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Ubiquitin-specific peptidase 24 (USP24), a member of the deubiquitinase family, plays an important role in tumor regulation. However, the role of USP24 in Hepatocellular carcinoma（HCC）is unknown. The aim of our study was to explore the role of USP24 in HCC to seek new therapeutic targets for HCC. In this study, we found that USP24 was aberrantly upregulated in HCC tissues and predicted poor prognosis. USP24 markedly promoted HCC proliferation and progression in vitro and in vivo. Mechanistically, USP24 binds to tumor necrosis factor receptor-associated factor 2(TRAF2) and inhibits its degradation, thereby promoting the accumulation of TRAF2. Upregulation of TRAF2 activated protein kinase B/nuclear factor kappa-B (AKT/ NF-κB) signaling pathway and promoted HCC cell survival. In addition, USP24 positively correlated with programmed cell death ligand 1(PD-L1) expression in HCC, highlighting the clinical significance of USP24 activation in tumor immune evasion. Deletion of USP24 enhanced the tumor-killing ability of CD8 + T cells. Deletion of USP24 combined with anti-PD-1 antibody significantly enhanced the efficacy of HCC immunotherapy. Taken together, USP24 can be employed as a promising target to restrain tumor growth and increase the efficacy of HCC immunotherapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Cullin-based RING ligases (CRLs) comprise the largest family of ubiquitin E3 ligases. CRL activity is tightly regulated by cullin neddylation, which has been associated with various diseases. Although inhibitors of CRLs neddylation have been reported, there is a lack of small molecules that can selectively target individual cullins. Here, we identified a natural product, liquidambaric acid (LDA), with relatively selective inhibition properties against cullin (Cul) 2 neddylation, and found that its target, Tumor Necrosis Factor receptor-associated factor 2 (TRAF2) was required for the activity. TRAF2 associates with the Cul2 neddylation complex and regulates the machinery assembly, especially that of E2 (UBC12) and E3 (RBX1) enzymes. In addition, we demonstrated that by intervention of the associations between TRAF2 and the neddylation machinery, LDA disturbed NEDD8 transfer from E1 to E2, therefore blocking Cul2 neddylation. Taken together, we show that TRAF2 plays a positive role in neddylation cascades, and we have identified a small molecule capable of selective modulation of cullin neddylation., (© 2024 Federation of European Biochemical Societies.)

The cGAS-STING pathway plays an essential role in the activation of tumor immune cells. Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants with potential carcinogenicity, and their exposure is associated with the development of colorectal cancer. However, the impacts of genetic factors in the cGAS‒STING pathway and gene‒environment interactions on colorectal cancer remain understudied. We used logistic regression models and interaction analysis to evaluate the impact of genetic variants on colorectal cancer risk and gene‒environment interactions. We analysed the expression patterns of candidate genes based on the RNA-seq data. Molecular biology experiments were performed to investigate the impact of PAHs exposure on candidate gene expression and the progression of colorectal cancer. We identified the susceptibility locus rs3750511 in the cGAS‒STING pathway, which is associated with colorectal cancer risk. A negative interaction between TRAF2 rs3750511 and PAHs exposure was also identified. Single-cell RNA-seq analysis revealed significantly elevated expression of TRAF2 in colorectal cancer tissues compared with normal tissues, especially in T cells. BPDE exposure increased TRAF2 expression and the malignant phenotype of colorectal cancer cells. The treatment also further increased the expression of the TRAF2 downstream gene NF-κB and decreased the expression of Caspase8. Our results suggest that the genetic variant of rs3750511 affects the expression of TRAF2, thereby increasing the risk of colorectal cancer through interaction with PAHs. Our study provides new insights into the influence of gene‒environment interactions on the risk of developing colorectal cancer., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: TRAF-interacting protein (TRAIP) is a RING-type E3 ubiquitin ligase, which has been implicated in various cellular processes and participated in various cancers as an oncogene. However, the function and potential mechanism of TRAIP in prostate cancer (PCa) have not been investigated so far., Methods: Public TGCA data were used to evaluate the expression profile of TRAIP in prostatic tumors. The relative expression of TRAIP and TRAF2 in PCa tissues and tumor cell lines was detected by qPCR, western blot, and IHC staining. Next, TRAIP knockdown and overexpression plasmids were constructed and transfected into PCa cell lines. Moreover, cell proliferation, invasion, migration, and apoptosis were measured by colony formation, Transwell, wound healing, and flow cytometry assays. Subsequently, cell cycle and signaling pathway-related proteins were tested by western blot. Finally, the effect of TRAIP on PCa was measured based on the nude mouse xenograft model., Results: TRAIP was significantly upregulated in PCa tissues and tumor cell lines. In addition, TRAIP promoted cell proliferation, invasion, and migration of PCa cell lines. Such an oncogenic property was mediated by the cell cycle arrest and the inhibition of apoptosis, as indicated by different functional assays and the expression of cell cycle and apoptosis regulatory proteins in cultured cells. Moreover, TRAIP combined with TRAF2 to activate PI3K/AKT pathway. Finally, TRAIP depletion suppressed the growth of tumors and cell proliferation in vivo., Conclusions: Our study first revealed that TRAIP promoted tumor progression and identified it as a potential therapeutic target for PCa patients in the future., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

In the aftermath of acute kidney injury (AKI), surviving proximal tubule epithelia repopulate injured tubules to promote repair. However, a portion of cells fail to repair [termed failed-repair proximal tubule cells (FR-PTCs)] and exert ongoing proinflammatory and profibrotic effects. To better understand the molecular drivers of the FR-PTC state, we reanalyzed a mouse ischemia-reperfusion injury single-nucleus RNA-sequencing (snRNA-seq) atlas to identify Traf2 and Nck interacting kinase ( Tnik ) to be exclusively expressed in FR-PTCs but not in healthy or acutely injured proximal tubules after AKI (2 and 6 wk) in mice. We confirmed expression of Tnik protein in injured mouse and human tissues by immunofluorescence. Then, to determine the functional role of Tnik in FR-PTCs, we depleted TNIK with siRNA in two human renal proximal tubule epithelial cell lines (primary and immortalized hRPTECs) and analyzed each by bulk RNA-sequencing. Pathway analysis revealed significant upregulation of inflammatory signaling pathways, whereas pathways associated with differentiated proximal tubules such as organic acid transport were significantly downregulated. TNIK gene knockdown drove reduced cell viability and increased apoptosis, including differentially expressed poly(ADP-ribose) polymerase ( PARP ) family members, cleaved PARP-1 fragments, and increased annexin V binding to phosphatidylserine. Together, these results indicate that Tnik upregulation in FR-PTCs acts in a compensatory fashion to suppress inflammation and promote proximal tubule epithelial cell survival after injury. Modulating TNIK activity may represent a prorepair therapeutic strategy after AKI. NEW & NOTEWORTHY The molecular drivers of successful and failed repair in the proximal tubule after acute kidney injury (AKI) are incompletely understood. We identified Traf2 and Nck interacting kinase ( Tnik ) to be exclusively expressed in failed-repair proximal tubule cells after AKI. We tested the effect of si TNIK depletion in two proximal tubule cell lines followed by bulk RNA-sequencing analysis. Our results indicate that TNIK acts to suppress inflammatory signaling and apoptosis in injured renal proximal tubule epithelial cells to promote cell survival.

