Protective Role of the Podocyte IL-15 / STAT5 Pathway in Focal Segmental Glomerulosclerosis.

Introduction: During glomerular diseases, podocyte-specific pathways can modulate the intensity of histological disease and prognosis. The therapeutic targeting of these pathways could thus improve the management and prognosis of kidney diseases. The Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathway, classically described in immune cells, has been recently described in detail in intrinsic kidney cells. Methods: We describe STAT5 expression in human kidney biopsies from patients with focal segmental glomerulosclerosis (FSGS) and studied mice with a podocyte-specific Stat5 deletion in experimental glomerular diseases. Results: Here, we show, for the first time, that STAT5 is activated in human podocytes in FSGS. In addition, podocyte-specific Stat5 inactivation aggravates the structural and functional alterations in a mouse model of FSGS. This could be due, at least in part, to an inhibition of autophagic flux. Finally, interleukin 15 (IL-15), a classical activator of STAT5 in immune cells, increases STAT5 phosphorylation in human podocytes, and its administration alleviates glomerular injury in vivo by maintaining autophagic flux in podocytes. Conclusion: Activating podocyte STAT5 with commercially available IL-15 represents a potential new therapeutic avenue for FSGS.

Toward Precision Medicine: Exploring the Landscape of Biomarkers in Acute Kidney Injury.

Acute kidney injury (AKI) remains a complex challenge with diverse underlying pathological mechanisms and etiologies. Current detection methods predominantly rely on serum creatinine, which exhibits substantial limitations in specificity and poses the issue of late-stage detection of kidney injury. In this review, we propose an up-to-date and comprehensive summary of advancements that identified novel biomarker candidates in blood and urine and ideal criteria for AKI biomarkers such as renal injury specificity, mechanistic insight, prognostic capacity, and affordability. Recently identified biomarkers not only indicate injury location but also offer valuable insights into a range of pathological processes, encompassing reduced glomerular filtration rate, tubular function, inflammation, and adaptive response to injury. The clinical applications of AKI biomarkers are becoming extensive and serving as relevant tools in distinguishing acute tubular necrosis from other acute renal conditions. Also, these biomarkers can offer significant insights into the risk of progression to chronic kidney disease CKD and in the context of kidney transplantation. Integration of these biomarkers into clinical practice has the potential to improve early diagnosis of AKI and revolutionize the design of clinical trials, offering valuable endpoints for therapeutic interventions and enhancing patient care and outcomes.

Kidney Transplantation in Patients With AA Amyloidosis: Outcomes in a French Multicenter Cohort.

RATIONALE & OBJECTIVE: Outcomes of kidney transplantation for patients with renal AA amyloidosis are uncertain, with reports of poor survival and high rates of disease recurrence. However, the data are inconclusive and mostly based on studies from the early 2000s and earlier. STUDY DESIGN: Retrospective multicenter cohort study. SETTING & PARTICIPANTS: We searched the French national transplant database to identify all patients with renal AA amyloidosis who underwent kidney transplantation between 2008 and 2018. EXPOSURES: Age, cause of amyloidosis, use of biotherapies, and C-reactive protein levels. OUTCOMES: Outcomes were all-cause mortality and allograft loss. We also reported amyloidosis allograft recurrence, occurrence of acute rejection episodes, as well as infectious, cardiovascular, and neoplastic disease events. ANALYTICAL APPROACH: Kaplan-Meier estimator for mortality and cumulative incidence function method for allograft loss. Factors associated with patient and allograft survival were investigated using a Cox proportional hazards model and a cause-specific hazards model, respectively. RESULTS: 86 patients who received kidney transplants for AA amyloidosis at 26 French centers were included. The median age was 49.4 years (IQR, 39.7-61.1). The main cause of amyloidosis was familial Mediterranean fever (37 cases; 43%). 16 (18.6%) patients received biotherapy after transplantation. Patient survival rates were 94.0% (95% CI, 89.1-99.2) at 1 year and 85.5% (77.8-94.0) at 5 years after transplantation. Cumulative incidences of allograft loss were 10.5% (4.0-17.0) at 1 year and 13.0% (5.8-20.1) at 5 years after transplantation. Histologically proven AA amyloidosis recurrence occurred in 5 transplants (5.8%). An infection requiring hospitalization developed in 55.8% of cases, and there was a 27.9% incidence of acute allograft rejection. Multivariable analysis showed that C-reactive protein concentration at the time of transplantation was associated with patient survival (HR, 1.01; 95% CI, 1.00-1.02; P=0.01) and allograft survival (HR, 1.68; 95% CI, 1.10-2.57; P=0.02). LIMITATIONS: The study lacked a control group, and the effect of biotherapies on transplantation outcomes could not be explored. CONCLUSIONS: This relatively contemporary cohort of patients who received a kidney transplant for AA amyloidosis experienced favorable rates of survival and lower recurrence rates than previously reported. These data support the practice of treating these patients with kidney transplantation for end-stage kidney disease. PLAIN-LANGUAGE SUMMARY: AA amyloidosis is a severe and rare disease. Kidney involvement is frequent and leads to end-stage kidney disease. Because of the involvement of other organs, these patients are often frail, which has raised concerns about their suitability for kidney transplantation. We reviewed all patients with AA amyloidosis nephropathy who underwent kidney transplantation in France in the recent era (2008-2018) and found that the outcomes after kidney transplantation were favorable, with 85.5% of patients still alive 5 years after transplantation, a survival rate that is comparable to the outcomes of patients receiving a transplant for other forms of kidney diseases. Recurrence of amyloidosis in the transplanted kidney was infrequent (5.8%). These data support the practice of kidney transplantation for patients with AA amyloidosis who experience kidney failure.

