Tryptophan Modulation in Cancer-Associated Cachexia Mouse Models.

Cancer cachexia is a multifactorial syndrome that interferes with treatment and reduces the quality of life and survival of patients. Currently, there is no effective treatment or biomarkers, and pathophysiology is not clear. Our group reported alterations on tryptophan metabolites in cachectic patients, so we aim to investigate the role of tryptophan using two cancer-associated cachexia syngeneic murine models, melanoma B16F10, and pancreatic adenocarcinoma that is KPC-based. Injected mice showed signs of cancer-associated cachexia as reduction in body weight and raised spleen weight, MCP1, and carbonilated proteins in plasma. CRP and Myostatin also increased in B16F10 mice. Skeletal muscle showed a decrease in quadriceps weight and cross-sectional area (especially in B16F10). Higher expression of atrophy genes, mainly Atrogin1, was also observed. Plasmatic tryptophan levels in B16F10 tumor-bearing mice decreased even at early steps of tumorigenesis. In KPC-injected mice, tryptophan fluctuated but were also reduced and in cachectic patients were significantly lower. Treatment with 1-methyl-tryptophan, an inhibitor of tryptophan degradation, in the murine models resulted in the restoration of plasmatic tryptophan levels and an improvement on splenomegaly and carbonilated proteins levels, while changes in plasmatic inflammatory markers were mild. After the treatment, CCR2 expression in monocytes diminished and lymphocytes, Tregs, and CD8+, were activated (seen by increased in CD127 and CD25 expression, respectively). These immune cell changes pointed to an improvement in systemic inflammation. While treatment with 1-MT did not show benefits in terms of muscle wasting and atrophy in our experimental setting, muscle functionality was not affected and central nuclei fibers appeared, being a feature of regeneration. Therefore, tryptophan metabolism pathway is a promising target for inflammation modulation in cancer-associated cachexia.

Shorter telomere length is associated with COVID-19 hospitalization and with persistence of radiographic lung abnormalities.

BACKGROUND: Age and comorbidity are the main determinants of COVID-19 outcome. Shorter leukocyte telomere length (TL), a hallmark of biological aging, has been associated with worse COVID-19 outcomes. We sought to determine TL in patients with severe COVID-19 requiring hospitalization to analyze whether clinical outcomes and post-COVID-19 manifestations are associated with shorter TL. RESULTS: We analyzed 251 patients with PCR-confirmed COVID-19, hospitalized in the first months of the pandemics. We determined TL in PBL at admission by quantitative-PCR (qPCR) analysis in patients. A healthy cohort from the same area with a similar age range (n = 169) was used to calculate TL Z-scores. After hospital discharge, 144 COVID-19 survivors were followed-up for persistent COVID-19 manifestations. A second TL determination was performed in a smaller group of 63 patients 1 year later and compared with baseline TL. Hospitalized COVID-19 patients had a decreased baseline age-adjusted TL Z-score compared to the reference group. No differences in Z-scores were observed in patients with different COVID-19 outcomes, classified as WHO ordinal scores. In 144 patients, followed for a median of 8 months, post-COVID manifestations were not associated to differences in TL. Persistence of lung radiographic abnormalities was associated with shorter baseline TL. In patients with a second TL determination, further telomere shortening (TS) was observed in 35% and telomere lengthening in 49%. Patients with further TS had suffered a more severe disease. CONCLUSION: Shorter TL is associated with COVID-19 hospitalization but not with hospital clinical outcomes nor with persistent post-COVID-19 manifestations. Delayed resolution of radiographic lung abnormalities was also associated with shorter TL.

Effective Natural Killer Cell Degranulation Is an Essential Key in COVID-19 Evolution.

