Tumor Necrosis Factor Receptor Gene Research Articles

Copy number variation and elevated genetic diversity at immune trait loci in Atlantic and Pacific herring

BackgroundGenome-wide comparisons of populations are widely used to explore the patterns of nucleotide diversity and sequence divergence to provide knowledge on how natural selection and genetic drift affect the genome. In this study we have compared whole-genome sequencing data from Atlantic and Pacific herring, two sister species that diverged about 2 million years ago, to explore the pattern of genetic differentiation between the two species.ResultsThe genome comparison of the two species revealed high genome-wide differentiation but with islands of remarkably low genetic differentiation, as measured by an FST analysis. However, the low FST observed in these islands is not caused by low interspecies sequence divergence (dxy) but rather by exceptionally high estimated intraspecies nucleotide diversity (π). These regions of low differentiation and elevated nucleotide diversity, termed high-diversity regions in this study, are not enriched for repeats but are highly enriched for immune-related genes. This enrichment includes genes from both the adaptive immune system, such as immunoglobulin, T-cell receptor and major histocompatibility complex genes, as well as a substantial number of genes with a role in the innate immune system, e.g. novel immune-type receptor, tripartite motif and tumor necrosis factor receptor genes. Analysis of long-read based assemblies from two Atlantic herring individuals revealed extensive copy number variation in these genomic regions, indicating that the elevated intraspecies nucleotide diversities were partially due to the cross-mapping of short reads.ConclusionsThis study demonstrates that copy number variation is a characteristic feature of immune trait loci in herring. Another important implication is that these loci are blind spots in classical genome-wide screens for genetic differentiation using short-read data, not only in herring, likely also in other species harboring qualitatively similar variation at immune trait loci. These loci stood out in this study because of the relatively high genome-wide baseline for FST values between Atlantic and Pacific herring.

B-220 Development of Reference Materials for Detection of Monkeypox

Abstract Background Monkeypox virus (MPXV) is a member of Poxviridae family and belongs to the genus of orthopoxvirus. The Monkeypox virus has a double stranded DNA genome of approximately 197 kb with almost 190 nonoverlapping ORFs. Monkeypox virus encodes all the enzymes needed for transcription and replication process of the viral genome. It has been hypothesized that progressive gene loss, primarily at the terminal end of the genome, has been the justification behind evolution of a new adaptive virus capable of attacking humans and causing serious illness. Monkeypox virus infection in human can lead to a smallpox-like illness with almost 11% fatality rate in individuals who are non-vaccinated. Monkeypox virus infects a wide range of mammalian cells without the need for specific host receptors, cell entry molecules and replication machinery. No specific approved treatment exists for Monkeypox. The symptoms could be managed by treating secondary bacterial infections, reducing symptoms, and providing supportive care. ST-246 (tecovirimat), FDA approved drug against Smallpox virus, has shown some efficacy against Monkeypox, and therefore, accurate diagnosis, usually through PCR-based testing is critical. LGC Clinical Diagnostics has developed AccuPlex Monkeypox Reference Material, which is a non-infectious, stable, and reproducible standard to aid nucleic acid testing for Monkeypox virus. Methods AccuPlex™ Monkeypox Reference Material consists of recombinant Adeno virus bearing the complete Monkeypox genes: J2L (TNF receptor gene), F8L (DNA polymerase gene), F3L (Double-stranded RNA-binding protein, inhibitor of IFN signaling), and N3R (orthopoxvirus MHC class I-like protein -OMCP). The design of the recombinant virus is based on the sequence of ON585038.1 USA strain from the recent outbreak. A digital PCR assay was developed using the E9L Non-variola (NVAR) Orthopox Generic Assay design described in Y. Li et al. /Journal of Clinical Virology 36 (2006) 194–203. Digital PCR testing using the BioRad QX-200 droplet digital PCR system was used to quantitate the viral copies/mL and guide formulation. The final product is targeted as a low positive control, approximately 2–3 times the detection limit of commercial PCR testing methods and is formulated in defibrinated human plasma. Results AccuPlex™ Monkeypox Reference Material gave an average value of 5.33E+03 copies/mL ± 6.34E+02 copies/mL by digital PCR. Along with the undiluted sample, 1:2 and 1:4 dilutions were made which were then tested on Quest Diagnostics Qualitative RT-PCR platform to verify performance and compatibility with the assay detection limits. All three samples were positively detected. Conclusion LGC Clinical Diagnostics has developed a stable, non-infectious reference material using recombinant Adenovirus technology for assays that detect Monkeypox DNA. Our study helps ensure that the control will be low positive for Monkeypox virus and could be used for monitoring the sensitivity of the assay. Data presented indicates the compatibility of the reference material with commercially available real time PCR assays. These materials may be used by labs for development, validation, training, and ongoing QC of Monkeypox detection assays and workflows.

