TNF Family Cytokines Research Articles

BAFF neutralization impairs the autoantibody-mediated clearance of dead adipocytes and aggravates obesity-induced insulin resistance.

B cell-activating factor (BAFF) is a critical TNF-family cytokine that regulates homeostasis and peripheral tolerance of B2 cells. BAFF overproduction promotes autoantibody generation and autoimmune diseases. During obesity, BAFF is predominantly produced by white adipose tissue (WAT), and IgG autoantibodies against adipocytes are identified in the WAT of obese humans. However, it remains to be determined if the autoantibodies formed during obesity affect WAT remodeling and systemic insulin resistance. Here, we show that IgG autoantibodies are generated in high-fat diet (HFD)-induced obese mice that bind to apoptotic adipocytes and promote their phagocytosis by macrophages. Next, using murine models of obesity in which the gonadal WAT undergoes remodeling, we found that BAFF neutralization depleted IgG autoantibodies, increased the number of dead adipocytes, and exacerbated WAT inflammation and insulin resistance. RNA sequencing of the stromal vascular fraction from the WAT revealed decreased expression of immunoglobulin light-chain and heavy-chain variable genes suggesting a decreased repertoire of B cells after BAFF neutralization. Further, the B cell activation and the phagocytosis pathways were impaired in the WAT of BAFF-neutralized mice. In vitro, plasma IgG fractions from BAFF-neutralized mice reduced the phagocytic clearance of apoptotic adipocytes. Altogether, our study suggests that IgG autoantibodies developed during obesity, at least in part, dampens exacerbated WAT inflammation and systemic insulin resistance.

TNF-like weak inducer of apoptosis inhibition is comparable to IL-13 blockade in ameliorating atopic dermatitis inflammation.

Targeting IL-13 is highly efficacious in patients with Th2-biased atopic dermatitis (AD), but inhibition of other inflammatory molecules might also limit disease. We investigated the importance of the TNF family cytokine TNF-like weak inducer of apoptosis (TWEAK; TNFSF12) to keratinocyte dysregulation and the pathogenesis of AD in mice and also tested if blocking TWEAK has a similar therapeutic effect as targeting IL-13. Conditional knockout mice lacking Fn14 (TNFRSF12A), the receptor for TWEAK, only in keratinocytes, were repetitively sensitized with house dust mite allergen and analyzed for AD-like skin inflammation. To determine the translational potential, wild-type mice with AD were therapeutically treated with anti-TWEAK and/or anti-IL-13 antibodies, and skin inflammation was assessed. Mice deficient in Fn14 in keratinocytes were resistant to developing maximal clinical features of AD, exhibiting reduced epidermal hyperplasia and dermal thickening, less skin infiltration of immune cells, and downregulated inflammatory gene expression. Moreover, therapeutic neutralization of TWEAK in wild-type mice with AD reduced all of the pathological features to a comparable extent as blocking IL-13. The activity of TWEAK in keratinocytes contributes to AD development, and neutralizing TWEAK represents a future potential therapeutic option in human AD similar to targeting IL-13.

ADSC-derived exosomes inhibit cbl-cathepsin K-dependent osteoclast activation in inflammatory arthritis

Abstract Rheumatoid arthritis (RA) is a chronic inflammatory disease that causes joint inflammation and bone destruction due to increased osteoclast differentiation and activity. Adipose-derived stem cells (ADSCs) are used to treat tissue injury because they are easy to harvest and exhibit immunomodulatory and regenerative capacity. Previous studies demonstrated that ADSC-derived exosomes (ADSC-Exo) protect articular cartilage from damage and exert anti-inflammation effects. Thus, we investigated the underlying mechanisms of ADSC-Exo in regulating osteoclast activation in inflammatory arthritis. RANKL is a TNF-family cytokine that binds to the RANK required for mature osteoclasts. However, whether ADSC-Exo is functionally involved in the inhibition of osteoclast formation via regulating RANK-mediated signaling in these pathological conditions is unclear. Our data revealed that ADSC-Exo ameliorated the RA severity and bone loss in vivo and inhibited osteoclast differentiation, RANK downstream signaling, NFATc1, cathepsin K, and cbl-b. Moreover, ADSC-Exo could significantly suppress the population of RANKL-expressing TH17 cells. In brief, ADSC-Exo protects articular cartilage and bone destruction from damage and ameliorates gait abnormality in CIA rats, which may be related to the inhibition of the RANK-signaling pathway in osteoclasts. As it is relatively feasible to obtain human adipose tissue, ADSC-Exo may be a potential therapy for RA. This study could comprehensively understand the immunomodulatory effects of ADSC-Exo on suppressing osteoclastogenesis. This finding is critical for understanding the bone homeostasis modulated by MSC-Exo and provides biological evidence of ADSC-Exo in inflammatory arthritis. Supported by grants from the Ministry of Science and Technology (111-2321-B-418-002, 111-2314-B-418-004), FEMH (2022-C-042 and 2022-C-123), NYCU-FEMH Cooperation grants (111DN12, 111DN22), NTUH-FEMH Cooperation (111-FTN0012).

