Innate Lymphoid Cells Groups 1 and 3 in the Epithelial Compartment of Functional Human Intestinal Allografts.

We examined intraepithelial lymphocytes (IELs) in 213 ileal biopsies from 16 bowel grafts and compared them with 32 biopsies from native intestines. During the first year posttransplantation, grafts exhibited low levels of IELs (percentage of CD103(+) cells) principally due to reduced CD3(+) CD8(+) cells, while CD103(+) CD3(-) cell numbers became significantly higher. Changes in IEL subsets did not correlate with histology results, isolated intestine, or multivisceral transplants, but CD3(-) IELs were significantly higher in patients receiving corticosteroids. Compared with controls, more CD3(-) IELs of the grafts expressed CD56, NKp44, interleukin (IL)-23 receptor, retinoid-related orphan receptor gamma t (RORγt), and CCR6. No difference was observed in granzyme B, and CD3(-) CD127(+) cells were more abundant in native intestines. Ex vivo, and after in vitro activation, CD3(-) IELs in grafts produced significantly more interferon (IFN)-γ and IL-22, and a double IFNγ(+) IL-22(+) population was observed. Epithelial cell-depleted grafts IELs were cytotoxic, whereas this was not observed in controls. In conclusion, different from native intestines, a CD3(-) IEL subset predominates in grafts, showing features of natural killer cells and intraepithelial ILC1 (CD56(+) , NKp44(+) , CCR6(+) , CD127(-) , cytotoxicity, and IFNγ secretion), ILC3 (CD56(+) , NKp44(+) , IL-23R(+) , CCR6(+) , RORγt(+) , and IL-22 secretion), and intermediate ILC1-ILC3 phenotypes (IFNγ(+) IL-22(+) ). Viability of intestinal grafts may depend on the balance among proinflammatory and homeostatic roles of ILC subsets.

Post-transplant monitoring of NK cell counts as a simple approach to predict the occurrence of opportunistic infection in liver transplant recipients.

BACKGROUND: Monitoring of peripheral blood lymphocyte subpopulation (PBLS) counts might be useful for estimating the risk of infection after liver transplantation (LT). METHODS: We prospectively measured total lymphocyte and PBLS counts at baseline and post-transplant months 1 and 6 in 92 LT recipients. PBLS were enumerated by single-platform 6-color flow cytometry technology. Areas under receiver operating characteristic (ROC) curves were used to evaluate the accuracy of different PBLS for predicting cytomegalovirus (CMV) disease and overall opportunistic infection (OI). Adjusted hazard ratios (aHRs) for both outcomes were estimated by Cox regression. RESULTS: After a median follow-up of 730.0 days, 29 patients (31.5%) developed 38 episodes of OI (including 22 episodes of CMV disease). The counts of CD3(+) , CD4(+) , and CD8(+) T cells, and CD56(+) CD16(+) natural killer (NK) cells at month 1 were significantly lower in patients subsequently developing OI. The NK cell count was the best predictive parameter (area under ROC curve for predicting CMV disease: 0.78; P-value = 0.001). Patients with an NK cell count <0.050 × 10(3) cells/µL had higher cumulative incidences of CMV disease (P-value = 0.001) and overall OI (P-value <0.001). In the multivariate models, an NK cell count <0.050 × 10(3) cells/µL at month 1 post transplantation remained as an independent risk factor for CMV disease (aHR: 5.54; P-value = 0.003) and overall OI (aHR: 7.56; P-value <0.001). CONCLUSION: Post-transplant kinetics of NK cell counts may be used as a simple and affordable proxy to the cell-mediated immunity status in LT recipients and to their associated risk of OI.

Hypocomplementemia in kidney transplant recipients: impact on the risk of infectious complications.

