Interleukin-33 and liver natural killer cells: A novel perspective on antitumor activity in liver fibrosis.

AIM: Liver fibrosis, heralding the potential progression to cirrhosis and hepatocellular carcinoma (HCC), compromises patient survival and augments post-hepatectomy recurrence. This study examined the detrimental effects of liver fibrosis on the antitumor functions of liver natural killer (NK) cells and the interleukin-33 (IL-33) signaling pathway. METHODS: Our investigation, anchored in both human physiologies using living and deceased donor livers and the carbon tetrachloride (CCl4)-induced mouse fibrosis model, aimed to show a troubling interface between liver fibrosis and weakened hepatic immunity. RESULTS: The Fibrosis-4 (FIB-4) index emerged as a salient, non-invasive prognostic marker, and its elevation correlated with reduced survival and heightened recurrence after HCC surgery even after propensity matching (n = 385). We established a strong correlation between liver fibrosis and liver NK cell dysfunction by developing a method for extracting liver NK cells from the liver graft perfusate. Furthermore, liver fibrosis ostensibly disrupted chemokines and promoted IL-33 expression, impeding liver NK cell antitumor activities, as evidenced in mouse models. Intriguingly, our results implicated IL-33 in diminishing the antitumor responses of NK cells. This interrelation, consistent across both mouse and human studies, coincides with clinical data suggesting that liver fibrosis predisposes patients to an increased risk of HCC recurrence. CONCLUSION: Our study revealed a critical relationship between liver fibrosis and compromised tumor immunity, emphasizing the potential interference of IL-33 with NK cell function. These insights advocate for advanced immunostimulatory therapies targeting cytokines, such as IL-33, aiming to bolster the hepatic immune response against HCC in the context of liver fibrosis.

Indicators of verbal creative thinking: results of a Delphi panel.

Introduction: Creativity is a fundamental competence that manifests itself in various domains of knowledge, including verbal creativity. The main aim of this study was to identify indicators of verbal creativity for the assessment of three writing tasks. Methods: Sixteen multidisciplinary and international creativity experts participated in a two-stage Delphi panel. The administered questionnaire asked about the measurement or non-measurement of eight indicators of verbal creative thinking in three tasks: problem posing, creative idea generation, and idea improvement. Originality is the most important indicator of creativity. The indicators identified in the first task were fluency, flexibility, originality, elaboration, and sensitivity to problems. The second task measures flexibility, originality, elaboration, opacity, and dynamic integration. In the third task, fluency, flexibility, originality, elaboration, dynamic integration, and refinement of ideas are considered. Results: The results of this study are key to progress in the field of measuring verbal creative thinking. Discussion: The identification of indicators of the construct called verbal creativity allows the determination of its components in order to be able to estimate the creative potential in this specific domain.

The IPTA Nashville consensus conference on post-transplant lymphoproliferative disorders after solid organ transplantation in children: IV-consensus guidelines for the management of post-transplant lymphoproliferative disorders in children and adolescents.

The International Pediatric Transplant Association convened an expert consensus conference to assess current evidence and develop recommendations for various aspects of care relating to post-transplant lymphoproliferative disorders (PTLD) after pediatric solid organ transplantation. This report addresses the outcomes of deliberations by the PTLD Management Working Group. A strong recommendation was made for reduction in immunosuppression as the first step in management. Similarly, strong recommendations were made for the use of the anti-CD20 monoclonal antibody (rituximab) as was the case for chemotherapy in selected scenarios. In some scenarios, there is uncoupling of the strength of the recommendations from the available evidence in situations where such evidence is lacking but collective clinical experiences drive decision-making. Of note, there are no large, randomized phase III trials of any treatment for PTLD in the pediatric age group. Current gaps and future research priorities are highlighted.

Epstein-Barr virus-associated post-transplant lymphoproliferative disorders in pediatric transplantation: A prospective multicenter study in the United States.