Prognostic Biomarkers in Kidney Transplantation: A Systematic Review and Critical Appraisal.

SIGNIFICANCE STATEMENT: Why are there so few biomarkers accepted by health authorities and implemented in clinical practice, despite the high and growing number of biomaker studies in medical research ? In this meta-epidemiological study, including 804 studies that were critically appraised by expert reviewers, the authors have identified all prognostic kidney transplant biomarkers and showed overall suboptimal study designs, methods, results, interpretation, reproducible research standards, and transparency. The authors also demonstrated for the first time that the limited number of studies challenged the added value of their candidate biomarkers against standard-of-care routine patient monitoring parameters. Most biomarker studies tended to be single-center, retrospective studies with a small number of patients and clinical events. Less than 5% of the studies performed an external validation. The authors also showed the poor transparency reporting and identified a data beautification phenomenon. These findings suggest that there is much wasted research effort in transplant biomarker medical research and highlight the need to produce more rigorous studies so that more biomarkers may be validated and successfully implemented in clinical practice. BACKGROUND: Despite the increasing number of biomarker studies published in the transplant literature over the past 20 years, demonstrations of their clinical benefit and their implementation in routine clinical practice are lacking. We hypothesized that suboptimal design, data, methodology, and reporting might contribute to this phenomenon. METHODS: We formed a consortium of experts in systematic reviews, nephrologists, methodologists, and epidemiologists. A systematic literature search was performed in PubMed, Embase, Scopus, Web of Science, and Cochrane Library between January 1, 2005, and November 12, 2022 (PROSPERO ID: CRD42020154747). All English language, original studies investigating the association between a biomarker and kidney allograft outcome were included. The final set of publications was assessed by expert reviewers. After data collection, two independent reviewers randomly evaluated the inconsistencies for 30% of the references for each reviewer. If more than 5% of inconsistencies were observed for one given reviewer, a re-evaluation was conducted for all the references of the reviewer. The biomarkers were categorized according to their type and the biological milieu from which they were measured. The study characteristics related to the design, methods, results, and their interpretation were assessed, as well as reproducible research practices and transparency indicators. RESULTS: A total of 7372 publications were screened and 804 studies met the inclusion criteria. A total of 1143 biomarkers were assessed among the included studies from blood ( n =821, 71.8%), intragraft ( n =169, 14.8%), or urine ( n =81, 7.1%) compartments. The number of studies significantly increased, with a median, yearly number of 31.5 studies (interquartile range [IQR], 23.8-35.5) between 2005 and 2012 and 57.5 (IQR, 53.3-59.8) between 2013 and 2022 ( P < 0.001). A total of 655 studies (81.5%) were retrospective, while 595 (74.0%) used data from a single center. The median number of patients included was 232 (IQR, 96-629) with a median follow-up post-transplant of 4.8 years (IQR, 3.0-6.2). Only 4.7% of studies were externally validated. A total of 346 studies (43.0%) did not adjust their biomarker for key prognostic factors, while only 3.1% of studies adjusted the biomarker for standard-of-care patient monitoring factors. Data sharing, code sharing, and registration occurred in 8.8%, 1.1%, and 4.6% of studies, respectively. A total of 158 studies (20.0%) emphasized the clinical relevance of the biomarker, despite the reported nonsignificant association of the biomarker with the outcome measure. A total of 288 studies assessed rejection as an outcome. We showed that these rejection studies shared the same characteristics as other studies. CONCLUSIONS: Biomarker studies in kidney transplantation lack validation, rigorous design and methodology, accurate interpretation, and transparency. Higher standards are needed in biomarker research to prove the clinical utility and support clinical use.

