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TNF signaling does not result in cell death unless multiple inhibitory signals are overcome, which can be accomplished by simultaneous signaling through IFNγ. In this issue, Deng and colleagues (http://doi.org/10.1083/jcb.202305026) dissect the mechanisms by which IFNγ signaling combines with TNF to mediate cell death through caspase-8, discussed by James E. Vince., (© 2024 Vince.)

Complement-C1q/tumor necrosis factor-related protein 3 (CTRP3) is an adipokine that primarily identified in 2003 and is an important member of CTRP family. CTRP3 is expressed in various tissues and cell types, and is highly conserved among different species. Multiple novel functions of CTRP3 have been reported recently as the further research on this protein develops. CTRP3 not only affects the proliferation of chondrocytes and the pathogenesis of osteoarthritis, but also regulates multiple physiological and pathological processes including the secretion of testosterone and adipokines, glucose and lipid metabolism, mitochondrial biogenesis, inflammatory response, cell apoptosis, angiogenesis, vascular calcification and ventricular remodeling. The present review mainly focuses on the research progresses on CTRP3, including its discovery, gene and protein structure, expression regulation, and biological functions. The progresses summarized may provide new clues for the further investigation of CTRP3.

The identification of the cellular origins of myofibroblasts has led to the discovery of novel pathways that potentially drive myofibroblast perpetuation in disease. Here, we further investigated the role of innate immune signaling pathways in this process. In mice, renal injury-induced activation of pericytes, which are myofibroblast precursors attached to endothelial cells, led to upregulated expression of TNF receptor superfamily member 12a, also known as fibroblast growth factor-inducible 14 (Fn14), by these cells. In live rat kidney slices, administration of the Fn14 ligand, TNF-related weak inducer of apoptosis (TWEAK), promoted pericyte-dependent vasoconstriction followed by pericyte detachment from capillaries. In vitro, administration of TWEAK activated and differentiated pericytes into cytokine-producing myofibroblasts, and further activated established myofibroblasts in a manner requiring canonical and noncanonical NF-κB signaling pathways. Deficiency of Fn14 protected mouse kidneys from fibrogenesis, inflammation, and associated vascular instability after in vivo injury, and was associated with loss of NF-κB signaling. In a genetic model of spontaneous CKD, therapeutic delivery of anti-TWEAK blocking antibodies attenuated disease progression, preserved organ function, and increased survival. These results identify the TWEAK-Fn14 signaling pathway as an important factor in myofibroblast perpetuation, fibrogenesis, and chronic disease progression., (Copyright © 2016 by the American Society of Nephrology.)

Objectives: Oestrogens have been shown to play key roles in the pathogenesis of SLE. The aim of this study was to investigate the roles and mechanisms of 17β-estradiol (E2) in TNF-like weak inducer of apoptosis (TWEAK) expression in LN., Methods: Peripheral blood mononuclear cells (PBMCs) obtained from LN patients were used for in vitro experiments, while female MRL/lpr and MRL/MpJ mice were used for in vivo studies. E2, ICI 182 780 [estrogen receptor (ER)-selective antagonist], methyl-piperidino-pyrazole (MPP, ERα-selective modulator), lentivirus (LV)-TWEAK-short hairpin RNA (shRNA) and LV-control-shRNA treatments were used in this study., Results: TWEAK mRNA expression in PBMCs was significantly increased following E2 treatment and downregulated after incubation with ICI 182 780 or MPP. Compared with sham-operated MRL/lpr mice, ovariectomized mice, treated with dimethyl sulphoxide vehicle alone, showed lower expression levels of renal TWEAK mRNA and protein. The expression of both mRNA and protein in ovariectomized mice was upregulated after E2 treatment and downregulated after ICI 182 780 or MPP co-treatment. Severe renal damage was observed in E2-treated ovariectomized mice, as were higher serum levels of IL-6, compared with dimethyl sulphoxide vehicle-treated ovariectomized mice. Co-treatment with LV-TWEAK-shRNA reversed these changes, and LV-control-shRNA treatment had no effect on them., Conclusion: Our results demonstrated that E2 plays an important role in the upregulation of TWEAK expression in LN, most likely through an ERα-dependent pathway, causing kidney damage. This provides a novel insight into the mechanisms of the E2-TWEAK signalling pathway in LN., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Our aim in this study was to determine soluble tumor necrosis factor (TNF)-like weak inducer of apoptosis (sTWEAK) and interleukin-17A (IL-17A) levels in celiac disease, and their association with the gluten diet and autoantibodies. Eighty patients with celiac diagnosis and 80 healthy control individuals with similar age, gender and body mass index to the patient group were included in the study. Serum sTWEAK and IL-17A levels were measured by the serum enzyme-linked immunosorbent assay kit. The median IL-17A (117.5 pg/mL vs. 56.7 pg/mL; P = 0.001) level in celiac patients was higher than in the control group, while the median sTWEAK (543 pg/mL vs. 643 pg/mL; P = 0.016) level in patients was determined to be lower. In the patient group, patients who complied with the gluten diet had a lower level of median IL-17A (98.1 pg/mL vs. 197.5 pg/mL; P = 0.034) and a higher level of sTWEAK (606 pg/mL vs. 522.8 pg/mL; P = 0.031) than those who did not adhere. Furthermore, the IL-17A level was higher and the sTWEAK level was lower in celiac patients with positive antibody than those with negative antibody. A positive correlation was determined among anti-gliadin antibody IgA, anti-gliadin antibody IgG, anti-tissue transglutaminase IgG levels and the IL-17A level, and a negative correlation was determined with the sTWEAK level. In celiac disease, the sTWEAK and IL-17A levels differ between patients who cannot adapt to the gluten diet and who are autoantibody positive, and patients who adapt to the diet and are autoantibody negative. We believe that sTWEAK and IL-17A are associated with the inflammation in celiac pathogenesis.

Uterine natural killer (uNK) cells are the most abundant lymphocyte population in the feto-maternal interface during early gestation, and uNK cells play a significant role in the establishment and maintenance of pregnancy-related vascularization, as well as in tolerance to the fetus. Tumour necrosis factor-like weak inducer of apoptosis (TWEAK) and its receptor, fibroblast growth factor-inducible molecule (Fn14), are involved in preventing local cytotoxicity and counterbalancing the cytotoxic function of uNK cells. Here, we studied the regulation of TWEAK/Fn14-mediated innate immunity in the uterus using a lipopolysaccharide (LPS)-induced model of abortion in pregnant mice. Specifically, we detected the expression of TWEAK and Fn14 in the uterus and in uNK cells following LPS treatment. Our results revealed that TWEAK and Fn14 are expressed by uNK cells in pregnant mice; in particular, it appears that the cytokine TWEAK is primarily derived from uNK cells. Interestingly, the down-regulation of TWEAK in uNK cells and the up-regulation of the Fn14 receptor in the uterus in LPS-treated mice may contribute to the disruption of decidual homeostasis by altering uNK cell cytotoxicity - ultimately leading to fetal rejection. In conclusion, the present study strongly suggests that the TWEAK-Fn14 axis in uNK cells is involved in maintaining the tolerance necessary for successful pregnancy., (© 2016 John Wiley & Sons Ltd.)

