Insulin-like growth factor-1 and insulin-like growth factor binding protein-3: impact on early haematopoietic reconstitution following allogeneic haematopoietic stem cell transplantation.

OBJECTIVES: The aim of the study was to investigate whether high endogenous levels of insulin-like growth factor-1 (IGF-1) and its binding protein-3 (IGFBP-3) were related to a faster reconstitution of different blood cell populations in the early phase after allogeneic myeloablative haematopoietic stem cell transplantation (HSCT). METHODS: We measured IGF-1 and IGFBP-3 by chemiluminescence during the first three weeks after transplantation in 35 adult patients undergoing myeloablative HSCT and calculated area under the curve divided by time (AUC/t) for each patient. RESULTS: Circulating levels of IGF-1 and IGFBP-3 correlated with counts of reticulocytes (rs  = 0.44, p = .011 and r = 0.41, p = .017, respectively) and thrombocytes (rs  = 0.38, p = .030 and rs  = 0.56, p = .0008) three weeks post-transplant. Furthermore, high IGFBP-3 levels correlated with absolute lymphocyte counts 3 weeks post-HSCT (rs  = 0.54, p = .012) and were associated with shorter time to neutrophil engraftment (rs  = -0.35, p = .043). Both IGF-1 and IGFBP-3 levels were associated with the number of circulating natural killer cells one month after HSCT (rs  = 0.42, p = .032 and rs  = 0.57, p = .0026). CONCLUSION: These data indicate that high levels of IGF-1 and IGFBP-3 relate to a faster haematopoietic reconstitution after HSCT and suggest a biological influence of these mediators in haematopoietic homeostasis in these patients.

High preharvest donor Foxp3 mRNA level predicts late relapse of acute lymphoblastic leukaemia after haematopoietic stem cell transplantation.

OBJECTIVES: The curative effect of allogeneic haematopoietic stem cell transplantation (HSCT) for acute leukaemia is due in part to the donor T cell-mediated graft-versus-leukaemia immune reaction (GvL). Several studies have suggested that donor CD25+CD4+Foxp3+regulator T cells (Tregs) may decrease graft-versus-host disease (GvHD) without abrogating GVL. This notion may need modification in acute lymphoblastic leukaemia (ALL). METHODS: Foxp3 mRNA level was measured by qPCR in preharvest donor blood CD4+ T cells. The study comprised 45 patients with ALL in 1st or 2nd CR who received myeloablative HSCT using T-replete bone marrow grafts. RESULTS: Relapse occurred in 17 patients median 363 days after HSCT. The relapse risk was estimated by Cox univariate and multivariate proportional hazard regression. The proportionality assumption was met by analysing the preharvest donor Foxp3 mRNA level as a time-dependent covariate. Early relapse was not modified by the Foxp3 mRNA level. However, a higher Foxp3 mRNA level was associated with a significantly increased relapse risk after day 363 after transplantation, compatible with inhibition of GvL. In contrast, a higher preharvest donor CD4+ T-cell concentration was associated with reduced relapse risk. CONCLUSION: A higher preharvest donor Foxp3 mRNA level may be predictive of late ALL relapse after HSCT.

Variation in interleukin 7 receptor alpha chain (IL7R) influences risk of multiple sclerosis.

Multiple sclerosis is a chronic, often disabling, disease of the central nervous system affecting more than 1 in 1,000 people in most western countries. The inflammatory lesions typical of multiple sclerosis show autoimmune features and depend partly on genetic factors. Of these genetic factors, only the HLA gene complex has been repeatedly confirmed to be associated with multiple sclerosis, despite considerable efforts. Polymorphisms in a number of non-HLA genes have been reported to be associated with multiple sclerosis, but so far confirmation has been difficult. Here, we report compelling evidence that polymorphisms in IL7R, which encodes the interleukin 7 receptor alpha chain (IL7Ralpha), indeed contribute to the non-HLA genetic risk in multiple sclerosis, demonstrating a role for this pathway in the pathophysiology of this disease. In addition, we report altered expression of the genes encoding IL7Ralpha and its ligand, IL7, in the cerebrospinal fluid compartment of individuals with multiple sclerosis.

