MicroRNAs as Potential Graft Rejection or Tolerance Biomarkers and Their Dilemma in Clinical Routines Behaving like Devilish, Angelic, or Frightening Elements.

Allograft rejection is a widespread complication in allograft recipients with chronic kidney disease. Undertreatment of subclinical and clinical rejection and later post-transplant problems are caused by an imperfect understanding of the mechanisms at play and a lack of adequate diagnostic tools. Many different biomarkers have been analyzed and proposed to detect and monitor these crucial events in transplant outcomes. In this sense, microRNAs may help diagnose rejection or tolerance and indicate appropriate treatment, especially in patients with chronic allograft rejection. As key epigenetic regulators of physiological homeostasis, microRNAs have therapeutic potential and may indicate allograft tolerance or rejection. However, more evidence and clinical validation are indispensable before microRNAs are ready for clinical prime time.

Genotypic Frequencies of Mutations Associated with Alpha-1 Antitrypsin Deficiency in Unrelated Bone Marrow Donors from the Murcia Region Donor Registry in the Southeast of Spain.

Alpha-1 antitrypsin (AAT1) deficiency (AAT1D) is an inherited disease with an increased risk of chronic obstructive pulmonary disease (COPD), liver disease, and skin and blood vessel problems. AAT1D is caused by mutations in the SERPINE1 gene (Serine Protease Inhibitor, group A, member 1). Numerous variants of this gene, the Pi system, have been identified. The most frequent allelic variants are Pi*M, Pi*S, and Pi*Z. The development of COPD requires both a genetic predisposition and the contribution of an environmental factor, smoking being the most important. Studies on this deficiency worldwide are very scarce, and it is currently considered a rare disease because it is underdiagnosed. The aim of this study was to analyze the genotypic frequencies of mutations associated with AAT1 deficiency in unrelated bone marrow donors from the donor registry of the Region of Murcia in southeastern Spain due to the high risk of presenting with different pathologies and underdiagnosis in the population. A total of 112 DNA-healthy voluntary unrelated bone marrow donors from different parts of the Region of Murcia were analyzed retrospectively. AAT1 deficiency patient testing involved an automated biochemical screening routine. The three main variants, Pi*M, Pi*Z, and Pi*S, were analyzed in the SERPINE1 gene. Our results showed a frequency of 3.12% of the Pi*Z (K342) mutation in over 224 alleles tested in the healthy population. The frequency of Pi*S (V264) was 11.1%. The frequency of the haplotype with the most dangerous mutation, EK342 EE264, was 4.46%, and the frequency of EK342 EV264 was 1.78% in the healthy population. Frequencies of other EE342 EV264-mutated haplotypes accounted for 18.7%. As for the EE342 VV264 haplotype, 0.89% of the total healthy population presented heterozygous for the EV264 mutation and one individual presented homozygous for the VV264 mutation. In conclusion, the frequencies of Pi mutations in the healthy population of the Region of Murcia were not remarkably different from the few studies reported in Spain. The genotype and haplotype frequencies followed the usual pattern. Health authorities should be aware of this high prevalence of the Pi*S allelic variant and pathological genotypes such as Pi*MZ and Pi*SZ in the healthy population if they consider screening the smoking population.

Analysis of null deletion polymorphism of glutathione S-transferase theta (GSTT-1), associated with anti-GSTT-1 antibodies development in transplantation.

Glutathione S-transferase theta 1 (GSTT1) is an enzyme involved in phase II biotransformation processes and a member of a multigene family of detoxifying and clearing reactive oxygen species. GSTT1 is polymorphic like other biotransforming enzymes, allowing variability in hepatic conjugation processes. Immunological recognition of the GSTT1 alloantigen, as evidenced by donor-specific antibodies formation, has previously been observed in recipients lacking GSTT1 protein (called GSTT1-, GSTT*0, null phenotype or homozygous for the GSTT1 deletion) who receive liver or kidney transplants from GSTT1+ donors and is a risk factor for the development of de novo hepatitis following liver transplants from a GSTT1 expressing donor. Antibodies against GSTT1 are demonstrated in patients who are GSTT1 null and received a transplant from a GSTT1+ donor. Understanding the local population frequency of the GSTT1 deletion is of value in understanding the potential clinical risk of developing post-transplant complications, which can be attributed to the nonexpression of GSTT1. A population of 173 healthy donors of the Murcia Region in Southeast Spain was evaluated for a null allele of GSTT1 (n = 173). DNA was extracted, and GSTT-1 null allele detection was performed by real-time polymerase chain reaction. The frequency of the null GSTT1 genotype (nonexpression or deletion of the homozygous polymorphism of the GSTT1 protein) was 17.9% (n = 31 null allele GSTT1/173 total individuals). Our data suggest that the frequency of null GSTT1 mutations in our population in Southeast Spain is 17.9%, lower than in other Caucasoid populations. This would convert our recipient population into more susceptible to nonlocal potential organ donors and less susceptible to local donors. All recipients bearing this GSTT1 deletion homozygous would be without the protein and triggering an alloantigen in the case of transplantation with a donor without deletion.

Early Cytomegalovirus Reactivation in Renal Recipients Is Associated with High Levels of B Cell Maturation Antigen Transcript Expression Prior to Transplantation.