Ethnopharmacological Relevance: Ankylosing spondylitis (AS) is a chronic rheumatic disease known for its insidious and refractory symptoms, primarily associated with immuno-inflammation in its early stages, that affects the self-perception of patients (SPP). The exploration of long noncoding RNA (lncRNA) in immuno-inflammation of AS has garnered considerable interest. Additionally, the effectiveness of traditional Chinese medicine Xinfeng Capsule (XFC) in mitigating immuno-inflammation in AS has also been observed. However, the specific mechanisms still need to be characterized., Aim of the Study: This study elucidated the mechanism of the lncRNA NONHSAT227927.1/TRAF2/NF-κB axis in the immuno-inflammation of AS and XFC in AS treatment., Methods: LncRNA NONHSAT227927.1 and mRNA expression were assessed utilizing real-time fluorescence quantitative PCR. Protein level was determined using Western blot, and cytokine expression was measured using ELISA. Furthermore, mass spectrometry was used to analyze the binding proteins of lncRNA and rescue experiments were conducted to validate the findings. Inconsistencies in clinical baseline data were addressed using propensity score matching. The association between the XFC effect and indicator changes was evaluated using the Apriori algorithm., Results: The study revealed a substantial elevation in the expression of lncRNA NONHSAT227927.1 and tumor necrosis factor receptor-associated factor 2 (TRAF2) in AS-peripheral blood mononuclear cells. Its expression was also notably reduced after XFC treatment. In addition to this, there was a positive correlation between lncRNA NONHSAT227927.1 and TRAF2 with clinical immuno-inflammatory indicators. On the other hand, they showed a negative association with the SPP indicators. In vitro experiments have demonstrated that lncRNA NONHSAT227927.1 activated the nuclear factor (NF)-κB-p65 pathway by promoting TRAF2 expression. This activation resulted in enhanced IL-6 and TNF-α levels and reduced IL-10 and IL-4 levels. Conversely, XFC decreased the expression of lncRNA NONHSAT227927.1 and TRAF2, inhibiting the stimulation of the NF-κB-p65 cascade and restoring balance to the cytokines. The association rule analysis results indicated a strong association between XFC and decreased levels of C-reactive protein, erythrocyte sedimentation rate, and immunoglobulin A. Furthermore, XFC was strongly associated with improved SPP indicators, including general health, vitality, mental health, and role-emotional., Conclusions: LncRNA NONHSAT227927.1 plays a pro-inflammatory role in AS. XFC treatment may reverse lncRNA NONHSAT227927.1 to suppress TRAF2-mediated NF-κB-p65 activation, which in turn suppresses immuno-inflammation and improves SPP, thereby making XFC a promising candidate for therapeutic applications in AS management., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

According to recent research, metabolic-associated fatty liver disease (MAFLD) has emerged as an important underlying etiology of hepatocellular carcinoma (HCC). However, the molecular mechanism of MAFLD-HCC is still unclear. Tumor necrosis factor receptor-associated factor 2 (TRAF2) is the key molecule to mediate the signal of inflammatory NF-κB pathway. This study aims to investigate the potential dysregulation of TRAF2 and its biological function in MAFLD-HCC. Huh7 TRAF2 -/- demonstrated increased tumor formation ability compared to huh7 TRAF2 +/+ when stimulated with transforming growth factor-β (TGF-β). The decisive role of TGF-β in the development of MAFLD-HCC was confirmed through the specific depletion of TGF-β receptor II gene in the hepatocytes (Tgfbr2ΔHep) of mice. In TRAF2 -/- cells treated with TGF-β, both the glycolysis rate and lipid synthesis were enhanced. We proved the signal of the mechanistic target of rapamycin complex 1 (mTORC1) could be activated in the presence of TGF-β, and was enhanced in TRAF2 -/- cells. The coimmunoprecipitation (co-IP) experiments revealed that TRAF2 fortified the Smurf2-mediated ubiquitination degradation of AXIN1. Hence, TRAF2 depletion resulted in increased Smad7 degradation induced by AXIN1, thus promoting the TGF-β signal. We also discovered that PLX-4720 could bind with AXIN1 and restrained the tumor proliferation of TRAF2 -/- in mice fed with high-fat diet (HFD). Our findings indicate that TRAF2 plays a significant role in the pathogenesis of MAFLD-HCC. The reduction of TRAF2 expression leads to the enhancement of the TGF-β-mTORC1 pathway by facilitating AXIN1-mediated Smad7 degradation., (© 2024 Federation of American Societies for Experimental Biology.)

STAM Binding Protein Like 1 (STAMBPL1), functions as a deubiquitinase (DUB) and plays a significant role in various types of cancers. However, its effect as a DUB participating in the HCC tumorigenesis and progression still unknown. In the study, the upregulation and strong prognosis value of STAMBPL1 were identified in HCC patients. Functionally, STAMBPL1 significantly promoted HCC cells proliferation and metastasis, and it interacts with TRAF2 and stabilize it via the deubiquitination at the K63 residue. The TRAF2 upregulation stabilized by STAMBPL1 overexpression transfers of P65 protein into the nucleus and activates the WNT/PI3K/ NF-kb signaling pathway. The 251-436 sites of STAMBPL1 particularly interact with the 294-496 sites of TRAF2, thereby exerting the function of DUB and removing the ubiquitin molecules attached to TRAF2. Our research unveiled a new function of STAMBPL1 in mediating TRAF2 deubiquitination and stabilization, thereby activating the WNT/PI3K/NF-kb signaling pathway, suggesting its potential as a novel biomarker and therapeutic target for HCC., (© 2024. The Author(s).)