NK degranulation plays an important role in the cytotoxic activity of innate immunity in the clearance of intracellular infections and is an important factor in the outcome of the disease. This work has studied NK degranulation and innate immunological profiles and functionalities in COVID-19 patients and its association with the severity of the disease. A prospective observational study with 99 COVID-19 patients was conducted. Patients were grouped according to hospital requirements and severity. Innate immune cell subpopulations and functionalities were analyzed. The profile and functionality of innate immune cells differ between healthy controls and severe patients; CD56dim NK cells increased and MAIT cells and NK degranulation rates decreased in the COVID-19 subjects. Higher degranulation rates were observed in the non-severe patients and in the healthy controls compared to the severe patients. Benign forms of the disease had a higher granzymeA/granzymeB ratio than complex forms. In a multivariate analysis, the degranulation capacity resulted in a protective factor against severe forms of the disease (OR: 0.86), whereas the permanent expression of NKG2D in NKT cells was an independent risk factor (OR: 3.81; AUC: 0.84). In conclusion, a prompt and efficient degranulation functionality in the early stages of infection could be used as a tool to identify patients who will have a better evolution.

Number and function of circulatory helper innate lymphoid cells are unaffected by immunosuppressive drugs used in solid organ recipients - a single centre cohort study.

In transplanted intestines, depletion of T cells together with long-term persistence of ILC is observed, suggesting ILC insensitivity to immunosuppressive drugs. To further analyze helper ILC (hILC) apparent resistance to therapy, cytotoxic ILC (NK cells), hILC subsets (ILC1, ILC2, and ILC precursors (ILCP)), and their signature cytokines (IFNγ, IL4 + IL13, and IL22) were analyzed in peripheral blood of kidney and liver transplant recipients. Early after transplantation (posTx), transplanted patients showed significantly lower Lin + and NK cells, whereas total hILC, ILC1, ILC2, and ILCP numbers were similar in patients and controls. Between paired pre- and posTx samples, Lin + cell and NK cell counts significantly decreased, whereas all three hILC counts and their cytokine production remained similar. ILC1, ILC2, and ILCP numbers were also similar in patients under thymoglobulin or basiliximab (BAS), patients without induction (only maintenance therapy) and controls. hILC showed lower TMG binding comparing to Lin + cells, reduced expression of CD25 (BAS target), and diminished calcineurin activity with undetectable calcineurin and FKBP12 (tacrolimus target). hILC counts were not related to delayed graft function or biopsy-proven acute rejection. Thus, hILC remain stable early after transplantation and seem unaffected by immunosuppressors, which may be related to reduced targets expression and low calcineurin activity.

Donor-Specific Antibodies in Pediatric Intestinal and Multivisceral Transplantation: The Role of Liver and Human Leukocyte Antigen Mismatching.

Rejection is one of the most important drawbacks for graft and patient survival in intestinal and multivisceral transplantation. However, there is no consensus on the diagnostic criteria for humoral rejection, and the literature about the role of donor-specific antibodies (DSA) on allograft outcome and the risk factors that contribute to their development is scant with contradictory results. The present study analyzes the role of DSA exclusively in a pediatric cohort of 43 transplants. Among our patients, 11.6% showed preformed DSA, but they did not correlate with more rejection or less allograft survival. Having previous transplants was the main sensitization factor with an odds ratio (OR) = 44.85 (P = 0.001). In total, 16.3% of recipients developed de novo donor-specific antibodies (dnDSA), mostly directed against human leukocyte antigen (HLA) class II, polyspecific and complement fixing. Additionally, the presence of dnDSA had a deleterious effect on graft rejection (hazard ratio [HR] = 11.00; P = 0.01) and survival (HR = 66.52; P < 0.001) in an observational period of 5 years after transplantation. The inclusion of the liver emerged as the main protective factor against dnDSA development with an OR = 0.07 (P = 0.007). The analysis of HLA compatibility at the serological and epitope level with the computational tools HLAMatchmaker and PIRCHE revealed no association between HLA mismatching and dnDSA. In conclusion, this study performed in pediatric recipients shows the deleterious effect of dnDSA on intestinal transplantation supported by the complement-fixing activity observed. Additionally, the liver inclusion in the allografts showed to be a protective factor against dnDSA generation.