The Potential of Therapeutic Targeting of the TWEAK-Fn14 pathway in Atopic Dermatitis

Abstract Atopic dermatitis (AD) is a chronic skin inflammatory condition exhibiting a markedly thickened epidermis. Keratinocytes, the major cells of the epidermis, are a key target of inflammation and several cytokines including IL-13 and IL-22 have receptors expressed on these cells. Because Fn14 (TNFRSF12A), the receptor for the TNF family cytokine TWEAK (TNFSF12), is also expressed on keratinocytes, we examined the inflammatory potential of TWEAK alone, and together with IL-13 and IL-22, in controlling AD-signature gene expression in using RNA-seq. Transcriptomic analysis showed that TWEAK strongly upregulated many AD genes in human keratinocytes, which partially overlapped with those induced by IL-13 and IL-22. TWEAK also acted synergistically with IL-13 and IL-22 to exaggerate induction of AD-related genes. Correspondingly, cKO mice lacking Fn14 only in keratinocytes were protected from developing pronounced allergen-induced AD-like skin inflammation. Furthermore, mice treated therapeutically with anti-TWEAK exhibited a significantly reduced AD phenotype including less epidermal thickening combined with a strongly reduced Th2 response in the skin. Moreover, the therapeutic efficacy of TWEAK inhibition was similar to that resulting from blockade of IL-13. The data suggest that TWEAK shares pathogenic activity with IL-13 and IL-22 and can be considered as a potential therapeutic target in AD similar to IL-13. Nil

Necroptosis of Lung Epithelial Cells Triggered by Ricin Toxin and Bystander Inflammation.

The ribosome-inactivating proteins include the biothreat agent, ricin toxin (RT). When inhaled, RT causes near complete destruction of the lung epithelium coincident with a proinflammatory response that includes TNF family cytokines, which are death-inducing ligands. We previously demonstrated that the combination of RT and TNF-related apoptosis inducing ligand (TRAIL) induces caspase-dependent apoptosis, while RT and TNF-α or RT and Fas ligand (FasL) induces cathepsin-dependent cell death in lung epithelial cells. We hypothesize that airway macrophages constitute a major source of cytokines that drive lung epithelial cell death. Here, we show that RT-induced apoptosis of the monocytic cell line, U937, leads to the bystander killing of the lung epithelial cell line, A549. U937 cells were treated with ricin. Following this, A549 cells were treated with supernatants from U937 cells and death was measured by WST-1 viability assay. Upon RT-induced U937 cell death, released RT and FasL contributed to A549 cell death. U937 cells also released nuclear protein HMGB1. The release of RT, FasL, and HMGB1 triggered A549 cell necroptosis, rather than cathepsin-dependent killing observed previously with RT and FasL. Reactive oxygen species (ROS) were produced in A549 cells due to HMGB1 ligation of the receptor for advanced glycation end products (RAGE). These findings demonstrate the potential for bystander necroptosis of lung epithelial cells during RT toxicosis which may perpetuate or increase the proinflammatory response.