BACKGROUND: Epstein-Barr virus (EBV)-associated post-transplant lymphoproliferative disorders (PTLD) is the most common malignancy in children after transplant; however, difficulties for early detection may worsen the prognosis. METHODS: The prospective, multicenter, study enrolled 944 children (≤21 years of age). Of these, 872 received liver, heart, kidney, intestinal, or multivisceral transplants in seven US centers between 2014 and 2019 (NCT02182986). In total, 34 pediatric EBV+ PTLD (3.9%) were identified by biopsy. Variables included sex, age, race, ethnicity, transplanted organ, EBV viral load, pre-transplant EBV serology, immunosuppression, response to chemotherapy and rituximab, and histopathological diagnosis. RESULTS: The uni-/multivariable competing risk analyses revealed the combination of EBV-seropositive donor and EBV-naïve recipient (D+R-) was a significant risk factor for PTLD development (sub-hazard ratio: 2.79 [1.34-5.78], p = .006) and EBV DNAemia (2.65 [1.72-4.09], p < .001). Patients with D+R- were significantly more associated with monomorphic/polymorphic PTLD than those with the other combinations (p = .02). Patients with monomorphic/polymorphic PTLD (n = 21) had significantly more EBV DNAemia than non-PTLD patients (p < .001) and an earlier clinical presentation of PTLD than patients with hyperplasias (p < .001), within 6-month post-transplant. Among non-liver transplant recipients, monomorphic/polymorphic PTLD were significantly more frequent than hyperplasias in patients ≥5 years of age at transplant (p = .01). CONCLUSIONS: D+R- is a risk factor for PTLD and EBV DNAemia and associated with the incidence of monomorphic/polymorphic PTLD. Intensive follow-up of EBV viral load within 6-month post-transplant, especially for patients with D+R- and/or non-liver transplant recipients ≥5 years of age at transplant, may help detect monomorphic/polymorphic PTLD early in pediatric transplant.

Application of Mass Cytometry Platforms to Solid Organ Transplantation.

Transplantation serves as the cornerstone of treatment for patients with end-stage organ disease. The prevalence of complications, such as allograft rejection, infection, and malignancies, underscores the need to dissect the complex interactions of the immune system at the single-cell level. In this review, we discuss studies using mass cytometry or cytometry by time-of-flight, a cutting-edge technology enabling the characterization of immune populations and cell-to-cell interactions in granular detail. We review the application of mass cytometry in human and experimental animal studies in the context of transplantation, uncovering invaluable contributions of the tool to understanding rejection and other transplant-related complications. We discuss recent innovations that have the potential to streamline and standardize mass cytometry workflows for application to multisite clinical trials. Additionally, we introduce imaging mass cytometry, a technique that couples the power of mass cytometry with spatial context, thereby mapping cellular interactions within tissue microenvironments. The synergistic integration of mass cytometry and imaging mass cytometry data with other omics data sets and high-dimensional data platforms to further define immune dynamics is discussed. In conclusion, mass cytometry technologies, when integrated with other tools and data, shed light on the intricate landscape of the immune response in transplantation. This approach holds significant potential for enhancing patient outcomes by advancing our understanding and facilitating the development of new diagnostics and therapeutics.

Influence of the chest wall on respiratory function at birth in near-term lambs.