High frequency of central memory regulatory T cells allows detection of liver recipients at risk of early acute rejection within the first month after transplantation.

Several studies have analyzed the potential of T regulatory cells (Treg cells) as biomarkers of acute rejection (AR). The aim of the present multicenter study was to correlate the percentage of peripheral Treg cells in liver graft recipients drawn at baseline up to 12 months after transplantation with the presence of AR. The percentage of central memory (cm) Treg cells (CD4(+)CD25(high)CD45RO(+)CD62L(+)) was monitored at pre-transplant and at 1 and 2 weeks, and 1, 2, 3 and 6 months and 1 year post-transplantation. The same validation standard operating procedures were used in all participating centers. Fifteen patients developed AR (23.4%). Hepatitis C virus recurrence was observed in 16 recipients, who displayed low peripheral blood cmTreg levels compared with patients who did not. A steady increase of cmTregs was observed during the first month after transplantation with statistically significant differences between AR and non-AR patients. The high frequency of memory Treg cells allowed us to monitor rejection episodes during the first month post-transplantation. On the basis of these data, we developed a prediction model for assessing risk of AR that can provide clinicians with useful information for managing patients individually and customizing immunosuppressive therapies.

5-gene differential expression predicts stability of human intestinal allografts.

In intestinal allografts, endoscopy and histology detect the injury once changes in the bowel wall architecture have occurred. We aimed to identify a molecular signature that could predict early deterioration, within histologically indistinguishable biopsies with "minimal changes" (MC) pathology. Sixty biopsies from 12 adult recipients were longitudinally taken during 8years post-transplant. They were classified as either stable (STA) or non-stable (NSTA) according to the prospectively recorded number, frequency and severity of rejection events of the allograft. In a discovery set of MC samples analyzed by RNA-Seq, 816 genes were differentially expressed in STA vs NSTA biopsies. A group of 5 genes (ADH1C, SLC39A4, CYP4F2, OPTN and PDZK1) correctly classified all NSTA biopsies in the discovery set and all STA biopsies from an independent set. These results were validated by qPCR in a new group of MC biopsies. Based on a logistic regression model, a cutoff of 0.28 predicted the probability of being a NSTA biopsy with 85% sensitivity and 69% specificity. In conclusion, by analyzing MC samples early after transplantation, the expression of a 5-gene set may predict the evolution of the bowel allograft. This prognostic biomarker may be of help to personalize care of the intestinal transplant recipient.

Isolated De Novo Antiendothelial Cell Antibodies and Kidney Transplant Rejection.

BACKGROUND: Studies analyzing the role of antiendothelial cell antibodies (AECAs) in large series of kidney transplant recipients are scarce, and HLA, MHC (major histocompatibility complex) class I-related chain A (MICA), and angiotensin II type 1 receptor have not been formally excluded as targets. STUDY DESIGN: Retrospective study of a cohort of kidney transplant recipients. SETTING & PARTICIPANTS: 324 kidney transplant recipients who were negative for anti-HLA, anti-MICA, and anti-angiotensin II type 1 receptor antibodies were tested for AECAs in pre- and posttransplantation serum samples. PREDICTORS: AECA-positive (preformed [pre+/post+] vs de novo [pre-/post+]) versus AECA-negative (pre-/post-) before or after transplantation. OUTCOMES: Patient mortality, transplant loss, and acute rejection events. RESULTS: 66 (20%) patients were AECA positive (39 [12%] preformed, 27 [8%] de novo) and 258 (80%) were AECA negative. During a follow-up of 10 years, 7 (18%) AECA pre+/post+ patients had rejections compared with 14 (52%) AECA pre-/post+ and 57 (22%) AECA pre-/post- recipients (OR, 3.80; P=0.001). AECA pre-/post+ status emerged as an independent risk factor for transplant rejection compared to the AECA pre-/post- group (OR, 5.17; P<0.001). However, AECA pre+/post+ and AECA pre-/post+ patients did not show higher risk for either patient death (ORs of 1.49 [P=0.7] and 1.06 [P=0.9], respectively) or transplant loss (ORs of 1.22 and 0.86, respectively; P for both = 0.8) compared to the AECA pre-/post- population. LIMITATIONS: Retrospective study. Posttransplantation sera were collected before or after rejection, entailing a nearly cross-sectional relationship between the exposure and outcome. Lack of identification of precise antigens for AECAs. CONCLUSIONS: De novo AECAs may be associated with rejection. These antibodies might serve as biomarkers of endothelium damage in kidney transplant recipients.

