular components of the pancreatic islets to which these immune responses are directed. These advances have led to the development of a number of immune markers for use in screening for individuals at risk for disease, and there is promise of antigen-specific immune intervention therapies to prevent diabetes in those identified as at risk. In this article, we review our current knowledge of autoantigens associated with IDDM and the implications this has on the prediction and prevention of the disease.

Autoantibodies↗
PubMed1996

Autoantibodies in insulin-dependent diabetes recognize distinct cytoplasmic domains of the protein tyrosine phosphatase-like IA-2 autoantigen.

Protein tyrosine phosphatase-like IA-2 is a major autoantigen in insulin-dependent diabetes. It has been identified as both a specificity of cytoplasmic islet cell Abs and one of the precursors of the 40- and 37-kDa tryptic fragment islet autoantigens. To characterize autoantibody binding to IA-2 and determine whether humoral autoimmunity extends to other tyrosine phosphatases, we analyzed serum reactivity in 100 patients with insulin-dependent diabetes against different in vitro translated portions of the IA-2 protein as well as the tyrosine phosphatase domains of HPTPbeta and HPTPdelta. All autoantibody reactivity was confined to the cytoplasmic portion of IA-2 (amino acids 601-979). At least four epitopes were found. These were contained within amino acids 605 to 620 and 605 to 682 of the juxtamembrane region and within amino acids 777 to 937 and 687 to 979 in the IA-2 tyrosine phosphatase-like domain. Footprinting studies confirmed the presence of multiple epitopes. Fifty-six percent of sera with IA-2 Abs bound epitopes within the juxtamembrane region, and 83% bound epitopes in the tyrosine phosphatase-like domain; 39% had Abs to both regions. No reactivity against the IA-2 ectodomain or the tyrosine phosphatase domains of HPTPbeta and HPTPdelta was found. These data suggest that the cytoplasmic region, in particular the tyrosine phosphatase-like domain, is the major target of IA-2 Abs in insulin-dependent diabetes, and that autoantibody reactivity is specific for IA-2 or IA-2-like molecules.

Adolescent↗
PubMed1996

Association of IA-2 autoantibodies with HLA DR4 phenotypes in IDDM.

Insulin, glutamate decarboxylase (GAD) and the protein tyrosine phosphatase-like molecule IA-2 are major targets of humoral autoimmunity in insulin-dependent diabetes mellitus (IDDM). These autoantibodies are heterogeneous with respect to age and patient human leukocyte antigen (HLA) phenotype. We have previously demonstrated that GAD and IA-2 antibodies potentially identify different subsets of IDDM patients. The aim of this study was to determine whether GAD and IA-2 autoantibodies were associated with different HLA DR phenotypes. We studied 160 patients with IDDM onset before age 16 years. At disease onset serum was tested for GAD and IA-2 antibodies by immunoprecipitation by in vitro-translated 35S-methionine labelled recombinant proteins. IA-2 antibodies were significantly associated with HLA DR4: 67 (86%) of 78 patients with HLA DR4 vs 31 (38%) of 82 non-DR4 patients had IA-2 antibodies (Pc < 0.0001) and IA-2 antibody levels were higher in patients with HLA DR4 (Pc < 0.0001). In contrast, GAD antibodies were more prevalent (Pc < 0.05) and antibody levels highest (Pc < 0.01) in patients with HLA DR3 phenotypes. These data provide further evidence that, in IDDM, production and titre of major autoantibody specificities are associated with HLA class II alleles.

Alleles↗
PubMed1995

Identification of protein tyrosine phosphatase-like IA2 (islet cell antigen 512) as the insulin-dependent diabetes-related 37/40K autoantigen and a target of islet-cell antibodies.

