The influence of tumor necrosis factor microsatellite polymorphisms on patient survival following hematopoietic stem cell transplantation

Aim

To investigate the influence of tumor necrosis factor (TNF) microsatellite polymorphisms on patient survival following hematopoietic stem cell transplantation.

Methods

We analyzed TNFa, TNFb, and TNFd microsatellites among 100 patients who underwent allogeneic hematopoietic stem cell transplantation from a human leukocyte antigen (HLA)-identical sibling donor at the Internal Clinic of the University Hospital Center Zagreb in the period 2001-2009. The analysis was performed using polymerase chain reaction amplification and electrophoresis on a polyacrylamide gel in an automated sequencer.

Results

There was no significant difference in patient survival with respect to the allele length at a given microsatellite. However, a significantly lower survival rate was noticed among patients who were positive for TNFa8 allele (P < 0.001) and a significantly higher survival rate among those who were positive for TNFa10 allele (P = 0.0220).

Conclusion

These results for the first time suggest an influence of TNFa microsatellite on patient survival following HSCT and indicate a need for further studies of this microsatellite.Hematopoietic stem cell transplantation (HSCT) is considered to be the most effective treatment for numerous diseases and disorders of hematological system despite its considerable side-effects and treatment-induced mortality. One of the most common complications of this procedure is acute graft vs host disease (GvHD), which can occur even in the cases when transplantation is performed from a human leukocyte antigen (HLA)-identical sibling donor and which affects up to 50% of the patients (1). GvHD is a consequence of tissue damage and subsequent bacterial infiltration due to the conditioning of patients prior to the HSCT. The resulting release of the proinflammatory cytokines then triggers the activation of donor''s T cells, which in turn leads to tissue damage of target organs and a possible multi-organ failure. The major role in this, so called cytokine storm, which constitutes both the induction and effector phase of the GvHD, is played by tumor necrosis factor α (TNFα) cytokine (2). The level of TNFα during the conditioning period has been associated with the severity of the acute GvHD (3).The role of polymorphisms located within the TNF gene cluster region in the etiology of various diseases that have shown an association with the HLA system has been extensively studied. A large number of single nucleotide polymorphisms and 6 microsatellites have been described in the TNF region to date. The first five microsatellites are referred to as TNFa, TNFb, TNFc, TNFd, and TNFe (4). The sixth microsatellite (TNFf) was described later (5). TNFc, TNFd, and TNFe are (GA)n repeats, whereas TNFa and TNFf are (GT)n and (CA)n repeats, respectively. The only exception of this dinucleotide repeat motif is the TNFb microsatellite, which is composed of mononucleotide tandem repeats (G/A)n. The most polymorphic TNF microsatellite is TNFa with 14 alleles, while 7 alleles have been reported so far for TNFb and TNFd loci each (4). The role of these microsatellites was implicated in the development of diseases such as rheumatoid arthritis, multiple sclerosis, or type 1 diabetes (6-8). However, in the transplantation field, the most intriguing aspect of these polymorphisms is that, according to several studies (9,10), the level of the TNFα secretion is influenced by the presence of some alleles at the TNF microsatellite loci. Taking into account the importance of this cytokine in the induction of GvHD, the information about the potential of the patient’s immune cells to produce it is of great value in the evaluation of risk for GvHD occurrence and prediction of HSCT outcome. The analysis of these microsatellites could therefore be applied in the detection of those patients who are at higher risk of developing GvHD after HSCT (11).The aim of this study was to investigate the importance of selected TNF microsatellites in the patient survival following HSCT. The choice of the loci included in the analysis (TNFa, TNFb, and TNFd) was guided by the above mentioned study (11), as well as by the level of polymorphism detected for a given locus in the Croatian population (12).     

Very Early Onset Inflammatory Bowel Disease Associated with Aberrant Trafficking of IL-10R1 and Cure by T Cell Replete Haploidentical Bone Marrow Transplantation

Purpose

Loss-of-function mutations in IL10 and IL10R cause very early onset inflammatory bowel disease (VEO-IBD). Here, we investigated the molecular pathomechanism of a novel intronic IL10RA mutation and describe a new therapeutic approach of T cell replete haploidentical hematopoietic stem cell transplantation (HSCT).

