HLA allele-specific expression : Methods, disease associations, and relevance in hematopoietic stem cell transplantation

Varying HLA allele-specific expression levels are associated with human diseases, such as graft versus host disease (GvHD) in hematopoietic stem cell transplantation (HSCT), cytotoxic T cell response and viral load in HIV infection, and the risk of Crohn's disease. Only recently, RNA-based next generation sequencing (NGS) methodologies with accompanying bioinformatics tools have emerged to quantify HLA allele-specific expression replacing the quantitative PCR (qPCR) -based methods. These novel NGS approaches enable the systematic analysis of the HLA allele-specific expression changes between individuals and between normal and disease phenotypes. Additionally, analyzing HLA allele-specific expression and allele-specific expression loss provide important information for predicting efficacies of novel immune cell therapies. Here, we review available RNA sequencing-based approaches and computational tools for NGS to quantify HLA allele-specific expression. Moreover, we explore recent studies reporting disease associations with differential HLA expression. Finally, we discuss the role of allele-specific expression in HSCT and how considering the expression quantification in recipient-donor matching could improve the outcome of HSCT.Peer reviewe

Single Cell Trapping by Superhydrophobic/Superhydrophilic Microarrays

Single cell trapping is demonstrated on superhydrophobic-superhydrophilic patterned microarrays. Superhydrophilic spots that are twice the size of the cells to be trapped are found to be optimal for obtaining single cell trapping. The single cell trapping is based on size exclusivity instead of heavy dilution of the cell suspension and relying on Poisson statistics. The superhydrophobic areas of the array are found to be very resistant toward unwanted adhesion of the cells and thus cleaning steps are not needed after deposition. Based on these properties, 20 mu m superhydrophilic spots are utilized for trapping two types of immune cells, primary peripheral blood mononuclear cells (PBMCs) and THP-1 cells. The cells are trapped from a 10 mu l cell suspension droplet dragged across the surface with different velocities. Single cell trapping efficiencies in the range of 10-30% are shown based on the cell type, the spot type, the seeding velocity and the cell suspension concentration.Peer reviewe

Coeliac disease re-screening among once seronegative at-risk relatives : A long-term follow-up study

Background Serological screening of the relatives of coeliac disease patients is widely endorsed. However, the need for and the optimal timing of possible re-testing of once seronegative at-risk individuals for coeliac disease remain unclear. Objective We investigated this issue by inviting a large cohort of previously screening-negative relatives of patients with coeliac disease to participate in a follow-up study. Methods Altogether 599 relatives of coeliac disease index patients not diagnosed with coeliac disease in a screening study carried out in 2006-2010 were asked about possible later diagnosis or re-tested with coeliac disease autoantibodies in 2017-2021. Besides incidence, the possible impact of various patient-related clinical factors and HLA haplotype on the later diagnosis or screening positivity was examined. Results Fifteen (2.5%) relatives were either diagnosed with a coeliac disease (n = 8) during the follow-up period or were found to be screening-positive in the re-testing (n = 7), giving a combined annual incidence of 221/100,000 person-years in all relatives and 336/100,000 among those carrying coeliac disease-associated HLA DQ2/DQ8. The new cases more often carried the high-risk (DQ2.5/2.5 or DQ2.5/2.2; 35.7% vs. 7.4%, respectively, p < 0.001) HLA and were younger at initial screening (23.3 vs. 40.5 years, p = 0.028) and - in spite of a negative screening outcome - had higher median transglutaminase antibody level in the first study than those not affected. There were no significant differences between the affected and non-affected relatives in other demographic data, degree of kinship with the index, current symptoms or frequency of chronic co-morbidities. Conclusion The incidence rate for later coeliac disease diagnosis or new seropositivity in relatives who had been tested once was 221/100,000 person-years in all and 336/100,000 among those carrying at-risk HLA genetics after similar to 10 years of follow-up. HLA-typing could help to target a subgroup of relatives who would benefit most from re-testing.Peer reviewe

Epigenetic and transcriptional analysis supports human regulatory T cell commitment at the CD4+CD8+thymocyte stage

The natural CD25 + FOXP3 + regulatory T cell (Treg) population is generated as a distinct lineage in the thymus, but the details of Treg development in humans remain unclear, and the timing of Treg commitment is also contested. Here we have analyzed the emergence of CD25 + cells at the CD4 + CD8 + double positive (DP) stage in the human thymus. We show that these cells share T cell receptor repertoire with CD25 + CD4 single-positive thymocytes, believed to be committed Tregs. They already have a fully demethylated FOXP3 enhancer region and thus display stable expression of FOXP3 and the associated Treg phenotype. Transcriptome analysis also grouped the DP CD25 + and CD4 CD25 + thymocytes apart from the CD25 - subsets. Together with earlier studies, our data are consistent with human Treg commitment already at the DP thymocyte stage. We suggest that the most important antigens and signals necessary for human Treg differentiation may be found in the thymic cortex.Peer reviewe

Deep infiltrating endometriosis affecting the urinary tract-surgical treatment and fertility outcomes in 2004-2013