Airway liquid is cleared into lung tissue after birth, which becomes edematous and forces the chest wall to expand to accommodate both the cleared liquid and incoming air. This study investigated how changing chest wall mechanics affects respiratory function after birth in near-term lambs with different airway liquid volumes. Surgically instrumented near-term lambs (139 ± 2 days) were randomized into Control (n = 7) or Elevated Liquid (EL; n = 6) groups. Control lambs had lung liquid drained to simulate expected volumes following vaginal delivery. EL lambs had airway liquid drained and 30 mL/kg liquid returned to simulate expected airway liquid volumes after elective cesarean section. Lambs were delivered, transferred to a Perspex box, and ventilated (30 min). Pressure in the box was adjusted to apply positive (7-8 cmH2O above atmospheric pressure) or negative (7-8 cmH2O below atmospheric pressure) pressures for 30 min before pressures were reversed. External negative pressures expanded the chest wall, reduced chest wall compliance (CCW) and increased lung compliance (CL) in Control and EL lambs. External positive pressures compressed the chest wall, increased CCW and reduced CL in Control and EL lambs. External negative pressure improved pulmonary oxygen exchange, reducing the alveolar-arterial difference in oxygen (AaDO2) by 69 mmHg (95% CI [13, 125]; P = 0.016) in Control lambs and by 300 mmHg (95% CI [233, 367]; P < 0.001) in EL lambs. In contrast, external positive pressures impaired pulmonary gas exchange, increasing the AaDO2 by 179 mmHg (95% CI [73, 285]; P = 0.002) in Control and by 215 mmHg (95% CI [89, 343]; P < 0.001) in EL lambs. The application of external thoracic pressures influences respiratory function after birth.NEW & NOTEWORTHY This study investigated how changes in chest wall mechanics influence respiratory function after birth. Our data indicate that the application of continuous external subatmospheric pressure greatly improves respiratory function in near-term lambs with respiratory distress, whereas external positive pressures impair respiratory function. Our findings indicate that, during neonatal resuscitation at birth, the forces applied to the chest wall should not be ignored as they can have a major impact on neonatal respiratory function.

The IPTA Nashville Consensus Conference on Post-Transplant lymphoproliferative disorders after solid organ transplantation in children: III - Consensus guidelines for Epstein-Barr virus load and other biomarker monitoring.

The International Pediatric Transplant Association convened an expert consensus conference to assess current evidence and develop recommendations for various aspects of care relating to post-transplant lymphoproliferative disorders after solid organ transplantation in children. In this report from the Viral Load and Biomarker Monitoring Working Group, we reviewed the existing literature regarding the role of Epstein-Barr viral load and other biomarkers in peripheral blood for predicting the development of PTLD, for PTLD diagnosis, and for monitoring of response to treatment. Key recommendations from the group highlighted the strong recommendation for use of the term EBV DNAemia instead of "viremia" to describe EBV DNA levels in peripheral blood as well as concerns with comparison of EBV DNAemia measurement results performed at different institutions even when tests are calibrated using the WHO international standard. The working group concluded that either whole blood or plasma could be used as matrices for EBV DNA measurement; optimal specimen type may be clinical context dependent. Whole blood testing has some advantages for surveillance to inform pre-emptive interventions while plasma testing may be preferred in the setting of clinical symptoms and treatment monitoring. However, EBV DNAemia testing alone was not recommended for PTLD diagnosis. Quantitative EBV DNAemia surveillance to identify patients at risk for PTLD and to inform pre-emptive interventions in patients who are EBV seronegative pre-transplant was recommended. In contrast, with the exception of intestinal transplant recipients or those with recent primary EBV infection prior to SOT, surveillance was not recommended in pediatric SOT recipients EBV seropositive pre-transplant. Implications of viral load kinetic parameters including peak load and viral set point on pre-emptive PTLD prevention monitoring algorithms were discussed. Use of additional markers, including measurements of EBV specific cell mediated immunity was discussed but not recommended though the importance of obtaining additional data from prospective multicenter studies was highlighted as a key research priority.

Dental caries prevalence and severity positively associate with AMY1 gene copy number.