Early renal graft function deterioration in recipients with preformed anti-MICA antibodies: partial contribution of complement-dependent cytotoxicity.

BACKGROUND: We previously reported that preformed anti-MHC class I-related chain A (MICA) antibodies increase the risk for renal graft rejection and enhance the deleterious effect of PRA(+) status early after transplantation. METHODS: We studied 727 kidney recipients. Days to reach optimal serum creatinine level, estimated glomerular filtration rate (eGFR) at Month 3 and chronic kidney disease (CKD) stages were recorded. Anti-MICA specificities and C1q binding were tested by solid-phase assay. Complement-dependent cytotoxicity (CDC) and flow cytometry (FC) cross-matches with HeLa and PMA/CD28-T-blasts were performed. RESULTS: PRA(+)MICA(+) recipients exhibited longer time to reach optimal serum creatinine level after transplantation (P = 0.005) and had the lowest eGFR at Month 3 (P = 0.006). PRA(+)MICA(+) status independently increased the risk for CKDT stage 5 at Month 3 [hazard ratio (HR) 4.92, P = 0.030]. Pre-transplant anti-MICA antibodies were polyspecific and showed stronger reactions when coexisting with anti-HLA antibodies (mean standard fluorescent intensity 112 157 ± 44 426 in HLA(+)MICA(+) sera versus 49 680 ± 33 116 in HLA(-)MICA(+) sera, P = 0.0006). Anti-AYVE supereplet reactivity was significantly higher in HLA(+)MICA(+) versus HLA(-)MICA(+) patients (P < 0.001) and significantly superior than anti-CMGWS supereplet within HLA(+)MICA(+) patients (P = 0.001). Three of 13 anti-MICA(+) pre-transplant sera were positive for the C1q binding assay; one of them (serum 3) exclusively recognized AYVE supereplet with a strong reactivity against MICA*027 antigen (same as MICA*008). Anti-MICA antibodies in anti-HLA-absorbed serum 3 bound native MICA molecules in MICA*008(+) HeLa and PMA/CD28-T-blasts and mediated cell death by activating complement. CONCLUSION: Preformed anti-MICA antibodies may occasionally be cytotoxic by fixing and activating complement. This way they might contribute to worse early kidney graft function.

Improving the Access of Highly Sensitized Patients to Kidney Transplantation From Deceased Donors: The Spanish PATHI Program With Allocation Based on the Virtual Crossmatch.

BACKGROUND: In 2015, the Spanish National Transplant Organization developed a prioritization system (Program for Access to Transplantation for Highly Sensitized Patients [PATHI]) to increase transplant options for patients with calculated panel-reactive antibodies (cPRAs) ≥98%, based on virtual crossmatch. We describe the experience with the implementation of PATHI and assess its efficacy. METHODS: PATHI registry was used to collect characteristics of donors and patients between June 15, 2015, and March 1, 2018. One-year graft and patient survival and acute rejection were also measured. A Cox model was used to identify factors related to patient death and graft loss and logistical regression for those associated with rejection. RESULTS: One thousand eighty-nine patients were included, and 272 (25%) were transplanted. Transplant rate by cPRA was 54.9%, 40.5%, and 12.8% in patients with cPRA98%, cPRA99%, and cPRA100%, respectively. One-year patient survival was 92.5%. Recipient age ≥60, time under dialysis >7 y, and delayed graft function were mortality risk factors. One-year graft survival was 88.7%. The factor related to graft loss was delayed graft function. The rejection rate was 22%. Factors related to rejection were sex, older recipients, and posttransplant donor-specific antibodies. CONCLUSIONS: A prioritization approach increases transplant options for highly sensitized patients with appropriate short-term postransplant outcomes. Along with other programs, PATHI may inspire other countries to adopt strategies to meet transplant needs of these patients.