Immune Factor, TNFα, Disrupts Human Brain Organoid Development Similar to Schizophrenia-Schizophrenia Increases Developmental Vulnerability to TNFα.

Schizophrenia (SZ) is a neurodevelopmental genetic disorder in which maternal immune activation (MIA) and increased tumor necrosis factor-α (TNF-α) may contribute. Previous studies using iPSC-derived cerebral organoids and neuronal cells demonstrated developmental malformation and transcriptional dysregulations, including TNF receptors and their signaling genes, common to SZ patients with diverse genetic backgrounds. In the present study, we examined the significance of the common TNF receptor dysregulations by transiently exposing cerebral organoids from embryonic stem cells (ESC) and from representative control and SZ patient iPSCs to TNF. In control iPSC organoids, TNF produced malformations qualitatively similar in, but generally less pronounced than, the malformations of the SZ iPSC-derived organoids. TNF and SZ alone disrupted subcortical rosettes and dispersed proliferating Ki67+ neural progenitor cells (NPC) from the organoid ventricular zone (VZ) into the cortical zone (CZ). In the CZ, the absence of large ramified pan-Neu+ neurons coincided with loss of myelinated neurites despite increased cortical accumulation of O4+ oligodendrocytes. The number of calretinin+ interneurons increased; however, they lacked the preferential parallel orientation to the organoid surface. SZ and SZ+TNF affected fine cortical and subcortical organoid structure by replacing cells with extracellular matrix (ECM)-like fibers The SZ condition increased developmental vulnerability to TNF, leading to more pronounced changes in NPC, pan-Neu+ neurons, and interneurons. Both SZ- and TNF-induced malformations were associated with the loss of nuclear (n)FGFR1 form in the CZ and its upregulation in deep IZ regions, while in earlier studies blocking nFGFR1 reproduced cortical malformations observed in SZ. Computational analysis of ChiPseq and RNAseq datasets shows that nFGFR1 directly targets neurogenic, oligodendrogenic, cell migration, and ECM genes, and that the FGFR1-targeted TNF receptor and signaling genes are overexpressed in SZ NPC. Through these changes, the developing brain with the inherited SZ genome dysregulation may suffer increased vulnerability to TNF and thus, MIA.

Molecular cloning, characterization, and expression of two TNFRs from the pearl oyster Pinctada fucata martensii

