HLA variations in patients with diffuse large B-cell lymphoma and association with disease risk and prognosis: a case-control study

IntroductionHuman leukocyte antigen (HLA) polymorphisms have been associated with the development of various autoimmune diseases, as well as malignant neoplasms. Non-Hodgkin lymphomas (NHLs) are a heterogenous group of lymphoid malignancies in which a genetic substrate has been established and is deemed to play a crucial role in disease pathogenesis. This study aimed to identify whether variations in the HLA gene region were associated with diffuse large B-cell lymphoma (DLBCL) risk and prognosis.MethodsWe defined HLA class I (HLA-A, HLA-B, HLA-C) and class II (HLA-DRB1, HLA-DQB1) alleles in 60 patients with DLBCL and compared the results to those found by 236 healthy adult donors from the bone marrow bank of Northern Greece. HLA typing was performed by two molecular methods, Sequence - Specific Oligonucleotide HLA typing (SSO) and Sequence - Specific Primer HLA typing (SSP), from white blood cells recovered from peripheral blood. The phenotypic frequencies of HLA-A, HLA-B, HLA-C, HLA-DRB1 and HLA-DQB1 between patients and controls were compared with the 2-sided Fisher’s exact test. Results with p-value <0.05 were considered statistically significant. Odds Ratios with 95% Confidence Intervals were calculated to further strengthen the results. The 2-sided Fisher’s exact test was also applied to alleles found only in one of the two groups, while the odds ratios together with the confidence intervals were corrected with Haldane-Anscombe method.ResultsAmong the studied HLA polymorphisms, the frequency HLA-C*12 allele was significantly lower in patients with DLBCL compared with control subjects (6.7% vs. 34.7%, OR = 0.16, 95% CI: 0.04–0.44). Frequency of HLA-B*39 was significantly lower in patients with DLBCL compared with controls, but due to the low frequency of this polymorphism in the studied population and small sample size, determinations regarding the significance of this findings were limited. Survival analysis revealed that the presence of HLA-C*12 was not associated with improved or worsened overall and progression-free survival. No statistically significant associations were observed in the phenotypic frequencies of HLA-A, HLA-DQB1, HLA-DRB1 and the rest of HLA-B alleles between the control and DLBCL groups.DiscussionCollectively, our results provide valuable insight regarding the role of HLA variations on DLBCL risk. Further studies are required to consolidate our findings and ascertain the clinical implications of these genetic variations on DLBCL management and prognosis

Hidden Patterns of Anti-HLA Class I Alloreactivity Revealed Through Machine Learning

Detection of alloreactive anti-HLA antibodies is a frequent and mandatory test before and after organ transplantation to determine the antigenic targets of the antibodies. Nowadays, this test involves the measurement of fluorescent signals generated through antibody–antigen reactions on multi-beads flow cytometers. In this study, in a cohort of 1,066 patients from one country, anti-HLA class I responses were analyzed on a panel of 98 different antigens. Knowing that the immune system responds typically to “shared” antigenic targets, we studied the clustering patterns of antibody responses against HLA class I antigens without any a priori hypothesis, applying two unsupervised machine learning approaches. At first, the principal component analysis (PCA) projections of intralocus specific responses showed that anti-HLA-A and anti-HLA-C were the most distantly projected responses in the population with the anti-HLA-B responses to be projected between them. When PCA was applied on the responses against antigens belonging to a single locus, some already known groupings were confirmed while several new crossreactive patterns of alloreactivity were detected. Anti-HLA-A responses projected through PCA suggested that three cross-reactive groups accounted for about 70% of the variance observed in the population, while anti-HLA-B responses were mainly characterized by a distinction between previously described Bw4 and Bw6 cross-reactive groups followed by several yet undocumented or poorly described ones. Furthermore, anti-HLA-C responses could be explained by two major cross-reactive groups completely overlapping with previously described C1 and C2 allelic groups. A second featurebased analysis of all antigenic specificities, projected as a dendrogram, generated a robust measure of allelic antigenic distances depicting bead-array defined cross reactive groups. Finally, amino acid combinations explaining major population specific crossreactive groups were described. The interpretation of the results was based on the current knowledge of the antigenic targets of the antibodies as they have been characterized either experimentally or computationally and appear at the HLA epitope registry