B-Cell Activating Factor secreted by neutrophils is a critical player in lung inflammation to cigarette smoke exposure

B cell activating factor (BAFF) is a TNF family cytokine mainly produced by adaptive immune cells in particular B and T lymphocytes. Depending on the context, innate immune cells can also produce BAFF. First of all, BAFF was known to play an important role in the maturation and survival of B lymphocytes in humoral immune response. Recent data indicate that BAFF may also participate in the regulation of innate immune responses and in the pathophysiology of pulmonary diseases. In models of mice chronically exposed to cigarette smoke (CS), BAFF plays a major role in the generation of pulmonary antinuclear antibodies and tertiary lymphoid follicles. However, the implication of BAFF in the development of innate immunity to cigarette smoking remains unknown. Wild-type or BAFF deficient mice were exposed to 3R4F cigarette smoke 3 times a day for 4 days. Mice were treated or not with mouse anti-GR1 or recombinant mouse BAFF during CS exposure. Broncho-alveolar lavage fluids (BALF) were done and cell infiltration was determined. Different cytokines and chemokines were measured in BALF and lung homogenates. Immunostaining on cytospin were done. Bone marrow derived neutrophils (BMDN) and Murine tracheal epithelial cells (MTEC) were stimulated in vitro by CSE (Cigarette Smoke Extract). Cell viability and BAFF levels were determined. Acute CS exposure promotes BAFF expression in airway recruited neutrophils. Immunostaining analysis revealed that neutrophils are the major source of BAFF. We confirmed in vitro that neutrophils secrete BAFF in response to CSE stimulation. Neutrophil depletion partially decreases BAFF expression in lung tissue and bronchoalveolar space suggesting additional sources of BAFF. Importantly, BAFF deficient mice displayed decreased airway neutrophil recruiting chemokines and neutrophil influx while the addition of exogenous BAFF significantly enhanced this CS-induced neutrophilic inflammation. These results demonstrate that BAFF is a key pro-inflammatory cytokine and that innate immune cells in particular neutrophils, are an unconsidered source of BAFF in early stages of CS-induced innate immunity.

Analysis of putative G-quadruplex forming sequences in inflammatory mediators and their potential as targets for treating inflammatory disorders

G-quadruplexes (G4s) are non-canonical secondary structures located in DNA and RNA which have demonstrable roles in the regulation of transcription and translation. G4s have received considerable interest as a drug target in cancer, given their ability to regulate the expression of proto-oncogenes and inhibit growth of cancer cells. However, their presence in the genes of inflammatory mediators has not been discussed to date. Therefore, we computationally investigated putative quadruplex-forming sequences (PQS) in the promoters and gene bodies of cytokines and chemokines. Here, we demonstrated that the promoters of IL-6, IL-12, IL-17, TGF-β, TNF, and β-chain family cytokines and XC and TAFA family chemokines display high PQS frequencies comparable to those observed in proto-oncogenes. Moreover, 47.82% of the gene promoters contained sequences with high propensity to form G4s. Furthermore, G4s can primarily be found within the GC-boxes and binding sites for specificity protein and Krϋppel-like transcription factors. However, they can also be found located in a further 59 sites involved in the binding of transcription factors involved in inflammation and immunity such as NF-κB1, RelA, RelB, IRF5, and NFAT5. We also identified that 72.17% and 70.43% of genes investigated contained sequences highly likely to form G4s in their coding and template strands, respectively. Exploring the regulatory roles of G4s in genes encoding inflammatory mediators could provide novel drug targets to modulate inflammation and treat inflammatory diseases.

TL1A Promotes Lung Tissue Fibrosis and Airway Remodeling.

Lung fibrosis and tissue remodeling are features of chronic diseases such as severe asthma, idiopathic pulmonary fibrosis, and systemic sclerosis. However, fibrosis-targeted therapies are currently limited. We demonstrate in mouse models of allergen- and bleomycin-driven airway inflammation that neutralization of the TNF family cytokine TL1A through Ab blocking or genetic deletion of its receptor DR3 restricted increases in peribronchial smooth muscle mass and accumulation of lung collagen, primary features of remodeling. TL1A was found as a soluble molecule in the airways and expressed on the surface of alveolar macrophages, dendritic cells, innate lymphoid type 2 cells, and subpopulations of lung structural cells. DR3 was found on CD4 T cells, innate lymphoid type 2 cells, macrophages, fibroblasts, and some epithelial cells. Suggesting in part a direct activity on lung structural cells, administration of recombinant TL1A into the naive mouse airways drove remodeling in the absence of other inflammatory stimuli, innate lymphoid cells, and adaptive immunity. Correspondingly, human lung fibroblasts and bronchial epithelial cells were found to express DR3 and responded to TL1A by proliferating and/or producing fibrotic molecules such as collagen and periostin. Reagents that disrupt the interaction of TL1A with DR3 then have the potential to prevent deregulated tissue cell activity in lung diseases that involve fibrosis and remodeling.

