Development of celiac disease-associated antibodies in offspring of parents with Type I diabetes

Aims/hypothesis. The aim of this study was to determine the frequency and temporal development of antibodies related to celiac disease in offspring of parents with Type I (insulin-dependent) diabetes mellitus. Methods. Sera from 913 offspring of parents with Type I diabetes prospectively followed from birth to the age of 8 years were tested for IgG-transglutaminase antibodies (IgG-tTGCAs), endomysial IgA antibodies (EMA) and gliadin antibodies. Results. We found tTGCAs in 32 (3.5 %) of the 913 relatives. Prevalence was related to age and reached 6.5 % at age 8 years. Endomysial IgA antibodies were detected in 44 % of the relatives with tTGCAs and 0.6 % of tTGCA negative relatives and were also most prevalent (5 %) in those aged 8 years. Both tTGCAs and EMAs were more frequent in relatives with the HLA DRB1^*03 DQA1*0501 DQB1^*02 haplotype (7.1 % and 7.2 %, respectively; p < 0.005). Anti-gliadin antibodies were common in both tTGCA positive (42 %) and negative (23 %) relatives, did not show a relation with age and were less prevalent in relatives with HLA DR3 (p < 0.05). There was no association between the presence of antibodies associated with celiac disease and islet autoantibodies in these relatives. Of the relatives 15 (1.6 %) had tTGCAs plus EMAs. In two of these, anti-gliadin antibodies were detected before the detection of tTGCAs and EMAs at the age of 9 months whereas none of the remainder had any antibodies associated with celiac disease before age 2 years. In three there were no detectable anti-gliadin antibodies in any of the samples tested. Celiac disease without clinical symptoms was diagnosed in 9 of 12 by intestinal biopsy. Conclusion/interpretation. A statistically significant proportion of relatives of patients with Type I diabetes have celiac disease-associated autoimmunity and the silent form of celiac disease early in life. These relatives should, therefore, be considered for celiac antibody screening. [Diabetologia (2000) 43: 1005–1011] Received: 20 January 2000 and in revised form: 26 April 2000     

老年缺血性脑血管病血瘀证患者的红细胞变形性及一氧化氮水平的检测及其意义

目的 :探讨老年缺血性脑血管病血瘀证患者的红细胞变形性及一氧化氮 ( NO)水平的变化。方法 :检测 5 5例老年缺血性脑血管病血瘀证患者的红细胞变形指数和 NO的水平。其中短暂性脑缺血发作 ( TIA) 2 5例 ,脑梗死 3 0例 ;另设 2 6例正常对照组作对比观察。结果 :血瘀证 TIA患者红细胞变形指数 ( 0 .4 67± 0 .14 5 )显著低于正常对照组 ( 0 .5 0 8± 0 .14 1) ,P<0 .0 5 ,脑梗死患者红细胞变形指数 ( 0 .4 43± 0 .15 6)降低更为明显( P<0 .0 1) ;NO水平 ,TIA患者与正常对照组比较无显著性差异 ( 79.10± 15 .3 7比 76.70± 17.10 ) ,而脑梗死患者 ( 88.5 0± 13 .68)显著升高。结论 :红细胞变形性改变及 NO水平升高在一定程度上均参与了老年缺血性脑血管病血瘀证的发生发展     

Autoantibodies to N-terminally truncated GAD improve clinical phenotyping of individuals with adult-onset diabetes: Action LADA 12

Aims/hypothesis

Adult-onset type 1 diabetes, in which the 65 kDa isoform of GAD (GAD65) is a major autoantigen, has a broad clinical phenotype encompassing variable need for insulin therapy. This study aimed to evaluate whether autoantibodies against N-terminally truncated GAD65 more closely defined a type 1 diabetes phenotype associated with insulin therapy.

Methods

Of 1114 participants with adult-onset diabetes from the Action LADA (latent autoimmune diabetes in adults) study with sufficient sera, we selected those designated type 1 (n?=?511) or type 2 diabetes (n?=?603) and retested the samples in radiobinding assays for human full-length GAD65 autoantibodies (f-GADA) and N-terminally truncated (amino acids 96–585) GAD65 autoantibodies (t-GADA). Individuals’ clinical phenotypes were analysed according to antibody binding patterns.

Results

Overall, 478 individuals were f-GADA-positive, 431 were t-GADA-positive and 628 were negative in both assays. Risk of insulin treatment was augmented in t-GADA-positive individuals (OR 4.69 [95% CI 3.57, 6.17]) compared with f-GADA-positive individuals (OR 3.86 [95% CI 2.95, 5.06]), irrespective of diabetes duration. Of 55 individuals who were f-GADA-positive but t-GADA-negative, i.e. with antibody binding restricted to the N-terminus of GAD65, the phenotype was similar to type 2 diabetes with low risk of progression to insulin treatment. Compared with these individuals with N-terminal GAD65-restricted GADA, t-GADA-positive individuals were younger at diagnosis (p?=?0.005), leaner (p?<?0.0001) and more often had multiple diabetes-associated autoantibodies (28.3% vs 7.3%; p?=?0.0005).