Proteins in the tumor necrosis factor receptor (TNFR) superfamily play significant roles in many physiological and pathological events, such as inflammation, apoptosis, autoimmunity, and organogenesis. Here, two TNFR gene homologs (PmTNFR1 and PmTNFR5) were identified in the pearl oyster Pinctada fucata martensii. The predicted PmTNFR1 and PmTNFR5 protein sequences were 406 and 533 amino acids long, respectively, and both possessed motifs characteristic of the TNFR family, including a TNFR homology domain (CRD), a transmembrane domain (TM), and death domains. However, the predicted amino acid sequences of PmTNFR1 and PmTNFR5 had low identity (~16–23%) with sequences of vertebrate TNFR family proteins. Furthermore, PmTNFR5 had a death domain at the C-terminal, indicating that this protein may be a novel member of the TNFR superfamily. Constitutive PmTNFR1 and PmTNFR5 mRNA expression was detected in all six pearl oyster tissues tested, with comparatively greater transcript abundance in the hepatopancreas and gill. The gene expression levels of PmTNFR1 and PmTNFR5, as well as those of downstream signaling molecules related to the NF-κB pathway (RIP, TRAF2, TRAF3, IKK, and NF-κB), were quantified in the gill after LPS challenge and in the hemocytes after nucleus insertion surgery using real-time PCR (qRT-PCR). We found that all genes were significantly upregulated at 6 h and 12 h post-injection, as well as at 15 d post-insertion. We used RNAi to inhibit the expression of the PmTNFR1 and PmTNFR5 genes. We then quantified the expression levels of PmTNFR1 and PmTNFR5, as well as downstream genes, using qRT-PCR. We found that RNAi inhibition of PmTNFR1 and PmTNFR5 downregulated the downstream genes (RIP, TRAF2, TRAF3, IKK, and NF-κB). Therefore, our results suggested that PmTNFR1 and PmTNFR5 mediate the NF-κB signaling pathway, and are closely related to immune defense, particularly allograft immunity, in the pearl oyster P. fucata martensii.

Effects of type 2 diabetes mellitus on gene expression profile of meibomian glands in mice

AbstractPurposeDiabetes mellitus (DM) is a major health problem and its complications cause blindness. Meibomian gland dysfunction and dry eye disease are also important ocular complications related with DM. According to our hypothesis, DM can exhibit significant altered gene expression in meibomian glands. To test our hypothesis, meibomian glands of leptin deficient spontaneous diabetic and non‐diabetic mice were extracted and gene expression profiles were analyzed with microarray.MethodsNine Lepob/ob spontaneous diabetic mice and nine non‐diabetic Balb/c mice (control group) were used. Blood glucose levels, tearfilm break‐up time and fluorescein scores were measured for 12 weeks. Meibomian glands were dissected and RNAs were isolated for microarray gene expression analysis. Probes with standard deviation more than 0.1 were filtered and used 40 452 of 45 281 probes for processing. Fold change analysis were performed and identified for affected genes, and impact of genes on proteins, pathways and gene ontologies were analyzed by using various databases.ResultsWe observed 172 up‐regulated and 118 down‐regulated genes in type‐2 diabetic mice when compared to non‐diabetic mice. Interestingly, expression of collagen type‐I, integrin beta‐I binding protein‐I, pyruvate dehydrogenase kinase, TNF receptor genes up‐regulated with DM; on other hand, IL‐33, cholecystokinin, plasminogen activator, IL‐1 and serine peptidase inhibitor genes down‐regulated significantly. Also, we have seen significant decrease in WNT signaling and pentose phosphate pathways related genes.ConclusionOur data shows these changes in gene expression caused by endocrine and immune mechanisms of type‐2 DM which result disrupted homeostasis of epithelial cells of MG. Increased expressions of apoptosis and inflammation related genes and their effects on related pathways have proven that meibomian glands were negatively affected from type‐2 DM.

Effects of Type 2 Diabetes Mellitus on Gene Expressions of Mouse Meibomian Glands

ABSTRACT Purpose Type 2 Diabetes mellitus (DM) is a major health problem and its ocular complications like orbital infections, cataract and diabetic retinopathy cause blindness. Meibomian gland (MG) dysfunction and dry eye disease are also important ocular complications of type 2 DM but not enough research has been conducted on these complications. Our hypothesis suggests type 2 DM can alter significant gene expressions of MG. In our study, MGs of leptin-deficient spontaneous diabetic and non-diabetic mice were extracted, and gene expression profiles were analyzed with microarray technology. Methods Mice were divided into two groups; nine Lep b/ob spontaneous diabetic mice as type 2 DM group and nine non-diabetic Balb/c mice as controls. Blood glucose levels, tearfilm break-up time and fluorescein scores were measured in both two groups for 12 weeks. MGs were dissected and RNAs were isolated for microarray gene expression analysis. We filtered probes with standard deviation of more than 0.1 and we used 40452 of 45281 probes for processing. We performed fold change analysis and identified which genes are affected, and we analyzed the impact of genes on proteins, pathways and gene ontologies by using various databases. Results We observed 172 up-regulated and 118 down-regulated genes in type 2 diabetic mice when compared to non-diabetic mice. Interestingly, expression of collagen type I, integrin beta-I binding protein-I, pyruvate dehydrogenase kinase, TNF receptor genes up-regulated with DM; on the other hand, IL-33, cholecystokinin, plasminogen activator, IL-1 and serine peptidase inhibitor genes down-regulated significantly. Also, we have seen a significant decrease in WNT signaling and pentose phosphate pathways-related genes. Conclusion Our data show these changes in gene expression caused by endocrine and immune mechanisms of type 2 DM which result disrupted homeostasis of epithelial cells of MG. Increased expressions of apoptosis and inflammation-related genes and their effects on related pathways have proven that MGs were negatively affected by type-2 DM.

