Donor-specific HLA antibodies in predicting crossmatch outcome: Comparison of three different laboratory techniques.

The virtual crossmatch, which is based on single antigen bead technology, is used in the prediction of crossmatch results. However, this assay differs in sensitivity and specificity from crossmatch methods. In our study, the results of physical crossmatches, performed with three different methods, were assessed against virtual crossmatch results. The aim was to determine the potential cut-off values for donor specific antibodies (DSA) that would predict the crossmatch results obtained by different methods. The results of different crossmatch techniques were correlated with the virtual crossmatch. The receiver operating characteristic (ROC) analysis revealed the Flow cytometric crossmatch (FCXM) and Luminex crossmatch (LXM) to be the most accurate, with area under curve (AUC) values of 0.861 and 0.805, respectively. While we found that the virtual crossmatch correlated well with all the crossmatch results, FCXM produced the best results (83% of the DSA detected). LXM outperformed the other tests in terms of the accuracy in separating class II DSA.

Description of four new HLA alleles in the Finnish population: A*03:283N, A*68:167, C*03:327, C*03:361.

New HLA alleles found in the Finnish population: A*03:283N, A*68:167, C*03:327 and C*03:361.

Platelet donor selection for HLA-immunised patients; the impact of donor-specific HLA antibody levels.

BACKGROUND: Approximately 20% of patients with a recurrently poor platelet transfusion increment show human leukocyte antigen (HLA) alloantibodies. The aim of this study was to analyse the impact of mean fluorescence intensity (MFI) levels of donor-specific HLA antibodies and the feasibility of the HLAMatchmaker algorithm in donor selection. STUDY DESIGN AND METHODS: A total of 270 HLA-typed platelet transfusion responses of 40 patients were included in the study. The patients' immunisation status was determined with Luminex-based methods, and HLA alloantibody strengths were defined as the MFI. For the Matchmaker eplet matching, the HLA-ABC Eplet Matching Version 2.1 was used. RESULTS: In 62% of the 270 transfusions, HLA antibodies against the transfused platelets were present, with a median cumulative MFI level of 2026 (range: 299-29 203). In multivariate analysis, a cumulative MFI level higher than 1000 emerged as an independent risk factor for a poor platelet transfusion increment, along with infection and the age of the product. CONCLUSION: The HLAMatchmaker algorithm alone is not a sufficient tool for donor selection. Donor selection based primarily on the levels of donor-specific HLA antibodies is a preferable practice.

Human leucocyte antigens B*08, DRB1*03 and DRB1*13 are significantly associated with autoimmune liver and biliary diseases in Finnish children.

AIM: The human leucocyte antigen (HLA) allele and haplotype frequencies of the Finnish population are unique because of the restricted and homogenous gene population. There are no published data on HLA genotype associations in paediatric autoimmune liver diseases in Scandinavia. This study characterised the HLA genotypes of children with autoimmune liver or biliary disease in Finland. METHODS: The study cohort comprised 19 paediatric patients (13 female) aged three years to 15 years treated for autoimmune liver or biliary disease at the Children's Hospital, Helsinki University Hospital, between 2000 and 2011, and followed up for four years and three months to 14.6 years. We genotyped HLA-B and HLA-DRB1 in the children, and the HLA antigen frequencies were compared with 19 807 records from the Finnish Bone Marrow Donor Registry. RESULTS: All paediatric patients with autoimmune liver or biliary disease had either autoimmune HLA haplotype B*08;DRB1*03 or DRB1*13. These were significantly more common among patients with autoimmune hepatitis, primary sclerosing cholangitis and autoimmune hepatitis/primary sclerosing cholangitis overlap syndrome than the Finnish control population. HLA RB1*04 was not found in the study cohort. CONCLUSION: Our study found that B*08, DRB1*03 and DRB1*13 were significantly associated with autoimmune liver and biliary diseases in Finnish paediatric patients.

Conflicting HLA assignment by three different typing methods due to the apparent loss of heterozygosity in the MHC region.