The interaction of the tumor necrosis factor receptor (TNFR) family member CD27 on naive CD8 + T (Tn) cells with homotrimeric CD70 on antigen-presenting cells (APCs) is necessary for T cell memory fate determination. Here, we examined CD27 signaling during Tn cell activation and differentiation. In conjunction with T cell receptor (TCR) stimulation, ligation of CD27 by a synthetic trimeric CD70 ligand triggered CD27 internalization and degradation, suggesting active regulation of this signaling axis. Internalized CD27 recruited the signaling adaptor TRAF2 and the phosphatase SHP-1, thereby modulating TCR and CD28 signals. CD27-mediated modulation of TCR signals promoted transcription factor circuits that induced memory rather than effector associated gene programs, which are induced by CD28 costimulation. CD27-costimulated chimeric antigen receptor (CAR)-engineered T cells exhibited improved tumor control compared with CD28-costimulated CAR-T cells. Thus, CD27 signaling during Tn cell activation promotes memory properties with relevance to T cell immunotherapy., Competing Interests: Declaration of interests C.A.J.-R., C.E.C., and S.R.R. are inventors on a patent (“engineered trimeric CD70 proteins and uses thereof”; WO2021072127A3) filed by Fred Hutchinson Cancer Center and licensed by Lyell Immunopharma. S.R.R. was a founder, has served as an advisor, and has patents licensed to Juno Therapeutics; S.R.R is a founder of and holds equity in Lyell Immunopharma and has served on the advisory boards for Adaptive Biotechnologies and Nohla., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The suppression of tumor proliferation via cellular senescence has emerged as a promising approach for anti-tumor therapy. Tumor necrosis factor receptor-associated factor 2 (TRAF2), an adaptor protein involved in the NF-κB signaling pathway and reactive oxygen species (ROS) production, has been implicated in hepatocellular carcinoma (HCC) proliferation. However, little is currently known about whether TRAF2 promotes HCC development by inhibiting cellular senescence. Replicative senescence model and IR-induced mouse model demonstrated that TRAF2 expression was decrease in senescence cells or liver tissues. Depletion of TRAF2 could inhibit proliferation and arrest the cell cycle via activating p53/p21 WAF1 and p16 INK4a /pRb signaling pathways in HCC cells and eventually lead to cellular senescence. Mechanistically, TRAF2 deficiency increased the expression of mitochondrial protein reactive oxygen species modulator 1 (ROMO1) and subsequently activated the NAD + /SIRT3/SOD2 pathway to promote the production of ROS and cause mitochondrial dysfunction, which eventually contributed to DNA damage response (DDR). Our findings demonstrate that TRAF2 deficiency inhibits the proliferation of HCC by promoting senescence. Therefore, targeting TRAF2 through various approaches holds therapeutic potential for treating HCC., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The Kelch-like ECH-associated protein 1 (KEAP1) - Nuclear factor erythroid 2 -related factor 2 (NRF2) pathway is the major transcriptional stress response system in cells against oxidative and electrophilic stress. NRF2 is frequently constitutively active in many cancers, rendering the cells resistant to chemo- and radiotherapy. Loss-of-function (LOF) mutations in the repressor protein KEAP1 are common in non-small cell lung cancer, particularly adenocarcinoma. While the mutations can occur throughout the gene, they are enriched in certain areas, indicating that these may have unique functional importance. In this study, we show that in the GSEA analysis of TCGA lung adenocarcinoma RNA-seq data, the KEAP1 mutations in R320 and R470 were associated with enhanced Tumor Necrosis Factor alpha (TNFα) - Nuclear Factor kappa subunit B (NFκB) signaling as well as MYC and MTORC1 pathways. To address the functional role of these hotspot mutations, affinity purification and mass spectrometry (AP-MS) analysis of wild type (wt) KEAP1 and its mutation forms, R320Q and R470C were employed to interrogate differences in the protein interactome. We identified TNF receptor associated factor 2 (TRAF2) as a putative protein interaction partner. Both mutant KEAP1 forms showed increased interaction with TRAF2 and other anti-apoptotic proteins, suggesting that apoptosis signalling could be affected by the protein interactions. A549 lung adenocarcinoma cells overexpressing mutant KEAP1 showed high TRAF2-mediated NFκB activity and increased protection against apoptosis, XIAP being one of the key proteins involved in anti-apoptotic signalling. To conclude, KEAP1 R320Q and R470C and its interaction with TRAF2 leads to activation of NFκB pathway, thereby protecting against apoptosis., Competing Interests: Declaration of competing interest The authors declare that there are no potential conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Expression of the costimulatory molecule CD40 on both B cells and dendritic cells (DCs) is required for induction of experimental autoimmune encephalomyelitis (EAE), and cell-autonomous CD40 expression on B cells is required for primary T-dependent (TD) Ab responses. We now ask whether the function of CD40 expressed by different cell types in these responses is mediated by the same or different cytoplasmic domains. CD40 has been reported to possess multiple cytoplasmic domains, including distinct TRAF6 and TRAF2/3 binding motifs. To elucidate the in vivo function of these motifs in B cells and DCs involved in EAE and TD germinal center responses, we have generated knock-in mice containing distinct CD40 cytoplasmic domain TRAF-binding site mutations and have used these animals, together with bone marrow chimeric mice, to assess the roles that these motifs play in CD40 function. We found that both TRAF2/3 and TRAF6 motifs of CD40 are critically involved in EAE induction and demonstrated that this is mediated by a role of both motifs for priming of pathogenic T cells by DCs. In contrast, the TRAF2/3 binding motif, but not the TRAF6 binding motif, is required for B cell CD40 function in TD high-affinity Ab responses. These data demonstrate that the requirements for expression of specific TRAF-binding CD40 motifs differ for B cells or DCs that function in specific immune responses and thus identify targets for intervention to modulate these responses., (Copyright © 2023 by The American Association of Immunologists, Inc.)

Background: Copine 1 (CPNE1) acts as a promoter in the progression of many kinds of cancers with the exception of pancreatic cancer (PC). This research is designed to probe the function of the CPNE1-tumor necrosis factor receptor-associated factor 2 (TRAF2) axis in PC., Methods: In vivo and in vitro models of PC were constructed, and a series of biological function tests, including MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], colony formation, flow cytometry, and immunohistochemistry, were performed., Results: The level of CPNE1 elevated dramatically in PC cells. Downregulation of CPNE1in PC cells resulted in the inhibition of colony formation and proliferation. In addition, the silencing of CPNE1 induced the G1/S arrest and apoptosis in PC cells. Additionally, TRAF2 positively interacted with CPNE1 in PANC cells. CPNE1 silencing also inhibited the growth of tumors in in vivo mouse models. Functional experiments revealed that the anti-tumor effect of CPNE1 silencing was counteracted by TRAF2 overexpression, and the tumor-promoting effect of TRAF2 overexpression was reversed by CPNE1 silencing., Conclusions: In summary, our findings indicate that the silencing of the CPNE1-TRAF2 axis restrains PC development., Competing Interests: The authors declare no conflict of interest., (© 2023 The Author(s). Published by IMR Press.)

