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Recently published studies have found an impaired immune response after SARS-CoV-2 vaccination in solid organ recipients. However, most of these studies have not assessed immune cellular responses in liver and heart transplant recipients. We prospectively studied heart and liver transplant recipients eligible for SARS-CoV-2 vaccination. Patients with past history of SARS-CoV-2 infection or SARS-CoV-2 detectable antibodies (IgM or IgG) were excluded. We assessed IgM/IgG antibodies and ELISpot against the S protein 4 weeks after receiving the second dose of the mRNA-1273 (Moderna) vaccine. Side effects, troponin I, liver tests and anti-HLA donor-specific antibodies (DSA) were also assessed. A total of 58 liver and 46 heart recipients received two doses of mRNA-1273 vaccine. Median time from transplantation to vaccination was 5.4 years (IQR 0.3-27). Sixty-four percent of the patients developed SARS-CoV-2 IgM/IgG antibodies and 79% S-ELISpot positivity. Ninety percent of recipients developed either humoral or cellular response (87% in heart recipients and 93% in liver recipients). Factors associated with vaccine unresponsiveness were hypogammaglobulinemia and vaccination during the first year after transplantation. Local and systemic side effects were mild or moderate, and none presented DSA or graft dysfunction after vaccination. Ninety percent of our patients did develop humoral or cellular responses to mRNA-1273 vaccine. Factors associated with vaccine unresponsiveness were hypogammaglobulinemia and vaccination during the first year after transplantation, highlighting the need to further protect these patients.

Extracorporeal photopheresis (ECP) is an immunomodulatory therapy based on the infusion of autologous cellular products exposed to ultraviolet light (UV) in the presence of a photosensitizer. The study evaluates the ECP efficacy as induction therapy in a full-mismatch kidney transplant rat model. Dark Agouti to Lewis (DA-L) kidney transplant model has been established. ECP product was obtained from Lewis rat recipients after DA kidney graft transplantation (LewDA). Leukocytes of those LewDA rats were exposed to 8-methoxy psoralen, and illuminated with UV-A. The ECP doses assessed were 10 × 106 and 100 × 106 cells/time point. Lewis recipients received seven ECP infusions. DA-L model was characterized by the appearance of donor-specific antibodies (DSA) and kidney function deterioration from day three after kidney transplant. The dysfunction progressed rapidly until graft loss (6.1 ± 0.5 days). Tacrolimus at 0.25 mg/kg prolonged rat survival until 11.4 ± 0.7 days (p = 0.0004). In this context, the application of leukocytes from LewDA sensitized rats accelerated the rejection (8.7 ± 0.45, p = 0.0012), whereas ECP product at high dose extended kidney graft survival until 26.3 ± 7.3 days, reducing class I and II DSA in surviving rats. ECP treatment increases kidney graft survival in full-mismatch rat model of acute rejection and is a suitable immunomodulatory therapy to be explored in kidney transplantation.

Recipients of simultaneous liver-kidney transplantations (SLKTs) have a lower risk of rejection compared with recipients of kidney transplants alone. However, there is disagreement about the impact of pretransplant anti-human leukocyte antigen sensitization on patient and kidney graft survival in the long term. The aim of the study was to evaluate the impact of the recipient immunological risk and comorbidities in renal graft outcomes on SLKT. We reviewed the SLKTs performed in our center from May 1993 until September 2017. Patient and graft survival were analyzed according to the immunological risk, comorbidities, liver and kidney rejection episodes, immunosuppression, and infections. A total of 20 recipients of SLKT were considered in the high immunological risk (HIR) group, and 68 recipients were included in the low immunological risk (LIR) control group. The prevalence of hepatitis C virus infection, second renal transplant, and time on dialysis prior to transplantation were significantly higher in the HIR group. The incidence of acute kidney rejection was higher in the HIR group (P

This Guide for Living Donor Kidney Transplantation (LDKT) has been prepared with the sponsorship of the Spanish Society of Nephrology (SEN), the Spanish Transplant Society (SET), and the Spanish National Transplant Organization (ONT). It updates evidence to offer the best chronic renal failure treatment when a potential living donor is available. The core aim of this Guide is to supply clinicians who evaluate living donors and transplant recipients with the best decision-making tools, to optimise their outcomes. Moreover, the role of living donors in the current KT context should recover the level of importance it had until recently. To this end the new forms of incompatible HLA and/or ABO donation, as well as the paired donation which is possible in several hospitals with experience in LDKT, offer additional ways to treat renal patients with an incompatible donor. Good results in terms of patient and graft survival have expanded the range of circumstances under which living renal donors are accepted. Older donors are now accepted, as are others with factors that affect the decision, such as a borderline clinical history or alterations, which when evaluated may lead to an additional number of transplantations. This Guide does not forget that LDKT may lead to risk for the donor. Pre-donation evaluation has to centre on the problems which may arise over the short or long-term, and these have to be described to the potential donor so that they are able take them into account. Experience over recent years has led to progress in risk analysis, to protect donors' health. This aspect always has to be taken into account by LDKT programmes when evaluating potential donors. Finally, this Guide has been designed to aid decision-making, with recommendations and suggestions when uncertainties arise in pre-donation studies. Its overarching aim is to ensure that informed consent is based on high quality studies and information supplied to donors and recipients, offering the strongest possible guarantees.