The differentiation and proper function of thymic epithelial cells (TECs) depend on various tumor necrosis factor superfamily (TNFSF) signals that are needed to maintain the thymic stromal microenvironment. Nevertheless, the direct transcriptional effects of these signals on TECs remain unclear. To address this issue, we stimulated murine embryonic thymus tissue with selected TNFSF ligands and performed a gene expression profiling study. We show that Aire expression is a direct and specific effect of RANKL stimulation, whereas LTβ and TNFα are major inducers of chemokines in the thymic stroma and we propose differential NF-κB binding as one possible cause of these gene expression patterns. Our work provides further insight into the complex molecular pathways that shape the thymic microenvironment and maintain central tolerance., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Aim: The interaction between TNF-like weak inducer of apoptosis (TWEAK, Tnfsf12) and the receptor, fibroblast growth factor-inducible 14 (Fn14), regulates vascular damage through different mechanisms, including inflammation. Oxidative stress plays a major role in inflammation and the development of atherosclerosis, but the relationship between TWEAK and oxidative stress is, however, poorly understood., Methods and Results: In this study, we found that TWEAK and Fn14 are co-localized with the NADPH subunits, p22phox and Nox2, in human advanced atherosclerotic plaques. Using primary human macrophages and a murine macrophage cell line, we demonstrate that TWEAK promotes ROS production and enhances NADPH oxidase activity. Hence, we show a direct involvement of the TWEAK-Fn14 axis in oxidative stress, as genetic silencing of Fn14 or Nox2 abrogates the TWEAK-induced ROS production. Furthermore, our results point at Rac1 as an upstream mediator of TWEAK during oxidative stress. Finally, using an in vivo murine model we confirmed the major role of TWEAK in oxidative stress, as genetic silencing of Tnfsf12 in an ApoE(-/-) background reduces the number of DHE and 8-hydroxydeoxyguanosine-positive macrophages by 50%., Conclusions: Our results suggest that TWEAK regulates vascular damage by stimulating ROS production in an Nox2-dependent manner. These new insights into the TWEAK/Fn14 axis underline their potential use as therapeutic targets in atherosclerosis., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2015. For permissions please email: journals.permissions@oup.com.)

Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK, TNFSF12) and its sole receptor Fn14, belonging to the TNF ligand and receptor superfamilies respectively, are involved in cell survival and cytokine production. The role of TWEAK/Fn14 interactions in the pathogenesis of cutaneous lupus has not been explored. TWEAK treatment of murine PAM212 keratinocytes stimulated the secretion of RANTES via Fn14 and promoted apoptosis. Parthenolide, but not wortmanin or the MAPK inhibitor PD98059, significantly decreased production of RANTES, indicating that this effect of TWEAK is mediated via NF-κB signaling. UVB irradiation significantly upregulated the expression of Fn14 on keratinocytes in vitro and in vivo and increased RANTES production. MRL/lpr Fn14 knockout (KO) lupus mice were compared with MRL/lpr Fn14 wild-type (WT) mice to evaluate for any possible differences in the severity of cutaneous lesions and the presence of infiltrating immune cells. MRL/lpr Fn14 KO mice had markedly attenuated cutaneous disease as compared with their Fn14 WT littermates, as evidenced by the well-maintained architecture of the skin and significantly decreased skin infiltration of T cells and macrophages. Our data strongly implicate TWEAK/Fn14 signaling in the pathogenesis of the cutaneous manifestations in the MRL/lpr model of spontaneous lupus and suggest a possible target for therapeutic intervention.

Skeletal muscle is responsible for the majority of glucose disposal in body. Impairment in skeletal muscle glucose handling capacity leads to the state of insulin resistance. The TNF-like weak inducer of apoptosis (TWEAK) cytokine has now emerged as a major regulator of skeletal muscle mass and function. However, the role of TWEAK in skeletal muscle metabolic function remains less understood. Here, we demonstrate that with progressive age, skeletal muscle-specific TWEAK-transgenic (TWEAK-Tg) mice gain increased body weight (∼16%) and fat mass (∼64%) and show glucose intolerance and insulin insensitivity. TWEAK-Tg mice also exhibit adipocyte hypertrophy in the epididymal fat. Oxygen uptake, voluntary physical activity, and exercise capacity were significantly reduced in TWEAK-Tg mice compared with controls. Overexpression of TWEAK inhibited (∼31%) 5' AMP-activated protein kinase (AMPK) and reduced (∼31%) the levels of glucose transporter type 4 (GLUT4) without affecting the Akt pathway. TWEAK also inhibited insulin-stimulated glucose uptake (∼32%) and repressed the levels of GLUT4 (∼50%) in cultured myotubes from C57BL6 mice. TWEAK represses the levels of Krüppel-like factor 15; myocyte enhancer factor 2, and peroxisome proliferator-activated receptor-γ coactivator-1α, which are required for the activation of the GLUT4 locus. Collectively our study demonstrates that elevated levels of TWEAK in skeletal muscle cause metabolic abnormalities. Inhibition of TWEAK could be a potential approach to prevent weight gain and type 2 diabetes., (© FASEB.)

Objective: Brain damage and ischemia often trigger cortical spreading depression (CSD), which aggravates brain damage. The proinflammatory cytokine tumor necrosis factor (TNF) is significantly upregulated during brain damage, but it is unknown whether TNF influences spreading depression in cerebral cortex in vivo. This question is important because TNF not only furthers inflammatory reactions but might also be neuroprotective. Here we tested the hypothesis that TNF affects CSD, and we explored the direction in which CSD is modified by TNF., Methods: CSD, elicited by pressure microinjection of KCl, was recorded in anesthetized rats and mice. TNF was administered locally into a trough, providing local TNF treatment of a cortical area. For further analysis, antibodies to TNF receptor (TNFR) 1 or 2 were applied, or CSD was monitored in TNFR1 and TNFR2 knockout mice. γ-Aminobutyric acid (GABA)A receptors were blocked by bicuculline. Immunohistochemistry localized the cortical expression of TNFR1 and TNFR2., Results: Local application of TNF to the cortex reduced dose-dependently the amplitude of CSD. This effect was prevented by blockade or knockout of TNFR2 but not by blockade or knockout of TNFR1. TNFR2 was localized at cortical neurons including parvalbumin-positive inhibitory interneurons, and blockade of GABAA receptors by bicuculline prevented the reduction of CSD amplitudes by TNF., Interpretation: We identified a functional link between TNF and CSD. TNF activates TNFR2 in cortical inhibitory interneurons. The resulting release of GABA reduces CSD amplitudes. In this manner, TNF might be neuroprotective in pathological conditions., (© 2014 American Neurological Association.)

Peritoneal dialysis (PD) is complicated by peritonitis episodes that cause loss of mesothelium and eventually sclerosing peritonitis. An improved understanding of the molecular contributors to peritoneal injury and defense may increase the therapeutic armamentarium to optimize peritoneal defenses while minimizing peritoneal injury. There is no information on the expression and function of the cytokine TWEAK and its receptor Fn14 during peritoneal injury. Fn14 expression and soluble TWEAK levels were measured in human PD peritoneal effluent cells or fluids with or without peritonitis. Fn14 expression was also analyzed in peritoneal biopsies from PD patients. Actions of intraperitoneal TWEAK were studied in mice in vivo. sTWEAK levels were increased in peritoneal effluent in PD peritonitis. Effluent sTWEAK levels correlated with the number of peritoneal macrophages (r=0.491, p=0.002). Potential TWEAK targets that express the receptor Fn14 include mesothelial cells and macrophages, as demonstrated by flow cytometry of peritoneal effluents and by analysis of peritoneal biopsies. Peritoneal biopsy Fn14 correlated with mesothelial injury, fibrosis and inflammation, suggesting a potential deleterious effect of TWEAK/Fn14. In this regard, intraperitoneal TWEAK administration to mice promoted peritoneal inflammation characterized by increased peritoneal effluent MCP-1, Fn14 and Gr1+ macrophages, increased mesothelial Fn14, MCP-1 and CCL21 expression and submesothelial tissue macrophage recruitment. Taken together these data suggest that the TWEAK/Fn14 system may promote inflammation and tissue injury during peritonitis and PD.