The usefulness of monitoring of complement levels in predicting the occurrence of infection in kidney transplant (KT) recipients remains largely unknown. We prospectively assessed serum complement levels (C3 and C4) at baseline and at months 1 and 6 in 270 patients undergoing KT. Adjusted hazard ratios (aHRs) for infection in each posttransplant period were estimated by Cox regression. The prevalence of C3 hypocomplementemia progressively decreased from 21.5% at baseline to 11.6% at month 6 (p = 0.017), whereas the prevalence of C4 hypocomplementemia rose from 3.7% at baseline to 9.2% at month 1 (p = 0.004). Patients with C3 hypocomplementemia at month 1 had higher incidences of overall (p = 0.002), bacterial (p = 0.004) and fungal infection (p = 0.019) in the intermediate period (months 1-6). On multivariate analysis C3 hypocomplementemia at month 1 emerged as a risk factor for overall (aHR 1.911; p = 0.009) and bacterial infection (aHR 2.130; p = 0.014) during the intermediate period, whereas C3 hypocomplementemia at month 6 predicted the occurrence of bacterial infection (aHR 3.347; p = 0.039) in the late period (>6 month). A simple monitoring strategy of serum C3 levels predicts the risk of posttransplant infectious complications in KT recipients.

Monitoring of immunoglobulin levels identifies kidney transplant recipients at high risk of infection.

We aimed to analyze the incidence, risk factors and impact of hypogammaglobulinemia (HGG) in 226 kidney transplant (KT) recipients in which serum immunoglobulin (Ig) levels were prospectively assessed at baseline, month 1 (T(1) ), and month 6 (T(6) ). The prevalence of IgG HGG increased from 6.6% (baseline) to 52.0% (T(1) ) and subsequently decreased to 31.4% (T(6) ) (p < 0.001). The presence of IgG HGG at baseline (odds ratio [OR] 26.9; p = 0.012) and a positive anti-HCV status (OR 0.17; p = 0.023) emerged as risk factors for the occurrence of posttransplant IgG HGG. Patients with HGG of any class at T(1) had higher incidences of overall (p = 0.018) and bacterial infection (p = 0.004), bacteremia (p = 0.054) and acute pyelonephritis (p = 0.003) in the intermediate period (months 1-6). Patients with HGG at T(6) had higher incidences of overall (p = 0.004) and bacterial infection (p < 0.001) in the late period (>6 month). A complementary log-log model identified posttransplant HGG as an independent risk factor for overall (hazard ratio [HR] 2.03; p < 0.001) and bacterial infection (HR 2.68; p < 0.0001). Monitoring of humoral immunity identifies KT recipients at high risk of infection, offering the opportunity for preemptive immunoglobulin replacement therapy.

[Molecular remission of chronic myeloid leukaemia in a patient with hepatitis and a second kidney transplant]. / Remisión molecular de una leucemia mieloide crónica en un paciente con segundo trasplante renal y hepatitis.

Chronic myeloid leukemia (CML) is a myeloproliferative disorder characterized by clonal expansion of cells in the myeloid line, expressing the BCR-ABL fusion protein responsible for the oncogenic effect of CML. The current frontline therapy in CML is the BCR-ABL tyrosine kinase inhibitor, Imatinib. Although this drug has been shown to improve survival in CML patients, its role in the context of a transplant setting has not been widely described in the literature. We report on the long term molecular remission of CML in a 55 year old man with a second renal transplant who is hepatitis C virus positive, and has associated cardiovascular and immunological risk factors.

Follicular helper T cells and humoral response in organ transplantation.

Antibody mediated rejection has been recognized as an important contributor to long-term graft loss in most solid organ transplants. Current immunosuppressive regimes are not capable of preventing anti-HLA antibody formation and eventual damage to the graft, and there is a need to develop drugs directed against novel targets to avoid graft allorecognition. In this review we introduce follicular helper T cells (Tfh), a subtype of lymphocyte specialized in helping B cells to differentiate into plasmablasts and produce class-switched antibodies. We focus on the role of Tfh in solid organ transplantation, what is known about Tfh and the production of alloantibodies, how current immunosuppressive therapies affect Tfh and what new molecules could be used to target Tfh in transplantation, with the goal of improving graft survival.

Longitudinal analysis of immune function in the first 3 years of life in thymectomized neonates during cardiac surgery.