Background: Recurrence of immunoglobulin (Ig)A nephropathy (rIgAN) is a growing cause of kidney allograft dysfunction. This study was aimed at investigating factors associated with rIgAN and the subsequent progression to end-stage renal disease (ESRD). Methods: Retrospective study including consecutive patients with IgA nephropathy (IgAN) who received a kidney transplant in our center between 1992 and 2016 and had a renal biopsy by clinical indication. The date of detection of chronic kidney disease (CKD) 5 was used as renal outcome. Results: Eighty-six kidney transplants were performed in patients with IgAN, 38 (44%) were from living donors (related n = 26). rIgAN was diagnosed in 23 allografts (27%). Renal function and proteinuria at the end of the follow-up period were worst in the rIgAN patients compared to those without rIgAN (2.2 vs. 1.4 mg/dL, p = 0.014, and 1.16 vs. 0.49 g/day, p = 0.005, respectively). Risk of rIgAN and progression to CKD 5 decreased with patient’s age (hazard ratio [HR] 0.95, 95% CI 0.92–0.98, p = 0.002, and HR 0.97, 95% CI 0.83–0.97, p = 0.008 per year, respectively). Patients with rIgAN had a higher risk of progression to CKD 5 (HR 6.7, 95% CI 1.3–35.7, p = 0.025). Full donor-recipient mismatch in the human leukocyte antigen (HLA)-B loci decreased the risk of rIgAN (HR 0.22, 95% CI 0.06–0.76, p = 0.017). Conclusions: rIgAN was an independent risk factor for ESRD after renal allograft. Younger age increased the risk of rIgAN and CKD 5. Conversely, HLA-B mismatching was a potential protective factor for rIgAN of this glomerular disease.

Living donor kidney transplantation (LDKT) is the best treatment for end-stage renal disease. In this setting, a significant percentage of transplants are not undertaken because of medical and nonmedical reasons of both donors and recipients. However, the impact of these discards in a transplant program has not been identified thoroughly so far. Our objective was to clarify key reasons for exclusion of LDKTs and the consequences for the discarded transplant candidates in the following 5 years.Analysis of donors' and recipients' characteristics of 781 couples evaluated in our hospital from January 2005 to December 2013. The consequences of discards in transplant candidates were analyzed in the cohort 2012 to 2013 (n = 106) and followed up until October 2018.In our study group, 402 (51.5%) LDKT couples were successfully donated, and 379 (48.5%) were excluded. Donor and transplant recipient candidates discarded were older at the evaluation (55.07 ± 12.14 years vs 51.73 ± 10.93 years, P .001; 48.81 ± 14.05 years vs 44.62 ± 13.91 years, P .001, respectively). The most frequent reason for kidney discard was medical contraindication found in the potential donor (47.5%; low eGFR, diabetes mellitus, impaired glucose tolerance, high blood pressure, cardiovascular pathology casually found during evaluation, and proteinuria). Of the discarded candidates from 2012 to 2013, 36.8% received a deceased donor kidney transplant, 17% a LDKT with another donor, 7.5% stayed on the waiting list, 18.9% died, 3.8% were excluded from the waiting list, and 14.2% were lost to follow-up.In most cases, transplantation was not undertaken because of donor pathology. Fifty-three percent of the discarded patients were eventually transplanted, with a 31.4% probability to receive an organ from another living donor.

Even though manufacturers claim that the dermal fillers are nontoxic and nonimmunogenic, adverse events may occur. Clinically and histologically, most of the late onset adverse events present as an inflammatory response. To assess whether HLA polymorphisms are associated with late-onset inflammatory adverse events related to dermal fillers. A total of 211 patients were included, of whom 129 experienced late-onset inflammatory adverse events to different fillers (Inflammation group) and 82 who did not (Reference group). Patients completed a standardized questionnaire and provided a blood sample or oral swap for HLA testing. The study population consisted of 188 (89%) women and 23 (11%) men. The two study groups were similar in the distributions of filler type, location of injecting, allergy, autoimmune disease, gender, age, ethnicity, and smoking status. Of the 211 patients in the sample, 25 had the combination of HLA subtype-B*08 and HLA subtype-DRB1*03. This was 16.3% of the inflammatory group and 4.9% of the reference group. This combination of HLA subtypes was associated with an almost 4-fold increase in the odds of developing immune mediated adverse events (odds ratio = 3.79, 95% CI 1.25-11.48). Genetic polymorphisms such as HLA combinations may identify patients at risk of developing late onset immune mediated adverse events to dermal fillers.

According to preliminary data, seroconversion after mRNA SARS‐CoV‐2 vaccination might be unsatisfactory in Kidney Transplant Recipients (KTRs). However, it is unknown if seronegative patients develop at least a cellular response that could offer a certain grade of protection against SARS‐CoV‐2. To answer this question, we prospectively studied 148 recipients of either kidney (133) or kidney‐pancreas (15) grafts with assessment of IgM/IgG spike (S) antibodies and ELISpot against the nucleocapside (N) and the S protein at baseline and two weeks after receiving the second dose of the mRNA‐1273 (Moderna) vaccine. At baseline, 31 patients (20.9%) had either IgM/IgG or ELISpot positivity and were considered to be SARS‐CoV‐2‐pre‐immunized, while 117 (79.1%) patients had no signs of either cellular or humoral response and were considered SARS‐CoV‐2‐naïve. After vaccination, naïve patients who developed either humoral or cellular response were finally 65.0%, of which 29.9% developed either IgG or IgM and 35.0% S‐ELISpot positivity. Factors associated with vaccine unresponsiveness were diabetes and treatment with anti‐thymocytes globulins during the last year. Side effects were consistent with that of the pivotal trial and no DSAs developed after vaccination. In conclusion, mRNA‐1273 SARS‐CoV‐2 vaccine elicits either cellular or humoral response in almost two thirds of KTRs.