Complement-activating donor-specific anti-HLA antibodies in solid organ transplantation: systematic review, meta-analysis, and critical appraisal.

Introduction: Several studies have investigated the impact of circulating complement-activating anti-human leukocyte antigen donor-specific antibodies (anti-HLA DSAs) on organ transplant outcomes. However, a critical appraisal of these studies and a demonstration of the prognostic value of complement-activating status over anti-HLA DSA mean fluorescence intensity (MFI) level are lacking. Methods: We conducted a systematic review, meta-analysis and critical appraisal evaluating the role of complement-activating anti-HLA DSAs on allograft outcomes in different solid organ transplants. We included studies through Medline, Cochrane, Scopus, and Embase since inception of databases till May 05, 2023. We evaluated allograft loss as the primary outcome, and allograft rejection as the secondary outcome. We used the Newcastle-Ottawa Scale and funnel plots to assess risk of bias and used bias adjustment methods when appropriate. We performed multiple subgroup analyses to account for sources of heterogeneity and studied the added value of complement assays over anti-HLA DSA MFI level. Results: In total, 52 studies were included in the final meta-analysis (11,035 patients). Complement-activating anti-HLA DSAs were associated with an increased risk of allograft loss (HR 2.77; 95% CI 2.33-3.29, p<0.001; I²=46.2%), and allograft rejection (HR 4.98; 95% CI 2.96-8.36, p<0.01; I²=70.9%). These results remained significant after adjustment for potential sources of bias and across multiple subgroup analyses. After adjusting on pan-IgG anti-HLA DSA defined by the MFI levels, complement-activating anti-HLA DSAs were significantly and independently associated with an increased risk of allograft loss. Discussion: We demonstrated in this systematic review, meta-analysis and critical appraisal the significant deleterious impact and the independent prognostic value of circulating complement-activating anti-HLA DSAs on solid organ transplant risk of allograft loss and rejection.

Distinct association between chronic Epstein-Barr virus infection and T cell compartments from pediatric heart, kidney, and liver transplant recipients.

Chronic Epstein-Barr virus (EBV) infection after pediatric organ transplantation (Tx) accounts for significant morbidity and mortality. The risk of complications, such as posttransplant lymphoproliferative disorders, in high viral load (HVL) carriers is the highest in heart Tx recipients. However, the immunologic signatures of such a risk have been insufficiently defined. Here, we assessed the phenotypic, functional, and transcriptomic profiles of peripheral blood CD8+/CD4+ T cells, including EBV-specific T cells, in 77 pediatric heart, kidney, and liver Tx recipients and established the relationship between memory differentiation and progression toward exhaustion. Unlike kidney and liver HVL carriers, heart HVL carriers displayed distinct CD8+ T cells with (1) up-regulation of interleukin-21R, (2) decreased naive phenotype and altered memory differentiation, (3) accumulation of terminally exhausted (TEX PD-1+T-bet-Eomes+) and decrease of functional precursors of exhausted (TPEX PD-1intT-bet+) effector subsets, and (4) transcriptomic signatures supporting the phenotypic changes. In addition, CD4+ T cells from heart HVL carriers displayed similar changes in naive and memory subsets, elevated Th1 follicular helper cells, and plasma interleukin-21, suggesting an alternative inflammatory mechanism that governs T cell responses in heart Tx recipients. These results may explain the different incidences of EBV complications and may help improve the risk stratification and clinical management of different types of Tx recipients.

Interleukin-21 promotes Type-1 activation and cytotoxicity of CD56dimCD16bright natural killer cells during kidney allograft antibody-mediated rejection showing a new link between adaptive and innate humoral allo-immunity.