Patterns of 1,748 Unique Human Alloimmune Responses Seen by Simple Machine Learning Algorithms

Allele specific antibody response against the polymorphic system of HLA is the allogeneic response marker determining the immunological risk for graft acceptance before and after organ transplantation and therefore routinely studied during the patient’s workup. Experimentally, bead bound antigen- antibody reactions are detected using a special multicolor flow cytometer (Luminex). Routinely for each sample, antibody responses against 96 different HLA antigen groups are measured simultaneously and a 96-dimensional immune response vector is created. Under a common experimental protocol, using unsupervised clustering algorithms, we analyzed these immune intensity vectors of anti HLA class II responses from a dataset of 1,748 patients before or after renal transplantation residing in a single country. Each patient contributes only one serum sample in the analysis. A population view of linear correlations of hierarchically ordered fluorescence intensities reveals patterns in human immune responses with striking similarities with the previously described CREGs but also brings new information on the antigenic properties of class II HLA molecules. The same analysis affirms that “public” anti-DP antigenic responses are not correlated to anti DR and anti DQ responses which tend to cluster together. Principal Component Analysis (PCA) projections also demonstrate ordering patterns clearly differentiating anti DP responses from anti DR and DQ on several orthogonal planes. We conclude that a computer vision of human alloresponse by use of several dimensionality reduction algorithms rediscovers proven patterns of immune reactivity without any a priori assumption and might prove helpful for a more accurate definition of public immunogenic antigenic structures of HLA molecules. Furthermore, the use of Eigen decomposition on the Immune Response generates new hypotheses that may guide the design of more effective patient monitoring tests

Detection of the α determinant mutations by partial sequencing of the S gene of hepatitis B virus in chronic carriers