Conclusions/interpretation

In individuals with adult-onset diabetes, presence of N-terminally truncated GAD65 autoantibodies is associated with the clinical phenotype of autoimmune type 1 diabetes and predicts insulin therapy.

     

Identification and assessment of frailty in older patients with chronic myeloid leukemia and myelofibrosis,and indications for tyrosine kinase inhibitor treatment

The incidence of cancer, including myeloproliferative neoplasms (MPNs), is projected to increase significantly due to the growing proportion of people aged >?65 years. These older individuals are a heterogeneous population in terms of fitness, comorbidity, and psychological reserve. Therefore, age per se does not always provide an accurate indication of condition in patients with cancer. Frailty has been proposed as an alternative measure of vulnerability that might better indicate which patients can tolerate standard cancer treatment and those who may benefit from treatment adjustment. A number of methods can be used to assess frailty in older patients with hematological malignancies, including the Cardiovascular Health Study Frailty Screening Measure, the FRAIL (Fatigue, Resistance, Ambulation, Illnesses, and Loss of weight) questionnaire, the Clinical Frailty Scale (CFS), and the Gérontopôle Frailty Screening Tool. In addition to physical frailty, comorbidity and quality of life should also be included in the assessment. Prior to the introduction of tyrosine kinase inhibitors (TKIs), age was considered a marker of poor prognosis in patients with MPNs. In contrast, data show that age is not necessarily a contraindication for TKI use. In CML, the efficacy of TKIs has been shown to be independent of age. The JAK1/2 inhibitor ruxolitinib also seems to be effective across a range of patient ages. Available data suggest that chronological age itself should not necessarily be a contraindication for many new therapies in patients with MPNs, and that frailty does provide a better measure of vulnerability. There is a need for specific methods to assess frailty in patients with MPNs, particularly the context of effective new treatment options, such as TKIs and ruxolitinib.     

Validation of a differential PCR and an ELISA procedure in studying HER-2/neu status in breast cancer

HER-2/neu oncogene status and total cellular p185^HER-2 content were simultaneously analyzed in 415 invasive breast-cancer specimens by differential PCR and ELISA respectively. Mathematical analysis of the data led us to establish a cut-off value of 1.7 for the ratio between the intensity of the HER-2/neu gene band and the reference gene band, to consider the HER-2/neu gene amplified, and of 260 fmol/mg protein, to consider p185^HER-2 over-expressed. Of the 415 tumors studied, 15% showed a diverse degree of HER-2/neu gene amplification. Of these tumors, 87% showed over-expression of the p185^HER-2. Of the remaining 352 specimens that did not display HER-2/neu gene amplification, 97% showed no p185^HER-2 over-expression (p < 0.0001). In 40 selected samples with a p185^HER-2 level lower than 260 fmol/mg protein, the degree of p185^HER-2 phosphorylation was very low or undetectable. Conversely, 38 of 46 selected tumors with a p185^HER-2 level higher than 260 fmol/mg protein exhibited a considerable degree of p185^HER-2 phosphorylation (p < 0.0001). Our data suggest that: (i) differential PCR and ELISA, which are relatively simple procedures, give similar information on HER-2/neu status in breast cancer; and (ii) given the large series analyzed, the cutoff values established can be considered as safe values for determining whether, in a given tumor, the HER-2/neu oncogene is amplified or p185^HER-2 is over-expressed. © 1996 Wiley-Liss, Inc.     

Combined testing of antibody titer and affinity improves insulin autoantibody measurement: Diabetes Antibody Standardization Program

The aim of the workshop was to assess whether four laboratories could reproducibly measure insulin autoantibody (IAA) affinity in coded sera from non-diabetic relatives of patients with type 1 diabetes, newly diagnosed patients, and healthy blood donors, and whether combining affinity with autoantibody titer could improve concordance and performance of IAA assays. IAA affinity was measured by competitive binding using constant amounts of Tyr14A [125I] human insulin and increasing quantities of unlabeled human insulin. There was high concordance between laboratories in distinguishing high, moderate, and low affinity IAA, although IAA binding to insulin varied with assay format. Multiple islet autoantibody-positive and patient sera were identified by high affinity IAA regardless of laboratory-designated IAA status. Combining affinity and titer significantly improved sensitivity, specificity, and concordance of IAA measurement. This workshop has demonstrated that different laboratories are able to reproduce IAA affinity results and that considering IAA affinity is likely to improve the diagnostic performance of IAA assays.     