Background and Aims The introduction of more sensitive and specific tools as solid phase immunoasssays has improved the ability to detect HLA antibodies. The techniques based on solid phase immunoassays in Luminex system can be performed either as screening assays (with pooled antigen panels that use bead populations coated in affinity-purified class I or class II HLA molecules) or to assess the specificity with single-antigen bead (SAB) assays (single cloned allelic HLA antigen).The sensitivity of each assay Is different, and their results may be discordant. Our objective was to evaluate the potential discrepancies between tests in a cohort of patients on our waiting list with no previous sensitizing events, for their potential influence in the access to transplantation. Method Observational study of 184 patients included in the kidney-transplant waiting list on March 31st, 2019. All possible sensitizing events (pregnancy, transfusion and previous transplants) were registered. HLA antibodies where analyzed both by screening and SAB tests. Results We limited the observations to males (64.7% of the cohort), as most women (95.4%) had a known sensitizing event, not being possible to discard possible pregnancies in the remaining. Of men, 46.2% had a sensitizing event and 63.8% did not. All 63 unsensitized patients showed a negative screening test, and 49 (77,78%) had at least one SAB done being positive in 73.4% (36/49). The positive SAB yielded a cPRA >10% in 30 cases (between 21-85%), mostly (27/30) due to HLA class II antibodies (88.8% with at least one DQ antibody) with a median MFI of 1571 (range:752-17650). A thorough review of specificities showed that 81% DQB1* and 89.6% DQA1* antibodies did not show consistency in all beads included for the same allele. Conclusion Screening and SAB assays show a significant discordancy in unsensitized men included in the waiting list for kidney transplantation. Automated detection of antibodies by SAB without a proper specialized interpretation may wrongly limit access to transplantation.

ObjectivesWe investigated the associations with HLA and microtubule-associated protein tau (MAPT) H1 haplotype in anti-IgLON5 disease, a recently identified disorder characterized by gait instability, brainstem dysfunction, and a prominent sleep disorder in association with IgLON5 antibodies and pathologic findings of a novel neuronal-specific tauopathy.MethodsWe compared the HLA alleles and MAPT H1/H1 genotype of 35 patients with anti-IgLON5 with healthy controls. The on-line server tool NetMHCIIpan 3.1 was used to predict the IgLON5 peptide binding to HLA Class II molecules.ResultsThe HLA-DRB1*10:01-DQB1*05:01 haplotype was overrepresented in patients with anti-IgLON5 disease (OR = 54.5; 95% CI: 22.2–133.9, p < 0.0001). In addition, HLA-DQA was genotyped in 27 patients, and 25 (92.6%) of them had DQ molecules composed by DQA1*01 and DQB1*05 chains compared with 148/542 (27.3%) controls (OR = 43.9; 95% CI: 10.4–185.5, p < 0.0001). Patients DRB1*10:01 positive developed more frequently sleep or bulbar symptoms than those carrying other HLA alleles (70.0% vs 26.7%; p = 0.011). Prediction algorithms identified 2 IgLON5 peptides (1 located in the signal sequence) that showed strong binding to HLA-DRB1*10:01 and other HLA-DRB1, but not to HLA-DQA and HLA-DQB molecules. The MAPT H1/H1 homozygous genotype was present in 20/24 (83.3%) anti-IgLON5 Caucasian patients compared with 54/116 (46.5%) healthy controls (p = 0.0007).ConclusionsThe robust association of anti-IgLON5 disease with distinct HLA Class II molecules supports a primary autoimmune origin. The significant association of MAPT H1 haplotype also suggests that an underlying neurodegenerative process could be involved in anti-IgLON5 disease.

AIM Acute antibody-mediated rejection (ABMR) after kidney transplantation (KT) is associated with poor allograft survival. Current therapies for ABMR are able to deplete B-lymphocytes but do not target plasma cells. Bortezomib is a proteasome inhibitor that can eliminate plasma cells and has demonstrated utility in the treatment of ABMR. METHODS A retrospective study was carried out from 2010 to 2014, including all patients with ABMR refractory to conventional treatment who received bortezomib. Bortezomib (1.3 mg/m(2) ) was administered intravenously on days 1, 4, 8, and 11. Renal function, graft survival, follow-up biopsies, and donor-specific antibodies (DSA) were recorded. RESULTS We identified seven patients. Of these, high immunological risk was found in 6 of 7, preformed DSA were found in 5 of 7, flow cytometry crossmatch was positive in 4 of 7, and desensitization before KTx was provided in 6 of 7 patients. ABMR was diagnosed at a median of 90 days (8-167) post-KT. After bortezomib therapy, renal function improved or stabilized in 5 of 7 patients and progressively deteriorated in 2 of 7, leading to haemodialysis after 7 and 11 months, respectively. Follow-up kidney biopsies showed persistence of ABMR in 2 of 7, chronic active ABMR 3 of 7 and inactive chronic lesions in 2 of 7. DSA titres significantly decreased after treatment (P = 0.028). All patients experienced mild adverse events. After a follow-up of 22 ± 18 months, three grafts were lost (42%) and four remained functioning. CONCLUSION Bortezomib could be useful as an adjuvant therapy for ABMR refractory to conventional treatment with acceptable mid-term outcomes in these severe cases. More research is needed to develop strategies to better preserve graft function after refractory ABMR.

Background Chronic active antibody-mediated rejection (c-aABMR) is an important cause of allograft failure and graft loss in long-term kidney transplants. Methods To determine the efficacy and safety of combined therapy with rituximab, plasma exchange (PE) and intravenous immunoglobulins (IVIG), a cohort of patients with transplant glomerulopathy (TG) that met criteria of active cABMR, according to BANFF’17 classification, was identified. Results We identified 62 patients with active c-aABMR and TG (cg ≥ 1). Twenty-three patients were treated with the combination therapy and, 39 patients did not receive treatment and were considered the control group. There were no significant differences in the graft survival between the two groups. The number of graft losses at 12 and 24 months and the decline of eGFR were not different and independent of the treatment. A decrease of eGFR≥13 ml/min between 6 months before and c-aABMR diagnosis, was an independent risk factor for graft loss at 24 months (OR = 5; P = 0.01). Infections that required hospitalization during the first year after c-aABMR diagnosis were significantly more frequent in treated patients (OR = 4.22; P = 0.013), with a ratio infection/patient-year of 0.65 and 0.20 respectively. Conclusions Treatment with rituximab, PE, and IVIG in kidney transplants with c-aABMR did not improve graft survival and was associated with a significant increase in severe infectious complications. Trial registration Agencia Española de Medicametos y Productos Sanitarios (AEMPS): 14566/RG 24161. Study code: UTR-INM-2017-01. Electronic supplementary material The online version of this article (10.1186/s12882-018-1057-4) contains supplementary material, which is available to authorized users.