Methods

Clinical data were collected by chart review. Genotypes of IL10 and IL10R genes were determined by Sanger sequencing. Expression and function of mutated IL-10R1 were assessed by quantitative PCR, Western blot analysis, enzyme-linked immunosorbent assays, confocal microscopy, and flow cytometry.

Results

We identified a novel homozygous point mutation in intron 3 of the IL10RA (c.368-10C > G) in three related children with VEO-IBD. Bioinformatical analysis predicted an additional 3′ splice site created by the mutation. Quantitative PCR analysis showed normal mRNA expression of mutated IL10RA. Sequencing of the patient’s cDNA revealed an insertion of the last nine nucleotides of intron 3 as a result of aberrant splicing. Structure-based modeling suggested misfolding of mutated IL-10R1. Western blot analysis demonstrated a different N-linked glycosylation pattern of mutated protein. Immunofluorescence and FACS analysis revealed impaired expression of mutated IL-10R1 at the plasma membrane. In the absence of HLA-identical donors, T cell replete haploidentical HSCT was successfully performed in two patients.

Conclusions

Our findings expand the spectrum of IL10R mutations in VEO-IBD and emphasize the need for genetic diagnosis of mutations in conserved non-coding sequences of candidate genes. Transplantation of haploidentical stem cells represents a curative therapy in IL-10R-deficient patients, but may be complicated by non-engraftment.     

Interleukin-9 release from human kidney grafts and its potential protective role in renal ischemia/reperfusion injury

Objective and design

The pathophysiology of ischemia/reperfusion (I/R) injury is dominated by an inflammatory response. In the identification of new therapeutic agents, the role of individual cytokines may be essential. Interleukin (IL)-9 is a pleiotropic cytokine recently identified to be involved in various immune responses. In this study, the role of IL-9 in renal I/R injury was assessed.

Methods

We performed repeated direct measurements of arteriovenous IL-9 concentration differences over the reperfused graft in human kidney transplantation.

Results

Substantial renal IL-9 release was observed from deceased donor kidneys (P = 0.006). In contrast, living donor kidneys, which have a more favourable clinical outcome, did not release IL-9 during early reperfusion (P = 0.78). Tissue expression of IL-9 did not change upon reperfusion in both living and deceased human donor kidneys. To assess the role of IL-9 in I/R injury, an experimental study comprising IL-9 inhibition in mice undergoing renal I/R was performed. Although there was no difference in kidney function, structural damage was significantly aggravated in anti-IL-9 treated mice.

Conclusions

Deceased donor grafts show a substantial IL-9 release upon reperfusion in clinical kidney transplantation. However, inhibition of IL-9 aggravated kidney damage, suggesting a regulating or minor role of IL-9 in clinical I/R injury.     

Plasmacytoid dendritic cells are increased in cerebrospinal fluid of untreated patients during multiple sclerosis relapse

Background

Toll-like receptor 2 (TLR2) represents a reasonable functional and positional candidate gene for Alzheimer's disease (AD) as it is located under the linkage region of AD on chromosome 4q, and functionally is involved in the microglia-mediated inflammatory response and amyloid-β clearance. The -196 to -174 del polymorphism affects the TLR2 gene and alters its promoter activity.

Methods

We recruited 800 unrelated Northern Han Chinese individuals comprising 400 late-onset AD (LOAD) patients and 400 healthy controls matched for gender and age. The -196 to -174 del polymorphism in the TLR2 gene was genotyped using the polymerase chain reaction (PCR) method.

Results

There were significant differences in genotype (P = 0.026) and allele (P = 0.009) frequencies of the -196 to -174 del polymorphism between LOAD patients and controls. The del allele was associated with an increased risk of LOAD (OR = 1.31, 95% CI = 1.07-1.60, Power = 84.9%). When these data were stratified by apolipoprotein E (ApoE) ε4 status, the observed association was confined to ApoE ε4 non-carriers. Logistic regression analysis suggested an association of LOAD with the polymorphism in a recessive model (OR = 1.64, 95% CI = 1.13-2.39, Bonferroni corrected P = 0.03).