The purpose of this study is to evaluate the effects of neonatal thymectomy in the functional capacity of the immune system. We selected a group of 23 subjects, who had undergone thymectomy in their first 30 days of life, during an intervention for congenital heart disease. Several parameters of the immune system were evaluated during their first 3 years of life. Lymphocyte populations and subpopulations (including naive, memory and effector subpopulations), T cell receptor (TCR) Vbeta repertoire, response of T cells following in vitro stimulation by mitogen, quantification of immunoglobulins, TCR excision circles (TRECS) and interleukin (IL)-7 were measured. We found that neonatal thymectomy produces long-term diminution in total lymphocyte counts, especially in naive CD4+ and CD8+ T cells. Additionally, TRECS were decreased, and plasma IL-7 levels increased. A statistically significant negative correlation was found between absolute CD4+ T cells and IL-7 (r = -0.470, P = 0.02). The patients did not suffer more infectious events than healthy control children, but thymectomy in neonates resulted in a significant decrease in T lymphocyte levels and TRECS, consistent with cessation of thymopoiesis. This could produce a compromise in immune function later in life, especially if the patients suffer T cell depletion and need a reconstitution of immune function.

Evaluation of three fully automated immunoassay systems for detection of IgA anti-beta 2-glycoprotein I antibodies.

INTRODUCTION: In recent years, we have been witnessing increased clinical interest in the determination of IgA anti-beta 2-glycoprotein I (aB2GPI) antibodies as well as increased demand for this test. Some ELISA-based diagnostic systems for IgA aB2GPI antibodies detection are suboptimal to detect it. The aim of our study was to determine whether the diagnostic yield of modern detection systems based on automatic platforms to measure IgA aB2GPI is equivalent to that of the well-optimized ELISA-based assays. METHODS: In total, 130 patients were analyzed for IgA aB2GPI by three fully automated immunoassays using an ELISA-based assay as reference. The three systems were also analyzed for IgG aB2GPI with 58 patients. RESULTS: System 1 was able to detect IgA aB2GPI with good sensitivity and kappa index (99% and 0.72, respectively). The other two systems had also poor sensitivity (20% and 15%) and kappa index (0.10 and 0.07), respectively. On the other hand, kappa index for IgG aB2GPI was >0.89 in the three systems. CONCLUSION: Some analytical methods to detect IgA aB2GPI are suboptimal as well as some ELISA-based diagnostic systems. It is important that the scientific community work to standardize analytical methods to determine IgA aB2GPI antibodies.

Study of ß2-Glycoprotein I Polymorphisms in Patients With Chronic Renal Failure as a Predisposing Factor for the Development of Anti-ß2-Glycoprotein I Auto-Antibodies.

BACKGROUND: Immunoglobulin (Ig)A anti-ß2-glycoprotein I (aB2GP1) antibodies are associated with thrombotic events, cardiovascular morbidity, and death in dialysis patients. About 30% of patients with chronic renal disease are positive for IgA aB2GP1; however, the origin of these antibodies is unknown. It has been speculated that dialysis membranes, age, or etiology of renal base disease are possible precipitating factors, although these factors do not appear to be the source of antibodies. B2GP1 is a protein of 326 amino acids grouped into five domains. Eight polymorphisms have been described; the most important are Val/Leu247, which appears to predispose aB2GP1 antibody production in patients with anti-phospholipid syndrome, and Trp/Ser316, which appears to have protective antibody production of aB2GP1. METHODS: DNA samples from 92 patients with renal failure on hemodialysis were randomly collected with a 1:1 ratio for the positivity for IgA aB2GP1. Forty-six samples were positive for IgA aB2GP1 (group 1) and 46 negative for IgA aB2GP1 (group 2). All samples were anonymized to study polymorphism Val/Leu247 and polymorphism Trp/Ser316. RESULTS: No significant differences were observed between those who were positive or negative for IgA aB2GP1 in patients with renal failure treated with hemodialysis and the polymorphism located in codons 247 and 316. CONCLUSIONS: The two groups of patients have the same prevalence in polymorphisms 247 and 316, and therefore there appears not to be a genetic predisposition in our population. New trigger factors must be studied.

Month of birth, HLA-DRB1*15 locus and risk of multiple sclerosis in offspring. / Mes de nacimiento, HLA-DRB1 y riesgo de esclerosis multiple en la descendencia.