A personalised delisting strategy enables successful kidney transplantation in highly sensitised patients with preformed donor-specific anti HLA antibodies.

This study investigates kidney transplant outcomes in highly sensitised patients after implementing a delisting strategy aimed at enabling transplantation despite preformed donor-specific antibodies (preDSA), with the goal of reducing acute antibody-mediated rejection (aAMR) risk. Fifty-three sensitised recipients underwent kidney transplant after delisting prohibited HLA antigens, focusing initially in low MFI antibodies (<5000), except for anti-HLA-DQ. If insufficient, higher MFI antibodies were permitted, especially for those without an immunogenic eplet pattern assigned. Delisting of Complement-fixing antibodies (C1q+) was consistently avoided. Comparison cohorts included 53 sensitised recipients without DSA (SwoDSA) and 53 non-sensitised (NS). The average waiting time prior to delisting was 4.4 ± 1.8 years, with a reduction in cPRA from 99.7 ± 0.5 to 98.1 ± 0.7, followed by transplantation within 7.2 ± 8.0 months (analysed in 34 patients). Rejection rates were similar among preDSA, SwoDSA, and NS groups (16%, 8%, and 11%, respectively; p = 0.46). However, aAMR was higher in the preDSA group (12%, 4%, and 2%, respectively; p = 0.073), only presented in recipients with DSA of MFI >5000. The highest MFI DSA were against HLA-DP (Median: 10796 MFI), with 50% of preDSA aAMR cases due to anti-DP antibodies (n = 3). Graft survival rates at 1 and 5 years in preDSA group were 94%, and 67%, comparable to SwoDSA (94%, and 70%; p = 0.69), being significantly higher in the NS group (p = 0.002). The five-year recipient survival rate was 89%, comparable to SwoDSA and NS groups (p = 0.79). A delisting strategy enables safe kidney transplant in highly sensitised patients with preDSA, with a slight increase in aAMR and comparable graft and patient survivals to non-DSA cohorts.

Androgen receptor polyQ alleles and COVID-19 severity in men: A replication study.

BACKGROUND: Ample evidence indicates a sex-related difference in severity of COVID-19, with less favorable outcomes observed in men. Genetic factors have been proposed as candidates to explain this difference. The polyglutamine (polyQ) polymorphism in the androgen receptor gene has been recently described as a genetic biomarker of COVID-19 severity. OBJECTIVE: To test the association between the androgen receptor polyQ polymorphism and COVID-19 severity in a large cohort of COVID-19 male patients. MATERIALS AND METHODS: This study included 1136 male patients infected with SARS-CoV-2 as confirmed by positive PCR. Patients were retrospectively and prospectively enrolled from March to November 2020. Patients were classified according to their severity into three categories: oligosymptomatic, hospitalized and severe patients requiring ventilatory support. The number of CAG repeats (polyQ polymorphism) at the androgen receptor was obtained by PCR and patients were classified as either short (<23 repeats) or long (≥23 repeats) allele carriers. The association between polyQ alleles (short or long) and COVID-19 severity was assessed by Chi-squared (Chi2 ) and logistic regression analysis. RESULTS: The mean number of polyQ CAG repeats was 22 (±3). Patients were classified as oligosymptomatic (15.5%), hospitalized (63.2%), and severe patients (21.3%) requiring substantial respiratory support. PolyQ alleles distribution did not show significant differences between severity classes in our cohort (Chi2 test p > 0.05). Similar results were observed after adjusting by known risk factors such as age, comorbidities, and ethnicity (multivariate logistic regression analysis). DISCUSSION: Androgen sensitivity may be a critical factor in COVID-19 disease severity. However, we did not find an association between the polyQ polymorphism and the COVID-19 severity. Additional studies are needed to clarify the mechanism underlying the association between androgens and COVID-19 outcome. CONCLUSIONS: The results obtained in our study do not support the role of this polymorphism as biomarker of COVID-19 severity.