TRAIL therapy and prospective developments for cancer treatment

Tumor Necrosis Factor (TNF) Related Apoptosis-Inducing Ligand (TRAIL), an immune cytokine of TNF-family, has received much attention in late 1990s as a potential cancer therapeutics due to its selective ability to induce apoptosis in cancer cells. TRAIL binds to cell surface death receptors, TRAIL-R1 (DR4) and TRAIL-R2 (DR5) and facilitates formation of death-inducing signaling complex (DISC), eventually activating the p53-independent apoptotic cascade. This unique mechanism makes the TRAIL a potential anticancer therapeutic especially for p53-mutated tumors. However, recombinant human TRAIL protein (rhTRAIL) and TRAIL-R agonist monoclonal antibodies (mAb) failed to exert robust anticancer activities due to inherent and/or acquired resistance, poor pharmacokinetics and weak potencies for apoptosis induction. To get TRAIL back on track as a cancer therapeutic, multiple strategies including protein modification, combinatorial approach and TRAIL gene therapy are being extensively explored. These strategies aim to enhance the half-life and bioavailability of TRAIL and synergize with TRAIL action ultimately sensitizing the resistant and non-responsive cells. We summarize emerging strategies for enhanced TRAIL therapy in this review and cover a wide range of recent technologies that will provide impetus to rejuvenate the TRAIL therapeutics in the clinical realm.

TNF Family Cytokines Induce Distinct Cell Death Modalities in the A549 Human Lung Epithelial Cell Line when Administered in Combination with Ricin Toxin.

Ricin is a member of the ribosome-inactivating protein (RIP) family of toxins and is classified as a biothreat agent by the Centers for Disease Control and Prevention (CDC). Inhalation, the most potent route of toxicity, triggers an acute respiratory distress-like syndrome that coincides with near complete destruction of the lung epithelium. We previously demonstrated that the TNF-related apoptosis-inducing ligand (TRAIL; CD253) sensitizes human lung epithelial cells to ricin-induced death. Here, we report that ricin/TRAIL-mediated cell death occurs via apoptosis and involves caspases -3, -7, -8, and -9, but not caspase-6. In addition, we show that two other TNF family members, TNF-α and Fas ligand (FasL), also sensitize human lung epithelial cells to ricin-induced death. While ricin/TNF-α- and ricin/FasL-mediated killing of A549 cells was inhibited by the pan-caspase inhibitor, zVAD-fmk, evidence suggests that these pathways were not caspase-dependent apoptosis. We also ruled out necroptosis and pyroptosis. Rather, the combination of ricin plus TNF-α or FasL induced cathepsin-dependent cell death, as evidenced by the use of several pharmacologic inhibitors. We postulate that the effects of zVAD-fmk were due to the molecule’s known off-target effects on cathepsin activity. This work demonstrates that ricin-induced lung epithelial cell killing occurs by distinct cell death pathways dependent on the presence of different sensitizing cytokines, TRAIL, TNF-α, or FasL.

IL-9high GATA3low ILC2s are potent inducers of allergic airway inflammation

Abstract IL-9 is key driver of chronic and allergic inflammation at mucosal surfaces, with important roles in the activation and survival of mast cells and type 2 innate lymphoid cells (ILC2s). IL-9-reporter mice have established T cells (Th9) and ILC2s as major sources of IL-9 in vivo. The discovery that TGF-β and IL-4 cooperate for induction of Th9 cells has been an important breakthrough. However, the signals that induce IL-9high ILC2s remain incompletely characterized. Here, we show that IL-33, a critical activator of ILC2s, with important roles in type-2 innate immunity and allergic inflammation (Cayrol, Duval et al., Nature Immunology 2018) synergizes with TNF-family cytokine TL1A to induce prodigious amounts of IL-9 secretion by ILC2s. Unbiased global proteomic analyses revealed that IL-9 was the most-induced protein in ILC2s treated with IL-33 and TL1A. More than 99% of ILC2s expressed IL-9 intracellularly after co-stimulation with IL-33 and TL1A. ILC2s are known to produce very large amounts of IL-5 and IL-13, but ILC2s producing such high levels of IL-9 have not been described previously. Interestingly, IL-9 production by ILC2s was transient and associated with a phenotypic shift characterized by upregulation of IRF4, pSTAT5 and downregulation of ICOS, KLRG1 as well as master transcription factor GATA-3 and its target genes (IL-9R, ST2). Lastly, we detected an expansion of IL-9high ILC2s in the lungs of mice after IL-33/TL1A inhalation and adoptive transfer experiments revealed that IL-9high ILC2s are potent inducers of allergic airway inflammation in vivo. Thus, our study supports the rising plasticity of ILC2s and their critical role in the induction of type-2 inflammatory responses in the lungs.