INTRODUCTION: A relationship among April births, HLA-DRB1*15:01 genotype and risk of multiple sclerosis (MS) has been described. We aim to determine this association in our cohort of Spanish MS patients. SUBJECTS AND METHODS: We genotyped HLA-DRB1*15:01 allele in 326 MS patients and 226 controls (non-neurological disease patients) by SSP-PCR and compared month of birth with local births during the same period. RESULTS: MS patients carrying HLA-DRB1*15 allele were more frequently born in December (10.3% HLA-DRB1*15+ vs. 3.8% HLA-DRB1*15-; p = 0.019). Controls carrying HLA-DRB1*15 allele were less frequently born in December than non-carrier controls (0% HLA-DRB1*15+ vs. 10.3% HLA-DRB1*15-; p = 0.028). Thus, December was confirmed as the common month of birth for HLA-DRB1*15-non-carrier controls and MS HLA-DRB1*15-carrier patients. CONCLUSIONS: Month of birth, HLA-DRB1 genotype and risk of MS are associated. In Spain, this association was found in December, supporting the potential interaction of a seasonal risk factor in winter, inside/close to HLA-DRB1*15 locus, during pregnancy or after birth.

TITLE: Mes de nacimiento, HLA-DRB1 y riesgo de esclerosis multiple en la descendencia.Introduccion. El haplotipo HLA-DRB1*1501 es el marcador genetico que se ha asociado con un riesgo tres veces mayor de padecer esclerosis multiple (EM) en caucasicos occidentales. Recientemente se ha sabido que hay una asociacion entre el mes de nacimiento en abril, el genotipo HLA-DRB1 y el riesgo de EM en paises del norte de Europa y Canada. Esto apoya la teoria de que debe haber una interaccion entre un factor de riesgo estacional con un locus cercano al HLA-DRB1*15 durante la gestacion o cerca del posparto. Sujetos y metodos. Se realizo el genotipado de la presencia y subtipo de HLA-DRB1*1501 en 326 pacientes de dos centros espa˜oles y en 226 controles sin patologia neurologica. Se compararon los meses de nacimiento de la muestra de pacientes con los nacimientos mensuales locales en los mismos periodos. Resultados. Comparados los pacientes con EM que eran portadores del alelo HLA-DRB1*15 (10,3%) frente a los pacientes no portadores (3,8%), significativamente mas pacientes nacian en diciembre (p = 0,0185). Tambien se confirmaba el mismo mes de nacimiento de diciembre entre sanos portadores frente a no portadores de HLA-DRB1*15 y entre pacientes portadores de HLA-DRB1*15 frente a sanos. Conclusiones. El mes de nacimiento, el genotipo HLA-DRB1*15 y el riesgo de presentar EM estan asociados. A diferencia de los resultados obtenidos en paises del norte de Europa, donde esta asociacion se ha encontrado en el mes de abril, en Espa˜a es en diciembre. Se demuestra la interaccion de un factor de riesgo estacional en invierno en el locus HLA-DRB1*15 o cercano a este durante la gestacion o tras el nacimiento.

Frequency in Spanish population of familial complement factor 2 type I deficits and associated HLA haplotypes.

We present two familial cases of complement factor 2 (C2) type I deficiency. Probands had experienced severe pyogenic bacteria infections in childhood and had undetectable levels of C2 and very low level of CH50. Both children were homozygous for the deletion of 28 bp in exon 6 of the C2 gene. Human leukocyte antigen (HLA) typing in family 1 had the commonly reported associations, but family 2 demonstrated a new association of the mutated C2 gene to HLA-A*3101, -Cw*0602, -B*1801, and -DRB1*0901. In addition, for the first time, the frequency of the 28-bp deletion of C2 and its HLA haplotypic associations have been analyzed in a sample of the Spanish population containing 790 haplotypes and 105 phenotypes. Cw*0602 is frequently found in Spanish haplotypes linked to the C2 mutated gene instead of the commonly reported -Cw*1203. The presence of heterozygous or homozygous individuals for the C2 deletion with low levels of IgD in both families supports the existence of a putative dominant susceptibility gene for IgD deficiency in haplotype HLA-B18, -S042, and -DR2. The frequency of the C2 28-bp deletion in heterozygosis is 1.4% (gene frequency 0.007) in Spanish healthy controls, similar to that reported in other white populations.