Age-dependent association of clonal hematopoiesis with COVID-19 mortality in patients over 60 years.

Clonal hematopoiesis, especially that of indeterminate potential (CHIP), has been associated with age-related diseases, such as those contributing to a more severe COVID-19. Four studies have attempted to associate CHIP with COVID-19 severity without conclusive findings. In the present work, we explore the association between CHIP and COVID-19 mortality. Genomic DNA extracted from peripheral blood of COVID-19 patients (n = 241 deceased, n = 239 survivors) was sequenced with the Myeloid Solutions™ panel of SOPHiA Genetics. The association between clonality and age and clonality and mortality was studied using logistic regression models adjusted for sex, ethnicity, and comorbidities. The association with mortality was performed with patients stratified into four groups of age according to the quartiles of the distribution: 60-74 years, 75-84 years, 85-91 years, and 92-101 years. Clonality was found in 38% of the cohort. The presence of CHIP variants, but not the number, significantly increased with age in the entire cohort of COVID-19 patients, as well as in the group of survivors (p < 0.001). When patients were stratified by age and the analysis adjusted, CHIP classified as pathogenic/likely pathogenic was significantly more represented in deceased patients compared with survivors in the group of 75-84 years (34.6% vs 13.7%, p = 0.020). We confirmed the well-established linear relationship between age and clonality in the cohort of COVID-19 patients and found a significant association between pathogenic/likely pathogenic CHIP and mortality in patients from 75 to 84 years that needs to be further validated.

Antibody-removal therapies for de novo DSA in pediatric intestinal recipients: Why, when, and how? A single-center experience.

Background: Donor-specific anti-HLA antibodies (DSA) impact negatively on the outcome of intestinal grafts. Although the use of antibody-removal therapies (ART) is becoming more frequent in the last few years, issues regarding their timing and effectiveness remain under discussion. Methods: In the present study, we report our experience with eight ART procedures (based on plasmapheresis, intravenous immunoglobulin, and rituximab) in eight pediatric intestinal and multivisceral transplants with de novo DSA (dnDSA). Results: ART were performed when dnDSA appeared in two contexts: (1) concomitant with rejection (acute or chronic) or (2) without rejection or any other clinical symptom. Complete DSA removal was observed in seven out of eight patients, showing an effectiveness of 88%. In the group treated for dnDSA without clinical symptoms, the success rate was 100%, with complete DSA removal and without rejection afterward. A shorter time between DSA detection and ART performance appeared as a significant factor for the success of the therapy (p = 0.0002). DSA against HLA-A and DQ alleles were the most resistant to ART, whereas anti-DR DSA were the most sensitive. In addition, the 8-year allograft survival rate in recipients undergoing ART was similar to that in those without DSA, being significantly lower in non-treated DSA-positive recipients (p = 0.013). Conclusion: The results confirm the effectiveness of ART in terms of DSA removal and allograft survival and encourage its early use even in the absence of clinical symptoms.

Presence of rare potential pathogenic variants in subjects under 65 years old with very severe or fatal COVID-19.

Rare variants affecting host defense against pathogens could be involved in COVID-19 severity and may help explain fatal outcomes in young and middle-aged patients. Our aim was to report the presence of rare genetic variants in certain genes, by using whole exome sequencing, in a selected group of COVID-19 patients under 65 years who required intubation or resulting in death (n = 44). To this end, different etiopathogenic mechanisms were explored using gene prioritization-based analysis in which genes involved in immune response, immunodeficiencies or blood coagulation were studied. We detected 44 different variants of interest, in 29 different patients (66%). Some of these variants were previously described as pathogenic and were located in genes mainly involved in immune response. A network analysis, including the 42 genes with candidate variants, showed three main components, consisting of 25 highly interconnected genes related to immune response and two additional networks composed by genes enriched in carbohydrate metabolism and in DNA metabolism and repair processes. In conclusion, we have detected candidate variants that may potentially influence COVID-19 outcome in our cohort of patients. Further studies are needed to confirm the ultimate role of the genetic variants described in the present study on COVID-19 severity.