Virological data on chronic hepatitis B virus (HBV) infection in Greece are limited. The aim of this study was to investigate the HBV genotypes, surface antigen (HBsAg) subtypes, and HBsAg ‘‘a’’ determinant mutations among patients infected chronically with HBV. Serum samples from 135 HBsAg positive patients were tested. Serologic (HBsAg, anti-HBs, anti-HBc (total), anti-HBc IgM, HBeAg, and anti-HBe), virologic (HBV-DNA quantitation) and biochemical markers (serum alanine aminotransferase/ALT and aspartate aminotransferase/ AST) were analyzed. HBV genotypes and HBsAg subtypes were determined by partial sequencing of the S gene. Genotyping was performed by using the National Center for Biotechnology Information online Genotyping tool and phylogenetic analysis. Nucleotide sequences were aligned pair wise with ClustalW and phylogenetic trees were constructed by the neighbor-joining method. Sequences were also used to predict HBV HBsAg subtypes. In six patients (4%), simultaneous presence of HBsAg and anti-HBs was determined, whereas 47 patients (35%) were HBeAg positive, 84 (62.5%) were anti-HBe positive, and three patients (2,2%) were characterized by the simultaneous presence of HBeAg and anti-HBe. Mean ALT was 238 IU/L (standard deviation=576.84), and HBV-DNA levels ranged from 1,01 x 10⁵ to 1,2 x 10⁷ IU/ml. Genotype D was predominant (98%), with viral groups D/ ayw2 (73%) and D/ayw3 (27%). Group A/adw accounted for 0,7% of cases. Genotypes B and C were found exclusively in the Chinese immigrants (1,4%). Single or multiple point mutations were found in 35 cases (26%). Some of the most common mutations occurred at amino acid positions 129, 133, 134, 144, 145, including the ‘‘vaccine escape’’ mutation G145R. Mutations analysis revealed that amino acid substitutions did not affect detection by commercial immunoassays, except for one case.Τα δεδομένα μοριακής επιδημιολογίας για την ηπατίτιδα Β που αφορούν τον Ελληνικό πληθυσμό είναι περιορισμένα. Σκοπός της παρούσας μελέτης ήταν ο προσδιορισμός των γονοτύπων, των υποτύπων του επιφανειακού αντιγόνου HBsAg και των μεταλλαγών στον α-αντιγονικό καθοριστή του επιφανειακού αντιγόνου σε χρόνιους φορείς της ηπατίτιδας Β. Εξετάσθηκαν οι οροί αίματος 135 φορέων θετικών για το επιφανειακό αντιγόνο. Προσδιορίστηκαν οι ορολογικοί δείκτες (HBsAg, antiHBs, anti-HBc (ολικό), anti-HBc IgM, HBeAg και anti-HBe) το ιικό φορτίο (HBVDNA ποσοτικός προσδιορισμός) και οι βιοχημικοί δείκτες (αλανινοαμινοτρασφεράση/ALT και ασπαρτική αμινοτρασφεράση/ AST) στον ορό των παραπάνω ασθενών. Από την ανάλυση της αλληλουχίας νουκλεοτιδίων τμήματος του γονιδίου που κωδικοποιεί την πρωτεΐνη S προσδιορίστηκαν οι γονότυποι των HBV στελεχών των παραπάνω ασθενών όπως και οι HBsAg υπότυποι. Η γονοτυπική ανάλυση έγινε με τη χρήση του γονοτυπικού εργαλείου (Genotyping tool) που είναι ελεύθερα διαθέσιμο στην ιστοσελίδα της Εθνικής Βιβλιοθήκης του Εθνικού Κέντρου για την Βιοτεχνολογική Πληροφόρηση των Η.Π.Α. (NCBI) και με φυλογενετική ανάλυση. Έγινε ανά ζεύγος σύγκριση των αλληλουχιών με αντίστοιχες αλληλουχίες στελεχών αναφοράς του ίδιου γονοτύπου με τη βοήθεια του λογισμικού ClustalW ενώ ο σχεδιασμός των φυλογενετικών δέντρων πραγματοποιήθηκε με τη μέθοδο neighbor-joining. Η ανάλυση των αλληλουχιών χρησιμοποιήθηκε επίσης και για τον προσδιορισμό των HBsAg υποτύπων. Σε έξι ασθενείς (4%) ανιχνεύθηκαν συγχρόνως τόσο το HBsAg όσο και το anti-HBs αντίσωμα, ενώ 47 ασθενείς (35%) ήταν HBeAg θετικοί, 84 (62,5%) ήταν θετικοί για anti-HBe αντίσωμα ενώ σε 3 ασθενείς (2,2%) υπήρχε ταυτόχρονη παρουσία του HBeAg και του anti-HBe. Η μέση τιμή της ALT ήταν 238IU/L με SD=576,84 και τα επίπεδα του ιικού φορτίου κυμάνθηκαν στα 1,01 x 10⁵ έως 1,2 x 10⁷ IU/ml. Ο γονότυπος D ήταν ο κυρίαρχος (98%) με τον υπότυπο D/ayw2 να φθάνει το 73% και τον D/ayw3 το 27%. Ο γονότυπος A με υπότυπο adw αφορούσε το 0,7%. Οι γονότυποι B και C βρέθηκαν αποκλειστικά σε μετανάστες από την Κίνα στο 1,4%. Απλές ή πολλαπλές σημειακές μεταλλαγές βρέθηκαν σε 35 ασθενείς (26%). Μερικές από τις πιο συχνές μεταλλαγές ήταν στις θέσεις αμινοξέων 129, 133, 134, 144 και 145, συμπεριλαμβανομένης και της μεταλλαγής διαφυγής του εμβολίου G145R. Από την ανάλυση των μεταλλαγών φάνηκε ότι οι υποκαταστάσεις αμινοξέων στις διάφορες θέσεις του α-αντιγονικού καθοριστή δεν επηρέασαν τη δυνατότητα ανίχνευσης του επιφανειακού αντιγόνου HBsAg από όλα τα στελέχη πλην ενός