Celiac disease (CD) is a complex autoimmune disorder caused by ingestion of gluten in genetically susceptible individuals. Different genetic risk factors have been identified, but virtually all patients are human leukocyte antigen (HLA)-DQ2 and/or HLA-DQ8 positive. We describe a new, fast, accurate and simple real-time polymerase chain reaction (PCR)-based assay for the genotyping and homozygosity analysis of the CD-related HLA alleles. The assay overcomes the major limitations of protocols currently in use, allowing HLA-DQ2/DQ8 genotyping by using only three real-time PCR reactions. For the appraisal of DQ2 homozygosity, only one more reaction is needed. These reactions are easily automated and suitable for large screening studies in diagnostic procedures, as it is demonstrated by their successful application in our HLA diagnostic laboratory. Finally, we assessed the clinical relevance of this real-time PCR-based assay by studying a cohort of fully characterized patients. As expected, all CD patients had at least one of the CD-associated alleles, and the highest CD risk was indicated by the presence of the HLA-DQ2.5 heterodimer (HLA-DQA1*05-DQB1*02) with HLA-DQB1*02 in homozygosity.

Psoriasis is a common inflammatory skin disease with key immunological and genetic components. Recruitment of leukocytes into the skin is a central step in its pathogenesis, mediated by cytokines. Among the cytokines expressed in psoriatic lesions, C-C chemokine ligand 4 (CCL4) and C-C chemokine ligand 4-like (CCL4L) chemokines appear to be pivotal elements for the skin recruitment of proinflammatory cells. The aim of this study is to evaluate the relationship between CCL4L polymorphisms (including single-nucleotide polymorphisms (SNPs) and copy number variation (CNV)) and the course and prognosis of psoriasis. We analyzed the CNV and the rs4796195 SNP in 211 psoriatic patients and 234 controls; sera from both populations were also quantified for CCL4/CCL4L protein. Our results showed that a high CNV (≥3 copies) is associated with psoriasis severity, whereas moderate disease correlated with a lower CNV (≤2 copies); specifically, the CCL4L1 allele frequency is higher in severe psoriasis, whereas CCL4L2 is more frequent in patients with a milder disease. In addition, we found a positive correlation between the CNV and sera protein levels. Our results suggest that CCL4L genotyping could not only allow a better understanding of the psoriatic pathogenesis but could also be used as a prognostic tool, even helping to modulate the efficacy of treatments.newline JID JOURNAL CLUB ARTICLE: For questions, answers, and open discussion about this article, please go to http://www.nature.com/jid/journalclub

Lung transplantation (LT) has become an accepted therapy for selected patients with advanced lung disease. One of the main limitations to successful LT is rejection of the transplanted organ where chemokines are pivotal mediators. Here, we test the relationship between copy number variation (CNV) in the CCL4L chemokine gene and rejection risk in LT patients (n=161). Patients with no acute rejection showed a significantly lower mean number of CCL4L copies than patients that showed acute rejection (1.66 vs 1.96, P=0.014), with an even greater number of gene copies seen in patients with more than one episode of acute rejection (1.66 vs 2.30, P=0.001). Additionally, patients withor =2 CCL4L copies had a significantly higher risk of acute rejection compared with patients that had 0-1 CCL4L copies (odds ratio 2.65; 95% confidence interval, 1.33-5.28; P=0.0046). A combined analysis of CCL4L CNV and the rs4796195 CCL4L single nucleotide polymorphism demonstrated that the effect of CCL4L copy number in acute rejection is mainly because of the number of copies of the CCL4L1 allelic variant. This finding constitutes the first report of CNV as a correlate factor in allograft rejection.

Acute antibody-mediated rejection (ABMR) after kidney transplantation (KT) is associated with poor allograft survival. Current therapies for ABMR are able to deplete B-lymphocytes but do not target plasma cells. Bortezomib is a proteasome inhibitor that can eliminate plasma cells and has demonstrated utility in the treatment of ABMR.A retrospective study was carried out from 2010 to 2014, including all patients with ABMR refractory to conventional treatment who received bortezomib. Bortezomib (1.3 mg/m(2) ) was administered intravenously on days 1, 4, 8, and 11. Renal function, graft survival, follow-up biopsies, and donor-specific antibodies (DSA) were recorded.We identified seven patients. Of these, high immunological risk was found in 6 of 7, preformed DSA were found in 5 of 7, flow cytometry crossmatch was positive in 4 of 7, and desensitization before KTx was provided in 6 of 7 patients. ABMR was diagnosed at a median of 90 days (8-167) post-KT. After bortezomib therapy, renal function improved or stabilized in 5 of 7 patients and progressively deteriorated in 2 of 7, leading to haemodialysis after 7 and 11 months, respectively. Follow-up kidney biopsies showed persistence of ABMR in 2 of 7, chronic active ABMR 3 of 7 and inactive chronic lesions in 2 of 7. DSA titres significantly decreased after treatment (P = 0.028). All patients experienced mild adverse events. After a follow-up of 22 ± 18 months, three grafts were lost (42%) and four remained functioning.Bortezomib could be useful as an adjuvant therapy for ABMR refractory to conventional treatment with acceptable mid-term outcomes in these severe cases. More research is needed to develop strategies to better preserve graft function after refractory ABMR.