Conclusions

Our data suggest that the -196 to -174 del/del genotype of TLR2 may increase risk of LOAD in a Northern Han Chinese population.     

NLRP3 inflammasome activation and interleukin-1β release in macrophages require calcium but are independent of calcium-activated NADPH oxidases

Objective and design

We studied the involvement of calcium and calcium-activated NADPH oxidases in NLRP3 inflammasome activation and IL-1β release to better understand inflammasome signaling in macrophages.

Material or subjects

Human volunteer blood donors were recruited to isolate monocytes to differentiate them into macrophages. Wild-type or DUOX1-deficient C57/B6 mice were used to prepare bone marrow-derived macrophages.

Treatment

Murine or human macrophages were treated in vitro with NLRP3 inflammasome agonists (ATP, silica crystals) or calcium agonists (thapsigargin, ionomycin) in calcium-containing or calcium-free medium.

Methods

Intracellular calcium changes were followed by measuring FURA2-based fluorescence. Gene expression changes were measured by quantitative real-time PCR. Protein expression was assessed by western blotting. Enzymatic activity was measured by fluorescence caspase-1 activity assay. IL-1β release was determined by ELISA. ELISA data were analyzed by ANOVA and Tukey’s post hoc test.

Results

Our data show that calcium is essential for IL-1β release in human macrophages. Increases in cytosolic calcium alone lead to IL-1β secretion. Calcium removal blocks caspase-1 activation. Human macrophages express Duox1, a calcium-regulated NADPH oxidase that produces reactive oxygen species. However, Duox1-deficient murine macrophages show normal IL-1β release.

Conclusions

Human macrophage inflammasome activation and IL-1β secretion requires calcium but does not involve NADPH oxidases.     

Further evidence that mutations in INS can be a rare cause of Maturity-Onset Diabetes of the Young (MODY)

Background

Interleukin (IL)-1β is a potent proinflammatory cytokine markedly overexpressed in the brains of patients with Alzheimer's disease (AD), and also involved in development of atherosclerosis and coronary artery disease. Caspase-1 (CASP1), formerly called IL-1β converting enzyme (ICE), mediates the cleavage of the inactive precursor of IL-1β into the biologically active form. CASP1 genetic variation (G+7/in6A, rs501192) has been associated with susceptibility to myocardial infarction and cardiovascular death risk. We examined the contribution of this gene to the susceptibility for AD.

Methods

We examined genetic variations of CASP1 by genotyping haplotype tagging SNPs (htSNPs) (rs501192, rs556205 and rs530537) in a group of 628 Spanish AD cases and 722 controls.

Results

There were no differences in the genotypic, allelic or haplotypic distributions between cases and controls in the overall analysis or after stratification by age, gender or APOE ε4 allele.

Conclusion

Our negative findings in the Spanish population argue against the hypothesis that CASP1 genetic variations are causally related to AD risk.     

Correlating Interleukin-12 Stimulated Interferon-γ Production and the Absence of Ectodermal Dysplasia and Anhidrosis (EDA) in Patients with Mutations in NF-κB Essential Modulator (NEMO)

Objective

Patients with hypomorphic mutations in Nuclear Factor-κB Essential Modulator (NEMO) are immunodeficient (ID) and most display ectodermal dysplasia and anhidrosis (EDA). We compared cytokine production by NEMO-ID patients with and without EDA.

Methods

PBMCs of NEMO-ID patients, four with EDA carrying E315A, C417R, D311N and Q403X, and three without EDA carrying E315A, E311_L333del and R254G, were cultured with PHA, PHA plus IL-12p70, LPS, LPS plus IFN-γ, TNF and IL-1β. The production of various cytokines was measured in the supernatants. Fifty-nine healthy individuals served as controls.

Results

PBMCs of NEMO-ID patients without EDA produce subnormal amounts of IFN-γ after stimulation with PHA, but normal amounts of IFN-γ after PHA plus IL-12p70. In contrast, IFN-γ production by patients with EDA was low in both cases. Patients with EDA also generate lower PHA-stimulated IL-10 and IL-1β than controls, whereas the production of these cytokines by patients without EDA was normal.

