Sinikka Koskinen

Association of IFIH1 and other autoimmunity risk alleles with selective IgA deficiency

To understand the genetic predisposition to selective immunoglobulin A deficiency (IgAD), we performed a genome-wide association study in 430 affected individuals (cases) from Sweden and Iceland and 1,090 ethnically matched controls, and we performed replication studies in two independent European cohorts. In addition to the known association of HLA with IgAD, we identified association with a nonsynonymous variant in IFIH1 (rs1990760G>A, P = 7.3 × 10−10) which was previously associated with type 1 diabetes and systemic lupus erythematosus. Variants in CLEC16A, another known autoimmunity locus, showed suggestive evidence for association (rs6498142C>G, P = 1.8 × 10−7), and 29 additional loci were identified with P < 5 × 10−5. A survey in IgAD of 118 validated non-HLA autoimmunity loci indicated a significant enrichment for association with autoimmunity loci as compared to non-autoimmunity loci (P = 9.0 × 10−4) or random SNPs across the genome (P < 0.0001). These findings support the hypothesis that autoimmune mechanisms may contribute to the pathogenesis of IgAD.

Long-term follow-up of health in blood donors with primary selective IgA deficiency

A 20-year health follow-up study of 159 initially healthy blood donors with a severe deficiency of serum IgA (<0.05×10−3 g/L) and of 45 donors with decreased serum IgA (0.05×10−3−0.8 g/L) was carried out. The findings indicate that persons with a severe deficiency of and decreased serum IgA who are healthy as young adults have an increased susceptibility to pneumonia and recurrent episodes of other respiratory infections and a higher risk of developing autoimmune diseases in middle age. Vitiligo, autoimmune hypothyreosis, milk intolerance, and possible rheumatoid arthritis were associated with severe IgA deficiency, but otherwise different degrees of IgA deficiency seem to be similar with respect to the appearance of diseases. Regardless of the fact that a total of 163 (80%) of the 204 IgA-deficient subjects had episodes of infections, drug allergy, or autoimmune or atopic disease, the finding of primary, selective IgA deficiency in a healthy adult per se does not seem to predict severe life-threatening illnesses at least during 20 years of life.

Fetal intracranial haemorrhages caused by fetal and neonatal alloimmune thrombocytopenia: an observational cohort study of 43 cases from an international multicentre registry

Objective To characterise pregnancies where the fetus or neonate was diagnosed with fetal and neonatal alloimmune thrombocytopenia (FNAIT) and suffered from intracranial haemorrhage (ICH), with special focus on time of bleeding onset. Design Observational cohort study of all recorded cases of ICH caused by FNAIT from the international No IntraCranial Haemorrhage (NOICH) registry during the period 2001–2010. Setting 13 tertiary referral centres from nine countries across the world. Participants 37 mothers and 43 children of FNAIT pregnancies complicated by fetal or neonatal ICH identified from the NOICH registry was included if FNAIT diagnosis and ICH was confirmed. Primary and secondary outcome measures Gestational age at onset of ICH, type of ICH and clinical outcome of ICH were the primary outcome measures. General maternal and neonatal characteristics of pregnancies complicated by fetal/neonatal ICH were secondary outcome measures. Results From a total of 592 FNAIT cases in the registry, 43 confirmed cases of ICH due to FNAIT were included in the study. The majority of bleedings (23/43, 54%) occurred before 28 gestational weeks and often affected the first born child (27/43, 63%). One-third (35%) of the children died within 4 days after delivery. 23 (53%) children survived with severe neurological disabilities and only 5 (12%) were alive and well at time of discharge. Antenatal treatment was not given in most (91%) cases of fetal/neonatal ICH. Conclusions ICH caused by FNAIT often occurs during second trimester and the clinical outcome is poor. In order to prevent ICH caused by FNAIT, at-risk pregnancies must be identified and prevention and/or interventions should start early in the second trimester.