Microbiome in Chronic Kidney Disease

The gut microbiome is a complex collection of microorganisms with discrete characteristics and activities. Its important role is not restricted to food digestion and metabolism, but extends to the evolution, activation and function of the immune system. Several factors, such as mode of birth, diet, medication, ageing and chronic inflammation, can modify the intestinal microbiota. Chronic kidney disease (CKD) seems to have a direct and unique effect, as increased urea levels result in alteration of the gut microbiome, leading to overproduction of its metabolites. Therefore, potentially noxious microbial uremic toxins, which have predominantly renal clearance, including p-cresyl sulfate, indoxyl sulfate and N-oxide of trimethylamine [Trimethylamine-N-Oxide (TMAO)], accumulate in human’s body, and are responsible not only for the clinical implications of CKD, but also for the progression of renal failure itself. Certain changes in gut microbiome are observed in patients with end stage renal disease (ESRD), either when undergoing hemodialysis or after kidney transplantation. The purpose of this review is to summarize the changes of gut microbiome and the protein bound uremic toxins which are observed in CKD and in different kidney replacement strategies. In addition, we attempt to review the connection between microbiome, clinical implications and immune response in CKD

COVID-19 Infection and Response to Vaccination in Chronic Kidney Disease and Renal Transplantation: A Brief Presentation

Chronic kidney disease (CKD) is associated with phenotypic and functional changes in the immune system, followed by detrimental clinical consequences, such as severe infections and defective response to vaccination. Two years of the pandemic, due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have undoubtedly changed the world; however, all efforts to confront infection and provide new generation vaccines tremendously improved our understanding of the mechanisms of the immune response against infections and after vaccination. Humoral and cellular responses to vaccines, including mRNA vaccines, are apparently affected in CKD patients, as elimination of recent thymic emigrant and naïve lymphocytes and regulatory T-cells, together with contraction of T-cell repertoire and homeostatic proliferation rate, which characterized CKD patients are responsible for impaired immune activation. Successful renal transplantation will restore some of these changes, although several epigenetic changes are irreversible and even accelerated by the induction of immunosuppression. Response to vaccination is definitely impaired among both CKD and RT patients. In the present review, we analyzed the differences in immune response after vaccination between these patients and healthy individuals and depicted specific parameters, such as alterations in the immune system, predisposing to this deficient response

Type of ANCA May Be Indispensable in Distinguishing Subphenotypes of Different Clinical Entities in ANCA-Associated Vasculitis

The traditional nomenclature system for classifying antineutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV) based on clinical phenotype describes granulomatosis with polyangiitis (GPA), eosinophilic granulomatosis with polyangiitis (EGPA) and microscopic polyangiitis (MPA) as distinct clinical entities. This classification has proved its expedience in clinical trials and everyday clinical practice; yet, a substantial overlap in clinical presentation still exists and often causes difficulties in prompt definition and clinical distinction. Additionally, new insights into the AAV pathogenesis point out that PR3 and MPO-AAV may not represent expressions of the same disease spectrum but rather two distinct disorders, as they display significant differences. Thus, it is supported that a classification based on ANCA serotype (PR3-ANCA, MPO-ANCA or ANCA-negative) could be more accurate and also closer to the nature of the disease compared to the phenotype-based one. This review aims to elucidate the major differences between PR3 and MPO-AAV in terms of epidemiology, pathogenesis, histological and clinical manifestations and response to therapeutic approaches

Cellular and Humoral Responses in Dialysis Patients after Vaccination with the BNT162b2 or mRNA-1273 Vaccines