OBJECTIVE: To establish the possible relation between total caries (TC) and caries severity (CS) with the AMY1 gene copy number (AMY1GCN). MATERIALS AND METHODS: This was an observational, cross-sectional, population-based, and association study with 303 participants. Each participant underwent a complete anamnesis and stomatological check-up, and peripheral blood was obtained to extract gDNA. TC and CS were determined as the number of caries at the dental exploration and the number of dental surfaces affected by caries, respectively, and AMY1GCN was determined by qPCR. RESULTS: We found an elevated caries prevalence (92.7%); TC and CS were 8 ± 10 and 10 ± 13 (median ± IR). There were higher TC and CS in those participants with AMY1GCN above the mean value (0.02 and 0.01 p values, respectively). A positive correlation between TC and CS with AMY1GCN (0.11 and 0.125 r values, 0.03 and 0.01 p values, respectively) was found, in addition to an association between TC and CS with AMY1GCN (1.5 and 1.6 OR values, 0.48 and 0.26 p values, respectively). CONCLUSION: TC and CS were positively related to the AMY1GCN. CLINICAL RELEVANCE: Dental caries has a high prevalence and a multifactorial etiology and has been related to a genetic component. Indeed, the salivary enzyme alpha-amylase could play a significant role in caries susceptibility, considering that its codifying gene (AMY1) can show variation in its gene copy number. This can be considered an important factor for the development of caries at a genetic level.

Effect of Cellular Senescence in Disease Progression and Transplantation: Immune Cells and Solid Organs.

Aging of the world population significantly impacts healthcare globally and specifically, the field of transplantation. Together with end-organ dysfunction and prolonged immunosuppression, age increases the frequency of comorbid chronic diseases in transplant candidates and recipients, contributing to inferior outcomes. Although the frequency of death increases with age, limited use of organs from older deceased donors reflects the concerns about organ durability and inadequate function. Cellular senescence (CS) is a hallmark of aging, which occurs in response to a myriad of cellular stressors, leading to activation of signaling cascades that stably arrest cell cycle progression to prevent tumorigenesis. In aging and chronic conditions, senescent cells accumulate as the immune system's ability to clear them wanes, which is causally implicated in the progression of chronic diseases, immune dysfunction, organ damage, decreased regenerative capacity, and aging itself. The intimate interplay between senescent cells, their proinflammatory secretome, and immune cells results in a positive feedback loop, propagating chronic sterile inflammation and the spread of CS. Hence, senescent cells in organs from older donors trigger the recipient's alloimmune response, resulting in the increased risk of graft loss. Eliminating senescent cells or attenuating their inflammatory phenotype is a novel, potential therapeutic target to improve transplant outcomes and expand utilization of organs from older donors. This review focuses on the current knowledge about the impact of CS on circulating immune cells in the context of organ damage and disease progression, discusses the impact of CS on abdominal solid organs that are commonly transplanted, and reviews emerging therapies that target CS.

High-dimensional profiling of pediatric immune responses to solid organ transplantation.

Solid organ transplant remains a life-saving therapy for children with end-stage heart, lung, liver, or kidney disease; however, ∼33% of allograft recipients experience acute rejection within the first year after transplant. Our ability to detect early rejection is hampered by an incomplete understanding of the immune changes associated with allograft health, particularly in the pediatric population. We performed detailed, multilineage, single-cell analysis of the peripheral blood immune composition in pediatric solid organ transplant recipients, with high-dimensional mass cytometry. Supervised and unsupervised analysis methods to study cell-type proportions indicate that the allograft type strongly influences the post-transplant immune profile. Further, when organ-specific differences are considered, graft health is associated with changes in the proportion of distinct T cell subpopulations. Together, these data form the basis for mechanistic studies into the pathobiology of rejection and allow for the development of new immunosuppressive agents with greater specificity.

Mutations in latent membrane protein 1 of Epstein-Barr virus are associated with increased risk of posttransplant lymphoproliferative disorder in children.