The role of Natural killer (NK) cells during kidney allograft antibody-mediated rejection (ABMR) is increasingly recognized, but an in-depth characterization of mechanisms that contribute to such immune response is still under investigation. Here, we characterized phenotypic, functional, and transcriptomic profiles of peripheral blood circulating and allograft infiltrating CD56dimCD16bright NK cells during anti-HLA donor-specific antibody (DSA)+ ABMR. Cross-sectional analyses performed in 71 kidney transplant recipients identified a unique phenotypic circulating CD56dimCD16bright NK cell cluster expanded in DSA+ ABMR. This cluster co-expressed high levels of the interleukin-21 Receptor (IL-21R); Type-1 transcription factors T-bet and EOMES, CD160 and natural killer group 2D cytotoxic and activating co-stimulatory receptors. CD160+ IL-21R+ NK cells correlated with elevated plasma IL-21, Ki-67+ ICOS+ (CD278) IL-21-producing circulating T follicular helper cells, enhanced Type-1 pro-inflammatory cytokines, NK cell cytotoxicity, worse microvascular inflammation and graft loss. Single-cell transcriptomic analysis of circulating NK cells delineated an expanded cluster in DSA+ ABMR characterized by elevated pro-inflammatory/cytotoxic pathways, IL-21/STAT3 signaling, and leukocyte trans-endothelial migration pathways. Infiltration of CD160+ IL-21R+ NK cells with similar transcriptomic profile was detected in DSA+ ABMR allograft biopsies, potentially contributing to allograft injury. Thus, the IL-21/IL-21R axis, linking adaptive and innate humoral allo-immunity, or NK cells may represent appealing immunotherapy targets in DSA+ ABMR.

Atypical presentation of Mycobacterium xenopi pulmonary infection in a kidney transplant recipient: A case report and literature review.

Background: Mycobacterium xenopi is one of the most common pathogens responsible for non-tuberculosis mycobacteria (NTM) pulmonary diseases, which are associated with poor prognosis in immunocompromised patients. Case presentation: We report the unusual case of a 44-year-old kidney transplant recipient with multiple pulmonary nodules revealing M. xenopi pulmonary disease with atypical presentation. A three drug-regimen containing moxifloxacin, ethambutol and azithromycin was prescribed, with careful monitoring of the immunosuppressive therapy. The outcome was favorable. Discussion and conclusion: Although infrequent in kidney transplant recipients, NTM can cause pulmonary infection several years after transplantation. Treatment of M. xenopi infection relies on a multidrug regimen with at least 3 antimycobacterial drugs. Drug-drug interactions between immunosuppressive treatments and rifamycins require careful dose adjustment and monitoring to avoid graft rejection.

Clinical recommendations for posttransplant assessment of anti-HLA (Human Leukocyte Antigen) donor-specific antibodies: A Sensitization in Transplantation: Assessment of Risk consensus document.

Although anti-HLA (Human Leukocyte Antigen) donor-specific antibodies (DSAs) are commonly measured in clinical practice and their relationship with transplant outcome is well established, clinical recommendations for anti-HLA antibody assessment are sparse. Supported by a careful and critical review of the current literature performed by the Sensitization in Transplantation: Assessment of Risk 2022 working group, this consensus report provides clinical practice recommendations in kidney, heart, lung, and liver transplantation based on expert assessment of quality and strength of evidence. The recommendations address 3 major clinical problems in transplantation and include guidance regarding posttransplant DSA assessment and application to diagnostics, prognostics, and therapeutics: (1) the clinical implications of positive posttransplant DSA detection according to DSA status (ie, preformed or de novo), (2) the relevance of posttransplant DSA assessment for precision diagnosis of antibody-mediated rejection and for treatment management, and (3) the relevance of posttransplant DSA for allograft prognosis and risk stratification. This consensus report also highlights gaps in current knowledge and provides directions for clinical investigations and trials in the future that will further refine the clinical utility of posttransplant DSA assessment, leading to improved transplant management and patient care.

Sensitization in transplantation: Assessment of Risk 2022 Working Group Meeting Report.