Alterations of the Immunologic Co-Stimulator B7 and TNFR Families Correlate with Hepatocellular Carcinoma Prognosis and Metastasis by Inactivating STAT3.

Blockade of the immunosuppressive checkpoint receptors cytotoxic T-lymphocyte-associated protein 4 (CTLA4) or programmed death 1 (PD-1) and its cognate ligand, programmed death 1 ligand (PD-L1), has altered the landscape of anti-tumor immunotherapy. B7 family and tumor necrosis factor receptor (TNFR) superfamily play a crucial role in T cell activation, tolerance, and anergy through co-stimulatory and inhibitory signal transduction. Investigating the immune molecular landscapes of the B7 and TNFR families is critical in defining the promising responsive candidates. Herein, we performed comprehensive alteration analysis of the B7 and TNFR family genes across six hepatocellular carcinoma (HCC) datasets with over 1000 patients using cBioPortal TCGA data. About 16% of patients had both B7 and TNFR gene alterations. TNFR gene amplifications were relatively more common (1.73–8.82%) than B7 gene amplifications (1.61–2.94%). Analysis of 371 sequenced samples revealed that all genes were upregulated: B7 and TNFR mRNA were upregulated in 23% of cases (86/371) and 28% of cases (105/371), respectively. Promoter methylation analysis indicated an epigenetic basis for B7 and TNFR gene regulation. The mRNA levels of B7 and TNFR genes were inversely correlated with promoter methylation status. B7-H6 expression was significantly associated with worse overall survival, and B7-H6 mRNA was increased gradually in cases with gene copy number alterations. B7-H6 overexpression was associated with aggressive clinicopathologic features and poor prognosis in HCC. Downregulation of B7-H6 in HCC cells significantly inhibited cell adhesion, proliferation, migration, and invasion. Knockdown of B7-H6 in HCC cells inhibited tumor growth and metastasis in vivo. B7-H6 promoted HCC metastasis via induction of MMP-9 expression and STAT3 activation. B7-H6 and STAT3 performed functional overlapping roles on enhancing the MMP-9 promoter activity in HCC cells. These results suggest that alterations of the immunologic co-stimulator B7 and TNFR families correlate with HCC metastasis and prognosis, and especially B7-H6 plays a critical role in promoting metastasis of HCC.

Mechanisms of Retinal Damage after Ocular Alkali Burns

Alkali burns to the eye constitute a leading cause of worldwide blindness. In recent case series, corneal transplantation revealed unexpected damage to the retina and optic nerve in chemically burned eyes. We investigated the physical, biochemical, and immunological components of retinal injury after alkali burn and explored a novel neuroprotective regimen suitable for prompt administration in emergency departments. Thus, invivo pH, oxygen, and oxidation reduction measurements were performed in the anterior and posterior segment of mouse and rabbit eyes using implantable microsensors. Tissue inflammation was assessed by immunohistochemistry and flow cytometry. The experiments confirmed that the retinal damage is not mediated by direct effect of the alkali, which is effectively buffered by the anterior segment. Rather, pH, oxygen, and oxidation reduction changes were restricted to the cornea and the anterior chamber, where they caused profound uveal inflammation and release of proinflammatory cytokines. The latter rapidly diffuse to the posterior segment, triggering retinal damage. Tumor necrosis factor-α was identified as a key proinflammatory mediator of retinal ganglion cell death. Blockade, by either monoclonal antibody or tumor necrosis factor receptor gene knockout, reduced inflammation and retinal ganglion cell loss. Intraocular pressure elevation was not observed in experimental alkali burns. These findings illuminate the mechanism by which alkali burns cause retinal damage and may have importance in designing therapies for retinal protection.