Epstein-Barr virus (EBV)-positive posttransplant lymphoproliferative disorder (PTLD) results in significant morbidity and mortality in pediatric transplant recipients. Identifying individuals at an increased risk of EBV-positive PTLD could influence clinical management of immunosuppression and other therapies, improving posttransplant outcomes. A 7-center prospective, observational clinical trial of 872 pediatric transplant recipients evaluated the presence of mutations at positions 212 and 366 of EBV latent membrane protein 1 (LMP1) as an indicator of risk of EBV-positive PTLD (clinical trials: NCT02182986). DNA was isolated from peripheral blood of EBV-positive PTLD case patients and matched controls (1:2 nested case:control), and the cytoplasmic tail of LMP1 was sequenced. Thirty-four participants reached the primary endpoint of biopsy-proven EBV-positive PTLD. DNA was sequenced from 32 PTLD case patients and 62 matched controls. Both LMP1 mutations were present in 31 of 32 PTLD cases (96.9%) and in 45 of 62 matched controls (72.6%) (P = .005; OR = 11.7; 95% confidence interval, 1.5, 92.6). The presence of both G212S and S366T carries a nearly 12-fold increased risk of development of EBV-positive PTLD. Conversely, transplant recipients without both LMP1 mutations carry a very low risk of PTLD. Analysis of mutations at positions 212 and 366 of LMP1 can be informative in stratifying patients for risk of EBV-positive PTLD.

The IPTA Nashville consensus conference on Post-Transplant lymphoproliferative disorders after solid organ transplantation in children: II-consensus guidelines for prevention.

The International Pediatric Transplant Association (IPTA) convened an expert consensus conference to assess current evidence and develop recommendations for various aspects of care relating to post-transplant lymphoproliferative disorder after solid organ transplantation in children. In this report from the Prevention Working Group, we reviewed the existing literature regarding immunoprophylaxis and chemoprophylaxis, and pre-emptive strategies. While the group made a strong recommendation for pre-emptive reduction of immunosuppression at the time of EBV DNAemia (low to moderate evidence), no recommendations for use could be made for any prophylactic strategy or alternate pre-emptive strategy, largely due to insufficient or conflicting evidence. Current gaps and future research priorities are highlighted.

Host microRNAs are decreased in pediatric solid-organ transplant recipients during EBV+ Post-transplant Lymphoproliferative Disorder.

Post-transplant lymphoproliferative disorder (PTLD) is a serious complication of solid organ transplantation. Predisposing factors include primary Epstein-Barr virus (EBV) infection, reactivation of EBV in recipient B cells, and decreased T cell immunity due to immunosuppression. In our previous studies EBV infection was demonstrated to markedly alter the expression of host B cell microRNA (miR). Specifically, miR-194 expression was uniquely suppressed in EBV+ B cell lines from PTLD patients and the 3'untranslated region of IL-10 was determined to be targeted by miR-194. Although EBV has been shown to regulate host miR expression in B cell lymphoma cell lines, the expression of miRs in the circulation of patients with EBV-associated PTLD has not been studied. The objective of this study was to determine if changes in miR expression are associated with EBV+ PTLD. In this study, we have shown that miR-194 is significantly decreased in EBV+PTLD tumors and that additional miRs, including miRs-17, 19 and 106a are also reduced in EBV+PTLD as compared to EBV-PTLD. We quantitated the levels of miRs-17, 19, 106a, 155, and 194 in the plasma and extracellular vesicles (EV; 50-70 nm as determined by nanoparticle tracking analysis) from pediatric recipients of solid organ transplants with EBV+ PTLD+ that were matched 1:2 with EBV+ PTLD- pediatric transplant recipients as part of the NIH-sponsored Clinical Trials in Organ Transplantation in Children, (CTOTC-06) study. Levels of miRs-17, 19, 106a, and 194 were reduced in the plasma and extracellular vesicles (EV) of EBV+ PTLD+ group compared to matched controls, with miRs-17 (p = 0.034; plasma), miRs-19 (p = 0.029; EV) and miR-106a (p = 0.007; plasma and EV) being significantly reduced. Similar levels of miR-155 were detected in the plasma and EV of all pediatric SOT recipients. Importantly, ~90% of the cell-free miR were contained within the EV supporting that EBV+ PTLD tumor miR are detected in the circulation and suggesting that EVs, containing miRs, may have the potential to target and regulate cells of the immune system. Further development of diagnostic, mechanistic and potential therapeutic uses of the miRs in PTLD is warranted.