The outbreak of SARS-CoV-2 has raised considerable concern about the detrimental effects it can induce in public health, with the interest of the scientific community being focused on the development of preventive and therapeutic approaches. Patients with end-stage renal disease (ESRD) are amongst vulnerable populations for critical illness owing to the presence of other comorbidities, their defective immune system, and their inability of self-isolation. To date, vaccination constitutes the most promising method to manage viral dispersion. Therefore, it is particularly important to investigate the effectiveness of available vaccines against SARS-CoV-2 in this risk group. Here, we summarize initial experience regarding the humoral and cellular immune responses elicited in dialysis patients after completion of the recommended vaccination regimen, as well as after booster dose administration, with one of the two mRNA vaccines, namely, BNT162b2 and mRNA-1273. In conclusion, a significantly diminished and delayed immune pattern was observed in ESRD patients compared to healthy population, with a peak in antibody titers occurring 3–5 weeks after the second dose. A booster dose significantly augmented the immune response in dialysis patients with either mRNA-based vaccine. Variables adversely correlating with the weak immunogenicity observed in dialysis patients include immunosuppressive therapy, older age, comorbidities, longer time in hemodialysis treatment, and higher body mass index. On the contrary, previous COVID-19 infection and administration of the mRNA-1273 vaccine are deemed to induce a more favorable immune response. Further investigation is needed to thoroughly understand the efficacy of mRNA-based vaccines in hemodialysis patients and define predictive factors that can influence it

Cellular and Humoral Responses in Dialysis Patients after Vaccination with the BNT162b2 or mRNA-1273 Vaccines

The outbreak of SARS-CoV-2 has raised considerable concern about the detrimental effects it can induce in public health, with the interest of the scientific community being focused on the development of preventive and therapeutic approaches. Patients with end-stage renal disease (ESRD) are amongst vulnerable populations for critical illness owing to the presence of other comorbidities, their defective immune system, and their inability of self-isolation. To date, vaccination constitutes the most promising method to manage viral dispersion. Therefore, it is particularly important to investigate the effectiveness of available vaccines against SARS-CoV-2 in this risk group. Here, we summarize initial experience regarding the humoral and cellular immune responses elicited in dialysis patients after completion of the recommended vaccination regimen, as well as after booster dose administration, with one of the two mRNA vaccines, namely, BNT162b2 and mRNA-1273. In conclusion, a significantly diminished and delayed immune pattern was observed in ESRD patients compared to healthy population, with a peak in antibody titers occurring 3–5 weeks after the second dose. A booster dose significantly augmented the immune response in dialysis patients with either mRNA-based vaccine. Variables adversely correlating with the weak immunogenicity observed in dialysis patients include immunosuppressive therapy, older age, comorbidities, longer time in hemodialysis treatment, and higher body mass index. On the contrary, previous COVID-19 infection and administration of the mRNA-1273 vaccine are deemed to induce a more favorable immune response. Further investigation is needed to thoroughly understand the efficacy of mRNA-based vaccines in hemodialysis patients and define predictive factors that can influence it

ANCA-Associated Vasculitis May Result as a Complication to Both SARS-CoV-2 Infection and Vaccination

In the last two years, our world experienced one of the most devastating and fast-exploding pandemic, due to the wide spread of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The scientific community managed to develop effective vaccines, the main weapons to shield the immune system and protect people. Nevertheless, both SARS-CoV-2 infection and the vaccination against it have been associated with the stimulation of inflammatory cells such as T and B lymphocytes that results in a cytokine storm, endothelial inflammation and vascular injury, which can lead to different types of vasculitis. We present the first case of de novo MPO-ANCA-associated vasculitis, which developed shortly after SARS-CoV-2 vaccination, adequately responded to treatment, and subsequently relapsed after COVID-19 infection. With this case, we indicate an etiological connection between viral infection and disease development, as well as the possibility of a common immune mechanism between SARS-CoV-2 infection and vaccination, that can stimulate vascular events and lead to vasculitis. There have been several case reports of de novo vasculitis, affecting large, medium, or small vessels, following either infection or vaccination against COVID-19, during the pandemic outbreak. We summarize previous reports and also analyze proposed pathogenic mechanisms between SARS-CoV-2 and vasculitis