High-Density SNP Mapping of the HLA Region Identifies Multiple Independent Susceptibility Loci Associated with Selective IgA Deficiency

Selective IgA deficiency (IgAD; serum IgA<0.07 g/l) is the most common form of human primary immune deficiency, affecting approximately 1∶600 individuals in populations of Northern European ancestry. The polygenic nature of IgAD is underscored by the recent identification of several new risk genes in a genome-wide association study. Among the characterized susceptibility loci, the association with specific HLA haplotypes represents the major genetic risk factor for IgAD. Despite the robust association, the nature and location of the causal variants in the HLA region remains unknown. To better characterize the association signal in this region, we performed a high-density SNP mapping of the HLA locus and imputed the genotypes of common HLA-B, -DRB1, and -DQB1 alleles in a combined sample of 772 IgAD patients and 1,976 matched controls from 3 independent European populations. We confirmed the complex nature of the association with the HLA locus, which is the result of multiple effects spanning the entire HLA region. The primary association signal mapped to the HLA-DQB1*02 allele in the HLA Class II region (combined P = 7.69×10−57; OR = 2.80) resulting from the combined independent effects of the HLA-B*0801-DRB1*0301-DQB1*02 and -DRB1*0701-DQB1*02 haplotypes, while additional secondary signals were associated with the DRB1*0102 (combined P = 5.86×10−17; OR = 4.28) and the DRB1*1501 (combined P = 2.24×10−35; OR = 0.13) alleles. Despite the strong population-specific frequencies of HLA alleles, we found a remarkable conservation of these effects regardless of the ethnic background, which supports the use of large multi-ethnic populations to characterize shared genetic association signals in the HLA region. We also provide evidence for the location of association signals within the specific extended haplotypes, which will guide future sequencing studies aimed at characterizing the precise functional variants contributing to disease pathogenesis.

Long-term persistence of selective IgA deficiency in healthy adults

A follow-up study of 204 healthy blood donors with IgA deficiency, identified between 1971 and 1980, was carried out. Sera were initially screened by a double diffusion method and 182 were retested by a more sensitive haemagglutination inhibition method. A reexamination was performed in 1990 and, again, in 1991–1992 using an enzyme immunoassay (EIA) developed for the measurement of very low concentrations of IgA. The median follow-up period was 19 years, and in 159 (78%) subjects no serum IgA could be detected in any of the measurements. In 42 (21%) subjects, serum IgA was detectable (>0.18 mg/L), but the level was below the lower limit of the reference range for adults (800 mg/L) and remained relatively constant. Three subjects showed minute amounts of IgA by EIA (0.2–3 mg/L) in one of the follow-up samples in 1990–1992, but the level was below the detection limit of the EIA (0.05 mg/L) in the other sample. Thus, not only does primary IgA deficiency appear to be permanent, but also lower than normal serum IgA levels remain the same in healthy adults.

Long-term follow-up of anti-IgA antibodies in healthy IgA-deficient adults

A follow-up study of anti-IgA antibodies in 159 healthy blood donors with severe deficiency of serum IgA (<0.05 mg/L) and in 45 donors with decreased serum IgA levels (0.05–799 mg/L), identified in 1971–1980, was carried out. Initially anti-IgA antibodies were determined by a hemagglutination (HA) method and two reexaminations were done in 1990–1992 by an enzyme immunoassay. The median follow-up period was 19 years, during which anti-IgA level was changed considerably in only four persons, increased in two, and high level antibodies (>1/1000 by HA) appeared in two. In reexaminations anti-IgA antibodies were found in 30 (19%) subjects with severe IgA deficiency and the antibody levels remained relatively constant in those who had high and medium antibody levels. Anti-IgA antibodies were not found in subjects with decreased, but detectable serum IgA. Thus it seems that only those healthy adults who have severe IgA deficiency develop anti-IgA antibodies and their anti-IgA levels remain fairly constant Of the 159 subjects with severe IgA deficiency, 66 had a history of IgA exposure, but no correlation to anti-IgA development was noted.