Malnutrition and immune cell subsets in children undergoing kidney transplantation.

BACKGROUND: Malnutrition, including obesity and undernutrition, among children is increasing in prevalence and is common among children on renal replacement therapy. The effect of malnutrition on the pre-transplant immune system and how the pediatric immune system responds to the insult of both immunosuppression and allotransplantation is unknown. We examined the relationship of nutritional status with post-transplant outcomes and characterized the peripheral immune cell phenotypes of children from the Immune Development of Pediatric Transplant (IMPACT) study. METHODS: Ninety-eight patients from the IMPACT study were classified as having obesity, undernutrition, or normal nutrition-based pre-transplant measurements. Incidence of infectious and alloimmune outcomes at 1-year post-transplantation was compared between nutritional groups using Gray's test and Fine-Gray subdistribution hazards model. Event-free survival was estimated by Kaplan-Meier method and compared between groups. Differences in immune cell subsets between nutritional groups over time were determined using generalized estimating equations accounting for the correlation between repeated measurements. RESULTS: We did not observe that nutritional status was associated with infectious or alloimmune events or event-free survival post-transplant. We demonstrated that children with obesity had distinct T-and B-cell signatures relative to those with undernutrition and normal nutrition, even when controlling for immunosuppression. Children with obesity had a lower frequency of CD8 Tnaive cells 9-month post-transplant (p < .001), a higher frequency of CD4 CD57 + PD1- T cells, and lower frequencies of CD57-PD1+ CD8 and CD57-PD1- CD8 T cells at 12-month transplant (p < .05 for all). CONCLUSIONS: Children with obesity have distinct immunophenotypes that may influence the tailoring of immunosuppression.

Human IL-10-producing B cells have diverse states that are induced from multiple B cell subsets.

Regulatory B cells (Bregs) suppress immune responses through the secretion of interleukin-10 (IL-10). This immunomodulatory capacity holds therapeutic potential, yet a definitional immunophenotype for enumeration and prospective isolation of B cells capable of IL-10 production remains elusive. Here, we simultaneously quantify cytokine production and immunophenotype in human peripheral B cells across a range of stimulatory conditions and time points using mass cytometry. Our analysis shows that multiple functional B cell subsets produce IL-10 and that no phenotype uniquely identifies IL-10+ B cells. Further, a significant portion of IL-10+ B cells co-express the pro-inflammatory cytokines IL-6 and tumor necrosis factor alpha (TNFα). Despite this heterogeneity, operationally tolerant liver transplant recipients have a unique enrichment of IL-10+, but not TNFα+ or IL-6+, B cells compared with transplant recipients receiving immunosuppression. Thus, human IL-10-producing B cells constitute an induced, transient state arising from a diversity of B cell subsets that may contribute to maintenance of immune homeostasis.

Understanding the processes in a historically relevant thermal and mineral spring water by using mixing and inverse geochemical models.

The thermal mineral water of Peñón de los Baños spa (Mexico City) has been used for over 500 years starting in pre-Hispanic times and is famous for the treatment of various pathologies. It has a temperature of 45 °C, which is rich in HCO3-, and its main trace elements are B, Li and Fe, which confers healing effects. Concerns about the sustainability of this important spa have motivated this study to understand the thermal system, possible hydraulic and hydrochemical changes over time and its implications. Stable water isotope data indicate that the thermal water sources originate from local precipitation at Sierra de las Cruces with a recharge elevation of approximately 2770 m above sea level. The recharged water percolates through volcanic and carbonate rock formations and ascends via fault structure conduits, where it eventually is extracted 25 km downstream in Peñon de los Baños. During the gravity-driven deep circulation of up to 4.9 km, the groundwater is heated up to 136-160 °C. A comparison of past and current water levels and water chemical analyses indicates a water table drop and few variations in the chemical composition, confirming the presence of anthropic impact on water quality. Due to the heavy groundwater extractions in Mexico City, the spring water flow has ceased, and water must be pumped now from a 203-m deep well. In addition, the concentration of bicarbonate, sodium and chloride has been reduced by half since the onset of groundwater development. The therapeutic effects of this thermal mineral water are at risk due to the alteration of the chemical signature. However, new and different therapeutical uses may prevent a future deterioration or closure of this historically important thermal spa. It is crucial to establish a monitoring program of the thermal mineral water and reducing or minimizing nearby urban extractions which tap the regional flow component to preserve the properties of the thermal water.

