High soluble HLA-DQB2 levels in posttransplant serum are associated with kidney graft dysfunction.

HLA-DQB2 is a gene of limited polymorphism, with unknown function that presents at least two transcript variants: v1, which encodes the full-length beta-chain, and v2, which lacks exon 4 and could give rise to a soluble protein. We previously showed a strong correlation between high v2 expression in preimplantation biopsies (PIB) of kidneys from young (18- to 49-year olds) but not from old, deceased donors and 1-year posttransplant low (estimated glomerular filtration rate < 45 ml/min/1.73 m2 ) graft function (GF). In this study, we aimed to investigate the impact of posttransplant soluble HLA-DQB2 (sDQB2) serum levels, v1 expression in PIB, and recipient HLA-DQB2 rs7453920 A/G polymorphism on GF. sDQB2 was evaluated by enzyme-linked immunosorbent assay in sera from 114 recipients, collected at least 1 year (median 2.1 years) after transplantation. Higher sDQB2 levels were observed in recipients of kidneys from young, but not from old, donors that had a ≥30% decline in GF within 1 year after blood collection for sDQB2 determination. Among the 15 recipients of kidneys from young donors with sDQB2 ≥ 1.52 ng/ml, 40% presented a ≥30% decline in GF, whereas this occurred in none of the 43 recipients with lower sDQB2 levels (p = 0.007; OR: 36.5). Expression of HLA-DQB2 variant 1, measured by reverse transcription-polymerase chain reaction (RT-PCR) in 92 PIB from young or old donors, did not significantly differ between transplants with high or low 4-year GF. HLA-DQB2 rs7453920 single nucleotide polymorphism (SNP) frequencies did not significantly differ between recipients with low or high 4-year GF. We conclude that HLA-DQB2 variant 1 expression in PIB and recipient rs7453920 SNP polymorphism are not associated with graft outcome. On the other hand, the association, in transplants of kidneys from young donors, between high posttransplant serum sDQB2 levels and decline in GF is a very interesting finding that deserves a validation study in a larger cohort.

HLA-A homozygosis is associated with susceptibility to COVID-19.

The purpose of this single center retrospective study was to investigate the relationship between HLA and ABO polymorphisms and COVID-19 susceptibility and severity in kidney transplant recipients. It included 720 recipients who had COVID-19 and 1680 controls composed by recipients in follow-up who did not contact the transplantation center for COVID-19 symptoms, up to the moment of their inclusion in the study. HLA-A, -B, and -DRB1 allele groups and ABO frequencies were compared between recipients with COVID-19 (all cases, or separately mild/moderate and severe disease) and controls. The HLA association study was conducted in two case-control series and only associations that showed a p-value <0.05 in both series were considered. No HLA association regarding COVID-19 occurrence or severity met this criterion. Homozygosity at HLA-A locus was associated with COVID-19 susceptibility (odds ratio 1.4) but not severity. Blood groups A and O were associated with susceptibility and resistance to COVID-19, respectively. COVID-19 severity was associated only with older age and cardiac disease, in a multivariate analysis. We conclude that an influence of HLA on COVID-19 susceptibility is supported by the association with homozygosity at HLA-A locus but that there is no evidence for a role of any particular HLA-A, -B, or -DRB1 polymorphism. Thus, we suggest that what matters is the overall capability of an individual's HLA molecules to present SARS-CoV-2 peptides to T cells, a factor that might have a great influence on the breadth of the immune response.

Impact of Combinations of Donor and Recipient Ages and Other Factors on Kidney Graft Outcomes.