Skeletal muscle wasting attributed to inactivity has significant adverse functional consequences. Accumulating evidence suggests that peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) and TNF-like weak inducer of apoptosis (TWEAK)-Fn14 system are key regulators of skeletal muscle mass in various catabolic states. While the activation of TWEAK-Fn14 signaling causes muscle wasting, PGC-1α preserves muscle mass in several conditions, including functional denervation and aging. However, it remains unknown whether there is any regulatory interaction between PGC-1α and TWEAK-Fn14 system during muscle atrophy. Here we demonstrate that TWEAK significantly reduces the levels of PGC-1α and mitochondrial content (∼50%) in skeletal muscle. Levels of PGC-1α are significantly increased in skeletal muscle of TWEAK-knockout (KO) and Fn14-KO mice compared to wild-type mice on denervation. Transgenic (Tg) overexpression of PGC-1α inhibited progressive muscle wasting in TWEAK-Tg mice. PGC-1α inhibited the TWEAK-induced activation of NF-κB (∼50%) and dramatically reduced (∼90%) the expression of atrogenes such as MAFbx and MuRF1. Intriguingly, muscle-specific overexpression of PGC-1α also prevented the inducible expression of Fn14 in denervated skeletal muscle. Collectively, our study demonstrates that TWEAK induces muscle atrophy through repressing the levels of PGC-1α. Overexpression of PGC-1α not only blocks the TWEAK-induced atrophy program but also diminishes the expression of Fn14 in denervated skeletal muscle.

Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) is a member of the TNF super family that controls many cellular activities including proliferation, migration, differentiation, apoptosis, and inflammation by binding to fibroblast growth factor-inducible 14 (Fn14), a highly inducible cell surface receptor. Recent studies have indicated that TWEAK-Fn14 axis signaling may contribute to chronic autoimmune diseases. TWEAK expression via microglia in cortical lesions, presence of TWEAK(+) macrophages in inflamed leptomeninges, and absence of TWEAK/Fn14 expression in healthy brain implicates importance of this pathway in pathogenesis of multiple sclerosis lesions. TWEAK-Fn14 axis blockade has also shown promise in various multiple sclerosis animal models. Stimulation of the TWEAK/Fn14 pathway can result in activation of both canonical and noncanonical NF-κB signaling and could also stimulate mitogen-activated protein kinase (MAPK) signaling pathways. Here, we have reviewed evidence of the possible role of TWEAK-Fn14 axis in pathophysiology of multiple sclerosis and experimental autoimmune encephalomyelitis (EAE) via neuroinflammation, tissue remodeling, blood-brain barrier (BBB) disruption, neurodegeneration, and astrogliosis.

Skeletal muscle (SM) regeneration after injury is impaired by excessive inflammation. Particularly, the inflammatory cytokine tumour necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) is a potent inducer of skeletal muscle wasting and fibrosis. In this study we investigated the role of Nrf2, a major regulator of oxidative stress defence, in SM ischemia/reperfusion (I/R) injury and TWEAK induced atrophy. We explored the time-dependent expression of TWEAK after I/R in SM of Nrf2-wildtype (WT) and knockout (KO) mice. Nrf2-KO mice expressed significant higher levels of TWEAK as compared to WT mice. Consequently, Nrf2-KO mice present an insufficient regeneration as compared to Nrf2-WT mice. Moreover, TWEAK stimulation activates Nrf2 in the mouse myoblast cell line C2C12. This Nrf2 activation inhibits TWEAK induced atrophy in C2C12 differentiated myotubes. In summary, we show that Nrf2 protects SM from TWEAK-induced cell death in vitro and that Nrf2-deficient mice therefore have poorer muscle regeneration.

Cell turnover is a fundamental feature in metazoans. Cells can die passively, as a consequence of severe damage to their structural integrity, or actively, owing to a more confined biological disruption such as DNA damage. Passive cell death is uncontrolled and often harmful to the organism. In contrast, active cell death is tightly regulated and serves to support the organism's life. Apoptosis-the primary form of regulated cell death-is relatively well defined. Necroptosis-an alternative, distinct kind of regulated cell death discovered more recently-is less well understood. Apoptosis and necroptosis can be triggered either from within the cell or by extracellular stimuli. Certain signaling components, including several death ligands and receptors, can regulate both processes. Whereas apoptosis is triggered and executed via intracellular proteases called caspases, necroptosis is suppressed by caspase activity. Here we highlight current understanding of the key signaling mechanisms that control regulated cell death.

Fibrosis is a major hallmark of progressive kidney disease. The cellular mechanisms that lead to kidney tissue fibrosis are complex and include, for example, increased inflammation, increased oxidative stress, and proximal tubule cell death in the form of apoptosis or senescence. Recent studies have identified TWEAK, a tumor necrosis factor-like weak inducer of apoptosis, as a novel cytokine that mediates kidney inflammation in models of renal fibrosis. Inhibition of apoptosis via TWEAK inhibition has been shown to reduce kidney fibrosis. Recent studies using lineage tracing suggest that interstitial pericytes/perivascular fibroblasts differentiate into myofibroblasts and undergo proliferative expansion during fibrosis. Furthermore, increased expression of nuclear peroxisome proliferator-activated receptor-α in proximal tubules can directly reduce increased expression of transforming growth factor-β1 and interstitial inflammation in models of renal fibrosis, which suggests preservation of proximal tubule cell metabolism and integrity represents an important new therapeutic target. In this review, the current evidence and potential molecular mechanisms involved in the development of kidney fibrosis are discussed., (2014 S. Karger AG, Basel.)

The TNF superfamily member TWEAK has emerged as a pleiotropic cytokine that regulates many cellular functions that include immune/inflammatory activity, angiogenesis, cell proliferation, and fate. TWEAK through its inducible receptor, FGF-inducible molecule 14 (Fn14), can induce both beneficial and deleterious activity that has a profound effect on cell survival. Thus it is highly likely that TWEAK and Fn14 expressed by cells of the neurovascular unit help regulate and maintain vascular and tissue homeostasis. In this chapter we discuss the expression of TWEAK and Fn14 signaling in the cerebral microvascular pericyte. Pericytes are a highly enigmatic population of microvascular cells that are important in regulatory pathways that modulate physiological angiogenesis in response to chronic mild hypoxic stress. A brief introduction will identify the microvascular pericyte. A more detailed discussion of pericyte TWEAK signaling during adaptive angioplasticity will follow.