The immune molecular landscape of the B7 and TNFR immunoregulatory ligand–receptor families in head and neck cancer: A comprehensive overview and the immunotherapeutic implications

ABSTRACTThe B7 family and tumor necrosis factor receptor (TNFR) superfamily play a vital role in the T-cell co-stimulatory and co-inhibitory pathways, regulating T-cell activation, tolerance, and exhaustion; therapeutic modulation of these pathways is translated into effective new cancer treatments. Better understanding of the immune molecular landscapes of the B7 and TNFR families would guide head and neck immuno-oncology clinical research. We performed comprehensive molecular profiling of 10 B7 and 6 TNFR family members in head and neck cancer. Over 20% of patients had B7 and TNFR gene alterations. B7 gene amplifications were relatively more common (3–11%) than TNFR gene amplifications (0–5%). Analysis of 496 sequenced samples revealed that all genes were upregulated: B7 and TNFR mRNA were upregulated in 158 cases (> 30%) and 83 cases (∼15%), respectively. B7-H1 (PD-L1) mRNA upregulation was the most common (∼10%). Promoter methylation analysis indicated an epigenetic basis for B7 and TNFR gene regulation (especially B7-H1, which was relatively strongly correlated with promoter methylation). B7-H1 expression was significantly associated with worse overall survival, and its expression was increased in cases with gene amplifications. Human papillomavirus (HPV) status correlated significantly with B7-H1 alterations at genetic level. Almost half (47.1%) of HPV-negative patients had deep or shallow B7-H1 deletion; >90% of HPV-positive patients had diploid, copy number gain, or amplification of B7-H1. This is the first study elucidating the immune molecular landscapes of the B7 and TNFR families in head and neck cancer, providing a potential novel rationale for clinical investigations.

Different types of cell cycle- and apoptosis-related gene expressions alter in corticosteroid-, vincristine-, and melphalan-resistant u-266 multiple myeloma cell lines.

Objective: Deregulation of the cell cycle and apoptosis mechanisms in normal cells causes many problems, including cancer. In this study, a genome-wide expression analysis of cell cycle- and apoptosis-related genes in corticosteroid-, vincristine-, and melphalan-resistant U-266 multiple myeloma cell lines was conducted. Materials and Methods: Resistant U-266 sublines were induced by application of each drug by stepwise dose increments. Resistance gained by the cells was confirmed with XTT cytotoxicity assay and microarray analyses were carried out. Among the cell cycle- and apoptosis-related gene expressions, alterations of more than 2-fold were considered significant. Results: Cyclin E2 was drastically overexpressed in the vincristine-resistant subline and a general upregulation was observed for various cyclin-dependent kinases. Some of the cyclin-dependent kinase inhibitor encoding genes were downregulated in resistant sublines in general. Tumor necrosis factor receptor genes were generally downregulated in corticosteroid- and melphalan-resistant U-266 sublines. Different types of effector caspases were downregulated in all resistant sublines. Ceramide metabolism genes seemed to be changed in favor of survival, especially in the melphalan-resistant subline. Conclusion: This report shows that different types of chemotherapeutic drugs alter different apoptotic and cell cycle-related gene expressions and, as a result, may cause drug-resistant phenotypes in U-266 multiple myeloma cell lines. Among those gene expressions, the most drastic increase in cyclin E2 could be important for the survival of vincristine-resistant U-266 cell lines, whereas alteration of ceramide metabolism genes could be important in melphalan resistance.