Course and Clinical Significance of CD8+ T-Cell Counts in a Large Cohort of HIV-Infected Individuals.

OBJECTIVES: To examine trajectories of CD8(+) T-cell counts before and after combination antiretroviral therapy (cART) in human immunodeficiency virus (HIV)-infected individuals and associations with mortality. METHODS: CD8(+) T-cell counts were measured in 3882 HIV-infected individuals who received care in Copenhagen during 1995-2012. Reference values were obtained from 1230 persons from the background population. Mortality rate ratios were estimated by Poisson regression. RESULTS: CD8(+) T-cell counts were elevated during untreated HIV infection and remained elevated through 10 years of cART. A slight drop of 130 cells/µL (interquartile range, -160 to 410 cells/µL) in the median CD8(+) T-cell count was observed after cART initiation. CD8(+) T-cell counts stabilized at approximately 900 cells/µL (95th percentile of the background population, 835 cells/µL). Markedly elevated CD8(+) T-cell counts at cART initiation were associated with a poor increase in the CD4(+) T-cell count (relative risk, 2.22; 95% confidence interval [CI], 1.42-3.48). Individuals with a CD8(+) T-cell count of <500 cells/µL 1 year after cART initiation had an increased mortality rate (mortality rate ratio, 1.73; 95% CI, 1.29-2.32) and a higher proportion of deaths attributable to AIDS-related conditions, compared with individuals with CD8(+) T-cell counts of ≥500 cells/µL. After receiving cART for 10 years, a CD8(+) T-cell count of >1500 cells/µL was associated with increased non-AIDS-related mortality (mortality rate ratio, 1.82; 95% CI, 1.09-3.22), compared with a CD4(+) T-cell count of 500-1500 cells/µL. CONCLUSIONS: CD8(+) T-cell counts are elevated during HIV infection and do not normalize despite long-term cART. Low CD8(+) T-cell counts are associated with increased AIDS-related mortality. Marked elevations in CD8(+) T-cell counts after long-term cART are associated with increased non-AIDS-related mortality.

GWAS of follicular lymphoma reveals allelic heterogeneity at 6p21.32 and suggests shared genetic susceptibility with diffuse large B-cell lymphoma.

Non-Hodgkin lymphoma (NHL) represents a diverse group of hematological malignancies, of which follicular lymphoma (FL) is a prevalent subtype. A previous genome-wide association study has established a marker, rs10484561 in the human leukocyte antigen (HLA) class II region on 6p21.32 associated with increased FL risk. Here, in a three-stage genome-wide association study, starting with a genome-wide scan of 379 FL cases and 791 controls followed by validation in 1,049 cases and 5,790 controls, we identified a second independent FL-associated locus on 6p21.32, rs2647012 (OR(combined)  = 0.64, P(combined)  = 2 × 10(-21)) located 962 bp away from rs10484561 (r(2)<0.1 in controls). After mutual adjustment, the associations at the two SNPs remained genome-wide significant (rs2647012:OR(adjusted)  = 0.70, P(adjusted)  =  4 × 10(-12); rs10484561:OR(adjusted)  = 1.64, P(adjusted)  = 5 × 10(-15)). Haplotype and coalescence analyses indicated that rs2647012 arose on an evolutionarily distinct haplotype from that of rs10484561 and tags a novel allele with an opposite (protective) effect on FL risk. Moreover, in a follow-up analysis of the top 6 FL-associated SNPs in 4,449 cases of other NHL subtypes, rs10484561 was associated with risk of diffuse large B-cell lymphoma (OR(combined)  = 1.36, P(combined)  =  1.4 × 10(-7)). Our results reveal the presence of allelic heterogeneity within the HLA class II region influencing FL susceptibility and indicate a possible shared genetic etiology with diffuse large B-cell lymphoma. These findings suggest that the HLA class II region plays a complex yet important role in NHL.