Conclusion

Responses of PBMCs in NEMO-ID patients with EDA to PHA with and without IL-12p70 appear less robust than in NEMO-ID patients without EDA. This possibly indicates a better preserved NEMO function in our patients without EDA.     

Proinflammatory Cytokine Gene Polymorphisms in Irritable Bowel Syndrome

Introduction

Irritable bowel syndrome (IBS) is a multifactorial functional gastrointestinal disorder, characterized by recurrent abdominal pain and altered bowel habits. Proinflammatory cytokines can play an important role in intestinal inflammation, while their production is under genetic control.

Methods

This study was performed in a group of patients with IBS to analyze the genotype frequencies of a number polymorphic genes coding for proinflammatory cytokine (interleukin-6 (IL), tumor necrosis factor-alpha (TNF-α), and IL-1 group). Using polymerase chain reaction with sequence-specific primers method, the cytokine genes were amplified, and alleles and genotypes of 71 patients with IBS were detected on gel electrophoresis, and the results were compared with healthy control subjects.

Results

Results of the analyzed data showed that the frequencies IL-1R C allele at position Pst-I 1970 (P?=?0.017), IL-6 G allele at position ?174 (P?=?0.002), and TNF-α G allele at position ?238 (P?<?0.001) in the patient group were significantly higher than the control group. IL-6 GG genotype (?174) and TNF-α GG genotype (?238) in the patient group were also significantly overrepresented (P?<?0.001), while IL-6 CG genotype (?174) and TNF-α GA genotype (?238) were significantly decreased in the patients with IBS (P?<?0.001). The frequencies of IL-6 (?174, nt565) GG haplotype and TNF-α (?308, ?238) GG haplotype were also significantly higher in the patient group (P?<?0.001), whereas the frequencies of the haplotypes IL-6 CG and TNF-α GA were significantly decreased in the patients with IBS (P?<?0.001).

Conclusion

IL-6 and TNF-alpha proinflammatory cytokine gene polymorphisms could change individual susceptibility to IBS and might have a role in pathophysiology of disease.     

Mental health and resiliency following 44 months of terrorism: a survey of an Israeli national representative sample

Background

Genetic variation in the innate immune system of the host may play a role in determining the risk of developing infection, as well as outcome from infection.

Methods

Infectious complications were retrospectively determined in 293 (233 African-American (AA), 57 Caucasian and 3 Hispanic) mechanically ventilated very low birth weight (VLBW) infants (<1500 grams at birth) who were genotyped for the IL-6 -174 G/C, IL-10 -1082 G/A and CD14 -260 C/T single nucleotide polymorphisms (SNPs).

Results

The IL-6 -174C allele was associated with an increased incidence of late blood stream infection (BSI) in AA but not Caucasian infants. In AA infants with the C allele the incidence of late BSI was 20/29 (69%) compared to 94/204 (46%) in homozygous GG infants (RR 2.6, 95% CI: 1.1–6.0, p = 0.021). The IL-10 -1082A allele was associated with an increased incidence of late BSI. One or more episodes of late BSI developed in 14 (35%) of 40 infants with the GG genotype, 71 (49%) of 145 infants with the GA genotype and 63 (58%) of 108 infants with the AA genotype (p = 0.036). Infants with the A allele (AA or GA genotypes) had an incidence of late BSI that was 134/253 (53%) compared to 14/40 (35%) in homozygous GG infants (RR 2.1, 95% CI: 1.04–4.19, p = 0.035). The CD14 -260 C/T SNP did not alter the overall risk for BSI in ventilated VLBW infants. Multiple BSI episodes were more common in the TT genotype group (CC: 17%, CT: 11%, TT: 30%, p = 0.022). This effect was due to the strong effect of the TT genotype on the incidence of multiple BSI in AA infants (CC: 15%, CT: 11%, TT: 39%, p = 0.003).

Conclusion

The IL-6 -174 G/C, IL-10 -1082 G/A and CD14 -260 C/T SNPs may alter risk for BSI in ventilated VLBW infants.     

Associations between interleukin-10 polymorphisms and susceptibility to psoriasis: a meta-analysis

Objective

The aim of this study was to determine whether three specific interleukin-10 (IL-10) polymorphisms confer susceptibility to psoriasis.