PTEN gene & TNF-alpha in acute myocardial infarction

BackgroundPTEN gene triggers cells to undergo apoptosis and promotes myocardial dysfunction. Several TNF family cytokines are elevated during acute myocardial infarction (AMI). Their role in predicting subsequent prognosis in these setting remains poorly understood. We assessed serum levels of PTEN gene activity & TNF-α in acute ST elevation myocardial infarction and determined the impact of their levels on both left ventricular function and the clinical outcome in these patients. Methods and resultsSeventy patients with AMI and seventy persons as control group were subjected to: ECG, echocardiography, serum TNF-α and PTEN gene assessment. Patients were classified into: Group I (n = 32): All had left ventricular systolic failure. Group II (n = 38): without left ventricular systolic failure. Group I had a statistically significant higher serum levels of both TNF-α & PTEN gene activity as compared to group II. EF% at presentation was weakly correlated with serum levels of both markers in both groups. However at follow up, EF% in group I showed a significant negative correlations with both serum levels of TNF-α and PTEN gene activity (r = 0.77 & r = 0.67, respectively). During one year follow, 5 patients died of cardiovascular causes and 6 patients had recurrent hospitalization with heart failure. These patients had statistically significant increased serum levels of TNF-α & PTEN gene activity levels as compared by other patients. ConclusionsPatients with acute myocardial infarction had statistically significant increased serum levels of PTEN & TNF-α gene activity. Both markers predict worsening of left ventricular systolic functions, development of heart failure and death.

Serum Levels of APRIL Increase in Patients with Glioma, Meningioma and Schwannoma

Objective:Brain tumors are of high mortality and morbidity for which there is still no cure. The TNF family cytokine, A Proliferation Inducing Ligand (APRIL), is shown to help proliferation and development of tumor cells. We assessed serum levels of APRIL in patients with glioma, meningioma and schwannoma in comparison to healthy individuals.Methods:Peripheral blood samples of 68 patients with brain tumors, divided into three groups of gliomas (n=25), meningiomas (n=30) and schwannomas (n=13), as well as 45 healthy individuals were obtained. Serum samples were prepared and stored in -40°C until usage. Using a commercial ELISA method, APRIL concentration was measured in each serum sample. The obtained data were then analyzed using SPSS software.Results:APRIL serum levels were higher in all patients compared to the controls (P<0.001). Moreover, APRIL serum levels were higher in each of the tumor bearing groups (gliomas, meningiomas and schwannomas) in comparison to the controls (P<0.001, <0.001 and =0.001, respectively). Comparing APRIL between the patients groups showed no significant difference. Age and gender showed no significant correlation with serum APRIL levels, although the age of patients in glioma group was significantly lower than controls (P=0.017). The serum APRIL levels in gliomas with histological grade showed no difference, but in meningiomas, it was lower in tumors with higher grades (P= 0.011).Conclusion:Increased serum levels of APRIL in patients with meningioma and schwannoma as well as glioma may indicate a common role of this cytokine in brain tumors.