sVEGF-R1 in acute non-infectious toxicity syndromes after pediatric allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (HSCT) is challenged by acute non-infectious toxicities, including sinusoidal obstruction syndrome (SOS), engraftment syndrome (ES) and capillary leak syndrome (CLS) among others. These complications are thought to be driven by a dysfunctional vascular endothelium, but the pathophysiological mechanisms remain incompletely understood, and the diagnoses are challenged by purely clinical diagnostic criteria that are partly overlapping, limiting the possibilities for progress in this field. There is, however, increasing evidence suggesting that these challenges may be met through the development of diagnostic biomarkers to improve diagnostic accuracy of pathogenetically homogenous entities, improved pre-transplant risk assessment and the early identification of patients with increased need for specific treatment. Soluble vascular endothelial growth factor receptor-1 (sVEGF-R1) is emerging as an important biomarker of endothelial damage in patients with trauma and sepsis but has not been studied in HSCT. OBJECTIVES: To investigate sVEGF-R1 as a marker of endothelial damage in pediatric HSCT patients by exploring associations with SOS, CLS, ES, and acute graft-versus-host disease (aGvHD). METHODS: We prospectively included 113 children undergoing myeloablative HSCT and measured sVEGF-R1 in plasma samples obtained weekly during the early period of transplantation and 3 months post-transplant. RESULTS: All over, sVEGF-R1 levels were significantly increased from day +7 after graft infusion, peaking at day +30, most pronounced in patients receiving busulfan. Patients considered to be at increased risk of SOS and therefore commenced on prophylactic defibrotide had significantly elevated levels of sVEGF-R1 before start of conditioning (446 pg/mL vs. 281 pg/mL, p = 0.0035), and this treatment appeared to stabilize sVEGF-R1 levels compared to patients not treated with defibrotide. Thirteen (11.5%) children meeting the modified Seattle criteria for SOS at median day +8 (1-18), had significantly elevated sVEGF-R1 levels on day +14 (489 pg/mL vs. 327 pg/mL, p = 0.007). In contrast. sVEGF-R1 levels in the much broader group of patients (45.1%) meeting EBMT-SOS criteria, including patients with very mild disease, did not overall differ in sVEGF-R1 levels, but higher sVEGF-R1 levels were seen in EBMT-SOS patients with an increased need for diuretic treatment. Importantly, sVEGF-R1 levels were not associated with ES and CLS but were significantly increased on day +30 in patients with grade III-IV aGvHD (OR = 4.2 pr. quartile, p = 0.023). CONCLUSION: VEGF-R1 levels are found to be increased in pediatric patients developing SOS, reflecting the severity of morbidity. sVEGF-R1 were unassociated with both CLS and ES. The potential of sVEGF-R1 as a clinically useful biomarker for SOS should be further explored to improve pre-transplant SOS-risk assessment, SOS-severity grading, and to guide treatment.

Dysregulation of CD4+CD25+CD127lowFOXP3+ regulatory T cells in HIV-infected pregnant women.

Pregnancy represents a major challenge to immunologic tolerance. How the fetal "semiallograft" evades maternal immune attack is unknown. Pregnancy success may involve alteration of both central (thymic) and peripheral tolerance mechanisms. HIV infection is characterized by CD4(+) T-cell depletion, chronic immune activation, and altered lymphocyte subsets. We studied immunologic consequences of pregnancy in 20 HIV-infected women receiving highly active antiretroviral therapy (HAART), and for comparison in 16 HIV-negative women. Lymphocyte subsets, thymic output, and cytokine profiles were measured prospectively during pregnancy and postpartum. A significant expansion of CD4(+)CD25(+)CD127(low)FoxP3(+) regulatory T cells indicating alteration of peripheral tolerance was seen during second trimester, but only in HIV-negative women. HIV-infected women had lower CD4 counts, lower thymic output and Th-2 cytokines, and more immune activation at all time points compared with controls. Immune activation was decreased in HIV-infected patients during pregnancy. In contrast, CD4 counts were increased in both groups. In conclusion, the study does not indicate that pregnancy adversely affects the immunologic course of HIV infection. However, despite HAART during pregnancy, HIV-infected women display different immunologic profiles from HIV-negative women, which may have importance for the induction of fetal-maternal tolerance and in part explain the increased risk of abortion in HIV-infected women.