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) regulates apoptosis, proliferation and inflammation in renal epithelial cells and plays a role in acute kidney injury. However, there is little information on the chronic effects of TWEAK. We hypothesized that TWEAK may influence renal fibrosis and regulate kidney fibroblast biology, in part, through Ras pathway. We studied a chronic model of experimental unilateral ureteral obstruction in wild type and TWEAK deficient mice, and a murine model of systemic TWEAK overexpression. TWEAK actions were also explored in cultured renal and embryonic fibroblasts. TWEAK and TWEAK receptor expression was increased in the obstructed kidneys. The absence of TWEAK decreased early kidney tubular damage, inflammatory infiltrates and myofibroblast number. TWEAK deficient mice had decreased renal fibrosis 21days after obstruction, as assessed by extracellular matrix staining. In mice without prior underlying kidney disease, systemic overexpression of TWEAK induced kidney inflammation and fibrosis. In cultured fibroblasts, TWEAK induced proliferation through activation of the Ras/ERK pathway. TWEAK also activated nuclear factor κB (NFκB)-dependent inflammatory chemokine production in murine renal fibroblasts. In conclusion, lack of TWEAK reduces renal fibrosis in a model of persistent kidney insult and overexpression of TWEAK led to renal fibrosis. TWEAK actions on renal fibroblasts may contribute to the in vivo observations, as TWEAK promotes inflammatory activity and proliferation in fibroblast cultures., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Tumour necrosis factor (TNF) was originally described as a circulating factor that can induce haemorrhagic necrosis of tumours. It is now clear that TNF has many different functions in cancer biology. In addition to causing the death of cancer cells, TNF can activate cancer cell survival and proliferation pathways, trigger inflammatory cell infiltration of tumours and promote angiogenesis and tumour cell migration and invasion. These effects can be explained by the diverse cellular responses TNF can initiate through distinct signal transduction pathways, opening the way for more selective targeting of TNF signalling in cancer therapy., (Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Crystal-cell interaction has been reported as one of the most crucial steps in urinary stone formation. Hyperoxaluria-induced apoptotic changes in renal tubular epithelial cells is the end-stage of this interaction. We aimed to evaluate the possible pathways responsible in the induction of apoptosis within the involved cells by assessing the receptoral expression of three different pathways. 16 male Spraque-Dowley rats were divided into two groups: Group 1 (n:8) received only distilled water; Group 2 (n:8) received 0.75 % ethylene glycol (EG) in their daily water to induce hyperoxaluria for 2 weeks. After 24 h urine collection, all animals were euthenized and right kidneys were removed and fixed for immunohistochemical evaluation. Oxalate and creatinine levels (in 24 h-urine) and FAS, tumor necrosis factor (TNF), TNF-related apoptosis-inducing ligand (TRAIL) and TRAIL receptor-2 expressions (in tissue) have been assessed. In addition to TNF (p = 0.0007) expression; both FAS (p = 0.0129 ) and FASL (p = 0.032) expressions significantly increased in animals treated with EG. The expressions of TRAIL (p = 0.49) and TRAIL-R2 (p = 0.34) receptors did not change statistically after hyperoxaluria induction. Although a positive correlation with cytokine expression density and 24 h-urinary oxalate expression (mg oxalate/mg creatinine) has been assessed with TNF (p = 0.04, r = 0.82), FAS (p = 0.05, r = 0.80), FAS-L (p = 0.04, r = 0.82); no correlation could be demonstrated between TRAIL and TRAIL R2 expressions. Our results indicate that apoptosis induced by oxalate is possibly mediated via TNF and FAS pathways. However, TRAIL and TRAIL-R2 seemed to have no function in the cascade. Correlation with urinary oxalate levels did further strengthen the findings.

Background: Fn14 is a therapeutic target in various diseases., Results: Anti-Fn14 antibodies activate the alternative NFκB pathway but not other Fn14-related activities induced by soluble or membrane-bound TWEAK. FcγR-bound anti-Fn14 antibodies, however, activate the full spectrum of Fn14-associated activities., Conclusion: Anti-Fn14 antibodies elicit agonistic activities differing from those of the natural Fn14 ligand TWEAK., Significance: These findings influence the rationale of designing Fn14-targeted therapies. The Fn14-specific monoclonal antibodies PDL192 and P4A8, which are under consideration in clinical trials, showed no agonistic activity with respect to IL8 production and cell death induction. However, oligomerization with protein G or binding to Fcγ receptors converted both anti-Fn14 antibodies into potent agonists. TNF-like weak inducer of apoptosis (TWEAK), the ligand of Fn14, occurs naturally in two forms with partly different signaling capabilities, as a membrane-bound ligand and as a soluble trimeric molecule. Although membrane TWEAK strongly triggers all Fn14-associated pathways, soluble TWEAK predominately triggers the alternative nuclear factor κB (NFκB) pathway and enhances TNF-induced cell death but has only a poor effect on the classical NFκB pathway and chemokine production. Thus, the oligomerized and FcγR-bound anti-Fn14 mAbs mimicked the activity of membrane TWEAK. Notably, both anti-Fn14 antibodies significantly triggered p100 processing, the hallmark of the alternative NFκB pathway, and therefore resembled soluble TWEAK. In contrast to the latter, however, the anti-Fn14s showed no effect on TNF receptor 1-induced cell death and P4A8 even blocked the corresponding TWEAK response. Thus, we showed that Fn14 antibodies display an alternative NFκB pathway-specific agonistic activity but fail to phenocopy other activities of soluble TWEAK, whereas oligomerized or FcγR-bound Fn14 antibodies fully mimic the activity of membrane TWEAK. In view of the trivalent nature of the TWEAK-Fn14 interaction, this suggests that the alternative NFκB pathway is uniquely responsive already to Fn14 dimerization enabling antibodies to elicit an unnatural response pattern distinct from that of the naturally occurring Fn14 ligands.

This paper will explore the pathological process involved in dental resorption as well as its classifications and aetiology. The second subsequent paper will look at its diagnosis and management.

Background: The TNF ligand family member TWEAK exists as membrane and soluble forms and is involved in the regulation of various human inflammatory pathologies, through binding to its main receptor, Fn14. We have shown that the soluble form of TWEAK has a pro-neuroinflammatory effect in an animal model of multiple sclerosis and we further demonstrated that blocking TWEAK activity during the recruitment phase of immune cells across the blood brain barrier (BBB) was protective in this model. It is now well established that endothelial cells in the periphery and astrocytes in the central nervous system (CNS) are targets of TWEAK. Moreover, it has been shown by others that, when injected into mice brains, TWEAK disrupts the architecture of the BBB and induces expression of matrix metalloproteinase-9 (MMP-9) in the brain. Nevertheless, the mechanisms involved in such conditions are complex and remain to be explored, especially because there is a lack of data concerning the TWEAK/Fn14 pathway in microvascular cerebral endothelial cells., Methods: In this study, we used human cerebral microvascular endothelial cell (HCMEC) cultures as an in vitro model of the BBB to study the effects of soluble TWEAK on the properties and the integrity of the BBB model., Results: We showed that soluble TWEAK induces an inflammatory profile on HCMECs, especially by promoting secretion of cytokines, by modulating production and activation of MMP-9, and by expression of cell adhesion molecules. We also demonstrated that these effects of TWEAK are associated with increased permeability of the HCMEC monolayer in the in vitro BBB model., Conclusions: Taken together, the data suggest a role for soluble TWEAK in BBB inflammation and in the promotion of BBB interactions with immune cells. These results support the contention that the TWEAK/Fn14 pathway could contribute at least to the endothelial steps of neuroinflammation.

Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) is a member of the TNF superfamily of structurally related cytokines and is known to induce proliferation, migration, differentiation, apoptotic cell death, inflammation, and angiogenesis. These physiological processes are induced by the binding of TWEAK to fibroblast growth factor-inducible 14 (Fn14), a highly inducible cell-surface receptor that is linked to several intracellular signaling pathways, including the nuclear factor-κB (NF-κB) pathway. This review discusses the role of the TWEAK-Fn14 axis in several rheumatic diseases and the potential therapeutic benefits of modulation of the TWEAK-Fn14 pathway.