Overturning M2 phenotype macrophage polarization is a promising therapeutic strategy for gastric cancer (GC). Diosmetin (DIO) is a natural flavonoid with antitumor effect. The aim of this study was to investigate the effect of DIO on polarization of M2 phenotype macrophages in GC. THP-1 cells were induced to M2 phenotype macrophages and co-cultured with AGS cells. The effects of DIO were determined by flow cytometry, qRT-PCR, CCK-8, Transwell, and western blot. To explore the mechanisms, THP-1 cells were transfected with adenoviral vectors containing tumor necrosis factor receptor-associated factor 2 (TRAF2) or si-TRAF2. DIO (0, 5, 10, and 20 μM) restrained the M2 phenotype macrophage polarization. In addition, DIO (20 μM) reversed the increased viability and invasion of AGS cells induced by the co-culture of M2 macrophages. Mechanistically, TRAF2 knockdown inhibited the effect of M2 phenotype macrophages on AGS cells' growth and invasion. Furthermore, DIO (20 μM) was found to decrease TRAF2/NF-κB activity in GC cells. However, TRAF2 overexpressed reversed the inhibitory effect of DIO on the co-culture system. The in vivo study confirmed that DIO treatment (50 mg/kg) could repress the growth of GC. DIO treatment markedly reduced the expressions of Ki-67 and N-cadherin, and decreased the protein levels of TRAF2 and p-NF-κB/NF-κB. In conclusion, DIO inhibited the growth and invasion of GC cells by interfering with M2 phenotype macrophage polarization through repression of the TRAF2/NF-κB signaling pathway., (© 2023 Wiley Periodicals LLC.)

Deletion of TRAF2 or TRAF3 in B cells prolongs their survival. However, it remains unknown whether deletion of such factors affects B cells' ability to tolerate DNA damage, which can be induced by chemotherapeutics and cause apoptosis. Genetic alterations of TRAF2 or TRAF3 are observed in subsets of human B-cell lymphomas and B cell-specific deletion of TRAF3 led to lymphoma development in aged mice. However, it remains unknown whether double deficiency of TRAF2 and TRAF3 accelerates B-cell lymphomagenesis. Here, we showed that B cell-specific TRAF2/3 double deficient (B-TRAF2/3-DKO) B cells were remarkably more resistant to DNA damage-induced apoptosis via upregulating cIAP2 and XIAP, which in turn attenuates caspase-3 activation. Mechanistically, resistance to DNA damage-induced apoptosis required NF-κB2, which effects by upregulating XIAP and cIAP2 transcription. B-TRAF2/3-DKO mice exhibited a shorter lifespan and succumbed to splenomegaly and lymphadenopathy. Unexpectedly, the incidence of B-cell lymphoma development in B-TRAF2/3-DKO mice was relatively rare (∼10%). Sequencing B cell receptor repertoire of diseased B cells revealed that TRAF2/3 deficiency caused abnormal oligoclonal or clonal expansion of B cells. While a fraction of mutant B cells (25-43%) from aged diseased mice harbored recurrent chromosomal translocations, primary B cells isolated from young B-TRAF2/3-DKO mice had no detectable chromosomal alterations, suggesting that TRAF2/3 deficiency per se does not cause evident genomic instability in B cells. Chemo-resistant TRAF3-deficient B-cell lymphomas were sensitized to chemotherapeutic drugs by blocking IAP activity using IAP antagonist. We conclude that double deficiency of TRAF2 and TRAF3 does not accelerate B-cell lymphomagenesis. Our studies provide insight into mechanisms regulating DNA damage-induced apoptosis and may help develop effective therapies targeting mutant B-cell lymphomas using IAP antagonist., (© 2023. The Author(s).)

Background: The management of advanced clear cell renal cell carcinoma (ccRCC) remains a major challenge in clinical practice, and the construction of more reliable prognostic prediction models and the further elucidation of key molecular mechanisms of tumor progression are topics in urgent need of in-depth investigation., Methods: We used CIBERSORT to estimate the proportion of 22 tumor-infiltrating immune cell types in the TCGA-KIRC cohort. Weighted gene co-expression network analysis, least absolute shrinkage and selection operator regression analysis were used to build risk prediction models. Expression patterns and clinical significance of TRAF2 were determined through bioinformatics analysis, real-time qPCR, Western Blot, immunohistochemistry. GSEA analysis, transmission electron microscopy, 2D/3D colony formation assay, cell migration and invasion assay, and tube-formation assay were used to investigate the underlying function and mechanism of the TRAF2/M2 macrophage/autophagy axis., Results: We constructed a novel prognostic prediction model based on M2 macrophage-related genes, which was identified as an accurate, independent and specific prognostic risk model for ccRCC patients. A reliable nomogram was constructed to predict 1-, 3-, and 5-year overall survival for patients with ccRCC. As one of the constituent genes of the risk model, TRAF2 was determined to be upregulated in ccRCC and associated with poor clinical prognosis. We found that TRAF2 promotes malignant progression of ccRCC by regulating macrophage polarization, migration and angiogenesis. Mechanistically, we found that TRAF2 promotes the polarization of M2 macrophages, and this chemotaxis is achieved in an autophagy-dependent pathway. Orthotopic tumor growth assay results revealed that TRAF2 plays a key role as a promotor of ccRCC growth and metastasis., Conclusions: In conclusion, this risk model is highly predictive of prognostic in ccRCC patients, which is expected to promote improved treatment evaluation and comprehensive management of ccRCC. Moreover, our findings reveal that the TRAF2/M2 macrophage/autophagy axis plays a key regulatory role in the malignant progression of ccRCC, and suggest that TRAF2 is a potential novel therapeutic target for advanced ccRCC., (© 2023. The Author(s).)