Methods

A meta-analysis was conducted on the associations between the IL-10-1082 G/A, -592 C/A polymorphisms and haplotypes of the IL-10-1082 G/A, -592 C/A, -819 C/T polymorphisms and psoriasis.

Results

A total of eight studies involving 1,018 psoriasis patients and 1,186 controls were considered in the meta-analysis. No association was found between psoriasis and the IL-10-1082 G allele in all study subjects [odds ratio (OR)?=?1.098, 95?% confidence interval (CI)?=?0.923?1.306, p?=?0.291] or between this allele and psoriasis in Europeans (OR?=?0.990, 95?% CI?=?0.809?1.214, p?=?0.925), but a significant association was found in Asians (OR?=?1.785, 95?% CI?=?1.144?2.76, p?=?0.011). Three polymorphisms at the promoter region of IL-10 (-1082 G/A, -819 C/T, -592 C/A) are known to be in complete linkage disequilibrium, but no association was found between the haplotype and psoriasis.

Conclusions

This meta-analysis shows that the IL-10-1082 G/A polymorphism confers susceptibility to psoriasis in Asians, and suggests that the IL-10 promoter -1082 polymorphism has an ethnicity-specific effect.     

Long-Term Persistence of Limited HTLV-I Tax-specific Cytotoxic T Cell Clones in a Patient with Adult T Cell Leukemia/Lymphoma after Allogeneic Stem Cell Transplantation

Purpose

Adult T cell leukemia/lymphoma (ATL) is a highly aggressive malignancy of T cells caused by human T cell lymphotropic virus type 1 (HTLV-1). Recent clinical studies have suggested that allogeneic stem cell transplantation (HSCT) improves the clinical course of ATL by harnessing a graft-versus-ATL effect, and that donor-derived HTLV-1 Tax-specific CD8⁺ cytotoxic T cells (CTLs) contribute to the graft-versus-ATL effect after HSCT. However, little is known about the immunological characteristics of Tax-specific CTLs in ATL patients who underwent HSCT.

Methods

We serially analyzed frequencies, differentiation, functions and clonal dynamics of Tax-specific CTLs in paired samples of peripheral blood (PB) and bone marrow (BM) from an ATL patient after HSCT at the single-cell level. We used flowcytometric and single-cell T cell receptor (TCR) repertoire analysis methods without culture steps.

Results

Donor-derived Tax-specific CTLs effectively suppressed HTLV-1 replication in both PB and BM at least during chronic graft-versus-host disease after HSCT. Furthermore, Tax-specific CTLs had comparable properties between BM and PB, except for preferential accumulation in BM rather than PB. Tax-specific CTLs persistently existed as less-differentiated CD45RA^-CCR7^- effector memory CTLs based on predominant phenotypes of CD27⁺, CD28^+/? and CD57^+/?. Our approach using single-cell TCR repertoire analysis method showed highly restricted oligoclonal responses of Tax-specific CTLs, and TCR BV7- or BV30- expressing two predominant CTL clones persistently existed and maintained strong cytotoxic activities against HTLV-1 in both PB and BM over three years after HSCT.

Conclusions

These findings about Tax-specific CTLs provide insights into future directions for studies on immunotherapy against ATL.     

Decreased CD200R expression on monocyte-derived macrophages correlates with Th17/Treg imbalance and disease activity in rheumatoid arthritis patients

Objectives

CD200 is expressed on various cell types, including T cells, while the CD200 receptor (CD200R) is expressed on myeloid cells such as monocytes-derived macrophages (MDMs). The CD200–CD200R interaction has been shown to play an important role in the prevention of autoimmune disease. Thus, we hypothesized that CD200/CD200R1 is involved in the pathogenesis of rheumatoid arthritis (RA).

Methods

In total, 35 RA patients and 17 healthy controls (HCs) were enrolled in this study. CD200/CD200R1 expression and Th17/Treg were examined by flow cytometry. Serum levels of interleukin (IL)-2, interferon-γ (IFN-γ), IL-4 and IL-10 were detected by ELISA. Disease activity was evaluated according to the C-reactive protein (CRP) levels, erythrocyte sedimentation rates (ESR) and 28-joint disease activity score (DAS28) scores.