HLA-A alleles and infectious mononucleosis suggest a critical role for cytotoxic T-cell response in EBV-related Hodgkin lymphoma.

A proportion of classical Hodgkin lymphoma (HL) is believed to be causally related to infection with the ubiquitous lymphotropic EBV. The determining factors for development of EBV-related HL remain poorly understood, but likely involve immunological control of the viral infection. Accordingly, markers of the HLA class I region have been associated with risk of EBV-related HL. To study the host genetic component of EBV-related HL further, we investigated the lymphoma's association with HLA-A*01 and HLA-A*02 simultaneously in the setting of infectious mononucleosis (IM), a risk factor for EBV-related HL, in a case-series analysis including 278 EBV-related and 656 EBV-unrelated cases of HL. By logistic regression, HLA-A*01 alleles [odds ratio (OR) per allele, 2.15; 95% CI, 1.60-2.88] were associated with increased and HLA-A*02 alleles (OR per allele, 0.70; 95% CI, 0.51-0.97) with decreased risk of EBV-related HL. These allele-specific associations corresponded to nearly 10-fold variation in risk of EBV-related HL between HLA-A*01 and HLA-A*02 homozygotes. History of IM was also associated with risk of EBV-related HL (OR, 3.40; 95% CI, 1.74-6.66). The association between history of IM and EBV-related HL was not seen in the presence of HLA-A*02 because this allele appeared to neutralize the effect of IM on EBV-related HL risk. Our findings suggest that HLA class I-restricted EBV-specific cytotoxic T-cell responses and events in the early immune response to EBV infection in IM play critical roles in the pathogenesis of EBV-related HL.

Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods.

Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and/or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative immunophenotype and antigen modulation) that highlight important methodological pitfalls. These findings demonstrate that with sufficient experience, flow cytometry is reliable for minimal residual disease monitoring in B-cell precursor acute lymphoblastic leukemia, although rare cases require supplementary PCR-based monitoring.

Higher recipient pre-transplant FOXP3 mRNA expression is associated with acute leukaemia relapse after HSCT.

The effect of higher FOXP3 mRNA expression by recipient pre-transplant CD4+ T cells on leukaemia relapse was analysed in a series of 106 patients who received allogeneic haematopoietic stem cell transplantation after myeloablative conditioning with or without antithymocyte globulin (ATG) due to acute leukaemia in 1st or 2nd complete remission. FOXP3 mRNA was measured by qPCR in purified CD4+ T cells from blood obtained before conditioning. Higher FOXP3 mRNA expression was associated with an increased relapse risk when conditioning included ATG (n = 43, hazard ratio [HR] 11.0 [2.50-48.4], p = 0.00001). No effect was observed in patients not receiving ATG (HR 0.95 [0.53-1.81]).

Association of HY-restricting HLA class II alleles with pregnancy outcome in patients with recurrent miscarriage subsequent to a firstborn boy.

Healthy females, pregnant with a boy, generate immune responses against male-specific minor histocompatibility (HY) antigens. The clinical importance of these responses is evident in stem cell transplantation. Birth of a boy prior to a series of miscarriages reduces the chance of a subsequent live birth. This study explores the putative impact of known HY-presenting HLA alleles on future pregnancy outcome in women with at least three consecutive miscarriages following a birth [secondary recurrent miscarriage (SRM)]. HLA-A, -B, -DRB1, DRB3-5 and DQB1 genotyping was performed in 358 SRM patients and in 203 of their children born prior to the miscarriages. The subsequent live birth in women with boys prior to the miscarriages compared with girls is lower in women with HY-restricting HLA class II alleles [odds ratio (OR): 0.17 (0.1-0.4), P = 0.0001]. One HY-restricting HLA class II allele in women with firstborn boys significantly reduces the chances of a live birth [OR: 0.46 (0.2-0.9), P = 0.02]. Two HY-restricting HLA class II alleles further reduced this chance [OR: 0.21 (0.1-0.7), P = 0.02]. HY-restricting HLA class II did not reduce the chances of a live birth in SRM women with firstborn girls. HY-restricting HLA class II alleles are associated with a decreased chance of a live birth in SRM women with firstborn boys. These findings strongly indicate an aberrant maternal immune reaction against fetal HY-antigens in SRM. The results may shed light on the as-yet unknown immunological causes of SRM and may help understand the successful maternal acceptance of the fetal semi-allograft.