Purpose: CD40 is upregulated in the retinas of diabetic mice, drives pro-inflammatory molecule expression, and promotes diabetic retinopathy. The role of CD40 in diabetic retinopathy in humans is unknown. Upregulation of CD40 and its downstream signaling molecules TNF receptor associated factors (TRAFs) is a key feature of CD40-driven inflammatory disorders. We examined the expression of CD40, TRAF2, and TRAF6 as well as pro-inflammatory molecules in retinas from patients with diabetic retinopathy., Methods: Posterior poles from patients with diabetic retinopathy and non-diabetic controls were stained with antibodies against von Willebrand factor (labels endothelial cells), cellular retinaldehyde-binding protein (CRALBP), or vimentin (both label Müller cells) plus antibodies against CD40, TRAF2, TRAF6, ICAM-1, CCL2, TNF-α, and/or phospho-Tyr783 phospholipase Cγ1 (PLCγ1). Sections were analyzed by confocal microscopy., Results: CD40 expression was increased in endothelial and Müller cells from patients with diabetic retinopathy. CD40 was co-expressed with ICAM-1 in endothelial cells and with CCL2 in Müller cells. TNF-α was detected in retinal cells from these patients, but these cells lacked endothelial/Müller cell markers. CD40 in Müller cells from patients with diabetic retinopathy co-expressed activated phospholipase Cγ1, a molecule that induces TNF-α expression in myeloid cells in mice. CD40 upregulation in endothelial cells and Müller cells from patients with diabetic retinopathy was accompanied by TRAF2 and TRAF6 upregulation., Conclusions: CD40, TRAF2, and TRAF6 are upregulated in patients with diabetic retinopathy. CD40 associates with expression of pro-inflammatory molecules. These findings suggest that CD40-TRAF signaling may promote pro-inflammatory responses in the retinas of patients with diabetic retinopathy.

Background/aim: Compared to other breast cancer types, triple-negative breast cancer (TNBC) has historically had few treatment alternatives. Therefore, exploring and pinpointing potentially implicated genes could be used for treating and managing TNBC. By doing this, we will provide essential data to comprehend how the genes are involved in the apoptotic pathways of the cancer cells to identify potential therapeutic targets. Analysis of a single genetic alteration may not reveal the pathogenicity driving TNBC due to the high genomic complexity and heterogeneity of TNBC. Therefore, searching through a large variety of gene interactions enabled the identification of molecular therapeutic genes., Materials and Methods: This study used integrated bioinformatics methods such as UALCAN, TNM plotter, PANTHER, GO-KEEG and PPIs to assess the gene expression, protein-protein interaction (PPI), and transcription factor interaction of apoptosis-regulated genes., Results: Compared to normal breast tissue, gene expressions of BNIP3, TNFRSF10B, MCL1, and CASP4 were downregulated in UALCAN. At the same time, BIK, AKT1, BAD, FADD, DIABLO, and CASP9 was down-regulated in bc-GeneExMiner v4.5 mRNA expression (BCGM) databases. Based on GO term enrichment analysis, the cellular process (GO:0009987), which has about 21 apoptosis-regulated genes, is the top category in the biological processes (BP), followed by biological regulation (GO:0065007). We identified 29 differentially regulated pathways, including the p53 pathway, angiogenesis, apoptosis signaling pathway, and the Alzheimer's disease presenilin pathway. We examined the PPIs between the genes that regulate apoptosis; CASP3 and CASP9 interact with FADD, MCL1, TNF, TNFRSRF10A, and TNFRSF10; additionally, CASP3 significantly forms PPIs with CASP9, DFFA, and TP53, and CASP9 with DIABLO. In the top 10 transcription factors, the androgen receptor (AR) interacts with five apoptosis-regulated genes (p<0.0001; q<0.01), followed by retinoic acid receptor alpha (RARA) (p<0.0001; q<0.01) and ring finger protein (RNF2) (p<0.0001; q<0.01). Overall, the gene expression profile, PPIs, and the apoptosis-TF interaction findings suggest that the 27 apoptosis-regulated genes might be used as promising targets in treating and managing TNBC. Furthermore, from a total of 27 key genes, CASP2, CASP3, DAPK1, TNF, TRAF2, and TRAF3 were significantly correlated with poor overall survival in TNBC (p-value <0.05); they could play important roles in the progression of TNBC and provide attractive therapeutic targets that may offer new candidate molecules for targeted therapy., Conclusion: Our findings demonstrate that CASP2, CASP3, DAPK1, TNF, TRAF2, and TRAF3 were substantially associated with the overall survival rate (OS) difference of TNBC patients out of a total of 27 specific genes used in this study, which may play crucial roles in the development of TNBC and offer promising therapeutic interventions., (Copyright © 2023, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Non-alcoholic steatohepatitis (NASH) is a metabolic disorder that often leads to other severe liver diseases, yet treatment options are limited. Endoplasmic reticulum (ER) stress is an important pathogenetic mechanism of NASH and plays a key role in tandem steatosis as well as liver inflammation. This study aims to develop a progressive NASH model through sustained lipid accumulation and to elucidate its molecular mechanism through IRE1α/TRAF2 complex. Male SD rats were fed a high-fat diet (HFD) for 4, 8, and 12 weeks to induce progressive NASH. MRNA sequencing and PPI analysis were used to screen core genes. Transmission electron microscopy, immunofluorescence staining, ELISA, qRT-PCR, and Western blotting were used at each time point to compare differences between each index of progressive NASH at 4, 8, and 12 weeks. Sustained lipid accumulation led to structural disruption of the ER, a reduction in ER number, and an increase of lipid droplet aggregation in hepatocytes. Persistent lipid accumulation led to a persistent increase in mRNA and protein expression of the IRE1α/TRAF2 complex, IKK/IκB/NF-κB signaling pathway and ASK1/JNK1 signaling pathway, and TNF-α, IL-1β, and IL-6 also continued to increase. Persistent lipid accumulation led to a persistent exacerbation of ER stress and inflammation in progressive NASH via the IRE1α/TRAF2 complex.