Activated Regulatory T Cells Expressing CD4(+)CD25(high)CD45RO(+)CD62L(+) Biomarkers Could Be a Risk Factor in Liver Allograft Rejection.

Activated regulatory T cells (aTregs) are nowadays a hot topic in organ transplantation to establish their role during acute rejection (AR) episodes. The aim of this multi-center study was to monitor the frequency of aTregs within the first year after transplantation in a cohort of first-time liver transplant recipients enrolled from 2010 to 2012. aTregs frequency was analyzed by means of flow cytometry. Patients who had AR showed higher levels of aTregs during first year after transplantation in comparison with patients who did not have higher levels. High levels of aTregs in liver recipients might be used as a biomarker of AR; however, further studies must be done to address the potential role of aTregs as biomarkers of AR in liver transplantation.

Peripheral blood reduction of memory (CD29+, CD45RO+, and "bright" CD2+ and LFA-1+) T lymphocytes in Papillon-Lefèvre syndrome.

A Papillon-Lefèvre patient with characteristic chronic periodontal disease and palmoplantar keratoderma was studied over a 4-year period. An abnormal T-cell phenotype was steadily observed in peripheral blood; both low numbers of CD29+ and CD45RO+ cells and a low density surface expression of CD2 and LFA-1 molecules were found. T-cell activation through CD3, CD2 and ConA, PWM and IL-2 receptors was normal; however, there was impairment in the activation via CD28. CD2, LFA-1 and CD45 molecules were normal in charge and molecular weight. There was no tissue sequestering of T lymphocytes in periodontal lesions, but rather a relative T-cell reduction. It is suggested that an important decrease of the so-called "memory/hyperreactive" (CD45RO-positive) T cells does exist; therefore, hyperreactive T cells would not be available in sufficient numbers to leave the bloodstream through blood vessel endothelium, and the periodontium would be left without these important defenses and thus exposed to chronic infections. A disregulated factor affecting the transition from "naive" to "memory" T cells and the increase in certain surface molecules expression (i.e., CD2, LFA-1, CD29, and CD45RO) or the reversion from memory to naive T cells may be responsible for the disease pathogenesis. CD2 and LFA-1 molecule synthesis might be conjointly regulated on T lymphocytes.

New species-specific alleles at the primate MHC-G locus.

Six different ape MHC-G DNA sequences (four in humans: HLA-G*01011, HLA-G*II, HLA-G*0103, and HLA-G*IV; one in chimpanzees: Patr-G*I; and one in gorillas: Gogo-G*1) have been obtained. Only synonymous or conservative ("Thr"-to-"Ser") substitutions are allowed between the four human alleles. One allele of MHC-G exon-2 sequences has been found both in gorilla (Gorilla gorilla) and chimpanzee (Pan troglodytes). The Patr-G*I DNA sequence shows two nonsynonymous substitutions when compared with the human HLA-G*01011 sequence: "CGG"-to-"TGG" ("Arg"-to-"Trp") at codon 35 and "ATG"-to-"ATA" ("Met"-to-"Ile") at codon 76. One nonsynonymous "GAG"-to-"GGG" ("Glu"-to-"Gly") substitution is observed in the Gogo-G*I exon-2 DNA sequence, when compared with the human *01011 allele. None of these three different substitutions have been observed in humans and are, thus, considered species specific. Also, evidence is provided that the human HLA-G*II and G*0103 may have been originated after human speciation. Finally, phylogenetic relationships among the six MHC-G alleles, tamarins G-"like" alleles, and other human class I genes (both "classical" and "nonclassic") are discussed.

Immunogenetics of systemic lupus erythematosus in Spanish patients: differential HLA markers.