Polymorphisms of innate pattern recognition receptors, response to interferon-beta and development of neutralizing antibodies in multiple sclerosis patients.

BACKGROUND: Interferon-beta therapy of patients with relapsing-remitting multiple sclerosis involves repeated 'immunizations' with exogenous protein solutions. Innate pattern recognition receptors play an important role in immune responses towards foreign substances and may thus be related to treatment outcome. OBJECTIVE: To determine the genotypes at 42 single nucleotide polymorphism loci in selected pattern recognition receptors for 567 prospectively followed relapsing-remitting multiple sclerosis patients treated with recombinant interferon-beta, and test for relationships to several outcome parameters, including formation of interferon-beta neutralizing antibodies. RESULTS: The results suggest an association between the rs5743810 polymorphism (Ser249Pro) of TLR6 and development of neutralizing antibodies after 24 months of therapy in males (p = 0.00002), but not in females (p = 0.2). This association survived crude Bonferroni correction (p (corrected) = 0.02). Additional associations were observed in carriers of the TLR2-rs5743708 and NOD2-rs3135499 SNPs (time to relapse), the TLR7-rs179008 and NOD1-rs2075820 SNPs (time to disease progression) and the TLR4-rs7873784, TLR9-rs5743836, and NOD2-rs2066842 SNPs (frequency of neutralizing antibodies development). All of these, however, failed to survive correction for multiple testing. There were no significant differences between interferon-beta responders and non-responders for any of the investigated single nucleotide polymorphisms. CONCLUSIONS: The rs5743810 polymorphism of TLR6 may be involved in development of anti-interferon-beta antibodies in males, although further studies are required to firmly establish this.

Cytomegalovirus-specific CD8+ T-cell responses are associated with arterial blood pressure in people living with HIV.

People living with HIV (PLHIV) are at increased risk for cardiovascular disease (CVD), and immunity against cytomegalovirus (CMV) may be a contributing factor. We hypothesized that enhanced T-cell responses against CMV and CMV-IgG antibody-levels are associated with higher arterial blood pressure in PLHIV. We assessed serum CMV-IgG, systolic- (SBP) and diastolic- (DBP) blood pressure, pulse pressure (PP), traditional risk factors, activated CD8+ T-cells (CD38+HLA-DR+), senescent CD8+ T-cells (CD28-CD57+) and interleukin-6 (IL-6) in 60 PLHIV and 31 HIV-uninfected controls matched on age, gender, education and comorbidity. In PLHIV, expression of interleukin-2, tumor necrosis factor-α and interferon-γ was measured by intracellular-cytokine-staining after stimulation of T-cells with CMV-pp65 and CMV-gB. Associations between CMV-specific immune responses and hypertension, SBP, DBP or PP were assessed by multivariate logistic and linear regression models adjusted for appropriate confounders. The median age of PLHIV was 47 years and 90% were male. Prevalence of hypertension in PLHIV was 37% compared to 55% of HIV-uninfected controls. CMV-specific CD8+ T-cell responses were independently associated with higher PP (CMV-pp65; ß = 2.29, p = 0.001, CMV-gB; ß = 2.42, p = 0.001) in PLHIV. No significant differences were found with regard to individual measures of SBP and DBP. A possible weak association was found between CMV-IgG and hypertension (ß = 1.33, p = 0.049) after adjustment for age, smoking and LDL-cholesterol. HIV-related factors, IL-6, CD8+ T-cell activation or CD8+ T-cell senescence did not mediate the associations, and no associations were found between CMV-specific CD4+ T-cell responses and blood pressure in PLHIV. In conclusion, increased arterial blood pressure in PLHIV may be affected by heightened CMV-specific CD8+ T-cell responses.

Graft rejection after hematopoietic cell transplantation with nonmyeloablative conditioning.