Although a couple of studies have reported that mutant superoxide dismutase 1 (SOD1), one of the causative genes of familial amyotrophic lateral, interacts physically with lysyl-tRNA synthetase (KARS1) by a gain of function, there is limited evidence regarding the detailed mechanism about how the interaction leads to neuronal cell death. Our results indicated that the aminoacyl-tRNA synthetase-interacting multi-functional protein 2 (AIMP2) mediated cell death upon the interplay between mutant SOD1 and KARS1 in ALS. Binding of mutant SOD1 with KARS1 led to the release of AIMP2 from its original binding partner KARS1, and the free form of AIMP2 induced TRAF2 degradation followed by TNF-α-induced cell death. We also suggest a therapeutic application that overexpression of DX2, the exon 2-deleted antagonistic splicing variant of AIMP2 (AIMP2-DX2), reduced neuronal cell death in the ALS mouse model. Expression of DX2 suppressed TRAF2 degradation and TNF-α-induced cell death by competing mode of action against full-length AIMP2. Motor neuron differentiated form iPSC showed a resistance in neuronal cell death after DX2 administration. Further, intrathecal administration of DX2-coding adeno-associated virus (AAV) improved locomotive activity and survival in a mutant SOD1-induced ALS mouse model. Taken together, these results indicated that DX2 could prolong life span and delay the ALS symptoms through compensation in neuronal inflammation., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Cellular inhibitor of apoptosis-1 (cIAP1) is a signaling regulator with oncogenic properties. It is involved in the regulation of signaling pathways controlling inflammation, cell survival, proliferation, differentiation and motility. It is recruited into membrane-receptor-associated signaling complexes thanks to the molecular adaptor TRAF2. However, the cIAP1/TRAF2 complex exists, independently of receptor engagement, in several subcellular compartments. The present work strengthens the importance of TRAF2 in the oncogenic properties of cIAP1. cIAPs-deficient mouse embryonic fibroblasts (MEFs) were transformed using the HRas-V12 oncogene. Re-expression of cIAP1 enhanced tumor growth in a nude mice xenograft model, and promoted lung tumor nodes formation. Deletion or mutation of the TRAF2-binding site completely abolished the oncogenic properties of cIAP1. Further, cIAP1 mediated the clustering of TRAF2, which was sufficient to stimulate tumor growth. Our TRAF2 interactome analysis showed that cIAP1 was critical for TRAF2 to bind to its protein partners. Thus, cIAP1 and TRAF2 would be two essential subunits of a signaling complex promoting a pro-tumoral signal. cIAP1/TRAF2 promoted the activation of the canonical NF-κB and ERK1/2 signaling pathways. NF-κB-dependent production of IL-6 triggered the activation of the JAK/STAT3 axis in an autocrine manner. Inhibition or downregulation of STAT3 specifically compromised the growth of cIAP1-restored MEFs but not that of MEFs expressing a cIAP1-mutant and treating mice with the STAT3 inhibitor niclosamide completely abrogated cIAP1/TRAF2-mediated tumor growth. Altogether, we demonstrate that cIAP1/TRAF2 binding is essential to promote tumor growth via the activation of the JAK/STAT3 signaling pathway., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Aims/hypothesis: CD40 expressed in Müller cells is a central driver of diabetic retinopathy. CD40 causes phospholipase Cγ1 (PLCγ1)-dependent ATP release in Müller cells followed by purinergic receptor (P2X 7 )-dependent production of proinflammatory cytokines in myeloid cells. In the diabetic retina, CD40 and P2X 7 upregulate a broad range of inflammatory molecules that promote development of diabetic retinopathy. The molecular event downstream of CD40 that activates the PLCγ1-ATP-P2X 7 -proinflammatory cytokine cascade and promotes development of diabetic retinopathy is unknown. We hypothesise that disruption of the CD40-driven molecular events that trigger this cascade prevents/treats diabetic retinopathy in mice., Methods: B6 and transgenic mice with Müller cell-restricted expression of wild-type (WT) CD40 or CD40 with mutations in TNF receptor-associated factor (TRAF) binding sites were made diabetic using streptozotocin. Leucostasis was assessed using FITC-conjugated concanavalin A. Histopathology was examined in the retinal vasculature. Expression of inflammatory molecules and phospho-Tyr783 PLCγ1 (p-PLCγ1) were assessed using real-time PCR, immunoblot and/or immunohistochemistry. Release of ATP and cytokines were measured by ATP bioluminescence and ELISA, respectively., Results: Human Müller cells with CD40 ΔT2,3 (lacks TRAF2,3 binding sites) were unable to phosphorylate PLCγ1 and release ATP in response to CD40 ligation, and could not induce TNF-α/IL-1β secretion in bystander myeloid cells. CD40-TRAF signalling acted via Src to induce PLCγ1 phosphorylation. Diabetic mice in which WT CD40 in Müller cells was replaced by CD40 ΔT2,3 failed to exhibit phosphorylation of PLCγ1 in these cells and upregulate P2X 7 and TNF-α in microglia/macrophages. P2x 7 (also known as P2rx7), Tnf-α (also known as Tnf), Il-1β (also known as Il1b), Nos2, Icam-1 (also known as Icam1) and Ccl2 mRNA were not increased in these mice and the mice did not develop retinal leucostasis and capillary degeneration. Diabetic B6 mice treated intravitreally with a cell-permeable peptide that disrupts CD40-TRAF2,3 signalling did not exhibit either upregulation of P2X 7 and inflammatory molecules in the retina or leucostasis., Conclusions/interpretation: CD40-TRAF2,3 signalling activated the CD40-PLCγ1-ATP-P2X 7 -proinflammatory cytokine pathway. Src functioned as a link between CD40-TRAF2,3 and PLCγ1. Replacing WT CD40 with CD40 ΔT2,3 impaired activation of PLCγ1 in Müller cells, upregulation of P2X 7 in microglia/macrophages, upregulation of a broad range of inflammatory molecules in the diabetic retina and the development of diabetic retinopathy. Administration of a peptide that disrupts CD40-TRAF2,3 signalling reduced retinal expression of inflammatory molecules and reduced leucostasis in diabetic mice, supporting the therapeutic potential of pharmacological inhibition of CD40-TRAF2,3 in diabetic retinopathy., (© 2022. The Author(s).)

Ovarian cancer is the leading cause of death in gynecological malignancies worldwide. Our previous studies have proved that metformin inhibited the proliferation and invasion of ovarian cancer in vitro and in vivo. However, the underlying mechanisms have not been fully elucidated. Immunohistochemistry was carried out to detect the expression of tripartite motif-containing 37 (TRIM37), Ki-67, and MMP-9 in ovarian cancer and normal tissues. The influence of TRIM37 on the proliferation and invasion of ovarian cancer cells was verified by the real-time cellular analysis proliferation test, colony formation test, and Transwell assay. Western blot analysis and immunoprecipitation were used to detect the expression of the nuclear factor-κB (NF-κB) pathway and the interaction between TRIM37 and tumor necrosis factor receptor-associated factor 2 (TRAF2). Ubiquitination detection was carried out to detect the ubiquitination level of TRAF2. The present study revealed that TRIM37 expression was significantly increased in ovarian cancer tissues compared with normal control tissues, and its overexpression was closely associated with proliferation and metastasis. Metformin inhibited the NF-κB signaling pathway by downregulating TRIM37. Metformin also inhibited the ubiquitination of TRAF2 induced by TRIM37 overexpression. Metformin inhibits the proliferation and invasion of ovarian cancer cells by suppressing TRIM37-induced TRAF2 ubiquitination., (© 2022 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

ADP-heptose activates the protein kinase ALPK1 triggering TIFA phosphorylation at Thr9, the recruitment of TRAF6 and the subsequent production of inflammatory mediators. Here, we demonstrate that ADP-heptose also stimulates the formation of Lys63- and Met1-linked ubiquitin chains to activate the TAK1 and canonical IKK complexes, respectively. We further show that the E3 ligases TRAF6 and c-IAP1 operate redundantly to generate the Lys63-linked ubiquitin chains required for pathway activation, which we demonstrate are attached to TRAF6, TRAF2 and c-IAP1, and that c-IAP1 is recruited to TIFA by TRAF2. ADP-heptose also induces activation of the kinase TBK1 by a TAK1-independent mechanism, which require TRAF2 and TRAF6. We establish that ALPK1 phosphorylates TIFA directly at Thr177 as well as Thr9 in vitro. Thr177 is located within the TRAF6-binding motif and its mutation to Asp prevents TRAF6 but not TRAF2 binding, indicating a role in restricting ADP-heptose signalling. We conclude that ADP-heptose signalling is controlled by the combined actions of TRAF2/c-IAP1 and TRAF6., (© 2022 The Author(s).)