INFLUENZA INFLAMMATION BIOMARKERS FEATURES

Aim. Analysis of inflammation biomarkers using reverse transcription with real time PCR (RT-PCR-RT) and multiplex immunofluorescent analysis xMAP with magnetic beads for the influenza infection. Materials and methods. Analysis of nasopharyngeal swabs, lymphocytes and blood sera of 10 patients with influenza and 10 donors was performed during the first 2 days of the disease by means of RT-PCR-RT and xMAP using the kit «37-plex» (BioRad). Results.The influenza virus A was revealed in 4 samples, the influenza virus B — in 6 swabs without mixed infections with other respiratory viruses. Analysis of the interferons (IFN) showed IFNα gene expression activation in patients’ lymphocytes but both the detection rate and the concentrations of IFNβ, IFNγ and IFNλ RNA were similar for patients and healthy donors. Among 37 inflammation biomarkers the concentrations of 7 proteins were enhanced including IFNα2, cytokines of TNF family (APRIL and BAFF), their soluble receptors sTNF-R1 and sTNF-R2, protein osteopontin and IL10. The concentrations of the complex of glycoprotein gp130 with the soluble receptor IL6 gp130/sIL-6Rβ and the matrix metalloprotease ММР-1 were reduced in patients’ sera. The polarization coefficient PI=[IL10]/[IFNγ]=0.53 for influenza samples suggested Th1 immune response. Conclusion. At the early stage of the influenza infection IFNα gene expression activation along with the induction of TNF family cytokines (APRIL and BAFF), their receptors (sTNF-R1 and sTNF-R2) and osteopontin as well as the inhibition of the complex gp130/sIL-6Rβ and metalloprotease ММР-1 were shown. Th1 immune response regulated by IL10 resulted in the recovery of the patients without complications.

Soluble RANKL contributes to osteoclast formation in adult mice but not ovariectomy-induced bone loss

Receptor activator of NFkB ligand (RANKL) is a TNF-family cytokine required for osteoclast formation, as well as immune cell and mammary gland development. It is produced as a membrane-bound protein that can be shed to form a soluble protein. We created mice harboring a sheddase-resistant form of RANKL, in which soluble RANKL is undetectable in the circulation. Lack of soluble RANKL does not affect bone mass or structure in growing mice but reduces osteoclast number and increases cancellous bone mass in adult mice. Nonetheless, the bone loss caused by estrogen deficiency is unaffected by the lack of soluble RANKL. Lymphocyte number, lymph node development, and mammary gland development are also unaffected by the absence of soluble RANKL. These results demonstrate that the membrane-bound form of RANKL is sufficient for most functions of this protein but that the soluble form does contribute to physiological bone remodeling in adult mice.

Macrophage migration inhibitory factor is required for NLRP3 inflammasome activation

Macrophage migration inhibitory factor (MIF) exerts multiple effects on immune cells, as well as having functions outside the immune system. MIF can promote inflammation through the induction of other cytokines, including TNF, IL-6, and IL-1 family cytokines. Here, we show that inhibition of MIF regulates the release of IL-1α, IL-1β, and IL-18, not by affecting transcription or translation of these cytokines, but via activation of the NLRP3 inflammasome. MIF is required for the interaction between NLRP3 and the intermediate filament protein vimentin, which is critical for NLRP3 activation. Further, we demonstrate that MIF interacts with NLRP3, indicating a role for MIF in inflammasome activation independent of its role as a cytokine. These data advance our understanding of how MIF regulates inflammation and identify it as a factor critical for NLRP3 inflammasome activation.

Cytomegalovirus blockade of TRAIL-mediated type 1 ILC defenses regulates viral persistence

Abstract Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) regulates cellular apoptosis and inflammation, and growing evidence indicates a key role in antiviral defense. Cytomegalovirus (CMV, a b-herpesvirus) establishes a lifelong persistent/latent infection that depends upon viral subversion of host innate and adaptive defenses. We have shown that both human and mouse CMV inhibit signaling by the TRAIL death receptors (DR) through the use of specific viral proteins. MCMV m166 blocks TRAIL-DR expression and promotes early viral replication by evading TRAIL-mediated natural killer (NK) cell defenses. After one week of systemic replication, MCMV spreads to the salivary glands (SG), the major mucosal organ responsible for horizontal transmission of the virus. We show that a MCMVΔm166 mutant replicates at lower absolute levels in the SG, but, surprisingly, persists for much longer times. Recent evidence suggests a role of type 1 Innate Lymphoid cells (ILC1) in control/clearance of MCMV. We investigated the role of the 2 main ILC1 populations (conventional and tissue resident (tr) NK cells) in controlling MCMV SG persistence. Depleting ILC1 prior to infection, or infection of TRAIL−/− mice, restored MCMVΔm166 SG replication to normal levels. In addition, SG trNK cells display a unique phenotype and express high levels of TRAIL during MCMV infection when compared to their conventional counterparts. Together our data reveal a key role for m166 inhibition of TRAIL-DR expression in promoting viral persistence by blocking ILC1 effector functions mediated by TRAIL, highlighting this TNF-family cytokine as an important factor regulating innate defenses in mucosal tissues.

Cleavage of TL1A Differentially Regulates Its Effects on Innate and Adaptive Immune Cells.