The Sensitization in Transplantation: Assessment of Risk workgroup is a collaborative effort of the American Society of Transplantation and the American Society of Histocompatibility and Immunogenetics that aims at providing recommendations for clinical testing, highlights gaps in current knowledge, and proposes areas for further research to enhance histocompatibility testing in support of solid organ transplantation. This report provides updates on topics discussed by the previous Sensitization in Transplantation: Assessment of Risk working groups and introduces 2 areas of exploration: non-human leukocyte antigen antibodies and utilization of human leukocyte antigen antibody testing measurement to evaluate the efficacy of antibody-removal therapies.

DSA in solid organ transplantation: is it a matter of specificity, amount, or functional characteristics?

PURPOSE OF REVIEW: The present review describes the clinical relevance of human leukocyte antigen (HLA) donor-specific antibodies (HLA-DSAs) as biomarkers of alloimmunity and summarizes recent improvements in their characterization that provide insights into immune risk assessment, precision diagnosis, and prognostication in transplantation. RECENT FINDINGS: Recent studies have addressed the clinical utility of HLA-DSAs as biomarkers for immune risk assessment in pretransplant and peritransplant, diagnosis and treatment evaluation of antibody-mediated rejection, immune monitoring posttransplant, and risk stratification. SUMMARY: HLA-DSAs have proved to be the most advanced immune biomarkers in solid organ transplantation in terms of analytical validity, clinical validity and clinical utility. Recent studies are integrating multiple HLA-DSA characteristics including antibody specificity, HLA class, quantity, immunoglobulin G subclass, and complement-binding capacity to improve risk assessment peritransplant, diagnosis and treatment evaluation of antibody-mediated rejection, immune monitoring posttransplant, and transplant prognosis evaluation. In addition, integration of HLA-DSAs to clinical, functional and histological transplant parameters has further consolidated the utility of HLA-DSAs as robust biomarkers and allows to build new tools for monitoring, precision diagnosis, and risk stratification for individual patients. However, prospective and randomized-controlled studies addressing the clinical benefit and cost-effectiveness of HLA-DSA-based monitoring and patient management strategies are required to demonstrate that the use of HLA-DSAs as biomarkers can improve current clinical practice and transplant outcomes.

mTOR signaling cascade: novel clinical implications in HLA and non-HLA antibody-mediated vasculopathies?

Antibodies targeting endothelial antigens represent major threats to the integrity of endothelial vasculature, contributing to development of organ transplant rejection and vasculopathies. In this issue, Catar et al. present the mechanisms by which non-human leukocyte antigen angiotensin II type 1 receptor and endothelin-1 type A receptor antibodies mediate impairment of endothelial repair via ß2-arrestin link to the mammalian target of rapamycin pathway. This commentary discusses the mechanisms of human leukocyte antigen and non-human leukocyte antigen antibody-endothelium interactions, and the clinical implications and challenges of the use of mammalian target of rapamycin signaling activation as biomarker and therapeutic target in vasculopathies.

Concomitant loss of regulatory T and B cells is a distinguishing immune feature of antibody-mediated rejection in kidney transplantation.

Although considerable advances have been made in understanding the cellular effector mechanisms responsible for donor-specific antibody generation leading to antibody-mediated rejection (ABMR), the identification of cellular regulators of such immune responses is lacking. To clarify this, we used high dimensional flow cytometry to concomitantly profile and track the two major subsets of regulatory lymphocytes in blood: T regulatory (TREG) and transitional B cells in a cohort of 96 kidney transplant recipients. Additionally, we established co-culture assays to address their respective capacity to suppress antibody responses in vitro. TREG and transitional B cells were found to be potent suppressors of T follicular helper-mediated B-cell differentiation into plasmablast and antibody generation. TREG and transitional B cells were both durably expanded in patients who did not develop donor-specific antibody post-transplant. However, patients who manifested donor-specific antibody and progressed to ABMR displayed a marked and persistent numerical reduction in TREG and transitional B cells. Strikingly, specific cell clusters expressing the transcription factor T-bet were selectively depleted in both TREG and transitional B-cell compartments in patients with ABMR. Importantly, the coordinated loss of these T-bet+CXCR5+TREG and T-bet+CD21- transitional B-cell clusters was correlated with increased and inflammatory donor specific antibody responses, more extensive microvascular inflammation and a higher rate of kidney allograft loss. Thus, our study identified coordinated and persistent defects in regulatory T- and B-cell responses in patients undergoing ABMR, which may contribute to their loss of humoral immune regulation, and warrant timely therapeutic interventions to replenish and sustain TREG and transitional B cells in these patients.