Background: Cytokines such as tumor necrosis factor-α (TNFα) have been implicated in cardiac dysfunction and toxicity associated with doxorubicin (DOX). Although TNFα can elicit different cellular responses, including survival or death, the mechanisms underlying these divergent outcomes in the heart remain cryptic. The E3 ubiquitin ligase TRAF2 (TNF receptor associated factor 2) provides a critical signaling platform for K63-linked polyubiquitination of RIPK1 (receptor interacting protein 1), crucial for nuclear factor-κB (NF-κB) activation by TNFα and survival. Here, we investigate alterations in TNFα-TRAF2-NF-κB signaling in the pathogenesis of DOX cardiotoxicity., Methods: Using a combination of in vivo (4 weekly injections of DOX 5 mg·kg -1 ·wk -1 ) in C57/BL6J mice and in vitro approaches (rat, mouse, and human inducible pluripotent stem cell-derived cardiac myocytes), we monitored TNFα levels, lactate dehydrogenase, cardiac ultrastructure and function, mitochondrial bioenergetics, and cardiac cell viability., Results: In contrast to vehicle-treated mice, ultrastructural defects, including cytoplasmic swelling, mitochondrial perturbations, and elevated TNFα levels, were observed in the hearts of mice treated with DOX. While investigating the involvement of TNFα in DOX cardiotoxicity, we discovered that NF-κB was readily activated by TNFα. However, TNFα-mediated NF-κB activation was impaired in cardiac myocytes treated with DOX. This coincided with loss of K63- linked polyubiquitination of RIPK1 from the proteasomal degradation of TRAF2. Furthermore, TRAF2 protein abundance was markedly reduced in hearts of patients with cancer treated with DOX. We further established that the reciprocal actions of the ubiquitinating and deubiquitinating enzymes cellular inhibitors of apoptosis 1 and USP19 (ubiquitin-specific peptidase 19), respectively, regulated the proteasomal degradation of TRAF2 in DOX-treated cardiac myocytes. An E3-ligase mutant of cellular inhibitors of apoptosis 1 (H588A) or gain of function of USP19 prevented proteasomal degradation of TRAF2 and DOX-induced cell death. Furthermore, wild-type TRAF2, but not a RING finger mutant defective for K63-linked polyubiquitination of RIPK1, restored NF-κB signaling and suppressed DOX-induced cardiac cell death. Last, cardiomyocyte-restricted expression of TRAF2 (cardiac troponin T-adeno-associated virus 9-TRAF2) in vivo protected against mitochondrial defects and cardiac dysfunction induced by DOX., Conclusions: Our findings reveal a novel signaling axis that functionally connects the cardiotoxic effects of DOX to proteasomal degradation of TRAF2. Disruption of the critical TRAF2 survival pathway by DOX sensitizes cardiac myocytes to TNFα-mediated necrotic cell death and DOX cardiotoxicity.

Background: Chronic inflammation due to Helicobacter pylori ( H. pylori ) infection promotes gastric carcinogenesis. Tumour necrosis factor-α (TNF-α), a key mediator of inflammation, induces cell survival or apoptosis by binding to two receptors (TNFR1 and TNFR2). TNFR1 can induce both survival and apoptosis, while TNFR2 results only in cell survival. The dysregulation of these processes may contribute to carcinogenesis., Aim: To evaluate the effects of TNFR1 and TNFR2 downregulation in AGS cells treated with H. pylori extract on the TNF-α pathway., Methods: AGS cell lines containing TNFR1 and TNFR2 receptors downregulated by specific shRNAs and nonsilenced AGS cells were treated with H. pylori extract for 6 h. Subsequently, quantitative polymerase chain reaction with TaqMan ® assays was used for the relative quantification of the mRNAs ( TNFA, TNFR1, TNFR2, TRADD, TRAF2, CFLIP, NFKB1, NFKB2, CASP8, CASP3 ) and miRNAs (miR-19a, miR-34a, miR-103a, miR-130a, miR-181c) related to the TNF-α signalling pathway. Flow cytometry was employed for cell cycle analysis and apoptosis assays., Results: In nonsilenced AGS cells, H. pylori extract treatment increased the expression of genes involved in cell survival and inhibited both apoptosis ( NFKB1, NFKB2 and CFLIP ) and the TNFR1 receptor. TNFR1 downregulation significantly decreased the expression of the TRADD and CFLIP genes, although no change was observed in the cellular process or miRNA expression. In contrast, TNFR2 downregulation decreased the expression of the TRADD and TRAF2 genes, which are both important downstream mediators of the TNFR1-mediated pathway, as well as that of the NFKB1 and CFLIP genes, while upregulating the expression of miR-19a and miR-34a . Consequently, a reduction in the number of cells in the G0/G1 phase and an increase in the number of cells in the S phase were observed, as well as the promotion of early apoptosis., Conclusion: Our findings mainly highlight the important role of TNFR2 in the TNF-α pathway in gastric cancer, indicating that silencing it can reduce the expression of survival and anti-apoptotic genes., Competing Interests: Conflict-of-interest statement: The authors declare that they have no conflict of interest., (©The Author(s) 2022. Published by Baishideng Publishing Group Inc. All rights reserved.)