The fusion of mononucleated muscle progenitor cells (myoblasts) into multinucleated muscle fibers is a critical aspect of muscle development and regeneration. We identified the noncanonical nuclear factor κB (NF-κB) pathway as a signaling axis that drives the recruitment of myoblasts into new muscle fibers. Loss of cellular inhibitor of apoptosis 1 (cIAP1) protein led to constitutive activation of the noncanonical NF-κB pathway and an increase in the number of nuclei per myotube. Knockdown of essential mediators of NF-κB signaling, such as p100, RelB, inhibitor of κB kinase α, and NF-κB-inducing kinase, attenuated myoblast fusion in wild-type myoblasts. In contrast, the extent of myoblast fusion was increased when the activity of the noncanonical NF-κB pathway was enhanced by increasing the abundance of p52 and RelB or decreasing the abundance of tumor necrosis factor (TNF) receptor-associated factor 3, an inhibitor of this pathway. Low concentrations of the cytokine TNF-like weak inducer of apoptosis (TWEAK), which preferentially activates the noncanonical NF-κB pathway, also increased myoblast fusion, without causing atrophy or impairing myogenesis. These results identify roles for TWEAK, cIAP1, and noncanonical NF-κB signaling in the regulation of myoblast fusion and highlight a role for cytokine signaling during adult skeletal myogenesis.

Background: The interaction of mast cells (MCs) with regulatory T cells through the OX40 ligand (OX40L):OX40 axis downregulates FcεRI-dependent immediate hypersensitivity responses both in vitro and in vivo. Little is known on OX40L-mediated intracellular signaling or on the mechanism by which OX40L engagement suppresses MC degranulation., Objective: We explored the role of OX40L engagement on IgE/antigen-triggered MCs both in vitro and in vivo., Methods: The soluble form of OX40 molecule was used to selectively trigger OX40L on MCs in vitro and was used to dissect OX40L contribution in an in vivo model of systemic anaphylaxis., Results: OX40L:OX40 interaction led to the recruitment of C-terminal src kinase into lipid rafts, causing a preferential suppression of Fyn kinase activity and subsequent reduction in the phosphorylation of Gab2, the phosphatidylinositol 3-OH kinase regulatory subunit p85, and Akt, without affecting the Lyn pathway. Dampening of Fyn kinase activity also inhibited RhoA activation and microtubule nucleation, key regulators of MC degranulation. The in vivo administration of a blocking antibody to OX40L in wild-type mice caused enhanced immediate hypersensitivity, whereas the administration of soluble OX40 to regulatory T-cell-depleted or OX40-deficient mice reduced MC degranulation., Conclusions: The engagement of OX40L selectively suppresses Fyn-initiated signals required for MC degranulation and serves to limit immediate hypersensitivity. Our data suggest that soluble OX40 can restore the aberrant or absent regulatory T-cell activity, revealing a previously unappreciated homeostatic role for OX40L in setting the basal threshold of MC response., (Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.)

The TWEAK/Fn14 pathway is a ligand/receptor pair of the TNFSF that has emerged as a prominent player in normal and pathological tissue remodeling. TWEAK/Fn14 pathway activation drives many processes relevant to autoimmune and inflammatory diseases. IBDs, including CD and UC, are chronic, relapsing inflammatory diseases of the GI tract. These diseases differ in their clinical, macroscopic, and histopathological presentation; however, pathological processes that prominently contribute, more or less in each case, include breakdown of the mucosal epithelial barrier, chronic inflammation, and tissue remodeling with fibrosis. TWEAK may promote the pathogenesis of IBD by signaling through Fn14, which can be up-regulated on IECs, thereby contributing to breakdown of the mucosal barrier; the induction of IEC-derived mediators that promote chronic inflammation and shape gut immunity against commensal flora; and delayed healing and fibrosis. TWEAK may also exert its action on endothelial and stromal cell types, including smooth muscle cells and fibroblasts, to promote chronic inflammation, dysregulated tissue repair, and fibrosis. Here, we review the data supporting an emerging role of the TWEAK/Fn14 pathway in autoimmune and inflammatory diseases, with a particular focus on IBD, and discuss how it interplays with other prominent pathways, including IL-13, TNF-α, and TGF-β, to aggravate and perpetuate the pathological processes underlying IBD.

Thrombocytopenia inhibits tumor growth and especially metastasis in mice, whereas additional depletion of NK cells reverts this antimetastatic phenotype. It has therefore been speculated that platelets may protect hematogenously disseminating tumor cells from NK-dependent antitumor immunity. Tumor cells do not travel through the blood alone, but are rapidly coated by platelets, and this phenomenon has been proposed to shield disseminating tumor cells from NK-mediated lysis. However, the underlying mechanisms remain largely unclear. In this study, we show that megakaryocytes acquire expression of the TNF family member glucocorticoid-induced TNF-related ligand (GITRL) during differentiation, resulting in GITRL expression by platelets. Upon platelet activation, GITRL is upregulated on the platelet surface in parallel with the α-granular activation marker P-selectin. GITRL is also rapidly mobilized to the platelet surface following interaction with tumor cells, which results in platelet coating. Whereas GITRL, in the fashion of several other TNF family members, is capable of transducing reverse signals, no influence on platelet activation and function was observed upon GITRL triggering. However, platelet coating of tumor cells inhibited NK cell cytotoxicity and IFN-γ production that could partially be restored by blocking GITR on NK cells, thus indicating that platelet-derived GITRL mediates NK-inhibitory forward signaling via GITR. These data identify conferment of GITRL pseudoexpression to tumor cells by platelets as a mechanism by which platelets may alter tumor cell immunogenicity. Our data thus provide further evidence for the involvement of platelets in facilitating evasion of tumor cells from NK cell immune surveillance.

The glucocorticoid-induced TNFR-related (GITR) protein is a coactivating receptor that is constitutively expressed on Treg cells and induced on activated T cells. To better under-stand the role of long-term GITR signaling, we generated a mouse that constitutively expresses GITR ligand (GITRL) on APCs that mimics the physiological distribution of GITRL in vivo. Despite a five-fold expansion of the Treg-cell pool, there is increased activation and depletion of naive T cells in the transgenic (Tg) mice, suggesting that the increased number of Treg cells cannot fully suppress T-cell activation. Interestingly, GITRL Tg mice have multiorgan lymphocytic infiltrates yet display no overt autoimmunity, indicating the existence of a compensatory immunoregulatory mechanism(s). In the spleens and tissue infiltrates ofGITRL Tg mice, we found increased numbers of Foxp3(-) IL-10-producing type 1 regulatory T (Tr-1)-like cells that suppress naïve T-cell proliferation in an IL-10-dependent fashion. Increased IL-27 production from Tg APCs and activation of c-Maf in the Tr1-like cells suggest a possible mechanism for their induction. Our results demonstrate that enhanced GITR/GITRL interactions have a pleiotropic role on the regulation of T-cell responses, which includes promoting the differentiation of Tr-1-like cells, which contribute to the maintenance of peripheral T-cell tolerance., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) and its receptor Fn14 exert pleiotropic effects, including regulation of myogenesis. Sporadic inclusion-body myositis (IBM) is the most common muscle disease of the elderly population and leads to severe disability. IBM mesoangioblasts, different from mesoangioblasts in other inflammatory myopathies, display a myogenic differentiation defect. The objective of the present study was to investigate TWEAK-Fn14 expression in IBM and other inflammatory myopathies and explore whether TWEAK modulation affects myogenesis in IBM mesoangioblasts. TWEAK, Fn14, and NF-κB expression was assessed by immunohistochemistry and Western blot in cell samples from both muscle biopsies and primary cultures. Mesoangioblasts isolated from samples of IBM, dermatomyositis, polymyositis, and control muscles were treated with recombinant human TWEAK, Fn14-Fc chimera, and anti-TWEAK antibody. TWEAK-RNA interference was performed in IBM and dermatomyositis mesoangioblasts. TWEAK levels in culture media were determined by enzyme-linked immunosorbent assay. In IBM muscle, we found increased TWEAK-Fn14 expression. Increased levels of TWEAK were found in differentiation medium from IBM mesoangioblasts. Moreover, TWEAK inhibited myogenic differentiation of mesoangioblasts. Consistent with this evidence, TWEAK inhibition by Fn14-Fc chimera or short interfering RNA induced myogenic differentiation of IBM mesoangioblasts. We provide evidence that TWEAK is a negative regulator of human mesoangioblast differentiation. Dysregulation of the TWEAK-Fn14 axis in IBM muscle may induce progressive muscle atrophy and reduce activation and differentiation of muscle precursor cells., (Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