OP0071 Tnf-Alpha, TNF Receptor, HLA-E and NKG2A Gene Polymorphisms and Response to Anti-TNF-Alpha Treatement in Rheumatoid Arthritis

BackgroundDespite the fact that therapy with TNF-alpha inhibitors constitutes a breakthrough in rheumatoid arthritis (RA) management, no improvement is still achieved in approximately 30% of cases.ObjectivesThe aim of the study...

Polymorphisms in the tumor necrosis factor receptor genes affect the expression levels of membrane-bound type I and type II receptors.

The level of TNF receptors on various cells of immune system and its association with the gene polymorphism were investigated. Determining the levels of membrane-bound TNFα receptors on peripheral blood mononuclear cells (PBMCs) was performed by flow cytometry using BD QuantiBRITE calibration particles. Soluble TNFα receptor (sTNFRs) levels were determined by ELISA and genotyping was determined by PCR-RFLP. Homozygous TT individuals at SNP −609G/T TNFRI (rs4149570) showed lower levels of sTNFRI compared to GG genotype carriers. Homozygous carriers of CC genotype at SNP −1207G/C TNFRI (rs4149569) had lower expression densities of membrane-bound TNFRI on intact CD14+ monocytes compared to individuals with the GC genotype. The frequency differences in the CD3+ and CD19+ cells expressing TNFRII in relation to SNP −1709A/T TNFRII (rs652625) in healthy individuals were also determined. The genotype CC in SNP −3609C/T TNFRII (rs590368) was associated with a lower percentage of CD14+ cells expressing TNFRII compared to individuals with the CT genotype. Patients with rheumatoid arthritis had no significant changes in the frequencies of genotypes. Reduced frequency was identified for the combination TNFRI −609GT + TNFRII −3609CC only. The polymorphisms in genes represent one of cell type-specific mechanisms affecting the expression levels of membrane-bound TNFα receptors and TNFα-mediated signaling.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Association between genetic variations in tumor necrosis factor receptor genes and survival of patients with T-cell lymphoma

The prognosis of T-cell lymphoma (TCL) has been shown to be associated with the clinical characteristics of patients. However, there is little knowledge of whether genetic variations also affect the prognosis of TCL. This study investigated the associations between single nucleotide polymorphisms (SNPs) in tumor necrosis factor receptor superfamily (TNFRSF) genes and the survival of patients with TCL. A total of 38 tag SNPs in 18 TNFRSF genes were genotyped using Sequenom platform in 150 patients with TCL. Kaplan-Meier survival estimates were plotted and significance was assessed using log-rank tests. Cox proportional hazard models were used to analyze each of these 38 SNPs with adjustment for covariates that might influence patient survival, including sex and international prognostic Index score. Hazard ratios (HRs) and their 95% confidence intervals (CIs) were calculated. Among the 38 SNPs tested, 3 were significantly associated with the survival of patients with TCL. These SNPs were located at LTβR (rs3759333C>T) and TNFRSF17 (rs2017662C>T and rs2071336C>T). The 5-year survival rates were significantly different among patients carrying different genotypes and the HRs for death between the different genotypes ranged from 0.45 to 2.46. These findings suggest that the SNPs in TNFRSF genes might be important determinants for the survival of TCL patients.

Enhancement of the survival of engrafted mesenchymal stem cells in the ischemic heart by TNFR gene transfectionThis paper is one of a selection of papers published in this special issue entitled “Second International Symposium on Recent Advances in Basic, Clinical, and Social Medicine” and has undergone the Journal's usual peer review process.