Graft rejection after hematopoietic cell transplantation (HCT) with nonmyeloablative conditioning is a rare but serious clinical problem. Graft rejection and salvage therapy in eight patients in a retrospective analysis of 124 consecutive patients is reported. The patients were conditioned with low-dose fludarabine and total body irradiation (TBI). The association of pretransplantation risk factors with rejection and the effect of chimerism and graft-versus-host disease on rejection were analyzed. Overall survival (OS) and progression free survival (PFS) were compared between patients with and without rejection. Retransplantation was performed with increased TBI conditioning for all patients, and with increased mycophenolate mofetil doses for recipients with HLA-identical sibling donors. No known pretransplantation risk factors were confirmed in this study. Rejection episodes were unevenly distributed over time. The storage temperature of the apheresis products was identified as a risk factor for rejection. Storage of the apheresis products at 5 degrees C diminished the risk of rejection. Low donor T cell chimerism at Day +14 significantly increased the risk of rejection. Seven patients were retransplanted. All but one engrafted successfully, but with decreased OS and PFS. Two patients received pentostatin infusion prior to donor lymphocyte infusions in unsuccessful attempts at reversing rejection. Storage temperature and donor chimerism had a significant effect on rejection. Following rejection, patients are at greater risk of dying from infections and progression/relapse of their malignancy. Retransplantation is feasible and well tolerated after HCT with nonmyeloablative conditioning and should be performed without delay in patients with imminent and manifest graft rejection.

Cytomegalovirus-specific T-cells are associated with immune senescence, but not with systemic inflammation, in people living with HIV.

In people living with HIV (PLWHIV), coinfection with cytomegalovirus (CMV) has been associated with inflammation, immunological ageing, and increased risk of severe non-AIDS related comorbidity. The effect of CMV-specific immune responses on systemic inflammation, immune activation and T-cell senescence was evaluated in 53 PLWHIV treated with combination antiretroviral therapy (cART). Activated-, terminally differentiated-, naïve-, and senescent T-cells were assessed by flow cytometry, and plasma levels of CMV IgG, interleukin-6, tumor necrosis factor-α, high-sensitivity C-reactive protein and soluble-CD14 were measured. In PLWHIV, expression of interleukin-2, tumor necrosis factor-α and interferon-γ was measured by intracellular-cytokine-staining after stimulation of T-cells with CMV-pp65, CMV-IE1, and CMV-gB. Increased CMV-specific T-cell responses were associated with a higher ratio of terminally differentiated/naïve CD8+ T-cells and with increased proportions of senescent CD8+ T-cells, but not with systemic inflammation or sCD14. Increased CMV-specific CD4+ T-cell responses were associated with increased proportions of activated CD8+ T-cells. In PLWHIV with expansion of CMV-specific T-cells or increased T-cell senescence, CMV-specific polyfunctionality was maintained. That the magnitude of the CMV-specific T-cell response was associated with a senescent immune phenotype, suggests that a dysregulated immune response against CMV may contribute to the immunological ageing often described in PLWHIV despite stable cART.

Donor Genotype in the Interleukin-7 Receptor α-Chain Predicts Risk of Graft-versus-Host Disease and Cytomegalovirus Infection after Allogeneic Hematopoietic Stem Cell Transplantation.