B cells are important for the regulation of autoimmune responses. In experimental autoimmune encephalomyelitis (EAE), B cells are required for spontaneous recovery in acute models. Production of IL-10 by regulatory B cells has been shown to modulate the severity EAE and other autoimmune diseases. Previously, we suggested that B cells regulated the number of CD4(+)Foxp3(+) T regulatory cells (Treg) in the CNS during EAE. Because Treg suppress autoimmune responses, we asked whether B cells control autoimmunity by maintenance of Treg numbers. B cell deficiency achieved either genetically (μMT) or by depletion with anti-CD20 resulted in a significant reduction in the number of peripheral but not thymic Treg. Adoptive transfer of WT B cells into μMT mice restored both Treg numbers and recovery from EAE. When we investigated the mechanism whereby B cells induce the proliferation of Treg and EAE recovery, we found that glucocorticoid-induced TNF ligand, but not IL-10, expression by B cells was required. Of clinical significance is the finding that anti-CD20 depletion of B cells accelerated spontaneous EAE and colitis. Our results demonstrate that B cells play a major role in immune tolerance required for the prevention of autoimmunity by maintenance of Treg via their expression of glucocorticoid-induced TNFR ligand.

Proinflammatory cytokine TWEAK has now emerged as a key mediator of skeletal muscle-wasting in many catabolic conditions. However, the mechanisms by which TWEAK induces muscle proteolysis remain poorly understood. Here, we have investigated the role of ubiquitin-proteasome system, autophagy, and caspases in TWEAK-induced muscle wasting. Addition of TWEAK to C2C12 myotubes stimulated the ubiquitination of myosin heavy chain (MyHC) and augmented the expression of E3 ubiquitin ligase MuRF1. Pretreatment of myotubes with proteasome inhibitors MG132 or lactacystin or knockdown of MuRF1 by RNAi blocked the TWEAK-induced degradation of MyHC and myotube atrophy. TWEAK increased the expression of several autophagy-related molecules. Moreover, the inhibitors of autophagy improved the levels of MyHC in TWEAK-treated myotubes. TWEAK also increased activity of caspases in C2C12 myotubes. Pan-caspase or caspase 3 inhibitory peptide inhibited the TWEAK-induced loss of MyHC and myotube diameter. Our study demonstrates that nuclear factor-kappa B (NF-κB) transcription factor is essential for TWEAK-induced expression of MuRF1 and Beclin1. Furthermore, our results suggest that caspases contribute, at least in part, to the activation of NF-κB in response to TWEAK treatment. Collectively, the present study provides novel insight into the mechanisms of action of TWEAK in skeletal muscle., (Copyright © 2011 Wiley Periodicals, Inc.)

The fibroblast growth factor-inducible 14 (Fn14) gene encodes a type I transmembrane protein that belongs to the tumor necrosis factor receptor superfamily and regulates multiple cellular processes in diverse physiological and pathological conditions, including cancer. Here, we describe an important role for Fn14 in regulating the growth of gastric cancer cells. Previous gene expression data analysis demonstrated that Fn14 was up-regulated in various tumor tissues, including gastric cancer. Using qRT-PCR, we showed that Fn14 was overexpressed in gastric tumor tissues compared to normal tissues. Furthermore, Fn14 expression levels were inversely correlated with gastric cancer patient survival. Using ectopic overexpression and shRNA-mediated knockdown of Fn14, we demonstrated that the expression level of Fn14 affected cell growth in gastric cancer. The effect of Fn14 on cell growth was mediated by the NF-κB activity and eventually by the transcriptional regulation of the anti-apoptotic Bcl-2 family gene (Bcl-xL). These results suggest that Fn14 may play an important role in gastric tumor growth by regulating NF-κB-mediated anti-apoptosis and that Fn14 may be a useful prognostic marker for gastric cancer., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Lymph node (LN) infiltration by neoplastic process involves important changes in lymph node immune microenvironment. In particular, regulatory T cells (Treg) seem to have a key role in altering the immunoediting function of the immune system which leads to the elusion of the tumor from immune surveillance. In this study, we evaluated the expression of T-cell markers in CD4+ and CD8+ subsets from tumor-positive and tumor-negative lymph nodes from the same, advanced stage breast cancer patient. The study was carried out on 3 patients and similar results were obtained. Flow cytometric analysis of CD8+ cells demonstrated a significant difference in the expression of CD25, CD45RA, CD45RO, and GITRL (Glucocorticoid-Induced TNF receptor-Related ligand). Flowcytometric analysis of CD4+ cells demonstrated a significant difference in the expression of GITR (Glucocorticoid-Induced TNF receptor-Related), CD25, FoxP3 (Forkhead box P3), CD28, and CD45RA. Multiple staining allowed the identification of two Treg subpopulations, CD4+ CD25 highGITR+ CD127-/low and CD4+ CD25 low GITR+ CD127+ cells, proving that both are increased in the positive nodes in comparison with the negative nodes from the same patient. We identified for the first time the CD4+ CD25 low GITR+ CD127+ Treg subpopulation in cancer, and the 2.6 fold increase in positive LN suggests that this Treg subpopulation could be a key player in metastasis. We also found GITRL expression in the CD8 lymphocytes, which may also contribute to the changes of metastatic lymph node microenvironment. These findings make both GITR and GITRL good possible co-candidates for future therapeutical intervention against metastasis and perhaps also as disease evolution biomarkers.

The idiopathic inflammatory myopathies (IM) represent a heterogeneous group of autoimmune diseases, of which dermatomyositis (DM), polymyositis (PM), and sporadic inclusion body myositis (IBM) are the most common. The crucial role played by tumor necrosis factor alpha (TNFα) in the IM has long been recognized. However, so far, 18 other members of the TNF superfamily have been characterized, and many of these have not yet received the attention they deserve. In this paper, we summarize current findings for all TNF cytokines in IM, pinpointing what we know already and where current knowledge fails. For each TNF family member, possibilities for treating inflammatory diseases in general and the IM in particular are explored.

Gluco-metabolic syndrome, oxidative stress and inflammation are common in chronic heart failure (CHF). Exercise training programmes are known to improve oxidative status, insulin sensitivity and endothelial function. In this study, the effects of walking on improving lipid and glucose metabolism in CHF patients, under statin treatment, were investigated. Fasting glucose, serum insulin, tumor necrosis factor-α (TNF-α), total antioxidant capacity (TAC), uric acid (UA), total cholesterol, triglycerides, high-density lipoproteins (HDL), direct low-density lipoproteins (LDL-dir), apolipoprotein-B, apolipoprotein-A1, and lipoprotein-a (Lp(a)) were monitored. Insulin resistance was depicted by fasting insulin resistance index (FIRI) (FIRI≥2.94±1.41). HDL significantly increased with walking and was positively correlated with the non-significant triglyceride decrease and significant Lp(a) decrease. Significant correlations were found in all CHF patients at baseline between FIRI and New York Heart Association (NYHA) functional class, ejection fraction, HDL, triglycerides and TNF-α. All non-diabetic CHF patients were characterized by insulin resistance. Serum insulin and fasting glucose significantly decreased with walking, while decrease in FIRI was positively associated with patients' adherence to the walking program. Elevated uric acid and TNF-α levels also significantly decreased. In conclusion, the present study demonstrated that moderate, unsupervised, everyday physical activity was able to ameliorate the lipid and glycemic profile of CHF patients, with simultaneous attenuation of inflammation and oxidative stress.