Autologous or allogeneic mesenchymal stem cells (MSCs) have been used as one of the potential cell sources for cellular cardiomyoplasty. The adverse microenvironment in acute myocardial infarction, however, is considered a deleterious factor for MSC transplantation and cell survival. Tumor necrosis factor (TNF)-alpha is an inflammatory mediator produced during ischemia that may affect the survival of MSCs. In this study, we investigated the enhancement of MSC survival by transfecting cells with the TNF receptor (TNFR) gene, leading to the overproduction of TNFR and the binding of TNF-alpha. Rats with acute myocardial infarction, induced by the occlusion of the left coronary artery, were transplanted with MSC or MSC-TNFR. After 2 weeks of acute myocardial infarction, cardiac function was assessed. Engrafted MSC survival and localization of TNF-alpha protein in infarction myocardium were evaluated. The levels of TNF-alpha and TNFR in the infarction zone were assessed. The results indicate that MSC-TNFR transplantation (1) improved left ventricular function; (2) enhanced engrafted MSC survival in the infarcted myocardium; (3) attenuated the level of TNF-alpha in serum and cardiac tissue; and (4) increased TNFR protein production in the infarcted myocardium. Our results showed that MSC modified by the TNFR gene improved cell viability and thereby has the potential to improve the efficiency of MSC transplantation therapy in the ischemic heart.

Autologous (T cell depleted) or allogeneic haematopoietic stem cell transplantation for treatment of recalcitrant, severe autoimmune/rheumatic disorders in children?

Autologous haematopoietic stem cell transplantation (HSCT) following immunosuppressive conditioning and T cell depletion of the graft is “a valuable option for individual” children with severe autoimmune/rheumatic disease recalcitrant to currently used treatments, although recent evidence of only transient benefit suggest this is not a curative procedure. Potentially curative allogeneic HSCT is “generally not recommended” and unacceptable for the paediatric rheumatology community, even for selected patients at the very severe end of the disease spectrum, as graft-versus-host disease (GVHD) and transplant-related mortality are considered too high a risk.1 Results Our data on autologous T cell-depleted HSCT in 6 children with juvenile idiopathic arthritis have been recently published2: four are long-term survivors, three in complete drug-free remission, one relapsed and two died. The authors compare these with the results of allogeneic HSCT in seven children with well-defined, severe and treatment-resistant autoimmune and autoinflammatory diseases: systemic lupus erythematosus (1), immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (3), autoimmune lymphoproliferative syndrome (2), complex periodic fever syndrome due to both mevalonic kinase and TNF-receptor gene mutations (1). Half received HSC from an HLA-matched sibling and half from an unrelated donor; 5/7 received modified myeloablative or reduced intensity conditioning. One child died; 6 achieved remission from their underlying disease: four are cured with 2–6 year follow-up, one with slipping donor chimerism requires further treatment and one is still Conclusion The benefit–risk ratio for allogeneic HSCT using contemporary protocols in specialised centres may have surpassed T cell-depleted autologous HSCT for severe autoimmune/rheumatic disease.

Low TNF-induced NF- B and p38 phosphorylation levels in leucocytes in tumour necrosis factor receptor-associated periodic syndrome

TNF receptor-associated periodic syndrome (TRAPS) is a systemic autoinflammatory disorder caused by mutations in the type 1 TNF receptor (TNFRSF1A) gene. Because the pathomechanism of TRAPS may involve aberrant TNF-mediated intracellular signalling, we examined phosphorylation levels of nuclear factor kappaB (NF-kappaB) and p38 in response to TNF in 10 patients with three different TNFRSF1A mutations (C73R, C88Y and F112I). Phosphorylation levels of NF-kappaB p65 and p38 were determined in fresh leucocytes stimulated with TNF (0-100 ng/ml) for 2.5-20 min and permeabilized for phospho-specific antibodies in a whole blood flow cytometry assay. As control agonists, we used bacterial lipopolysaccharide (LPS) and IFN-gamma, the latter mediating phosphorylation of the signal transducer and activator of transcription 1. Areas under curve values for dose-response and time course of NF-kappaB and p38 phosphorylation were calculated for the comparison of patients and reference subjects. NF-kappaB and p38 phosphorylation levels of monocytes, lymphocytes and neutrophils stimulated with TNF were significantly lower in TRAPS patients than in reference subjects. Phosphorylation levels induced by LPS, or by IFN-gamma, in patient and reference samples were comparable, indicating that the defect was confined to TNF-mediated signalling. In the three families studied, TRAPS was associated with low TNF-mediated signalling in leucocytes. This deficiency of the innate immune system may result in the activation of as yet unidentified compensatory regulatory mechanisms yielding the hyperinflammatory phenotype of TRAPS.