An Early Th1 Response Is a Key Factor for a Favorable COVID-19 Evolution.

The Th1/Th2 balance plays a crucial role in the progression of different pathologies and is a determining factor in the evolution of infectious diseases. This work has aimed to evaluate the early, or on diagnosis, T-cell compartment response, T-helper subsets and anti-SARS-CoV-2 antibody specificity in COVID-19 patients and to classify them according to evolution based on infection severity. A unicenter, randomized group of 146 COVID-19 patients was divided into four groups in accordance with the most critical events during the course of disease. The immunophenotype and T-helper subsets were analyzed by flow cytometry. Asymptomatic SARS-CoV-2 infected individuals showed a potent and robust Th1 immunity, with a lower Th17 and less activated T-cells at the time of sample acquisition compared not only with symptomatic patients, but also with healthy controls. Conversely, severe COVID-19 patients presented with Th17-skewed immunity, fewer Th1 responses and more activated T-cells. The multivariate analysis of the immunological and inflammatory parameters, together with the comorbidities, showed that the Th1 response was an independent protective factor for the prevention of hospitalization (OR 0.17, 95% CI 0.03-0.81), with an AUC of 0.844. Likewise, the Th1 response was found to be an independent protective factor for severe forms of the disease (OR 0.09, 95% CI: 0.01-0.63, p = 0.015, AUC: 0.873). In conclusion, a predominant Th1 immune response in the acute phase of the SARS-CoV-2 infection could be used as a tool to identify patients who might have a good disease evolution.

Rationale and Protocol of the Multimodality Evaluation of Antibody-Mediated Injury in Heart Transplantation (LEONE-HT) Observational Cross-Sectional Study.

INTRODUCTION: Heart transplant (HT) survival has barely improved in the last decades, which is unsatisfactory for many HT recipients. The development of anti-human leukocyte antigen (anti-HLA) antibodies in HT patients is associated with a cardiac allograft dysfunction. The mechanisms leading to this damage are unclear. The Multimodality Evaluation Of Antibody-Mediated Injury In Heart Transplantation (LEONE-HT) study aimed to thoroughly describe the damage inflicted on the myocardium by anti-HLA antibodies. METHODS AND ANALYSIS: The LEONE-HT study is a cohort study with a cross-sectional approach in which HT patients with positive anti-HLA antibodies are compared with coetaneous HT patients with negative anti-HLA antibodies. All patients will undergo a state-of-the-art multimodal assessment, including imaging techniques, coronary anatomy and physiology evaluations and histological and immunological analyses. The individual and combined primary outcomes of structural graft injuries and longitudinal secondary outcomes are to be compared between the exposed and non-exposed groups with univariate and multivariable descriptive analyses. ETHICS AND DISSEMINATION: The LEONE-HT study is carried out in accordance with the principles set out in the Declaration of Helsinki and the International Conference on Harmonization guidelines for good clinical practice and following national laws and regulations. The study design, objectives and participant centers have been communicated to clinicaltrials.gov (NCT05184426). The LEONE-HT study counts on the support of patient associations to disseminate the objectives and results of the research. This study was funded by the Spanish Ministry of Science and Innovation and the Spanish Society of Cardiology.

Phenotypic and functional evaluation of CD3+CD4-CD8- T cells in human CD8 immunodeficiency.