The majority of patients with insulin-dependent diabetes (IDDM) have Abs to 40- and/or 37-kDa tryptic fragments (37/40K-Abs) deriving from an unidentified islet cell membrane protein distinct from glutamate decarboxylase (GAD). Recently, autoantibodies against ICA512, which has identity with the protein tyrosine phosphatase-like protein IA2, were reported. In this study we have examined whether IA2/ICA512 is the Ag specificity of 37/40K-Abs, and one of the determinants of islet cell Abs (ICA) detected by immunofluorescence. Serum from 51 of 100 new onset IDDM patients immunoprecipitated 40- and/or 37-kDa insulinoma polypeptides, and 53 immunoprecipitated in vitro translated rIA2; 49 had both 37/40K-Abs and rIA2 Abs. There were strong correlations between the levels of Abs to rIA2 and both 40 kDa (r = 0.85, p < 0.0001) and 37 kDa (r = 0.70, p < 0.0001) insulinoma polypeptides. Trypsin treatment of immunoprecipitated rIA2 yielded 40- and 37-kDa fragments, and preincubation of sera with rIA2 completely inhibited binding to the insulinoma 40- and 37-kDa polypeptides. IA2 Ab levels also correlated with ICA titer in GAD-Ab negative sera, and preincubation with rIA2 reduced ICA staining intensity in sera with ICA and IA2 Abs, but not in sera with ICA in the absence of IA2 Abs. These results provide clear evidence for the identification of IA2/ICA512 as the precursor of the islet 40- and 37-kDa polypeptide autoantigens and as one of the ICA specificities. Combined detection of Abs to IA2 and GAD65 in a single radio-binding assay identified Abs in 88 of 100 IDDM patients, and potentially facilitates population screening for IDDM risk assessment.

Adolescent↗
PubMed1995

HLA DQA1-DQB1-TAP2 haplotypes in IDDM families: no evidence for an additional contribution to disease risk by the TAP2 locus.

The TAP2 gene, located in the HLA class II region, encodes a subunit of a transporter involved in the endogenous antigen-processing pathway, and has been suggested to contribute to the genetic risk for insulin-dependent diabetes (IDDM). In order to determine whether the TAP2 locus modulates the risk conferred by HLA DQ loci, HLA DQA1-DQB1-TAP2 haplotypes were analysed in 48 IDDM probands, their first degree relatives, and in 62 normal control subjects. A decreased frequency of the TAP2B allele was confirmed in this IDDM cohort (12 vs 28% in control subjects, pc < 0.05). Analysis of 73 informative meiotic events in IDDM and control families demonstrated a recombination fraction between HLA DQB1 and TAP2 loci of 0.041 (Log of the odds score = 16.5; p < 10(-8)) indicating strong linkage between these loci. Family haplotype analysis demonstrated linkage disequilibrium between TAP2 and HLA DQA1-DQB1, and showed that the reduced frequency of TAP2B was associated with its absence on the IDDM susceptible DQA1*0301-DQB1*0302 haplotype, its low frequency on DQA1*0501-DQB1*0201, and the association of TAP2B with DQA1*0101-DQB1*0501 haplotypes which were less frequent in IDDM patients. Comparison of transmitted with non-transmitted haplotypes in IDDM families showed a slight but not significant decrease in TAP2B allele frequency on transmitted (3 of 37) vs non-transmitted (2 of 9) HLA DQA1*0501-DQB1*0201 haplotypes. No other differences were observed. Twenty-four unrelated DQA1*0501-DQB1*0201 haplotypes from non-diabetic families had a TAP2B allele frequency (4%) similar to that in IDDM haplotypes.(ABSTRACT TRUNCATED AT 250 WORDS)

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

HLA-DQ screening for risk assessment of insulin dependent diabetes in northern Italy.

Genetic markers may be used to improve the prediction of insulin-dependent diabetes mellitus (type 1) in individuals with islet autoantibodies. In order to develop a risk assessment strategy for the Lombardy region of northern Italy based on genetic and immunological markers, we analyzed HLA DQA1 and DQB1 alleles in 60 type 1 probands and their first-degree relatives and 65 unrelated control subjects from the same area using polymerase chain reaction (PCR) and oligonucleotide probes. The major risk haplotypes were DQA1 *0501-DQB1*0201 (39.1% of diabetic vs. 8.9% of non-diabetic haplotypes) and DQA1 *0301-DQB1*0302(20% of diabetic vs 7.1% of non-diabetic haplotypes). Stratified analysis showed DQA1*0102-DQB1*0502 also to be associated with type 1 susceptibility when found together with DQA1*0501-DQB1*0201 or DQA1*0301-DQB1*0302. One type 1 patient had the type 1-protective DQA1*0102-DQB1*0602 haplotype. Overall, 88% of patients and 20% of unrelated control subjects had either DQA1*0501-DQB1*0201 or DQA1*0301-DQB1*0302 in the absence of DQA1*0102-DQB1*0602. These data suggest that typing for markers identifying these three haplotypes in the Lombardy population will achieve a sensitivity of almost 90% and exclude 80% of children from subsequent islet autoantibody testing.

Adolescent↗
PubMed1995

Islet autoantibody markers in IDDM: risk assessment strategies yielding high sensitivity.