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) is a cytokine of the TNF superfamily that activates the Fn14 receptor. TWEAK may regulate cell proliferation, cell death, cell differentiation, and inflammation. TWEAK and Fn14 are constitutively present in the kidney. Sources of TWEAK and Fn14 include intrinsic renal cells and infiltrating leukocytes. Basal Fn14 expression is low, but Fn14 is greatly upregulated during kidney injury. TWEAK contributes to kidney inflammation promoting chemokine secretion by renal cells through canonical and non-canonical NFκB activation. TWEAK also promotes tubular cell proliferation. However, TWEAK induces mesangial and tubular cell apoptosis under proinflammatory conditions. These data indicate that TWEAK is a multifunctional cytokine in the kidney, the actions of which are modulated by the cell microenvironment. Confirmation of the role of TWEAK in kidney injury came from functional studies in experimental animal models. The TWEAK/Fn14 pathway contributed to cell death and interstitial inflammation during acute kidney injury, to glomerular injury in lupus nephritis, to hyperlipidemia-associated kidney injury, and to tubular cell hyperplasia following unilateral nephrectomy. Circulating soluble TWEAK (sTWEAK) levels are a potential biomarker of adverse outcomes in chronic kidney disease and urinary sTWEAK is a potential biomarker of lupus nephritis activity. The available evidence suggests that TWEAK may provide diagnostic information and be a therapeutic target in renal injury. Its role in human kidney disease should be further explored.

Background and Objectives: Chronic kidney disease (CKD) conveys high mortality rates. Soluble TNF-like weak inducer of apoptosis (sTWEAK) and long pentraxin 3 (PTX3) are predictors of mortality in dialysis patients and determinants of endothelial dysfunction. Now, we hypothesize that both sTWEAK and PTX3 act as biomarkers of cardiovascular outcomes in nondialysis CKD patients., Design, Setting, Participants, & Measurements: Cross-sectional analysis in which flow-mediated dilation (FMD) and intima-media thickness (IMT) were assessed in 257 nondialysis stage 1 to 5 CKD patients (mean age, 52 ± 12 years; 130 men), together with biochemical measurements and sTWEAK and PTX3 assessments. Patients were followed for cardiovascular outcomes., Results: PTX3 and IMT increased, whereas FMD and sTWEAK decreased across CKD stages (P<0.001 for all). Both PTX3 and sTWEAK appeared as strong determinants of FMD in multivariate analysis. The univariate associations of sTWEAK and PTX3 with IMT were dependent on estimated GFR. After a median of 39 months (range, 2 to 43 months), 22 fatal and 57 nonfatal cardiovascular events occurred. In a Cox model excluding PTX3, decreasing sTWEAK concentration was associated with increased risk of cardiovascular events independently of basic confounders (age, gender, estimated GFR, C reactive protein, diabetes, and cardiovascular comorbidity) and FMD. In a model excluding sTWEAK, circulating levels of PTX3 were directly associated with cardiovascular outcomes independently of basic confounders, but this association was lost after adjustment for FMD., Conclusions: Both PTX3 and sTWEAK levels associated with the endothelial dysfunction observed with progressive kidney failure. Additionally, both biomarkers impacted the predictability of cardiovascular outcomes., (© 2011 by the American Society of Nephrology)

Aim: To express the mouse glucocorticoid-induced tumor necrosis factor receptor ligand (GITRL) protein with Bac-to-Bac baculovirus expression system., Methods: GITRL gene was obtained by double digestion using EcoR I and Sal I and cloned into the baculovirus transfer vector pFastBacHTA. Then the pFastBacHTA was transformed into competent 10BacTM E.coli cells. The transposition occurred between pFastBacHTA and bacmid and a recombinant bacmid was obtained. The positive clones were picked out and the recombinant bacmid was isolated, and then complete transfected into Tn cells for producing complete recombinant baculovirus. The baculoviral stock was amplified and the GITRL protein was expressed., Results: The presence of GITRL gene containing the recombinant bacmid was verified by PCR and gene sequencing. The cytopathic effect (CPE) displayed in the transfected Tn cells assumed that the transfection was successful. Western blot analysis showed that the molecular weight of mGITRL protein was about 20 KD., Conclusion: The GITRL protein is expressed successfully with Bac-to-Bac baculovirus expression system, which may lay the foundation for further study on biological activity and function of GITRL protein.

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by synovitis and progressive destruction of the joint cartilage and underlying bone, together with diverse extra-articular manifestations. Cytokines act as soluble effector mediators of the inflammatory process. Therapeutic neutralization with monoclonal antibodies against the pro-inflammatory cytokines TNF-alpha and interleukin 1 (IL-1) has shown a clear efficacy on inflammation and clinical manifestations of RA, although a percentage of patients do not respond. This review covers new relevant cytokines in the RA physiopathology and potential biomarkers of inflammation. The current challenge is to develop biomarkers that enable an earlier diagnosis, as well as prognostic markers and new therapeutic candidates. Combined administration of several of these cytokines could eventually address a personalized treatment approach for each patient., (Copyright © 2010 Elsevier España, S.L. All rights reserved.)

Background: TWEAK (Tumor necrosis factor like WEAK inducer of apoptosis) is highly expressed by different immune cells and triggers multiple cellular responses, including control of angiogenesis. Our objective was to investigate its role in the human endometrium during the implantation window, using an ex-vivo endometrial microhistoculture model. Indeed, previous results suggested that basic TWEAK expression influences the IL-18 related uNK recruitment and local cytotoxicity., Methodology/principal Findings: Endometrial biopsies were performed 7 to 9 days after the ovulation surge of women in monitored natural cycles. Biopsies were cut in micro-pieces and cultured on collagen sponge with appropriate medium. Morphology, functionality and cell death were analysed at different time of the culture. We used this ex vivo model to study mRNA expressions of NKp46 (a uNK cytotoxic receptor) and TGF-beta1 (protein which regulates uNK cytokine production) after adjunction of excess of recombinant IL-18 and either recombinant TWEAK or its antibody. NKp46 protein expression was also detailed by immunohistochemistry in selected patients with high basic mRNA level of IL-18 and either low or high mRNA level of TWEAK. The NKp46 immunostaining was stronger in patients with an IL-18 over-expression and a low TWEAK expression, when compared with patients with both IL-18 and TWEAK high expressions. We did not observe any difference for TWEAK expression when recombinant protein IL-18 or its antibody was added, or conversely, for IL-18 expression when TWEAK or its antibody was added in the culture medium. In a pro-inflammatory environment (obtained by an excess of IL-18), inhibition of TWEAK was able to increase significantly NKp46 and TGF-beta1 mRNA expressions., Conclusions/significance: TWEAK doesn't act on IL-18 expression but seems to control IL-18 related cytotoxicity on uNK cells when IL-18 is over-expressed. Thus, TWEAK appears as a crucial physiological modulator to prevent endometrial uNK cytotoxicity in human.