In vitro selection of resistance in Haemophilus influenzae by amoxicillin-clavulanate,cefpodoxime, cefprozil,azithromycin, and clarithromycin

Abilities of amoxicillin-clavulanate, cefpodoxime, cefprozil, azithromycin, and clarithromycin to select resistant mutants of Haemophilus influenzae were tested by multistep and single-step methodologies. For multistep studies, 10 random strains were tested: 5 of these were beta-lactamase positive. After 50 daily subcultures in amoxicillin-clavulanate, MICs did not increase more than fourfold. However, cefprozil MICs increased eightfold for one strain. Clarithromycin and azithromycin gave a >4-fold increase in 8 and 10 strains after 14 to 46 and 20 to 50 days, respectively. Mutants selected by clarithromycin and azithromycin were associated with mutations in 23S rRNA and ribosomal proteins L4 and L22. Three mutants selected by clarithromycin or azithromycin had alterations in ribosomal protein L4, while five had alterations in ribosomal protein L22. Two mutants selected by azithromycin had mutations in the gene encoding 23S rRNA: one at position 2058 and the other at position 2059 (Escherichia coli numbering), with replacement of A by G. One clone selected by clarithromycin became hypersusceptible to macrolides. In single-step studies azithromycin and clarithromycin had the highest mutation rates, while amoxicillin-clavulanate had the lowest. All resistant clones were identical to parents as observed by pulsed-field gel electrophoresis. The MICs of azithromycin for azithromycin-resistant clones were 16 to >128 micro g/ml, and those of clarithromycin for clarithromycin-resistant clones were 32 to >128 micro g/ml in multistep studies. For strains selected by azithromycin, the MICs of clarithromycin were high and vice versa. After 50 daily subcultures in the presence of drugs, MICs of amoxicillin-clavulanate and cefpodoxime against H. influenzae did not rise more than fourfold, in contrast to cefprozil, azithromycin, and clarithromycin, whose MICs rose to variable degrees.     

Activity of cefditoren against respiratory pathogens

The activity of cefditoren and six other cephalosporins was tested against 250 pneumococci, including strains resistant to macrolides and quinolones. Cefditoren gave the lowest MICs, with MIC(50) and MIC(90) values of < or =0.016/0.03, 0.125/0.5 and 0.5/2.0 mg/L for penicillin-susceptible, -intermediate and -resistant pneumococci, respectively. A time-kill study of 12 pneumococcal strains with varying drug susceptibilities showed that cefditoren, at 2 x MIC, gave 99% killing of all strains after 12 h, with 99.9% killing after 24 h. Other cephalosporins gave similar kill kinetics but at higher concentrations. Against 160 Haemophilus influenzae, cefditoren had the lowest MICs (MIC(50) and MIC(90) both < or =0.016 mg/L), irrespective of beta-lactamase production. Time-kill studies of cefditoren compared with five other oral cephalosporins showed that cefditoren, at 8 x MIC, was bactericidal against 8/9 strains and gave 90% killing of all strains at the MIC after 12 h. Activity was bactericidal (99.9% killing) after 24 h with all drugs tested. Multistep studies of four penicillin-susceptible, four penicillin-intermediate and four penicillin-resistant strains showed that cefditoren, co-amoxiclav and cefprozil did not select for resistant mutants after 50 subcultures, compared with cefuroxime and azithromycin, where resistant mutants were selected in two and nine strains, respectively. Single-step mutation studies showed that cefditoren, at the MIC, had the lowest frequency of spontaneous mutants compared with other drugs.     

Assessment of precision,concordance, specificity,and sensitivity of islet cell antibody measurement in 41 assays

Summary Forty-one assays were analysed at the 3rd International Workshop on the standardisation of islet cell antibodies. Analysis of precision demonstrated assays consistently detecting blind duplicates within one doubling dilution and capable of discriminating one doubling dilution differences in islet cell antibody concentration. Some assays, however, reported duplicates discrepantly by more than seven doubling dilutions, and consequently could not distinguish even large quantities of islet cell antibodies. Precision was best in assays from laboratories which had participated in all three Standardisation Workshops and was not dependent upon methodology. The use of the Juvenile, Diabetes Foundation reference islet cell antibody standard and standard curves reduced the scatter of results, and was best amongst assays with better precision. Twenty-seven assays reported all ten blood donor sera as negative. However, 14 assays did not, and specificity (negativity in health) was <50% in three assays. Low specificity was strongly associated with poor precision. The detection limit of assays ranged from <5 to 50 JDF units and was partially dependent upon methodology. Assays incorporating extended incubation had the lowest detection limits without a decrease in the specificity of the ten blood donor sera. Precise quantification is fundamental for the standardisation and comparability of islet cell antibodies. Precise quantitative assays have been identified and reference standards and common units established.