Loss of heterozygosity (LOH) has been reported to cause false human leukocyte antigen (HLA) homozygous typing results in pre-transplant patients suffering from haematological malignancies, who in fact are HLA heterozygous. This poses a challenge for histocompatibility testing, as a stem cell graft from a genuinely HLA homozygous donor to a mistyped patient may lead to acute life-threatening graft-vs-host disease. LOH in the HLA region on chromosome 6 is known to be quite common in solid tumours, helping malignant cells to escape T-cell surveillance, but the incidence in haematological malignancies is less well known and the estimates vary. Here we report LOH in the HLA region of five patients with haematological malignancy. We found considerable differences in sensitivity between the three different HLA typing methods used in our laboratory: SSP was clearly the most sensitive method for detecting the lost haplotype, followed by rSSO, while SBT was the least sensitive technique. A subsequent, retrospective genotyping of 65 HLA homozygous haematological patients by SSP method showed no mistyped LOH cases in our laboratory in the past 10 years. The frequency of HLA homozygosity was found to be similar between haematological patients and control groups. It is important for an HLA laboratory to be aware of the differences in various HLA typing techniques' sensitivity for detecting an under-represented haplotype between HLA typing techniques when genotyping patients with haematological diseases. It is advisable for HLA laboratories to have at least two different methods with different sensitivities in their repertoire to be able to retype samples when a false homozygous result is suspected.

HLA associations with tubulointerstitial nephritis with or without uveitis in Finnish pediatric population: a nation-wide study.

The human leukocyte antigen (HLA) genotype has been shown to associate with tubulointerstitial nephritis (TIN) and tubulointerstitial nephritis with uveitis syndrome (TINU). The association of HLA genes with TIN was examined in this nation-wide study. HLA genotyping was performed in 31 pediatric patients with biopsy-proven TIN. All patients were examined by an ophthalmologist to diagnose possible uveitis. Class II HLA genotypes of TIN patients were compared with the Finnish reference population. We found a significant association between the HLA alleles DQA1*04:01 [risk ratio (RR) 5.0, 95% confidence interval (CI) 2.0-11.2], DQB1*04:02 (RR 2.7, 95% CI 1.4-5.3), and DRB1*08 (RR 3.8, 95% CI 1.5-8.4) and TIN. Uveitis was found in 20/31 (64.5%) patients. HLA genotyping of the TINU patients showed additional risk HLA alleles: DQA1*01:04 (RR 6.1, 95% CI 1.5-17.8), and DRB1*14 (RR 8.2, 95% CI 2.2-22.1). The alleles DQA1*01:04 (RR 8.8, 95% CI 2.2-26.5), DQA1*04:01 (RR 3.2, 95% CI 1.2-7.3), and DRB1*14 (RR 12.0, 95% CI 3.2-33.0) were more frequent in patients with TIN and chronic uveitis than in reference population. The HLA class II haplotype DQA1*04:01/DQB1:04:02/DRB1*08 was the most common combination in our study population (58.1%). None of the patients had haplotype DQA1*04:01/DQB1*06:02/DRB1*15, which is common in Finland. HLA genotype did not predict the renal outcome. We found a strong association between certain HLA genotypes both in TIN and TINU patients. The TIN/TINU-associated HLA alleles appear to vary depending on study population.

HLA antigen, allele and haplotype frequencies and their use in virtual panel reactive antigen calculations in the Finnish population.

The human leukocyte antigen (HLA) antigen, allele and haplotype frequencies of the Finnish population are quite unique because of a rather restricted and homogeneous gene pool. This has a strong influence on finding suitable donors for transplant patients; hence knowledge about the HLA frequencies of the patient population is essential. Here we report the HLA antigen frequencies for a large population sample and show high resolution HLA allele frequencies for 11 loci, including the rarely typed DPA1 and DQA1 loci. Furthermore, the most common Finnish high resolution haplotypes are presented for five HLA loci. The study shows that there are fewer HLA haplotypes in the Finnish population compared with mixed populations, and the common Finnish HLA haplotypes are more frequent. Using HLA antibody identification and panel reactive antibody calculations we show that a virtual population-specific panel, combined with single antigen testing, gives a more accurate and reliable estimate of the reactivity of the recipient serum against potential solid organ donors within the Finnish population. The results can be directly used to improve donor search for patients waiting for stem cell transplantation and to allocate highly immunised patients accurately to acceptable mismatch programs.