HLA-DR3 antigen included in the compound phenotype B18BfF1 (but not the one linked to the B8BfS compound phenotype) was found to be significantly increased in our SLE patients. It is remarkable that in our Southern-Mediterranean population, B18BfF1DR3 individuals (but not B8BfSDR3) are prone to SLE with renal disease, in contrast with other Northern European and Caucasoid populations. Also, patients with autoantibodies to Ro/La have a significant increase of the B8DR3 compound phenotype. Production of autoantibodies against Ro alone was associated to DR2 and production of anti-Sm/nRNP to DR3 (either B18BfF1 or B8BfS associated) only in the subgroup without renal disease. The distinctive HLA and autoimmune associations to SLE with and without renal disease suggests that both clinical forms may not share a common identical pathogenesis.

Clinical value of a single determination of intracellular ATP levels in stimulated CD4+ T lymphocytes in pediatric patients with stable liver transplantation.

In the follow-up of transplanted patients under immunosupression, the functional assessment of CD4+ T cells activation by measuring intracellular ATP levels in vitro, using the Immuknow test give information on how immune system is functioning. Therefore, it has been reported that low levels of ATP correlate with the risk of opportunistic infection. Although limited, comprehensive results in adult recipients as well as in pediatric transplanted patients have been reported. Forty stable liver pediatric transplanted patients (mean age: 11.0 years [SD 5.65]), within at least 1 year of transplant were selected for a scheduled review, and an unique determination of Immuknow was performed. Average ATP levels were 317 ng/mL (200-400 ng/mL) which were similar to the values observed in adult population. ATP values among patients with monotherapy Cyclosporin A (CSA) or tacrolimus (TAC) were significantly higher (P = .005) than in patients with double immunosupressive therapy using either CSA/TAC + Mycophenolate Mofetil (MMF). In CSA treatment, there are significant differences (P = .0003) between monotherapy and double therapy, but in the case of TAC we did not find differences (P > .1). A single determination of levels of ATP on CD4+ lymphocytes, can provide additional information that could be used as a complementary test to guide immunosuppressive therapy in paediatric liver transplant recipients.

In vitro evaluation of new possible cell engraftment enhancers for cell transplantation.

Orthotopic liver transplantation (OLT) is the best treatment to restore liver function in liver failure. The low availability of organs has focused interest on the use of cell transplantation to restore liver function. However, this technique is limited because cells can not bind to liver parenchyma and die soon after perfusion. Pretransplant treatment with engraftment enhancers (EE) to increase vascular permeability may increase cell attachment. Using an endothelial cell culture to measure the loss of intercellular endothelial adhesion as a screening test, we evaluated the capacity of 15 monoclonal antibodies against adhesion molecules expressed on endothelial cells to act as EE showing that 3 antibodies (anti-CD54, efalizumab, and abciximab) act as EE by producing disruptions in the cell layer.

Prospective study on autoantibodies against apolipoprotein H (beta2GPI) in several clinical parameters from patients with terminal renal failure and functioning renal transplants.

Renal transplantation is the best therapeutic option for patients with end-stage renal disease. However, long-term results have not been very encouraging because of patient deaths due to cardiovascular disease and chronic transplant nephropathy (CTN), which includes vascular damage similar to arteriosclerosis injury. Several autoantibodies have been related to vasculopathy in the transplant such as anti-beta2GPI IgA, IgG, and IgM autoantibodies. We analyzed the levels of these autoantibodies among a cohort of 176 transplant recipients and a control group of 80 healthy subjects using enzyme-linked immunosorbent assay (ELISA). We collected data concerning the cardiovascular status of the patients, such as age, sex, diabetes mellitus, biopsy-confirmed CTN, schemic cardiopathy, cholesterol, triglycerides, and renal status by Modification of Diet in Renal Disease (MDRD) clearance and proteinuria. We also selected other characteristics, including hepatitis C virus infection and systolic/diastolic arterial pressures. The proportion of patients with high levels of IgG and IgM anti-beta2GPI autoantibodies did not differ from that observed in the control group, whereas the difference became significant in the case of anti-beta2GPI IgA autoantibodies (19.88% vs 1%). These results for the presence of anti-beta2GPI IgA autoantibodies were related to clinical data through a multivariate analysis, where the only parameter influenced by the presence of these autoantibodies seemed to be proteinuria, which in most cases was due to CTN.