Peer reviewe

Differences Between Familial and Sporadic Celiac Disease

Background It is not known if genetic background, characteristics at diagnosis, physical and psychological well-being, and adherence to a gluten-free diet are comparable between patients with familial or sporadic celiac disease. These issues were investigated in a follow-up study. Methods Altogether 1064 patients were analyzed for celiac disease-associated serology, predisposing HLA-DQ, and non-HLA genotypes. Medical data were collected from patient records and supplementary interviews. Current symptoms and quality of life were further evaluated with the Gastrointestinal Symptom Rating Scale (GSRS), the Psychological General Well-Being questionnaire (PGWB), and Short Form 36 (SF-36) questionnaires. Results Familial and sporadic groups differed (P <0.001) in the reason for diagnosis and clinical presentation at diagnosis, familial patients being more often screen-detected (26% vs. 2%,P <0.001) and having less often gastrointestinal (49% vs. 69%) and severe symptoms (47% vs. 65%). The groups were comparable in terms of histological damage, frequency of malabsorption, comorbidities, childhood diagnoses, and short-term treatment response. At the time of the study, familial cases reported fewer symptoms (21% vs. 30%,P = 0.004) and lower prevalence of all (78% vs. 86%,P = 0.007), neurological (10% vs. 15%,P = 0.013), and dermatological (9% vs. 17%,P = 0.001) comorbidities. Dietary adherence and GSRS scores were comparable, but familial cases had better quality of life according to PGWB and SF-36. High-risk genotype HLA-DQ2.5/DQ2.5 was more frequent among familial cases, and four non-HLA SNPs were associated with familial celiac disease. Conclusions Despite the greater proportion of high-risk genotypes, familial cases had milder symptoms at presentation than did sporadic cases. Worse experience of symptoms and poorer quality of life in sporadic disease indicate a need for intensified support.Peer reviewe

First-degree Relatives of Celiac Disease Patients Have Increased Seroreactivity to Serum Microbial Markers

Risk of celiac disease (CD) is increased in relatives of CD patients due to genetic and possible environmental factors. We recently reported increased seropositivity to anti-Saccharomyces cerevisiae (ASCA), Pseudomonas fluorescens-associated sequence (anti-I2) and Bacteroides caccae TonB-linked outer membrane protein (anti-OmpW) antibodies in CD. We hypothesized these markers also to be overrepresented in relatives. Seropositivity and levels of ASCA, anti-I2 and anti-OmpW were compared between 463 first-degree relatives, 58 untreated and 55 treated CD patients, and 80 controls. CD-associated human leukocyte antigen (HLA)-haplotypes and transglutaminase (tTGab) and endomysium (EmA) antibodies were determined. One or more of the microbial antibodies was present in 75% of relatives, 97% of untreated and 87% of treated CD patients and 44% of the controls. The relatives had higher median ASCA IgA (9.13 vs. 4.50 U/mL, p < 0.001), ASCA IgG (8.91 vs. 5.75 U/mL, p < 0.001) and anti-I2 (absorbance 0.74 vs. 0.32, p < 0.001) levels than controls. There was a weak, positive correlation between tTGab and ASCA (r = 0.31, p < 0.001). Seropositivity was not significantly associated with HLA. To conclude, seropositivity to microbial markers was more common and ASCA and anti-I2 levels higher in relatives of CD patients than controls. These findings were not associated with HLA, suggesting the role of other genetic and environmental factors

First-degree Relatives of Celiac Disease Patients Have Increased Seroreactivity to Serum Microbial Markers

Risk of celiac disease (CD) is increased in relatives of CD patients due to genetic and possible environmental factors. We recently reported increased seropositivity to anti-Saccharomyces cerevisiae (ASCA), Pseudomonas fluorescens-associated sequence (anti-I2) and Bacteroides caccae TonB-linked outer membrane protein (anti-OmpW) antibodies in CD. We hypothesized these markers also to be overrepresented in relatives. Seropositivity and levels of ASCA, anti-I2 and anti-OmpW were compared between 463 first-degree relatives, 58 untreated and 55 treated CD patients, and 80 controls. CD-associated human leukocyte antigen (HLA)-haplotypes and transglutaminase (tTGab) and endomysium (EmA) antibodies were determined. One or more of the microbial antibodies was present in 75% of relatives, 97% of untreated and 87% of treated CD patients and 44% of the controls. The relatives had higher median ASCA IgA (9.13 vs. 4.50 U/mL, p < 0.001), ASCA IgG (8.91 vs. 5.75 U/mL, p < 0.001) and anti-I2 (absorbance 0.74 vs. 0.32, p < 0.001) levels than controls. There was a weak, positive correlation between tTGab and ASCA (r = 0.31, p < 0.001). Seropositivity was not significantly associated with HLA. To conclude, seropositivity to microbial markers was more common and ASCA and anti-I2 levels higher in relatives of CD patients than controls. These findings were not associated with HLA, suggesting the role of other genetic and environmental factors

Influence of HLA-DQ2.5 Dose on Clinical Picture of Unrelated Celiac Disease Patients

The clinical phenotype of celiac disease varies considerably among patients and the dosage of HLA-DQ2.5 alleles has been suggested to be a contributing factor. We investigated whether HLA-DQ2.5 allele dosage is associated with distinct clinical parameters at the time of diagnosis and with patients’ response to a gluten-free diet. The final cohort included 605 carefully phenotyped non-related Finnish celiac disease patients grouped as having 0, 1 or 2 copies of HLA-DQ2.5. Clinical data at the time of diagnosis and during gluten-free diet were collected systematically from medical records and supplementary interviews. An increasing HLA-DQ2.5 dose effect was detected for celiac disease antibody positivity at diagnosis (p = 0.021) and for the presence of any first-degree relatives with celiac disease (p = 0.011 and p = 0.031, respectively). Instead, DQ2.5-negative patients were suffering most often from classical symptoms at diagnosis (p = 0.007 between HLA groups). In addition, during follow-up they were most often symptomatic despite a gluten-free diet (p = 0.002 between groups). Our results thus suggest that increasing HLA-DQ2.5 dose only has a minor effect on the clinical picture of celiac disease. However, HLA-DQ2.5-negative patients should not be overlooked in clinical practice and particular attention should be paid to this patient group during gluten-free diet