Celiac disease is usually associated with autoimmune disorders and has occasionally been reported in patients with inflammatory myopathies. Our aim was to determine the presence of celiac disease and antibodies associated with celiac disease in patients with inflammatory myopathies and to investigate their relationship. Serum antigliadin, anti-tissue transglutaminase, and antiendomysial antibodies were determined in 51 patients with inflammatory myopathies. HLA-DQ2 and -DQ8 alleles were studied to assess their complementary diagnostic value. Jejunal biopsy was performed in patients with moderate to high levels of antigliadin antibodies. Patients with jejunal histology consistent with celiac disease initiated a gluten-free diet. Seventeen patients (31%) were positive for antigliadin antibodies, which were significantly more frequent in patients with inclusion-body myositis than dermatomyositis (P < 0.001). Positive status to HLA-DQ2 and/or -DQ8 did not differ between antigliadin-positive (75% and 12.5%) or -negative (60% and 15%) patients. Three of five jejunal biopsies were diagnostic for celiac disease with histological normalization after a gluten-free diet. Thus, celiac disease is more prevalent in patients with inflammatory myopathies than in the general population. Positive status to HLA-DQ2 allele, which is known to be more frequent in patients with inflammatory myopathies, could explain the high prevalence of antigliadin antibodies in this population. The diagnostic value of HLA-DQ2 or -DQ8 haplotypes to detect celiac disease in patients with inflammatory myopathy is limited.

Plasma stromal cell-derived factor (SDF)-1 levels, SDF1-3'A polymorphism, and CXCR4(+) T lymphocytes in relation to resistance to human immunodeficiency virus (HIV)-1 infection and its progression were investigated in a study of HIV-positive patients, exposed but uninfected (EU) subjects, and healthy control subjects, all lacking CCR5 Delta 32 homozygosity. SDF1-3'A homozygosity was associated with low plasma SDF-1 levels in uninfected persons and was not related to long-term nonprogression. HIV-1 infection involved increased plasma SDF-1 levels, which were not attributable to any kind of chronic viral infection, because all EU hemophiliacs were hepatitis C virus-positive but had normal SDF-1 levels. High plasma SDF-1 levels and low CXCR4 expression on T lymphocytes was associated with long-term nonprogression, whereas in advancing disease expression of CXCR4 increased, accompanied by a decrease in plasma SDF-1 during the more advanced stages of HIV-1 infection. EU subjects with sexual exposure to HIV-1, but not EU hemophiliacs, showed an underpresentation of SDF1-3'A allele frequency, which was coupled with high plasma SDF-1 levels and low CXCR4 expression.

CD148, a receptor-like protein tyrosine phosphatase also known as HPTP-η/DEP-1, is involved in signal transduction in leucocytes and is thought to contribute to mechanisms of cellular differentiation. We have investigated the in situ expression of CD148 in various fresh-frozen tissues by immunohistology and analyzed its expression on subpopulations of activated peripheral blood leucocytes by flow cytometry. In lymphoid organs, CD148 was found to be widely expressed on B and T cells, granulocytes, macrophages, certain dendritic cells as well as mature thymocytes. The cellular level of CD148 was increased after in vitro activation of peripheral blood leucocytes. Comparative analysis of tissue samples from normal gut and from patients with active Crohn′s disease showed that leucocytes expressing CD148 are significantly upregulated in inflamed tissues and that a subset of these cells co-express the activation marker CD25. In non-lymphoid tissues, CD148 was found to be present on many epithelial cell types with glandular and/or endocrine differentiation as well as on fibrocytes, melanocytes and Schwann cells. CD148 expression was maintained also in malignant counterparts of such tissues. However, a marked loss of CD148 immunoreactivity was apparent in some of the investigated high-grade carcinomas. In summary, our results confirm a role of CD148 as a leucocyte activation marker. Among non-hematopoietic cells, CD148 is expressed by characteristic types of epithelial and non-epithelial cells. Downregulation of CD148 might promote dedifferentiation and autonomous growth of such cells in malignant tumors.

The leukocyte differentiation antigen CD50 (intercellular adhesion molecule-3, ICAM-3), mediates cell-cell adhesion through its ligand LFA-1 and is a transducting receptor molecule during T-cell activation. Since CD50 homologues in other species have not yet been identified, the role of this molecule can only be analyzed in human cell models. Thus, to better study CD50 function in T cells, we have obtained two CD50-negative T-cell clones, named CAMY.l and CAMY.2. These clones were derived from the Jurkat T-cell variant PPL.l. Data from analysis of protein expression, specific mRNA content and calcium mobilization assays have confirmed the absence of functional CD50 molecules on these two clones. Thus, CAMY.l and CAMY.2 show no CD50 expression by phenotypical and immunoprecipit-ation analysis. CD50-sperific mRNA content is undetectable by Northern blot analysis in these clones and, only, when RT-PCR was performed could specific mRNA be detected. Additionally, CD50 cross-linking on theses clones shows no increase in intracellular calcium. Transfection of CD50 cDNA on CAMY cells restores not only CD50 surface expression, but its functional ability to induce calcium mobilization, CD69 upregulation and cell morphological changes. The CAMY.l and CAMY.2 clones provide useful model systems to analyze CD50 function in T cells.