Identification of islet autoantigens offers the possibility that antibody tests other than islet cell antibodies may be used for assessing risk of insulin-dependent diabetes mellitus (IDDM). The aim of this study was to determine the combination of islet autoantibody markers that could identify most future cases of IDDM. Islet cell antibodies, antibodies to glutamic acid decarboxylase (GAD)65, 37,000/40,000 M(r) islet tryptic fragments, carboxypeptidase-H, and islet cell autoantigen (ICA)69 were measured in sera from 100 newly-diagnosed IDDM patients, 27 individuals prior to onset of IDDM, and 83 control subjects. Islet cell antibodies were detected in 88% of IDDM patients and 81% with pre-IDDM, GAD65 antibodies in 70% of IDDM patients and 89% with pre-IDDM, and antibodies to 37,000/40,000 M(r) islet tryptic fragments in 54% of IDDM patients and in 48% with pre-IDDM. The latter were found only in conjunction with islet cell antibodies and were more frequent in young onset cases. All 20 IDDM patients and the 3 pre-IDDM subjects who had islet cell antibodies without GAD65 antibodies had antibodies to 37,000/40,000 M(r) islet tryptic fragments, and all but one had disease onset before age 15 years. No sera strongly immunoprecipitated in vitro translated ICA69 or carboxypeptidase-H; 4% of patients had anti-ICA69 and 11% anti-carboxypeptidase-H levels above those of the control subjects. The findings suggest that none of the single antibody specificities are as sensitive as islet cell antibodies, but that a combination of GAD65 antibodies and antibodies to 37,000/40,000 M(r) islet tryptic fragments has the potential to identify more than 90% of future cases of IDDM.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1995

Disease sensitivity and specificity of 52 assays for glutamic acid decarboxylase antibodies. The Second International GADAB Workshop.

There is increasing interest in the use of glutamic acid decarboxylase antibodies (GADAbs) for identification of subjects at increased risk of developing insulin-dependent diabetes mellitus (IDDM). However, considerable variation exists between laboratories in the reported frequency of GADAb in various clinical situations, and disease sensitivity and specificity have not yet been compared between assays. An international workshop was held in which 101 coded freeze-dried sera, including 39 from subjects with newly diagnosed IDDM, 32 from healthy control subjects, 4 from nondiabetic subjects with Graves' disease, and 4 from islet cell antibody-positive subjects, were analyzed in 52 assays (radiobinding assay [RBA], 26; enzyme-linked immunosorbent assay [ELISA], 19; and enzymatic immunoprecipitation assay [EIP], 7). The mean sensitivity for RBAs (76.2%) was higher than for ELISAs (36.5%) and EIPs (49.9%) (P < 0.01). The mean specificity was similar for each assay format (RBA, 89.4%; ELISA. 89.4%; and EIP, 92.3%). The lower sensitivities of the ELISA and EIP were predominantly due to the inability of these assays to detect low levels of GADAb in IDDM. To convert results to standard units, standard curves were constructed using duplicate dilutions of the anti-glutamic acid decarboxylase monoclonal antibody MICA 3 and serum from a patient with stiff-man syndrome (SMS). Curves could be derived in 28 assays using the MICA 3 serum and in 29 using the SMS serum. The mean coefficients of variation between assays for disease and control samples were 45% when results were converted to MICA units, 77% for SMS units, and 76% for SD scores.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1995

[Leukocytic procoagulant activity and chemotherapeutic drugs: the comparative effects of cisplatin and carboplatin on lympho-/monocytic procoagulant activity in vitro].

It is well known that a number of chemotherapeutic agents are able to induce procoagulating activity not only in neoplastic cells but also in normal, monocyte/macrophage cells, and some of them, including cisplatin, even increase procoagulating activity of the factor already expressed, providing a further example of the possible co-participation of chemotherapy in the onset of thrombotic complications in cancer patients. Carboplatin is an analogue of cisplatin but differs strikingly in terms of its collateral effects, in particular being less oto- and nephrotoxic. To the authors' knowledge there are no data regarding the possible effect of carboplatin on lympho/monocyte procoagulating activity. This study shows that not only platin but also carboplatin is able to increment the levels of lympho/monocyte procoagulating activity in vitro, with a dose-dependent effect, and to synergize with bacterial endotoxin in increasing this leukocyte activity, although the synergic effect is significantly greater in the case of carboplatin. The importance of these findings at a practical and clinical and clinical level still remains to be defined, in particular in the light of the different pharmacokinetic behaviour of these two chemotherapeutic agents and in the context of those neoplastic diseases for which cisplatin and carboplatin treatment is most frequently used.

Blood Coagulation Factors↗