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) and its receptor, fibroblast growth factor-inducible 14 (TWEAK-Fn14), are widely expressed and are involved in both injury and repair. Hotta et al. now demonstrate an important role for Fn14 in the common clamp ischemia model of acute kidney injury. Their data suggest paracrine and autocrine effects in which TWEAK produced by tubule cells feeds back on them via upregulated Fn-14 receptors expressed downstream in the proximal tubule.

Vascular calcification is a major risk factor for cardiovascular morbidity and mortality. A full understanding of the signalling pathways mediating vascular calcification is crucial not just because of the importance of this pathology in disease, but also for exploring potential therapeutic targets. Clinically there is a need to develop therapies to prevent or even reverse calcification in situations of atherosclerosis, chronic kidney disease, diabetes, and aging. In this brief review, we intend to explore the initial triggers, which are commonly related to calcification in different disease scenarios and examine the downstream signalling pathways that instigate the process of vascular calcification. In particular, we try to dissect these pathways and also examine cross-talk between different signalling pathways. Our focus is the vascular smooth muscle cell (VSMC) as it is ultimately the phenotypic modulation of these cells that may drive the calcification process.

In patients with end-stage renal disease (ESRD), inflammation, and protein energy wasting (PEW) are two highly prevalent and interconnected entities, jointly exerting a deleterious effect on multiple other ESRD-specific pathological processes and eventually on patient outcome. With respect to the pathophysiology underlying this strong association, knowledge has been actively expanded over the past few years. As such, it is nowadays recognized that inflammation acts via direct, as well as indirect, pathways in its contribution to PEW. Directly, inflammation causes alterations in amino acid utilization, translating into increased catabolism and decreased anabolism of muscle tissue. Indirectly, inflammation may act via altered ghrelin and adipokine metabolism, adipose tissue distribution, and pathological neuroendocrine signaling, as well as coexistent depression in inducing anorexia and PEW. In addition, two relatively new inflammatory markers (pentraxin-3 and TNF-like weak inducer of apoptosis) have gained attention with respect to their roles in this specific context. The current review deals with recent updates in the literature on the aforementioned pathways connecting inflammation to PEW and subsequent mortality., (Copyright © 2011 S. Karger AG, Basel.)

Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is the causative agent of KS, the second most common AIDS-associated malignancy. KSHV expresses at least 18 different mature microRNAs (miRNAs) during latency. To identify cellular targets of KSHV miRNAs, we have analyzed a previously reported series of microarrays examining changes in cellular gene expression in the presence of KSHV miRNAs. Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) receptor (TWEAKR) was among the most consistently and robustly downregulated genes in the presence of KSHV miR-K12-10a (miR-K10a). Results from luciferase assays with reporter plasmids containing the 3' untranslated region (UTR) of TWEAKR suggest a targeting of TWEAKR by miR-K10a. The mutation of two predicted miR-K10a recognition sites within the 3' UTR of TWEAKR completely disrupts inhibition by miR-K10a. The expression of TWEAKR was downregulated in cells transfected with miR-K10a as well as during de novo KSHV infection. In a KS tumor-derived endothelial cell line, the downregulation of TWEAKR by miR-K10a resulted in reduced levels of TWEAK-induced caspase activation. In addition, cells transfected with miR-K10a showed less induction of apoptosis by annexin V staining and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assays. Finally, the downregulation of TWEAKR by miR-K10a in primary human endothelial cells resulted in a decrease in levels of expression of the proinflammatory cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) in response to TWEAK. These results identify and validate an important cellular target of KSHV miRNAs. Furthermore, we demonstrate that a viral miRNA protects cells from apoptosis and suppresses a proinflammatory response, which may have significant implications in the complex context of KS lesions.

Purpose: The pathway between the glucocorticoid-induced tumor necrosis factor receptor family-related protein (GITR) and GITR ligand (GITRL) has been shown to control the function of regulatory T cells (Treg). The present study was conducted to investigate the role of this pathway and Treg in establishing immune privilege status for corneal allografts., Methods: Corneas of C57BL/6 mice were orthotopically transplanted into the eyes of BALB/c mice, and graft survival was assessed. A separate set of BALB/c mice received an anterior chamber injection of C57BL/6 splenocytes, and induction of allo-specific anterior chamber-associated immune deviation was assessed. Recipients were intraperitoneally administrated anti-GITRL, anti-CD25 monoclonal antibodies (mAb), or control immunoglobulin (IgG). Expressions of GITRL, GITR, and Foxp3 in the allografts were assessed. In vitro, cornea pretreated with anti-GITRL mAb or control IgG was incubated with T cells, and destruction of corneal endothelial cells and the population of Foxp3(+)CD25(+)CD4(+) T cells were assessed., Results: GITRL was expressed constitutively in the cornea and iris-ciliary body. GITRL-expressing allografts were infiltrated with Foxp3+GITR+CD25+CD4(+) T cells. Blockade of GITRL did not affect allo-specific ACAID but led to infiltration of Foxp3(-)CD4(+) T cells and allograft rejection. Depletion of CD25+CD4(+) Treg also accelerated allograft rejection. Destruction of corneal endothelial cells by T cells was significantly enhanced in GITRL-blocked cornea compared with control cornea. Foxp3+CD25+CD4(+) T cells were increased after incubation with GITRL-expressing cornea, but not with GITRL-blocked cornea., Conclusions: Presence of Foxp3+CD25+CD4(+) Treg in the allograft is necessary for allograft survival. GITRL-dependent expansion of Treg within the cornea is one mechanism underlying immune privilege in corneal allografts.

Tumor necrosis factor (TNF) has long been recognized to promote malaria parasite killing, but also to contribute to the development of severe malaria disease. The precise molecular mechanisms that influence these different outcomes in malaria patients are not well understood, but the virulence and drug-resistance phenotype of malaria parasites and the genetic background and age of patients are likely to be important determinants. In the past few years, important roles for other TNF family members in host immune responses to malaria parasites and the induction of disease pathology have been discovered. In this review, we will summarize these more recent findings and highlight major gaps in our current knowledge. We will also discuss future research strategies that may allow us to better understand the sometimes subtle and intricate effects of TNF family molecules during malaria infection., ((c) 2010. Published by Elsevier Inc. All rights reserved.)

Impairment in the regeneration process is a critical determinant for skeletal muscle wasting in chronic diseases and degenerative muscle disorders. Inflammatory cytokines are known to cause significant muscle wasting, however, their role in myofiber regeneration is less clear. In this study we have investigated the role of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) in skeletal muscle regeneration in vivo. Our results show that expression levels of TWEAK and its receptor Fn14 are significantly increased in skeletal muscles of mice after injury. Genetic deletion of TWEAK increased the fiber cross-sectional area and levels of embryonic isoform of myosin heavy chain in regenerating tibial anterior muscle. Conversely, muscle-specific transgenic overexpression of TWEAK reduced the fiber cross-sectional area and levels of the embryonic myosin heavy chain in regenerating muscle. TWEAK induced the expression of several inflammatory molecules and increased interstitial fibrosis in regenerating muscle. Genetic ablation of TWEAK suppressed, whereas overexpression of TWEAK increased, the activation of nuclear factor-kappa B without affecting the activation of Akt or p38 kinase in regenerating myofibers. Primary myoblasts from TWEAK-null mice showed enhanced differentiation in vitro, whereas myoblasts from TWEAK-Tg mice showed reduced differentiation compared with wild-type mice. Collectively, our study suggests that TWEAK negatively regulates muscle regeneration and that TWEAK is a potential therapeutic target to enhance skeletal muscle regeneration in vivo.