Cytotoxic T lymphocyte (CTL) responses targeting specific HIV proteins, in particular Gag, have been associated with relative control of viral replication in vivo. However, Gag-specific CTL can also be detected in individuals who do not control the virus and it remains thus unclear how Gag-specific CTL may mediate the beneficial effects in some individuals but not in others. Here, we used a 10mer peptide set spanning HIV Gag-p24 to determine immunogen-specific T-cell responses and to assess functional properties including functional avidity and cross-reactivity in 25 HIV-1 controllers and 25 non-controllers without protective HLA class I alleles. Our data challenge the common belief that Gag-specific T cell responses dominate the virus-specific immunity exclusively in HIV-1 controllers as both groups mounted responses of comparable breadths and magnitudes against the p24 sequence. However, responses in controllers reacted to lower antigen concentrations and recognized more epitope variants than responses in non-controllers. These cross-sectional data, largely independent of particular HLA genetics and generated using direct ex-vivo samples thus identify T cell responses of high functional avidity and with broad variant reactivity as potential functional immune correlates of relative HIV control.

High levels of HIV-1 replication during the chronic phase of infection usually correlate with rapid progression to severe immunodeficiency. However, a minority of highly viremic individuals remains asymptomatic and maintains high CD4⁺ T cell counts. This tolerant profile is poorly understood and reminiscent of the widely studied nonprogressive disease model of SIV infection in natural hosts. Here, we identify transcriptome differences between rapid progressors (RPs) and viremic nonprogressors (VNPs) and highlight several genes relevant for the understanding of HIV-1-induced immunosuppression. RPs were characterized by a specific transcriptome profile of CD4⁺ and CD8⁺ T cells similar to that observed in pathogenic SIV-infected rhesus macaques. In contrast, VNPs exhibited lower expression of interferon-stimulated genes and shared a common gene regulation profile with nonpathogenic SIV-infected sooty mangabeys. A short list of genes associated with VNP, including CASP1, CD38, LAG3, TNFSF13B, SOCS1, and EEF1D, showed significant correlation with time to disease progression when evaluated in an independent set of CD4⁺ T cell expression data. This work characterizes 2 minimally studied clinical patterns of progression to AIDS, whose analysis may inform our understanding of HIV pathogenesis.

We report here the identification of a novel DQB1*06 allele, DQB1*0618, found in a bone marrow donor. The new allele was detected during routine DNA-based HLA typing by an ambiguous pattern of probe hybridization, obtained by polymerase chain reaction using sequence-specific oligonucleotides (PCR-SSO). Molecular cloning and sequencing confirmed that the new allele is identical to DQB1*0609 at exon 2 except for 3 nucleotide substitutions at positions 353, 356 and 367, also found in other alleles. These nucleotide changes may explain its anomalous reactivity.

In this report we describe the identification of a novel DRB1*04 allele, DRB1*0437, found in a Spanish individual. The routine HLA typing, in the context of bone marrow transplantation, by polymerase chain reaction-sequence-based typing (PCR-SBT) made possible the identification of this new allele. This allele is identical to DRB1*0402 except for a single nucleotide substitution at position 286 (A-->C), changing the encoded Isoleucine to a Leucine. The DRB1*0437 allele conserves the same two acidic residues at codons 70 and 71 that confer to DRB1*0402 its association to some autoimmune diseases.

In this report we describe the identification of a novel HLA-A*11 allele, HLA-A*1108, found in two individuals of a Spanish family. This new allele was detected during routine HLA typing by an atypical serological reactivity pattern and by inconclusive patterns obtained in DNA-based typing methods. The nucleotide sequence of exons 2 and 3 of HLA-A*1108 was identical to HLA-A*11011 except for two nucleotide substitutions at codons 152 (GCG-->GAG) and 156 (CAG-->CGG). These mutations change the non-charged amino acid alanine, at codon 152, to negative glutamic acid, and also non-charged glutamine, at codon 156, to positive arginine, which may explain its anomalous serological reactivity.

BACKGROUND: Human immunodeficiency virus (HIV) takes advantage of multiple host proteins to support its own replication. The gene ZNRD1 (zinc ribbon domain-containing 1) has been identified as encoding a potential host factor that influenced disease progression in HIV-positive individuals in a genomewide association study and also significantly affected HIV replication in a large-scale in vitro short interfering RNA (siRNA) screen. Genes and polymorphisms identified by large-scale analysis need to be followed up by means of functional assays and resequencing efforts to more precisely map causal genes. METHODS: Genotyping and ZNRD1 gene resequencing for 208 HIV-positive subjects (119 who experienced long-term nonprogression [LTNP] and 89 who experienced normal disease progression) was done by either TaqMan genotyping assays or direct sequencing. Genetic association analysis was performed with the SNPassoc package and Haploview software. siRNA and short hairpin RNA (shRNA) specifically targeting ZNRD1 were used to transiently or stably down-regulate ZNRD1 expression in both lymphoid and nonlymphoid cells. Cells were infected with X4 and R5 HIV strains, and efficiency of infection was assessed by reporter gene assay or p24 assay. RESULTS: Genetic association analysis found a strong statistically significant correlation with the LTNP phenotype (single-nucleotide polymorphism rs1048412; [Formula: see text]), independently of HLA-A10 influence. siRNA-based functional analysis showed that ZNRD1 down-regulation by siRNA or shRNA impaired HIV-1 replication at the transcription level in both lymphoid and nonlymphoid cells. CONCLUSION: Genetic association analysis unequivocally identified ZNRD1 as an independent marker of LTNP to AIDS. Moreover, in vitro experiments pointed to viral transcription as the inhibited step. Thus, our data strongly suggest that ZNRD1 is a host cellular factor that influences HIV-1 replication and disease progression in HIV-positive individuals.