A sensitive enzyme immunoassay for the measurement of low concentrations of IgA

A microtitre plate enzyme immunoassay (EIA) for determining low concentrations of IgA is described and validated for serum and plasma products. The measuring range of the assay was 3.3-150 micrograms/l. The predilution requirement of samples was matrix dependent, ranging from 1/16 for serum to 1/2 for 4% albumin. Dilute protein solutions required no predilution. The limits of detection were 50 micrograms/l for serum, 25 micrograms/l for intravenous immunoglobulin, 13 micrograms/l for 20% albumin, 7 micrograms/l for 4% albumin and 3 micrograms/l for washing solutions of blood cell components. Interassay coefficients of variation over the range of 3.4 mg to 1.5 g IgA/l ranged from 3.8 to 5.7%. Respective values for two low-level sera, containing 309 and 512 micrograms IgA/l, were 15.5% and 11.1%. Comparison of the EIA with a commercial radial immunodiffusion (RID) method showed that the results of the two assays correlated well ([EIA] = 0.877 x [RID] + 0.401 mg/l, r = 0.996, n = 20). This assay is also suitable for the large-scale screening of blood donors.

An enzyme immunoassay for the determination of anti-IgA antibodies using polyclonal human IgA.

An enzyme immunoassay (EIA) for screening and quantitation of serum anti-IgA antibodies of IgG class is described. This method is based on the use of purified polyclonal human serum IgA as the coating antigen and a commercial alkaline phosphatase-conjugated anti-human IgG as the detecting antibody. Nonspecific reactions were minimized by blocking vacant protein binding sites with bovine serum albumin and by using individual sample blanks. The IgA specificity of a positive antibody finding was confirmed by testing inhibition: pooled normal human serum inhibited the binding of specific antibodies by over 80%. The same degree of inhibition could also be demonstrated by a commercial myeloma IgA preparation and by the IgA used for coating but not by IgA-deficient serum (< 0.05 mg/l). On the basis of the mean anti-IgA antibody titre in EIA, a value of 12,000 arbitrary units of anti-IgA per litre (AU/l) was assigned to a patient serum used as standard in the assay. Anti-IgA results obtained by EIA and haemagglutination correlated well, which makes it possible to compare earlier HA results with those obtained now by EIA. The measuring range of the assay was 0.6-27 AU/l and the lowest quantifiable concentration 7 AU/l. The dilution requirement for serum was 1/16. The interassay coefficients of variation for control sera with antibody levels from 35 AU/l to 3770 AU/l varied from 9 to 12%.

The PAX5 gene: a linkage and mutation analysis in candidate human primary immunodeficiencies

The paired-box-containing (PAX) genes encode a family of developmentally regulated transcription factors that show a spatially and temporally restricted pattern of expression (for review see Strachan and Read 1994). Nine PAX genes have been identified so far in mouse and human and mapped to different chromosomal regions (Stapleton et al. 1993). Defects in several members of the PAX gene family have been associated with mouse developmental semidominant mutants such as undulated (Balling et al. 1988), Splotch (Epstein et al. 1991), and Small eye (Hill et al. 1991), and as well as human disorders such as aniridia (Jordan et al. 1992) and Waardenburgs syndrome (Tassabehji et al. 1992; Baldwin et al. 1992) that exhibit a dominant inheritance, In these Pax-associated phenotypes, only some expressing tissues are affected and to a variable extent. The PAX5 gene encodes a B-cell-specific activator protein (BSAP) that was found to bind to promoters of the CD19 gene (Kozm~ et al. 1992), B-cell-specific tyrosine kinase gene BLK (Zwollo et al. 1994), and to regulatory regions of the immunoglobulin (Ig) heavy chain locus, including sequences implicated in lg class switching (Liao et al. 1992; Singh and Birshtein 1993; Waters et al. 1989; Xu et al. 1992). The gene was mapped to mouse chromosome 4 using linkage analysis (Walther et al. 1991) and to human chromosomal region 9p13 using fluorescence in situ hybridization and somatic cell hybrid panel (Stapleton et al. 1993). Because targeted inactivation of the Pax5

Usefulness of maternal anti-HPA-1a antibody quantitation in predicting severity of foetomaternal alloimmune thrombocytopenia

To study the clinical usefulness of maternal anti‐HPA‐1a antibody levels in predicting severe foetomaternal alloimmune thrombocytopenia (FMAIT).