Mitochondrial Oxidation of the Cytoplasmic Reducing Equivalents at the Onset of Oxidant Stress in the Isoproterenol-Induced Rat Myocardial Infarction.

We have developed and characterized a model of isoproterenol (ISO)-induced myocardial necrosis, identifying three stages of cardiac damage: a pre-infarction (0-12 h), infarction (24 h), and post-infarction period (48-96 h). Using this model, we have previously found alterations in calcium homeostasis and their relationship with oxidant stress in mitochondria, which showed deficient oxygen consumption and coupled ATP synthesis. Therefore, the present study was aimed at assessing the mitochondrial ability to transport and oxidize cytoplasmic reducing equivalents (NADH), correlating the kinetic parameters of the malate-aspartate shuttle, oxidant stress, and mitochondrial functionality. Our results showed only discreet effects during the cardiotoxic ISO action on the endogenous malate-aspartate shuttle activity, suggesting that endogenous mitochondrial NADH oxidation capacity (Nohl dehydrogenase) was not affected by the cellular stress. On the contrary, the reconstituted system showed significant enhancement in maximal capacity of the malate-aspartate shuttle activity only at later times (post-infarction period), probably as a compensatory part of cardiomyocytes' response to the metabolic and functional consequences of the infarcted tissue. Therefore, these findings support the notion that heart damage associated with myocardial infarction suffers a set of sequential biochemical and metabolic modifications within cardiomyocytes, where mitochondrial activity, controlling the redox state, could play a relevant role.

Functional Consequences of Memory Inflation after Solid Organ Transplantation.

CMV is a major infectious complication following solid organ transplantation. Reactivation of CMV leads to memory inflation, a process in which CD8 T cells expand over time. Memory inflation is associated with specific changes in T cell function, including increased oligoclonality, decreased cytokine production, and terminal differentiation. To address whether memory inflation during the first year after transplantation in human subjects alters T cell differentiation and function, we employed single-cell-matched TCRαß and targeted gene expression sequencing. Expanded T cell clones exhibited a terminally differentiated, immunosenescent, and polyfunctional phenotype whereas rare clones were less differentiated. Clonal expansion occurring between pre- and 3 mo posttransplant was accompanied by enhancement of polyfunctionality. In contrast, polyfunctionality and differentiation state were largely maintained between 3 and 12 mo posttransplant. Highly expanded clones had a higher degree of polyfunctionality than rare clones. Thus, CMV-responsive CD8 T cells differentiated during the pre- to posttransplant period then maintained their differentiation state and functional capacity despite posttransplant clonal expansion.

Evolution of Cytomegalovirus-Responsive T Cell Clonality following Solid Organ Transplantation.

CMV infection is a significant complication after solid organ transplantation. We used single cell TCR αß sequencing to determine how memory inflation impacts clonality and diversity of the CMV-responsive CD8 and CD4 T cell repertoire in the first year after transplantation in human subjects. We observed CD8 T cell inflation but no changes in clonal diversity, indicating homeostatic stability in clones. In contrast, the CD4 repertoire was diverse and stable over time, with no evidence of CMV-responsive CD4 T cell expansion. We identified shared CDR3 TCR motifs among patients but no public CMV-specific TCRs. Temporal changes in clonality in response to transplantation and in the absence of detectable viral reactivation suggest changes in the repertoire immediately after transplantation followed by an expansion with stable clonal competition that may mediate protection.