Results

Compared with HCs, RA patients exhibited a significantly decreased level of CD200R1 on MDMs. CD200R1 expression correlated negatively with DAS28, ESR, and CRP levels. This abnormal expression was associated with Th17/Treg imbalance in the active RA patients. However, expression of CD200R1 was not correlated with Th1 (IL-2, IFN-γ) or Th2 (IL-4, IL-10) cytokine responses.

Conclusion

In this study, we demonstrate a significant correlation between CD200R1⁺ cells and disease severity in RA patients, thus indicating the relevance of the CD200/CD200R1 signaling pathway's potential involvement in the pathogenesis of RA.     

Association of interleukin-1 gene polymorphisms with multiple sclerosis: a meta-analysis

Background

Dysregulated levels of interleukin-1 (IL-1) were observed in patients with multiple sclerosis (MS). Previous studies have provided conflicting evidence implicating the IL-1 gene polymorphisms in MS risk.

Methods

A meta-analysis of 16 case–control studies involving 3,482 cases and 3,528 controls was conducted to evaluate this association.

Results

No association was found between the IL-1α ?889 (rs1800587), IL-1α +4,845 (rs17561), IL-1β ?511 (rs16944), IL-1β +3,953 (rs1143634), IL-1ra variable number tandem repeat (VNTR) polymorphisms and MS risk. However, in subgroup analyses for the IL-1ra VNTR polymorphism, we found that individuals carrying the 2 allele had a 32 % increased risk for bout-onset MS (relapsing remitting and secondary progressive MS) when compared to the LL homozygotes (OR = 1.32, 95 % CI = 1.06–1.66, P _z = 0.014).

Conclusion

Common variants in the IL-1 region are not associated with MS risk but our data suggest that the IL-1ra VNTR polymorphism might be associated with bout-onset MS subtype.     

Exercise alters the immune profile in Tg2576 Alzheimer mice toward a response coincident with improved cognitive performance and decreased amyloid

Background

Inflammation is associated with Aβ pathology in Alzheimer's disease (AD) and transgenic AD models. Previously, it has been demonstrated that chronic stimulation of the immune response induces pro-inflammatory cytokines IL-1β and TNF-α which contribute to neurodegeneration. However, recent evidence has shown that inducing the adaptive immune response reduces Aβ pathology and is neuroprotective. Low concentrations of IFN-γ modulate the adaptive immune response by directing microglia to differentiate to antigen presenting cells. Our objective was to determine if exercise could induce a shift from the immune profile in aged (17–19 months) Tg2576 mice to a response that reduces Aβ pathology.

Methods

TG (n = 29) and WT (n = 27) mice were divided into sedentary (SED) and exercised (RUN) groups. RUN animals were provided an in-cage running wheel for 3 weeks. Tissue was harvested and hippocampus and cortex dissected out. Quantitative data was analyzed using 2 × 2 ANOVA and student's t-tests.

Results

IL-1β and TNF-α were significantly greater in hippocampi from sedentary Tg2576 (TG_SED) mice than in wildtype (WT_SED) (p = 0.04, p = 0.006). Immune response proteins IFN-γ and MIP-1α are lower in TG_SED mice than in WT_SED (p = 0.03, p = 0.07). Following three weeks of voluntary wheel running, IL-1β and TNF-α decreased to levels indistinguishable from WT. Concurrently, IFN-γ and MIP-1α increased in TG_RUN. Increased CD40 and MHCII, markers of antigen presentation, were observed in TG_RUN animals compared to TG_SED, as well as CD11c staining in and around plaques and vasculature. Additional vascular reactivity observed in TG_RUN is consistent with an alternative activation immune pathway, involving perivascular macrophages. Significant decreases in soluble Aβ₄₀ (p = 0.01) and soluble fibrillar Aβ (p = 0.01) were observed in the exercised transgenic animals.

Conclusion

Exercise shifts the immune response from innate to an adaptive or alternative response. This shift in immune response coincides with a decrease in Aβ in advanced pathological states.     