The efficacy of allogeneic hematopoietic stem cell transplantation (HSCT) is challenged by acute and chronic graft-versus-host disease (aGVHD and cGVHD) and viral infections due to long-lasting immunodeficiency. Interleukin-7 (IL-7) is a cytokine essential for de novo T cell generation in thymus and peripheral T cell homeostasis. In this study, we investigated the impact of the single nucleotide polymorphism rs6897932 in the IL-7 receptor α-chain (IL-7Rα) which has previously been associated with several autoimmune diseases. We included 460 patients undergoing allogeneic HSCT after a myeloablative conditioning. Patients had a median age of 26.3 years (0.3-67.0 years), and 372 (80.9%) underwent HSCT for malignant diseases. Donors were matched sibling donors (n = 147), matched unrelated donors (n = 244) or mismatched unrelated donors (n = 69), and the stem cell source were either bone marrow (n = 329) or peripheral blood (n = 131). DNA from donors was genotyped for the IL-7Rα single nucleotide polymorphism (SNP) rs6897932 using an allele-specific primer extension assay (CC: n = 252, CT: n = 178, TT: n = 30). The donor T allele was associated with a higher risk of grades III-IV aGVHD (HR = 2.0, 95% CI = 1.1-3.8, P = 0.034) and with significantly increased risk of extensive cGVHD (HR = 2.0, 95% CI = 1.1-3.6, P = 0.025) after adjustment for potential risk factors. In addition, the TT genotype was associated with a higher risk of cytomegalovirus (CMV) infection post-transplant (HR = 2.4, 95% CI = 1.2-4.3, P = 0.0068). Numbers of T cells were significantly higher on day +60 in patients receiving a rs6897932 TT graft (CD3+: 109% increase, P = 0.0096; CD4+: 64% increase, P = 0.038; CD8+: 133% increase, P = 0.011). Donor heterozygosity for the T allele was associated with inferior overall survival (HR = 1.7, 95% CI = 1.2-2.3, P = 0.0027) and increased treatment-related mortality (HR = 2.3, 95% CI = 1.3-4.0, P = 0.0047), but was not associated with the risk of relapse (P = 0.35). In conclusion, the IL-7Rα rs6897932 genotype of the donor is predictive of aGVHD and cGVHD, CMV infection, and mortality following HSCT. These findings indicate that IL-7Rα SNP typing of donors may optimize donor selection and facilitate individualization of treatment in order to limit treatment-related complications.

Reconstitution of Th17, Tc17 and Treg cells after paediatric haematopoietic stem cell transplantation: Impact of interleukin-7.

Successful reconstitution of T lymphocytes after allogeneic haematopoietic stem cell transplantation (HSCT) is needed to establish the graft-versus-leukaemia effect and an effective anti-microbial defense, but the ratio between functionally different T-cell subsets needs to be balanced to avoid graft-versus-host disease (GVHD). IL-7 is essential for T-cell generation in the thymus and peripheral T-cell homeostasis. High IL-7 levels have been associated with impaired T-cell reconstitution, increased risk of acute GVHD and treatment-related mortality, but the underlying cellular mechanisms behind these associations have not been investigated previously. We hypothesized that increased levels of IL-7 post-transplant alters the balance between immune-regulatory T cell subsets during the post-transplant lymphocyte recovery towards a more pro-inflammatory profile. We quantified Th17 cells, Tc17 cells and Tregs in 29 children following HSCT. Th17 cell and Treg counts rose significantly from day +90 to +180 post-HSCT, and prior acute GVHD was associated with significant changes in the concentration of Tregs (9.4×106/L vs. 1.3×106/L, P=0.0052) and the Th17/Treg ratio (1.5 vs. 4.2, P=0.025). The plasma level of IL-7 at day +90 correlated inversely with Th17 cell counts (rs=-0.65, P=0.0002) and the proportion of Tc17 cells (rs=0.64, P=0.0005) at day +90, but not with Tregs. Furthermore, high IL-7 levels at day +7 were predictive of a less naïve T-cell phenotype at day +90. These findings add further evidence that IL-7 is a key regulatory factor that may tune the balance between functionally different T-cell subsets following HSCT.

Cytomegalovirus-Specific CD4+ T-cell Responses and CMV-IgG Levels Are Associated With Neurocognitive Impairment in People Living With HIV.