Adaptive immune cell responses as therapeutic targets in antibody-mediated organ rejection.

Humoral alloimmunity of organ transplant recipient to donor can lead to antibody-mediated rejection (ABMR), causing thousands of organ transplants to fail each year worldwide. However, the mechanisms of adaptive immune cell responses at the basis of humoral alloimmunity have not been entirely understood. In this review, we discuss how recent investigations have uncovered the key contributions of T follicular helper (TFH) and B cells and their coordinated actions in driving donor-specific antibody generation and immune progression towards ABMR. We show how recognition of the role of TFH-B cell interactions may allow the elaboration of improved clinical strategies for immune monitoring and the identification of novel therapeutic targets to tackle ABMR that will ultimately improve organ transplant survival.

Impact of the COVID-19 pandemic on publication dynamics and non-COVID-19 research production.

BACKGROUND: The COVID-19 pandemic has severely affected health systems and medical research worldwide but its impact on the global publication dynamics and non-COVID-19 research has not been measured. We hypothesized that the COVID-19 pandemic may have impacted the scientific production of non-COVID-19 research. METHODS: We conducted a comprehensive meta-research on studies (original articles, research letters and case reports) published between 01/01/2019 and 01/01/2021 in 10 high-impact medical and infectious disease journals (New England Journal of Medicine, Lancet, Journal of the American Medical Association, Nature Medicine, British Medical Journal, Annals of Internal Medicine, Lancet Global Health, Lancet Public Health, Lancet Infectious Disease and Clinical Infectious Disease). For each publication, we recorded publication date, publication type, number of authors, whether the publication was related to COVID-19, whether the publication was based on a case series, and the number of patients included in the study if the publication was based on a case report or a case series. We estimated the publication dynamics with a locally estimated scatterplot smoothing method. A Natural Language Processing algorithm was designed to calculate the number of authors for each publication. We simulated the number of non-COVID-19 studies that could have been published during the pandemic by extrapolating the publication dynamics of 2019 to 2020, and comparing the expected number to the observed number of studies. RESULTS: Among the 22,525 studies assessed, 6319 met the inclusion criteria, of which 1022 (16.2%) were related to COVID-19 research. A dramatic increase in the number of publications in general journals was observed from February to April 2020 from a weekly median number of publications of 4.0 (IQR: 2.8-5.5) to 19.5 (IQR: 15.8-24.8) (p < 0.001), followed afterwards by a pattern of stability with a weekly median number of publications of 10.0 (IQR: 6.0-14.0) until December 2020 (p = 0.045 in comparison with April). Two prototypical editorial strategies were found: 1) journals that maintained the volume of non-COVID-19 publications while integrating COVID-19 research and thus increased their overall scientific production, and 2) journals that decreased the volume of non-COVID-19 publications while integrating COVID-19 publications. We estimated using simulation models that the COVID pandemic was associated with a 18% decrease in the production of non-COVID-19 research. We also found a significant change of the publication type in COVID-19 research as compared with non-COVID-19 research illustrated by a decrease in the number of original articles, (47.9% in COVID-19 publications vs 71.3% in non-COVID-19 publications, p < 0.001). Last, COVID-19 publications showed a higher number of authors, especially for case reports with a median of 9.0 authors (IQR: 6.0-13.0) in COVID-19 publications, compared to a median of 4.0 authors (IQR: 3.0-6.0) in non-COVID-19 publications (p < 0.001). CONCLUSION: In this meta-research gathering publications from high-impact medical journals, we have shown that the dramatic rise in COVID-19 publications was accompanied by a substantial decrease of non-COVID-19 research. META-RESEARCH REGISTRATION: https://osf.io/9vtzp/ .

Mass Spectrometry-Based System for Identifying and Typing Norovirus Major Capsid Protein VP1.