As the availability of kidneys for transplantation continues to be outpaced by its growing demand, there has been an increasing utilization of older deceased donors in the last decades. Considering that definition of factors that influence deceased donor kidney transplant outcomes is important for allocation policies, as well as for individualization of post-transplant care, the purpose of this study was determine the risks for death censored graft survival and for patient survival conferred by older age of the donor in the context of the age of the recipient and of risk factors for graft and/or patient survival. The investigation was conducted in a single-center cohort of 5,359 consecutive first kidney transplants with adult deceased donors performed on non-prioritized adult recipients from January 1, 2002, to December 31, 2017. Death censored graft survival and patient survival were lower in older donors, whereas graft survival was higher and patient survival was lower in old recipients. The analyses of combinations of donor and recipient ages showed that death censored graft survival was lower in younger recipients in transplants from 18 to 59-year old donors, with standard or extended criteria, but no difference in graft survival was observed between younger and older recipients when the donor was ≥ 60-year old. Patient survival was higher in younger recipients in transplants with younger or older donors. Two to six HLA-A,B,DR mismatches, when compared to 0-1 MM, conferred risk for death-censored graft survival only in transplants from younger donors to younger recipients. Pre-transplant diabetes conferred risk for patient survival only in 50-59-year old recipients, irrespectively, of the age of the donor. Time on dialysis ≥ 10 years was a risk factor for patient survival in transplants with all donor-recipient age combinations, except in recipients with ≥ 60 years that received a kidney from an 18-49-year old donor. In conclusion, the results obtained in this study underline the importance of analyzing the impact of the age of the donor taking into consideration different scenarios.

Heightened expression of HLA-DQB1 and HLA-DQB2 in pre-implantation biopsies predicts poor late kidney graft function.

BACKGROUND: Accurate pre-transplant prediction of late graft function remains an unmet need in kidney transplantation. The aim of this study was to evaluate HLA genes expression levels in pre-implantation biopsies (PIB) of deceased donor kidneys as markers for long-term graft outcome. METHODS: HLA genes expression analysis was initially performed using microarray data of 53 PIB, previously generated by our laboratory. The validation analysis was performed by real-time PCR in 116 PIB from an independent cohort. RESULTS: The microarray data showed association between high expression levels of HLA class II genes, especially HLA-DQB1 and -DQB2, in kidneys from young (18 to 49-year-old) donors and poor (eGFR < 45 mL/min/1.73 m2) 1- and 5-year graft function. A subsequent study in an independent cohort, in which only HLA-DQB2 expression was evaluated, validated the association between increased HLA-DQB2 expression in PIB of kidneys from young donors and poor 1-year graft function: expression levels ≥0.0025 relative units conferred an odds ratio of 22.5, with positive and negative predictive values of 71.4% and 90.0%, respectively. CONCLUSION: Heightened expression of HLA-DQB1 and -DQB2 in PIB are promising tools for pre-transplant risk assessment of poor late graft function in transplants with kidneys from 18 to 49-year-old donors.

Unexpected links reflect the noise in networks.

BACKGROUND: Gene covariation networks are commonly used to study biological processes. The inference of gene covariation networks from observational data can be challenging, especially considering the large number of players involved and the small number of biological replicates available for analysis. RESULTS: We propose a new statistical method for estimating the number of erroneous edges in reconstructed networks that strongly enhances commonly used inference approaches. This method is based on a special relationship between sign of correlation (positive/negative) and directionality (up/down) of gene regulation, and allows for the identification and removal of approximately half of all erroneous edges. Using the mathematical model of Bayesian networks and positive correlation inequalities we establish a mathematical foundation for our method. Analyzing existing biological datasets, we find a strong correlation between the results of our method and false discovery rate (FDR). Furthermore, simulation analysis demonstrates that our method provides a more accurate estimate of network error than FDR. CONCLUSIONS: Thus, our study provides a new robust approach for improving reconstruction of covariation networks. REVIEWERS: This article was reviewed by Eugene Koonin, Sergei Maslov, Daniel Yasumasa Takahashi.

Predicting delayed kidney graft function with gene expression in preimplantation biopsies and first-day posttransplant blood.