A new autoantibody against a 155-kDa protein has been described in patients with myositis. We conducted a study to determine the occurrence and types of cancer occurring in a cohort of patients with polymyositis (PM) or dermatomyositis (DM) and analyzed the value of this autoantibody as a serologic marker of cancer-associated myositis (CAM). Serum samples from all patients were examined by protein immunoprecipitation assays with HeLa cell, to determine the presence of a 155-kDa protein band. HLA-DRB1 and DQA1 typing was performed by polymerase chain reaction-reverse sequence Specific oligonucleotide. Statistical analyses were carried out with the Mann Whitney U and Fisher exact tests. Associations were determined using odds ratios (ORs) with 95% confidence intervals (CI). Eighty-five patients with myositis (20 PM and 65 DM) were included. CAM was detected in 16 patients (19%), 14 with DM. The shawl sign rash was significantly more frequent in patients with CAM than in those without (p < 0.01). Adenocarcinoma was the most frequent type of cancer (87.5%). Anti-p 155 autoantibody was found in 1 of the 20 (5%) patients with PM and in 15 of the 65 (23%) patients with DM. A relationship between anti-p155 and CAM was found in DM patients (OR 23; 95% CI, 5.23-101.2). The HLA-DQA1*0102 allele was not found in any of the anti-p155-positive patients. The prevalence of CAM in our cohort was 19%. Autoantibody against p155 was highly related to CAM and could be a reliable marker of cancer in patients with DM.

In this report, we describe the identification of a novel DRB4*01 allele, DRB4*01033, found in two Spanish Caucasian individuals. The new allele was detected during routine HLA typing by an unusual pattern of amplification obtained by polymerase chain reaction using sequence-specific primers (PCR-SSP) that did not match with any of the previously described DRB4 alleles. In order to establish the polymorphism responsible for this pattern exons 2 and 3 of the DRB4 locus were amplified and directly sequenced. The new DRB4*01 allele is identical to DRB4*0103101 except for a single nucleotide substitution in codon 78 (TAC-->TAT). This nucleotide change does not cause an amino acid change as both triplets code for a tyrosine.

Objective.To examine the role of HLA-DRB1 and HLA-DQB1 alleles in the susceptibility to systemic sclerosis (SSc) and its clinical expression in a Spanish population.Methods.One hundred Spanish Caucasian patients with SSc and 130 controls were studied. Molecular HLA-DRB1 and HLA-DQB1 typing was performed by polymerase chain reaction (PCR) sequence-based typing and PCR sequence-specific oligonucleotide.Results.HLA-DRB1*11 was associated with genetic susceptibility to SSc, whereas HLA-DRB1*07 (HLA-DRB1*0701) showed a protective effect. A significant increase in the frequency of the DRB1*1104 allele was observed in patients with anti-topoisomerase I autoantibodies (anti-Topo I) while HLA-DRB1*01 and HLA-DQB1*05 alleles were significantly increased in patients with anti-centromere antibodies (ACA). The HLA-DRB1*11 allele was more frequent in patients with pulmonary fibrosis; however, no significant association with any HLA-DRB1 or DQB1 alleles was identified in patients with pulmonary arterial hypertension.Conclusion.HLA alleles play a role in genetic susceptibility to SSc in Spanish patients. Some alleles are more prevalent in patients with pulmonary fibrosis and in patients with certain SSc-specific autoantibodies (anti-Topo I and ACA).

In humans, the region configurations DR1, DR8, DR51, DR52 and DR53 are known to display copy number as well as allelic variation, rendering high resolution typing of HLA-DRB haplotypes cumbersome. Advantage was taken of microsatellite D6S2878, present in all DRB genes/pseudogenes with an intact exon 2-intron 2 segment. This DRB-STR is highly polymorphic in composition and length. Recently, it was proven that all exon 2 sequences could be linked to a certain DRB-STR that segregates with the respective DRB allele. Because haplotypes show differential copy numbers and compositions of exon 2- positive DRB genes/pseudogenes, unique DRB-STR patterns could be described that appear to be specific for a particular DRB haplotype. The aim of this workshop project was to approve and to qualify this simple typing protocol in a larger panel covering different European populations. All participants succeeded in correctly defining the DRB-STR amplicons varying from 135 to 222 base pair (bp) lengths. The panel of 101 samples covered 50 DRB alleles distributed over 37 different haplotypes as defined by exon 2 sequence-based typing. These haplotypes could be refined into 105 haplotypes by DRB-STR typing. Thus, discrimination of exon 2-identical DRB alleles was feasible, as well as the exact description of three different crossing-over events that resulted in the generation of hybrid DR region configurations. This typing procedure appears to be a quick and highly robust technique that can easily be performed by different laboratories, even without experience in microsatellite typing ; thus, it is suitable for a variety of researchers in diverse research areas.

In the field of organ transplantation, the state of thymic function has not been a major concern but data from bone marrow transplantation studies have unravel the persistence of some thymopoiesis in the adult and, more importantly, the possibility of reinducing it. Given the central role of the thymus in tolerance, these facts have stimulated the interest in the biology of the thymus in humans. Contemporarily, basic research has provided new tools, if imperfect, to monitor thymic function, that is, T-cell receptor excision circles, markers for lymphocytes recently emigrated from the thymus and new imaging techniques. The deployment of these new tools is already changing some paradigms and has now established that re-enactment of thymic activity in the course of bone marrow transplantation or in patients with human immunodeficiency virus on highly active anti-retroviral therapy is beneficial and that can be achieved in the adult. Clinical trials using thymopoiesis-stimulating factors are underway. On the other hand, the discovery that the thymus contains a broad representation of self-antigens and that this depends on the expression of the product of the gene AIRE by the medullary thymic epithelial cells opens the possibility of manipulating central tolerance. Current protocols inducing microchimerism to generate tolerance to solid organ grafts suggest that this could be a feasible therapeutic goal. Therefore, there are many signs indicating that a period of translational research applying the principles of thymic biology and central tolerance to transplantation has already started.