Polymorphisms in CCR6 Are Associated with Chronic Graft-versus-Host Disease and Invasive Fungal Disease in Matched-Related Hematopoietic Stem Cell Transplantation

Graft-versus-host disease (GVHD) and fungal infections are frequent complications after allogeneic hematopoietic stem cell transplantation (HSCT). Single nucleotide polymorphisms (SNPs) in genes of the immune system can influence the inflammatory cascade and T cell–driven alloimmune reactions after HSCT, and thus increasing the incidence of GVHD and infectious complications. Here, we investigated the effect of SNPs in IL-23R and CCR6 on posttransplantation outcome in 161 recipients of partially T cell–depleted HSCT. Remarkably, IL-23R SNPs were not associated with clinical outcome, but we found that disparities in the CCR6 tagSNP rs2301436 and SNP rs3093023 are independently associated with the occurrence of chronic GVHD (cGVHD) and invasive fungal disease. In multivariate analysis, patients receiving a transplant from a homozygous rs2301436 G allele donor showed less cGVHD (odds ratio [OR]: 0.16; P = .002), as was the case for a homozygous donor rs3093023 G allele (OR: 0.24; P = .005). In parallel, the GG genotype at rs2301436 in donors was associated with a higher incidence of invasive fungal disease at day 100 after HSCT (OR: 3.59; P = .008). This study shows that CCR6 SNPs can be used to predict clinical outcome, and that polymorphisms in the CCR6 gene may influence T cell–mediated immune reactions after HSCT.     

Anti-nociceptive and anti-edematogenic effects of glibenclamide in a model of acute gouty attack in rats

Objective and design

We investigated the effect of glibenclamide on inflammatory parameters in a model of acute gouty attack in rats.

Treatment

Intra-articular injection of 50 μl of monosodium urate (MSU) crystals (1.25 mg/site) was used to induce gout-related inflammation. The effects of glibenclamide (1–10 mg/kg, s.c.) or dexamethasone (8 mg/kg, s.c., positive control) were assessed on several inflammation parameters.

Methods

Spontaneous nociception assessment, edema measurement, total and differential leucocyte counts, interleukin (IL)-1β release, prostaglandin E₂ production and determination of blood glucose levels were analyzed. Peritoneal macrophages were incubated with MSU and levels of IL-1β were measured. Statistical significance was assessed by one- or two-way analysis of variance.

Results

Glibenclamide (3 mg/kg) or dexamethasone (8 mg/kg) prevented nociception and edema induced by MSU injection in rats. Glibenclamide did not affect leukocyte infiltration, IL-1β release and PGE₂ production, but only reduced IL-1β production by MSU-stimulated macrophages at very high concentration (200 μM). Dexamethasone significantly reduced leukocyte infiltration, IL-1β release and PGE₂ production. Glibenclamide reduced whereas dexamethasone increased blood glucose levels of MSU-injected rats.

Conclusions

Glibenclamide reduced nociception and edema, but not leukocyte infiltration, IL-1β release and PGE₂ production. However, its substantial effect on nociception and edema suggests that glibenclamide can be an interesting option as an adjuvant treatment for pain induced by acute attacks of gout.     

Expression Pattern of Serum Cytokines in Hepatitis B Virus Infected Patients with Persistently Normal Alanine Aminotransferase Levels

Purpose

About 60–80 % of chronic hepatitis B virus (HBV) carriers are characterized with persistently normal alanine transaminase (ALT). Differences of cytokine expression are associated with the prognosis of HBV infection. We investigated the expression pattern of 30 cytokines associated with anti-HBV immunity in patients with normal ALT.

Methods

Four patient groups (immune tolerance, inactive hepatitis B surface antigen carriers, resolved hepatitis B, and control; 10 subjects per group) were assigned. Thirty cytokines, including IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-7, IL-9, IL-10, IL-12p40, IL-12p70, IL-15, IL-17A, IL-17C, IL-21, IL-22, IL-23p19, IL-28A, IL-29, CCL5, CCL16, CCL20, CCL22, CXCL9, CXCL10, CXCL11, TNFRSF8, TNFRSF18, IL-6R, gp130, and TGF-β1, were measured using a human cytokine antibody array. Signal intensities were obtained by laser scanner. Protein-protein interactions were analyzed by STRING (Search Tool for the Retrieval of Interacting Genes/Proteins).