The purpose of this study was to investigate possible markers for predicting delayed graft function (DGF). To this end we analyzed, in pre-implantation biopsies (PIB) and in first-day post-Tx peripheral blood mononuclear cells (PBMC), the expression of five genes (ACSL4, CUBN, DEFB1, FABP3, GK) through real-time TaqMan PCR assays. These genes were selected from a large scale gene expression study in PIB. DEFB1, FABP3 and GK expression levels in PIB were lower in cases with DGF and, in a multivariate analysis which included these genes and clinical variables, only FABP3 expression remained independently associated with DGF. FABP3 expression lower than -1.32 units of relative expression conferred an odds ratio for DGF of 41.1. Compared to the PBMC of recipients without DGF, recipients with prolonged DGF (pDGF) had lower ACSL4 and higher DEFB1 expression levels. In a multivariate analysis, including PBMC gene expression levels of ACSL4, DEFB1 and TLR4 (data from a previous study with the same patients) and clinical variables, only TLR4 remained independently associated with pDGF. In summary, this study revealed FABP3 expression in PIB as a marker for DGF and disclosed new genes possibly involved in the pathogenesis of DGF.

Gene network reconstruction reveals cell cycle and antiviral genes as major drivers of cervical cancer.

Although human papillomavirus was identified as an aetiological factor in cervical cancer, the key human gene drivers of this disease remain unknown. Here we apply an unbiased approach integrating gene expression and chromosomal aberration data. In an independent group of patients, we reconstruct and validate a gene regulatory meta-network, and identify cell cycle and antiviral genes that constitute two major subnetworks upregulated in tumour samples. These genes are located within the same regions as chromosomal amplifications, most frequently on 3q. We propose a model in which selected chromosomal gains drive activation of antiviral genes contributing to episomal virus elimination, which synergizes with cell cycle dysregulation. These findings may help to explain the paradox of episomal human papillomavirus decline in women with invasive cancer who were previously unable to clear the virus.

Construct and Compare Gene Coexpression Networks with DAPfinder and DAPview.

BACKGROUND: DAPfinder and DAPview are novel BRB-ArrayTools plug-ins to construct gene coexpression networks and identify significant differences in pairwise gene-gene coexpression between two phenotypes. RESULTS: Each significant difference in gene-gene association represents a Differentially Associated Pair (DAP). Our tools include several choices of filtering methods, gene-gene association metrics, statistical testing methods and multiple comparison adjustments. Network results are easily displayed in Cytoscape. Analyses of glioma experiments and microarray simulations demonstrate the utility of these tools. CONCLUSIONS: DAPfinder is a new friendly-user tool for reconstruction and comparison of biological networks.

Differential expression of new LTA splice variants upon lymphocyte activation.

Lymphotoxin alpha (LTA) is a member of the TNF cytokine superfamily, produced principally by lymphocytes. It plays an important role in immune and inflammatory responses. Many TNF superfamily members have functionally important isoforms generated by alternative splicing but alternative splicing of LTA has never been studied. The known LTA protein is encoded by a transcript containing four exons. Here we report seven new LTA splice variants, three of them evolutionary conserved. We demonstrate their presence in cytoplasmic RNA suggesting that they could be translated into new LTA isoforms. We observed that their expression is differentially regulated upon activation of peripheral blood mononuclear cells and lymphocyte subpopulations (CD4+, CD8+, and CD19+). Our data suggest that the new LTA splice variants might play a role in the regulation of the immune response.

A novel strategy for defining haplotypes by selective depletion using restriction enzymes.

Various single nucleotide polymorphisms (SNPs) have been investigated regarding association with gene expression levels or human diseases. Although different SNPs within one gene are frequently analyzed individually, it is highly probable that in the majority of the cases, a precise combination of SNP alleles, i.e., haplotype, determines a functional trait. Methods commonly used for haplotype determination, involving studies in families, cloning, or somatic cell hybrids, are expensive and time-consuming. We herein suggest a novel and simple strategy for haplotype determination, involving selective haplotype depletion with a restriction enzyme, followed by sequencing. We studied 11 LTA gene polymorphisms in 102 Brazilian individuals, and we applied this novel methodology for haplotyping 67 out of 70 LTA heterozygous individuals. We concluded that the method is rapid and efficient, and, as it includes only simple and widespread-used techniques, it could be used in most of the laboratories without further investment in equipments. The wider usage of haplotyping could be important to clarify contradictory results frequently observed among studies that focus on a single SNP.