Cytomegalovirus (CMV) infection is the most frequent infection episode in kidney transplant (KT) recipients. Reactivation usually occurs in the first three months after transplantation and is associated with higher cellular and/or antibody-mediated rejection rates and poorer graft performance. CMV induces the expression of BAFF (B-cell-activating factor, a cytokine involved in the homeostasis of B cells), which communicates signals for survival and growth to B cells and virus-specific plasma cells via the R-BAFF (BAFF receptor), TACI (the calcium modulator, the cyclophilin ligand interactor), and BCMA (B cell maturation antigen) receptors. These molecules of the BAFF system have also been suggested as biomarkers for the development of alloantibodies and graft dysfunction. This prospective study included 30 CMV-IgG seropositive KT recipients. The expression levels of the genes BAFF-R, transmembrane activator and CAML interactor (TACI), and B cell maturation antigen (BCMA) in peripheral blood leukocytes (PBL) pre-KT were determined using qPCR. qPCR was also used to monitor CMV reactivation in the first three months following KT. The remainder of the KT recipients were classified as CMV- reactivation, and those with more than 500 copies/mL in at least one sample were classified as CMV+ reactivation. There were no discernible variations in the BAFF-R and TACI transcript expression levels. In the CMV+ group, we examined the relationship between the transcript levels and peak viremia. Peak viremia levels and BCMA transcript levels showed a strong correlation. BAFF-R and TACI expressions showed no measurable differences. In patients with early CMV reactivation, high BCMA receptor expression was associated with increased plasmablast, lymphocyte B cell class-switched levels (LBCS), and viral load. Our findings demonstrate that pre-KT BCMA transcript levels increased in KT recipients with early CMV reactivation. These transcript levels positively correlate with peak viremia and weakly with plasmablast and LBCS levels in PBLs.

Monitoring of Serological, Cellular and Genomic Biomarkers in Transplantation, Computational Prediction Models and Role of Cell-Free DNA in Transplant Outcome.

The process and evolution of an organ transplant procedure has evolved in terms of the prevention of immunological rejection with the improvement in the determination of immune response genes. These techniques include considering more important genes, more polymorphism detection, more refinement of the response motifs, as well as the analysis of epitopes and eplets, its capacity to fix complement, the PIRCHE algorithm and post-transplant monitoring with promising new biomarkers that surpass the classic serum markers such as creatine and other similar parameters of renal function. Among these new biomarkers, we analyze new serological, urine, cellular, genomic and transcriptomic biomarkers and computational prediction, with particular attention to the analysis of donor free circulating DNA as an optimal marker of kidney damage.

Evaluating the Link between BAFF System Gene Expression and Acute Rejection Development in Kidney Transplantation.

B-cell activating factor (BAFF) system signaling is critical for B-cell homeostasis, effector functions, and tolerance maintenance in transplants, but it has not been studied in kidney transplant recipients (KTRs). The aim was to analyze the changes in BAFF system expression in KTRs with/without acute rejection (AR/NAR). The BAFF system expression was analyzed by qPCR in 40 KTRs. A meta-analysis of BAFF system expression and histological renal damage was identified by the Chronic Allograft Damage Index (CADI) and performed from the GEO database. Proliferation-inducing ligand (APRIL) expression increased at three- and six-months post-KT (p = 0.014 and p < 0.001). B-cell maturation antigen (BCMA) expression increased at six-months post-KT (p = 0.038). BAFF expression remained stable in NAR-KTRs, but was increased in CADI concerning the No-CADI group at one year (p = 0.008). BCMA expression increased in the CADI group at one- (p = 0.001) and six-years post-KT (p = 0.024). At three months, the transmembrane activator and calcium modulator interactor (TACI) gene significantly elevated KTRs with DSAs (donor-specific antibody; p = 0.034). KTRs with DSAs significantly increase the B-cell activating factor receptor (R-BAFF; p = 0.021) and TACI (p = 0.018) between pre- and three-month post-KT. Changes in the expression of the BAFF system increase during post-KTR in the development of AR and chronic allograft damage, and could be an important pathological tool to detect and prevent kidney graft outcomes.

Computational Prediction of Biomarkers, Pathways, and New Target Drugs in the Pathogenesis of Immune-Based Diseases Regarding Kidney Transplantation Rejection.

Background: The diagnosis of graft rejection in kidney transplantation (KT) patients is made by evaluating the histological characteristics of biopsy samples. The evolution of omics sciences and bioinformatics techniques has contributed to the advancement in searching and predicting biomarkers, pathways, and new target drugs that allow a more precise and less invasive diagnosis. The aim was to search for differentially expressed genes (DEGs) in patients with/without antibody-mediated rejection (AMR) and find essential cells involved in AMR, new target drugs, protein-protein interactions (PPI), and know their functional and biological analysis. Material and Methods: Four GEO databases of kidney biopsies of kidney transplantation with/without AMR were analyzed. The infiltrating leukocyte populations in the graft, new target drugs, protein-protein interactions (PPI), functional and biological analysis were studied by different bioinformatics tools. Results: Our results show DEGs and the infiltrating leukocyte populations in the graft. There is an increase in the expression of genes related to different stages of the activation of the immune system, antigenic presentation such as antibody-mediated cytotoxicity, or leukocyte migration during AMR. The importance of the IRF/STAT1 pathways of response to IFN in controlling the expression of genes related to humoral rejection. The genes of this biological pathway were postulated as potential therapeutic targets and biomarkers of AMR. These biological processes correlated showed the infiltration of NK cells and monocytes towards the allograft. Besides the increase in dendritic cell maturation, it plays a central role in mediating the damage suffered by the graft during AMR. Computational approaches to the search for new therapeutic uses of approved target drugs also showed that imatinib might theoretically be helpful in KT for the prevention and/or treatment of AMR. Conclusion: Our results suggest the importance of the IRF/STAT1 pathways in humoral kidney rejection. NK cells and monocytes in graft damage have an essential role during rejection, and imatinib improves KT outcomes. Our results will have to be validated for the potential use of overexpressed genes as rejection biomarkers that can be used as diagnostic and prognostic markers and as therapeutic targets to avoid graft rejection in patients undergoing kidney transplantation.