Results

Significant differences of signal intensities were observed for IL-2, IL-4, IL-6, IL-7, IL-9, IL-10, IL-12p40, IL-12p70, IL-15, IL-21, IL-23p19, IL-28A, and IL-29. The lowest intensity was in controls. Among three HBV infection groups, significant differences were observed in IL-2, IL-4, IL-12p70, IL-15, IL-21, IL-23p19, and IL-29. The highest intensity was in the inactive group. All cytokines with significant differences were involved JAK-STAT signaling that up-regulate FOXP3, SOCS3 and MX1.

Conclusion

Differential expression of cytokines in JAK-STAT signaling is an important factor associated with prognosis of HBV infection. The elevation of γC cytokines, IL-12p70, IL-23p19, and IL-29 may promote spontaneous HBeAg seroconversion and HBV clearance.     

Successful Hematopoietic Stem Cell Transplantation in a Patient with LPS-Responsive Beige-Like Anchor (<Emphasis Type="Italic">LRBA</Emphasis>) Gene Mutation

Purpose

Autosomal recessive mutations in LRBA, encoding for LPS-responsive beige-like anchor protein, were described in patients with a common variable immunodeficiency (CVID)-like disease characterized by hypogammaglobulinemia, autoimmune cytopenias, and enteropathy. Here, we detail the clinical, immunological, and genetic features of a patient with severe autoimmune manifestations.

Methods

Whole exome sequencing was performed to establish a molecular diagnosis. Evaluation of lymphocyte subsets was performed for immunological characterization. Medical files were reviewed to collect clinical and immunological data.

Results

A 7-year-old boy, born to consanguineous parents, presented with autoimmune hemolytic anemia, hepatosplenomegaly, autoimmune thyroiditis, and severe autoimmune gastrointestinal manifestations. Immunological investigations revealed low immunoglobulin levels and low numbers of B and NK cells. Treatment included immunoglobulin replacement and immunosuppressive therapy. Seven years after disease onset, the patient developed severe neurological symptoms resembling acute disseminated encephalomyelitis, prompting allogeneic hematopoietic stem cell transplantation (HSCT) with the HLA-identical mother as donor. Whole exome sequencing of the patient uncovered a homozygous 1 bp deletion in LRBA (c.7162delA:p.T2388Pfs*7). Importantly, during 2 years of follow-up post-HSCT, marked clinical improvement and recovery of immune function was observed.

Conclusions

Our data suggest a beneficial effect of HSCT in patients with LRBA deficiency.

     

Elevated levels of Th17 cells and Th17-related cytokines are associated with disease activity in patients with inflammatory bowel disease

Objective

Interleukin-17(IL-17)-producing T helper(Th)17 cells are considered as a new subset of cells critical to the development of inflammatory bowel disease (IBD). We aimed to investigate the distribution of Th17 cells, the expressions of Th17-related cytokines (IL-17, IL-21 and IL-22) and their association with disease activity in IBD patients.

Methods

We collected intestinal tissue biopsies from 40 patients with active ulcerative colitis (UC), 20 patients with active Crohn’s disease (CD) and 20 healthy controls. The distribution of Th17 cells and expressions of Th17-related cytokines in colonic tissues were evaluated by a standard immunohistochemical procedure. Serum IL-17, IL-21 and IL-22 levels were determined by ELISA. Pearson’s and Spearman’s correlation analyses were performed to analyze the correlation between the number of Th17 cells, the expressions of Th17-related cytokines and disease activity index, endoscopic and histological grading, and CRP and PLT levels, respectively.

Results

Compared with healthy controls, the number of Th17 cells and the expressions of IL-17, IL-21 and IL-22 were significantly increased in active IBD patients (P < 0.05). In addition, Pearson’s and Spearman’s correlation analyses showed that the number of Th17 cells and the expressions of Th17-related cytokines were correlated with disease activity index, endoscopic and histological grading, CRP and PLT levels (P < 0.05).

Conclusions

Th17 cells and Th17-related cytokines (IL-17, IL-21 and IL-22) were increased in the intestinal mucosa in active IBD patients and may play an important role in disease activity and mucosal damage.