Norovirus-associated diseases are the most common foodborne illnesses worldwide. Polymerase chain reaction-based methods are the primary diagnostics for clinical samples; however, the high mutation rate of norovirus makes viral amplification and genotyping challenging. Technological advances in mass spectrometry (MS) make it a promising tool for identifying disease markers. Besides, the superior sensitivity of MS and proteomic approaches may enable the detection of all variants. Thus, this study aimed to establish an MS-based system for identifying and typing norovirus. We constructed three plasmids containing the major capsid protein VP1 of the norovirus GII.4 2006b, 2006a, and 2009a strains to produce virus-like particles for use as standards. Digested peptide signals were collected using a nano-flow ultra-performance liquid chromatography mass spectrometry (nano-UPLC/MSE) system, and analyzed by ProteinLynx Global SERVER and TREE-PUZZLE software. Results revealed that the LC/MSE system had an excellent coverage rate: the system detected more than 94% of amino acids of 3.61 femtomole norovirus VP1 structural protein. In the likelihood-mapping analysis, the proportions of unresolved quartets were 2.9% and 4.9% in the VP1 and S domains, respectively, which is superior to the 15.1% unresolved quartets in current PCR-based methodology. In summary, the use of LC/MSE may efficiently monitor genotypes, and sensitively detect structural and functional mutations of noroviruses.

The emerging field of non-human leukocyte antigen antibodies in transplant medicine and beyond.

The major medical advances in our knowledge of the human leukocyte antigen (HLA) system have allowed us to uncover several gaps in our understanding of alloimmunity. Although the non-HLA system has long sparked the interest of the transplant community, recognition of the role of immunity to non-HLA antigenic targets has only emerged recently. In this review, we will provide a comprehensive summary of the paradigm-changing concept of immunity to the non-HLA angiotensin II type 1 receptor (AT1R), discovered by Duska Dragun et al., that began from careful bedside clinical observations, to validated detection of anti-AT1R antibodies and lead to clinical intervention. This scientific approach has also allowed the recognition of broader pathogenicity of anti-AT1R antibodies across multiple organ transplants and in other human diseases, the integration of both non-HLA and HLA systems to understand their immunologic effects on organ allografts, and the identification of future directions for therapeutic intervention to modulate immunity to AT1R. Rationally designed successful interventions to target AT1R system provide an exemplar for other non-HLA antibodies to cross borders between medical specialties, will generate new avenues in translational research beyond transplantation, and will foster the development of new and reliable tools to improve our understanding of non-HLA immunity and ultimately allow us to improve patient care.

T-bet+CD27+CD21- B cells poised for plasma cell differentiation during antibody-mediated rejection of kidney transplants.

Alloimmune responses driven by donor-specific antibodies (DSAs) can lead to antibody-mediated rejection (ABMR) in organ transplantation. Yet, the cellular states underlying alloreactive B cell responses and the molecular components controlling them remain unclear. Using high-dimensional profiling of B cells in a cohort of 96 kidney transplant recipients, we identified expanded numbers of CD27+CD21- activated memory (AM) B cells that expressed the transcription factor T-bet in patients who developed DSAs and progressed to ABMR. Notably, AM cells were less frequent in DSA+ABMR- patients and at baseline levels in DSA- patients. RNA-Seq analysis of AM cells in patients undergoing ABMR revealed these cells to be poised for plasma cell differentiation and to express restricted IGHV sequences reflective of clonal expansion. In addition to T-bet, AM cells manifested elevated expression of interferon regulatory factor 4 and Blimp1, and upon coculture with autologous T follicular helper cells, differentiated into DSA-producing plasma cells in an IL-21-dependent manner. The frequency of AM cells was correlated with the timing and severity of ABMR manifestations. Importantly, T-bet+ AM cells were detected within kidney allografts along with their restricted IGHV sequences. This study delineates a pivotal role for AM cells in promoting humoral responses and ABMR in organ transplantation and highlights them as important therapeutic targets.

Targeting T Follicular Helper Cells to Control Humoral Allogeneic Immunity.

Humoral allogeneic immunity driven by anti-HLA donor-specific antibodies and antibody-mediated rejection (AMR) significantly impede prolonged survival of organ allografts after transplantation. Although the importance of T follicular helper (TFH) cells in controlling antibody responses has been long established, their role in directing donor-specific antibody generation leading to AMR was only recently appreciated in the clinical setting of organ transplantation. In this review, we provide a comprehensive summary of the current knowledge on the biology of human TFH cells as well as their circulating counterparts and describe their pivotal role in driving humoral alloimmunity. In addition, we discuss the intrinsic effects of current induction therapies and maintenance immunosuppressive drugs as well as of biotherapies on TFH cells and provide future directions and novel opportunities of biotherapeutic targeting of TFH cells that have the potential of bringing the prophylactic and curative treatments of AMR toward personalized and precision medicine.