BACKGROUND: Human CD8 immunodeficiency is characterized by undetectable CD8(+) lymphocytes and an increased population of CD4(-)CD8(-) (double negative) T lymphocytes. DESIGN AND METHODS: We hypothesized that the double negative subset corresponds to the cellular population that should express CD8 and is committed to the cytotoxic T lymphocyte lineage. To assess this, we determined the phenotype and function of peripheral blood mononuclear cells and/or magnetically isolated double negative T lymphocytes from two CD8-deficient patients. To analyze the expression and co-localization with different organelles, 293T cells were transfected with plasmids bearing wild-type or mutated CD8α. RESULTS: CD8α mutated protein was retained in the cytoplasm of transfected cells. The percentages of double negative cells in patients were lower than the percentages of CD8(+) T cells in healthy controls. Double negative cells mostly had an effector or effector memory phenotype whereas naïve T cells were under-represented. A low concentration of T-cell receptor excision circles together with a skewed T-cell receptor-V repertoire were observed in the double negative population. These data suggest that, in the absence of CD8 co-receptor, the thymic positive selection functions suboptimally and a limited number of mature T-cell clones would emerge from the thymus. In vitro, the double negative cells showed a mild defect in cytotoxic function and decreased proliferative capacity. CONCLUSIONS: It is possible that the double negative cells are major histocompatibility complex class-I restricted T cells with cytolytic function. These results show for the first time in humans that the presence of the CD8 co-receptor is dispensable for cytotoxic ability, but that it affects the generation of thymic precursors committed to the cytotoxic T lymphocyte lineage and the proliferation of mature cytotoxic T cells.

T-Helper Cell Subset Response Is a Determining Factor in COVID-19 Progression.

The immune response type organized against viral infection is determinant in the prognosis of some infections. This work has aimed to study Th polarization in acute COVID-19 and its possible association with the outcome through an observational prospective study. Fifty-eight COVID-19 patients were recruited in the Medicine Department of the hospital "12 de Octubre," 55 patients remaining after losses to follow-up. Four groups were established according to maximum degree of disease progression. T-helper cell percentages and phenotypes, analyzed by flow cytometer, and serum cytokines levels, analyzed by Luminex, were evaluated when the microbiological diagnosis (acute phase) of the disease was obtained. Our study found a significant reduction of %Th1 and %Th17 cells with higher activated %Th2 cells in the COVID-19 patients compared with reference population. A higher percent of senescent Th2 cells was found in the patients who died than in those who survived. Senescent Th2 cell percentage was an independent risk factor for death (OR: 13.88) accompanied by the numbers of total lymphocytes (OR: 0.15) with an AUC of 0.879. COVID-19 patients showed a profile of pro-inflammatory serum cytokines compared to controls, with higher levels of IL-2, IL-6, IL-15, and IP-10. IL-10 and IL-13 were also elevated in patients compared to controls. Patients who did not survive presented significantly higher levels of IL-15 than those who recovered. No significant differences were observed according to disease progression groups. The study has shown that increased levels of IL-15 and a high Th2 response are associated with a fatal outcome of the disease.

Immunosuppression minimization in kidney transplant recipients hospitalized for COVID-19.

BACKGROUND: Immunosuppressed patients such as kidney transplant recipients (KTs) have increased mortality risk in the setting of coronavirus disease 2019 (COVID-19). The role and management of chronic immunosuppressive therapies during COVID-19 must be characterized. METHODS: Herein, we report the follow-up of a cohort of 47 KTs admitted at two Spanish Kidney Transplant Units, who survived COVID-19. The impact of the management of immunosuppression during COVID-19 on graft function and immunologic events was evaluated. RESULTS: At least one immunosuppressive agent was withdrawn in 83% of patients, with antimetabolites being the most frequent. Steroids were generally not stopped and the dose was even increased in 15% of patients as part of the treatment of COVID-19. Although immunosuppressive drugs were suspended during a median time of 17 days, no rejection episodes or de novo donor-specific antibodies were observed up to 3 months after discharge, and no significant changes occurred in calculated panel reactive antibodies. Acute graft dysfunction was common (55%) and the severity was related to tacrolimus trough levels, which were higher in patients receiving antivirals. At the end of follow-up, all patients recovered baseline kidney function. CONCLUSIONS: Our observational study suggests that immunosuppression in KTs hospitalized due to COVID-19 could be safely minimized.