We report herein the identification of a new DRB1 allele using sequence-based typing. The new allele, DRB1*0106, was detected during routine HLA typing of a patient undergoing bone marrow transplantation. DRB1*0106 is identical to DRB1*0101 except for two codons, 71 (AGG-->GCG) and 86 (GGT-->GTG), changing the encoded arginine to alanine and glycine to valine. Both sequences were confirmed by polymerase chain reaction with sequence-specific primers (PCR-SSP). The polymorphism at codon 71 has not been, until now, identified in DRB1*01 alleles, although it is present in all the DRB1*15 alleles as well as DRB1*1309 and DRB1*1424.

We conducted the current study to determine the prevalence, incidence density, and clinical significance of thyroid disease in a series of 109 patients diagnosed with idiopathic inflammatory myopathy. We analyzed 109 patients diagnosed with idiopathic inflammatory myopathy during a 20-year period (1986-2006). Routine determination of thyroid profile, including hormone levels and antithyroid antibodies (antithyroid peroxidase and antithyroglobulin antibody systematically, and antithyroid-stimulating hormone receptor antibody when appropriate) was carried out at onset and at least once again during the follow-up period. Human leukocyte antigen (HLA) study was performed by polymerase chain reaction sequence-specific oligonucleotide typing. Six patients (5.5%) developed thyroid disease (hypo- or hyperthyroidism), with an incidence density of 8.7 new cases per 1000 person-years; 4 of the 6 cases were of autoimmune origin (3.6%). Thyroid dysfunction, whether autoimmune or not, was clearly related with onset or relapse of disease activity. Neoplastic disease was detected in 3 of 4 (75%) patients with autoimmune thyroid disease, a significantly higher frequency than was observed in the remaining patients in the series (12 of 103, 11.7%; p < 0.05; RR: 22.8; 95% confidence interval, 2.2-236.6). HLA DRB1*04 was found in 4 of the 6 patients with thyroid disease and inflammatory myopathy. Thyroid disorders are clinically relevant in patients with inflammatory myopathies. An association was observed between autoimmune thyroid disease and cancer. Thyroid assessment is strongly recommended in patients with idiopathic inflammatory myopathy and clinical relapse.

HLA genotyping by polymerase chain reaction (PCR) has some inherent labor-intensive and effort-demanding limitations. To overcome them, we have developed a real-time PCR with hybridization probes approach able to obtain a medium-low resolution HLA-B genotyping with fewer tubes and probes and with a shorter time requirement. Our strategy used 18 simultaneous reactions amplifying HLA-B alleles and an internal control. Monitorization of both amplifications in each tube is performed by the simultaneous application of two fluorescent resonance emission transfer probes: the first probe, different for each tube, is specific for the HLA-B locus and the second probe detects the control gene. A medium-low resolution (300 HLA-B allelic groups) typing is obtained for each sample by analyzing the melting curve patterns. Because some alleles may be determined without using the complete set of reactions, we present an alternative strategy: a first round of seven reactions and, according to the result, a second (or third) round of PCRs to solve the ambiguities. This method was validated in pretyped clinical samples and the results were completely concordant. Moreover, fewer ambiguous results were obtained. In summary, we present a new, faster, and more accurate method than currently used PCR techniques to type HLA-B alleles.

OBJECTIVES To assess whether an almost complete restoration of immune system can be achieved when antiretroviral therapy is initiated at very early stages of asymptomatic chronic HIV-1 infection. DESIGN T cell subsets and cell-mediated responses were analysed at baseline and after 12 months of either a double or a triple antiretroviral therapy in 26 asymptomatic HIV-1-infected patients with CD4 T cell counts > 500 x 10(6) cells/l and a baseline plasma viral load > 10000 copies/ml. RESULTS Triple therapy was significantly more effective in reducing plasma HIV RNA to undetectable levels, in returning CD4:CD8 ratio to nearly normal levels, in reducing activated cells (CD38) and in increasing naive (CD45RA+CD45RO-) and memory (CD45RA-CD45RO+) CD4 cells. Both double and triple therapies caused a clear decrease in memory (CD45RA-CD45RO+) CD8 cells as well as a significant increase in the CD28 subset of CD8 cells. At baseline, there was an important increase in cells producing interferon-gamma (IFNgamma) with no significant abnormalities in T lymphocytes producing interleukin 2 (IL-2), tumour necrosis factor alpha and interleukin 4. Both types of therapy reduced IFNgamma- and IL2-producing CD4 T lymphocytes while IFNgamma-producing CD8 cells remained increased. Even before therapy, these HIV-1-positive patients lacked significant abnormalities in the T cell responsiveness to polyclonal stimuli as well as in the secretion of CCR5 chemokines by peripheral blood mononuclear cells. CONCLUSIONS Initiating highly active antiretroviral therapy at very early stages of chronic HIV-1 infection allows rapid and almost complete normalization of T cell subsets and preservation of T cell functions. These early-treated patients could be excellent candidates for receiving additional HIV-specific immune-based therapies, which might be essential for the control of HIV infection.

Protein tyrosine phosphatases play an essential role in the control of leucocyte cell growth an differentiation. Recently a new receptor type membrane tyrosine phosphatase named CD148 has been identified. This molecule is present on the membrane of all the hematopoietic lineages as well as on several other cell types, mainly epithelial cells and its expression increases after cell activation. This molecule is able to act as a transducing molecule. Moreover, CD148 is able to modulate the signal transduction through the TCR/CD3 complex, in a manner similar to CD45. It has also been suggested that CD148 could be involved in mechanisms of differentiation and inhibition of cell growth. In addition, CD148 seems to be associated with a serine/threonine kinase in certain epithelial cell lines and leucocytes. Here, we review recent data on the expression and function of CD148 in both human, mouse and rat.