IL-1 or TNF receptor gene deletion delays onset of encephalopathy and attenuates brain edema in experimental acute liver failure

Previous reports suggested that brain-derived proinflammatory cytokines are involved in the pathogenesis of hepatic encephalopathy (HE) and brain edema in acute liver failure (ALF). To further address this issue, expression of interleukin-1beta (IL-1β), tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) mRNAs were measured in the brains of mice with acute liver failure resulting from exposure to azoxymethane. In addition, time to severe encephalopathy (coma) was assessed in mice lacking genes coding for interferon-gamma, the tumor necrosis factor receptor-1 or the interleukin-1 type 1 receptor. Interleukin-1beta, tumor necrosis factor-alpha and interferon-gamma expression were quantified using RT-PCR. Significant increases in interleukin-1beta and tumor necrosis factor-alpha mRNA were observed in the frontal cortex of azoxymethane-treated wild-type mice at coma stages of encephalopathy. Interferon-gamma, however, could not be detected in the brains of these animals. Onset of severe encephalopathy (coma) and brain edema in ALF mice were significantly delayed in interleukin-1 type 1 receptor or tumor necrosis factor receptor-1 knockout mice. Deletion of the interferon-gamma gene, on the other hand, had no significative effect on the neurological status or brain water content of acute liver failure mice. These results demonstrate that toxic liver injury resulting from exposure to azoxymethane is associated with selective induction of proinflammatory cytokines in the brain and that deletion of tumor necrosis factor receptor-1 or interlukin-1 type 1 receptor delays the onset of coma and brain edema in this model of acute liver failure. These findings further support a role for selective brain-derived cytokines in the pathogenesis of the cerebral complications in acute liver failure and suggest that anti-inflammatory strategies could be beneficial in their prevention.

Association between polymorphisms in tumor necrosis factor (TNF) and TNF receptor genes and circulating TNF, soluble TNF receptor levels, and bone mineral density in postmenopausal Korean women

The aim of the present study was to investigate the relationship of polymorphisms in the tumor necrosis factor (TNF)-alpha, TNF-beta, and TNF receptor (TNFR) genes to circulating TNF, soluble TNFR (sTNFR) levels, and bone mineral density (BMD) in women. The TNF-alpha G(-308)A, C(-857)T, C(-863)A, T(-1031)C, TNF-beta A252G, TNFRI A36G, TNFRII T676G, A1663G, A1668G, and C1690T polymorphisms were analyzed in 377 postmenopausal Korean women. The levels of serum TNF-alpha, TNF-beta, sTNFRI, sTNFRII, and bone turnover markers were measured. BMD was examined by dual energy x-ray absorptiometry. After adjustment for age, body mass index, and years since menopause, no significant differences in BMD of the lumbar spine and femoral neck and serum levels of bone turnover markers were detected, according to single polymorphisms in TNF and TNFR genes and combined polymorphisms. However, the frequencies of the TT genotype of TNF-alpha T(-1031)C polymorphism, the non-AA genotype of TNF-beta A252G polymorphism, and the GG genotype of TNFRII A1663G polymorphism were significantly higher in osteoporotic women than in women with normal BMD, respectively (P < 0.05). The TNFRII T676G polymorphism affected the serum sTNFRI and sTNFRII levels. The serum sTNFRII levels in the CC genotype of TNFRII C1690T polymorphism were the highest, with a G or C allele dose effect, and the TNFRII G676C/C1690T haplotype genotype also affected serum sTNFRII levels. TNF-alpha T(-1031)C, TNF-beta A252G, and TNFRII A1663G polymorphisms may be genetic factors for osteoporosis in Korean postmenopausal women, and the TNFRII T676G and C1690T polymorphisms and their combined polymorphism affected serum sTNFRII levels. The TNFRII T676G polymorphism also affected the serum sTNFRI levels.

Index of Suspicion

Index of Suspicion