A bioinformatics approach to ascertaining the rarity of HLA alleles.

A project of the 15th International Histocompatibility Workshop examined the rarity of human leukocyte antigen (HLA) alleles. A section was constructed in the website, www.allelefrequencies.net to contain this data from different sources. A mechanism to search the data was implemented for use by any individual.

The major horse allergen Equ c 1 contains one immunodominant region of T cell epitopes.

BACKGROUND: Despite the fact that most significant mammalian respiratory allergens are lipocalin proteins, information on the human T cell reactivity to these allergenic proteins is largely missing. OBJECTIVE: Knowing the T cell epitopes in allergens is a prerequisite for developing novel preparations for allergen immunotherapy. METHODS: Specific T cell lines were generated with recombinant Equ c 1 from the peripheral blood mononuclear cells (PBMCs) of 10 horse-allergic subjects. For determining T cell epitopes, the lines were stimulated with 16mer synthetic Equ c 1 peptides overlapping by 14 amino acids. The binding capacity of Equ c 1 peptides to human leucocyte antigen class II molecules was determined by the competitive ELISA. RESULTS: The major horse allergen Equ c 1 resembles two other lipocalin allergens, the major cow allergen Bos d 2 and the major dog allergen Can f 1, in that it is weakly stimulatory for the PBMCs of sensitized subjects. Moreover, the T cell epitopes of Equ c 1 are clustered in a few regions along the molecule, as is the case with Bos d 2 and Can f 1. Similar to Bos d 2, Equ c 1 contains one immunodominant epitope region at the carboxy-terminal end of the molecule. The T cell lines of eight horse-allergic subjects out of 10 showed strong reactivity to one or both of the two overlapping peptides, p143-158 and p145-160, in this region. The region probably contains two overlapping epitopes. CONCLUSION: The 18mer peptide p143-160 from the immunodominant region of Equ c 1 is a potential candidate for the peptide-based immunotherapy of horse-sensitized subjects.

Characterization a novel HLA-B40 allele with serological Bw4 motif, HLA-B*4047, in the Finnish population and confirmation of B*270503 allele.

We describe a novel HLA-B*40 allele assigned as B*4047*. The B*4047 allele was detected in a Finnish patient awaiting kidney transplantation. The patient had a "short" B60-like serological specificity with Bw4 association. After sequencing, the B*4047 allele was found to be identical to B*4001, except having five amino acid changes in exon 2, including the entire motif corresponding to Bw4 and w6 specificity. As a result of recombination or gene conversion, B*4047 has the Bw4 motif instead of expected Bw6. Screening of B40 alleles in the Finnish population revealed no other cases with this pattern, suggesting that this allele is rare. The sequence of B*270503 presented here provides the complete sequence for exons 2 and 3 for this allele. B*270503 allele differs from B*270502 by a single synonymous nucleotide substitution at non-variable position 489 in exon 3.

Wnts as kidney tubule inducing factors.

Since the discovery that inductive tissue interactions regulate nephrogenesis, one of the aims has been to identify the molecules that mediate this induction. The small size of embryonic tissue has limited the possibilities to identify the inducers biochemically, even though such efforts were directed to study, e.g. neural induction (for a comprehensive review, Saxén and Toivonen, Primary embryonic induction, Academic Press, London, 1962). The rapid progress in molecular biology made it possible to identify genes from minute amounts of tissue and provided techniques to generate recombinant proteins to assay their action in classic experimental systems. This led to the identification of some signals that are involved in primary and secondary inductive interactions during embryogenesis. Here, we will review evidence suggesting that secreted signaling molecules from the Wnt gene family mediate kidney tubule induction.