TNF superfamily cytokines play major roles in the regulation of adaptive and innate immunity. The TNF superfamily cytokine TL1A (TNFSF15), through its cognate receptor DR3 (TNFRSF25), promotes T cell immunity to pathogens and directly costimulates group 2 and 3 innate lymphoid cells. Polymorphisms in the TNFSF15 gene are associated with the risk for various human diseases, including inflammatory bowel disease. Like other cytokines in the TNF superfamily, TL1A is synthesized as a type II transmembrane protein and cleaved from the plasma membrane by metalloproteinases. Membrane cleavage has been shown to alter or abrogate certain activities of other TNF family cytokines; however, the functional capabilities of membrane-bound and soluble forms TL1A are not known. Constitutive expression of TL1A in transgenic mice results in expansion of activated T cells and promotes intestinal hyperplasia and inflammation through stimulation of group 2 innate lymphoid cells. Through the generation of membrane-restricted TL1A-transgenic mice, we demonstrate that membrane TL1A promotes expression of inflammatory cytokines in the lung, dependent upon DR3 expression on T cells. Soluble TL1A alone was unable to produce this phenotype but was still able to induce intestinal type 2 inflammation independently of T cells. These data suggest differential roles for membrane and soluble TL1A on adaptive and innate immune cells and have implications for the consequences of blocking these two forms of TL1A.

Influence ofTNF-α promoter variability on stage and grade in individuals with colorectal cancer.

Carcinogenesis is amultistep process in which inflammation plays animportant role. Tumour necrosis factor a (TNF-α) is acytokine that plays amajor role in inflammation. Activity oftheTNF cytokine family could influence progression ofcolorectal cancer (CRC). Theaim ofthestudy was to establish anassociation between TNF-α promoter variability and stage/grade in individuals with sporadic CRC. The study included 152 CRC patients and 107 healthy volunteers. Four single nucleotide polymorphisms (rs361525, rs1800629, rs1799724, and rs1799964) located at thepromoter ofTNFA gene were genotyped using commercially available TaqMan allelic discrimination assays by real-time PCR. CRC stage was described on thebasis ofpreoperative imaging studies and postoperative histopathological report. Thegrade was described on thebasis postoperative pathological examination ofthespecimen. In thecase ofrs361525, there was astatistically significant association with M-score (p = 0.0209). Rs361525 has significant association with tumour grade (p = 0.0260). We failed to demonstrate significant association between theother 3 SNPs and cancer grade. Rs361525 potentially could be under consideration when thesurvival rate and prognosis is assessed.

Regulation of RANKL-induced osteoclastogenesis by RING finger protein RNF114.

Normal bone remodeling is a continuous process orchestrated by bone-resorbing osteoclasts and bone-forming osteoblasts, which an imbalance in bone remodeling results in metabolic bone diseases. RANKL, a member of the TNF cytokine family, functions as a key stimulator for osteoclast differentiation and maturation. Here, we report that RNF114, previously identified as a psoriasis susceptibility gene, plays a regulatory role in the RANKL/RANK/TRAF6 signaling pathway that mediates osteoclastogenesis. Our results demonstrated that RNF114 expression was significantly down-regulated in mouse osteoclast precursor cells undergoing RANKL-induced osteoclast differentiation. RNF114 knockout did not affect development or viability of the subpopulation of bone marrow macrophages capable of differentiating into osteoclasts in culture. However, in the presence of RANKL, RNF114 knockout bone marrow macrophages exhibited enhanced cell proliferation and augmented osteoclast differentiation, as shown by an increased expression of mature osteoclast markers, increased osteoclastic TRAP activity and bone resorption. Conversely, ectopic expression of RNF114 inhibited CTSK expression, TRAP activity, and bone resorption in RANKL-treated pre-osteoclasts. RNF114 also suppressed RANKL-activated NFATc1 expression and NFAT-regulated promoter activity. RNF114 suppressed TRAF6-, but not TAK1/TAB2-mediated NF-κB activation downstream of RANKL/RANK. In particular, TRAF6 protein levels were down-regulated by RNF114, possibly via K48-mediated proteasome-dependent degradation. These data suggested that RNF114's inhibitory effect on RANKL-stimulated osteoclastogenesis was mediated by blocking RANK/TRAF6/NF-κB signal transduction. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:159-166, 2018.