BACKGROUND: Mechanisms leading to neurocognitive impairment (NCI) in people living with HIV (PLWHIV) on stable combination antiretroviral therapy (cART) remain unknown. We investigated the association between immunity against cytomegalovirus (CMV), HIV-specific variables, and NCI in PLWHIV on stable cART and with low comorbidity. METHODS: Fifty-two PLWHIV on stable cART and 31 HIV-uninfected controls matched on age, sex, education, and comorbidity were tested with a neurocognitive test battery, and CMV-immunoglobulin G (CMV-IgG) levels were measured. In PLWHIV, CMV-specific (CMV-pp65 and CMV-gB) CD4 and CD8 T-cell responses were measured using intracellular cytokine staining and flow cytometry. NCI was defined as a global deficit scale score (GDS score) ≥0.5. GDS scores and domain-specific scores defined severity of NCI. Logistic and linear multivariable regression analyses were used. RESULTS: NCI was detected in 30.8% of PLWHIV, and HIV was associated with an adjusted odds ratio (aOR) of 5.18 [95% confidence interval (CI): 1.15 to 23.41, P = 0.033] for NCI. In PLWHIV, higher CMV-specific CD4 T-cell responses increased the probability of NCI with an aOR of 1.68 (95% CI: 1.10 to 2.57) for CMV-pp65 or an aOR of 3.73 (95% CI: 1.61 to 16.98) for CMV-gB, respectively. Similar associations were not found with CMV-IgG or CMV-specific CD8 T cells, but when assessing severity of NCI, higher CMV-IgG (per 100 U/mL) was associated with worse GDS scores (ß = 0.08) (0.01-0.16), P = 0.044), specifically in the domain of speed of information processing (ß = 0.20 (0.04-0.36, P = 0.019). CONCLUSIONS: PLWHIV had increased risk of NCI. Excess risk may be associated with CMV-specific CD4 T-cell responses and CMV-IgG. Larger longitudinal studies investigating the impact of immunity against CMV on risk of NCI are warranted.

A functional polymorphism in the Eta-1 promoter is associated with allele specific binding to the transcription factor Sp1 and elevated gene expression.

Early T lymphocyte activator 1 (Eta-1), also known as Osteopontin, is a cytokine produced by macrophages and T lymphocytes. It is involved in the regulation of IL-12 and IL-10 expression in macrophages and stimulates the polarization of T cells to the Th1 subset. Three promoter polymorphisms of the human Eta-1 gene, -443T/C, -156delG/G, -66T/G, were investigated for possible influence on gene expression. Electrophoretic mobility shift assays (EMSA) with nuclear extract from the human myeloid leukaemia premonocyte cell line, THP-1, revealed sequence specific binding of the transcription factor Sp1 to the -66T allele but not the -66G allele, and haplotype -443C/-156G/-66T showed a marked increase in promoter activity of a luciferase reporter gene. Thus, a substitution of the T-base with G at position -66 in the Eta-1 promoter modulates the promoter activity of the Eta-1 gene, which might influence the Th1 versus Th2 balance. These observations are discussed in relation to a recently reported related observation on the same gene, and it is argued that discrepancies between reporter gene assays in the two studies may be due to the use of different cell lines and may reflect requirements for different transcription factors in cells involved in immune responses compared with other cells.

Gene variation in IL-7 receptor (IL-7R)α affects IL-7R response in CD4+ T cells in HIV-infected individuals.

Optimal CD4+ T cell recovery after initiating combination antiretroviral treatment (cART) in HIV infection reduces risk of morbidity and mortality. T-allele homozygosity ('TT') in the single nucleotide polymorphism, rs6897932(C/T), in the IL-7 receptor α (IL-7RA) is associated with faster CD4+ T cell recovery after cART initiation compared to C-allele homozygosity in rs6897932 ('CC'). However, underlying mechanisms are unknown. We aimed to examine potential mechanisms explaining the association between rs6897932 and CD4+ T cell recovery. Ten 'TT' and 10 'CC' HIV-infected individuals matched on gender, age, and nadir and current CD4+ T cell counts were included in a cross-sectional study. 'TT' individuals had higher proportion of CD4+ T cells expressing pSTAT5 compared to 'CC' individuals after stimulating with IL-7, especially when co-stimulated with soluble IL7-RA (sIL-7RA). Furthermore, 'TT' individuals had a higher proportion of proliferating CD4+ T cells after 7 days of culture with IL-7 + sIL-7RA compared to 'CC' individuals. No differences between 'TT' and 'CC' in binding of biotinylated IL-7 were found. In conclusion, increased signal transduction and proliferation in response to IL-7 was found in 'TT' compared to 'CC' HIV-infected individuals providing a mechanistic explanation of the effect of rs6897932 T-allele on CD4+ T cell recovery in HIV infection.