Breast cancer (BC) is the most common cancer in women worldwide, and it is one of the leading causes of cancer death in women. triple-negative breast Cancer (TNBC), a subtype of BC, is typically associated with the highest pathogenic grade and incidence in premenopausal and young African American (AA) women. Chemotherapy, the most common treatment for TNBC today, can lead to acquired resistance and ineffective treatment. Therefore, novel therapeutic approaches are needed to combat medication resistance and ineffectiveness in TNBC patients. Thymoquinone (TQ) is shown to have a cytotoxic effect on human cancer cells in vitro. However, TQ's mode of action and precise mechanism in TNBC disease in vitro have not been adequately investigated. Therefore, TQ's effects on the genetically different MDA-MB-468 and MDA-MB-231 human breast cancer cell lines were assessed. The data obtained show that TQ displayed cytotoxic effects on MDA-MB-468 and MDA-MB-231 cells in a time- and concentration-dependent manner after 24 h, with IC 50 values of 25.37 µM and 27.39 µM, respectively. Moreover, MDA-MB-231 and MDA-MB-468 cells in a scratched wound-healing assay displayed poor wound closure, inhibiting invasion and migration via cell cycle blocking after 24 h. TQ arrested the cell cycle phase in MDA-MB-231 and MDA-MB-468 cells. The three cell cycle stages in MDA-MB-468 cells were significantly affected at 15 and 20 µM for G0/G1 and S phases, as well as all TQ concentrations for G2/M phases. In MDA-MB-468 cells, there was a significant decrease in G0/G1 phases with a substantial increase in the S phase and G2/M phases. In contrast, MDA-MB-231 showed a significant effect only during the two cell cycle stages (S and G2/M), at concentrations of 15 and 20 µM for S phases and all TQ values for G2/M phases. The TQ effect on the apoptotic gene profiles indicated that TQ upregulated 15 apoptotic genes in MDA-MB-231 TNBC cells, including caspases, GADD45A, TP53, DFFA, DIABLO, BNIP3, TRAF2/3, and TNFRSF10A. In MDA-MB-468 cells, 16 apoptotic genes were upregulated, including TNFRSF10A, TNF, TNFRSF11B, FADD TNFRSF10B, CASP2, and TRAF2, all of which are important for the apoptotic pathway andsuppress the expression of one anti-apoptotic gene, BIRC5, in MDA-MB-231 cells. Compared to MDA-MB-231 cells, elevated levels of TNF and their receptor proteins may contribute to their increased sensitivity to TQ-induced apoptosis. It was concluded from this study that TQ targets the MDA-MB-231 and MDA-MB-468 cells differently. Additionally, due to the aggressive nature of TNBC and the lack of specific therapies in chemoresistant TNBC, our findings related to the identified apoptotic gene profile may point to TQ as a potential agent for TNBC therapy.

Endothelial activation plays an essential role in the pathogenesis of sepsis-induced acute lung injury, however, the detailed regulatory mechanisms remain largely unknown. Here, we reported that TRIM47, an E3 ubiquitin ligase of the tripartite motif-containing protein family, was highly expressed in vascular endothelial cells. TRIM47-deficient mice were effectively resistant to lipopolysaccharide (LPS)-induced acute lung injury and death by attenuating pulmonary inflammation. TRIM47 was upregulated during TNFα-induced endothelial activation in vitro. Knockdown of TRIM47 in endothelial cells inhibited the transcription of multiple pro-inflammatory cytokines, reduced monocyte adhesion and the expression of adhesion molecules, and suppressed the secretion of IL-1β and IL-6 in endothelial cells. By contrast, overexpression of TRIM47 promoted inflammatory response and monocyte adhesion upon TNFα stimulation. In addition, TRIM47 was able to activate the NF-κB and MAPK signaling pathways during endothelial activation. Furthermore, our experiments revealed that TRIM47 resulted in endothelial activation by promoting the K63-linked ubiquitination of TRAF2, a key component of the TNFα signaling pathway. Taken together, our studies demonstrated that TRIM47 as a novel activator of endothelial cells, promoted LPS-induced pulmonary inflammation and acute lung injury through potentiating the K63-linked ubiquitination of TRAF2, which in turn activates NF-κB and MAPK signaling pathways to trigger an inflammatory response in endothelial cells., (© 2022. The Author(s).)

The scaffold protein Tumor Necrosis Factor Receptor-Associated Factor 2 (TRAF2) has been reported to play a key role in the endoplasmic reticulum (ER) stress-induced activation of c-Jun N-terminal Kinase (JNK) and hence autophagy. Autophagy is a highly conserved catabolic process, whose dysregulation is involved in the pathogenesis of various human diseases, including cancer. We investigated the involvement of TRAF2 in autophagy regulation in the human leukemic HAP1 cell line, under both basal and ER stress conditions. In TRAF2-knockout HAP1 cell line (KO), the basal autophagic flux was higher than in the parental cell line (WT). Moreover, tunicamycin-induced ER stress stimulated JNK activation and autophagy both in WT and KO HAP1. On the other hand, re-expression of a TRAF2 C-terminal fragment (residues ,310-501), in a TRAF2-KO cellular background, rendered HAP1 cells unable to activate both JNK and autophagy upon ER stress induction. Of note, this apparent dominant negative effect of the C-terminal fragment was observed even in the absence of the endogenous, full-length TRAF2 molecule. Furthermore, the expression of the C-terminal fragment resulted in both protein kinase B (AKT) pathway activation and increased resistance to the toxic effects induced by prolonged ER stress conditions. These findings indicate that TRAF2 is dispensable for the activation of both JNK and autophagy in HAP1 cells, while the TRAF2 C-terminal domain may play an autonomous role in regulating the cellular response to ER stress., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Mammals TRAF2 played a dual role in several immune signaling transduction processes. In this study, TRAF2 was cloned from Nile tilapia, Oreochromis niloticus, which named OnTRAF2. The open reading frame was 1797 bp, encoding 598 amino acids. Amino acid alignment and phylogenetic analysis indicated that OnTRAF2 showed relatively low identify with other teleost TRAF2 proteins, with the exception of TRAF2s from Epinephelus coioides. In healthy tilapia, OnTRAF2 was expressed widely in all the examined tissues, which had highest expression level in the brain. After Streptococcus agalactiae infection, the expression level of OnTRAF2 was increased significantly at different times in several organs, implying that OnTRAF2 may be involved in host defense against S. agalactiae infection. The result of subcellular localization showed that OnTRAF2 presented in cytoplasm and nucleus of HEK293T cells. Additionally, overexpression of OnTRAF2 significantly decreased the transcriptional activity of the NF-κB reporter in HEK293T cells, yeast two-hybrid results revealed that OnTRAF2 had no interaction with E3 ubiquitin ligase OnNEDD4. These results indicated that OnTRAF2 played important function during bacterial infection, and negatively mediated the immune signaling transduction in Nile tilapia, while the mechanism need further study., (Copyright © 2022. Published by Elsevier Ltd.)

Spring viremia of carp virus (SVCV) causes severe morbidity and mortality in grass carp (Ctenopharyngodon idellus) in Europe, America and several Asian countries. We found that FKBP5 (FK506-binding protein 5) is an SVCV infection response factor; however, its role in the innate immune mechanism caused by SVCV infection remains unknown. This study cloned gcFKBP5 (grass carp FKBP5) and made its mimic protein structure for function discussion. We found that gcFKBP5 expression in the primary innate immune organs of grass carp, including intestine, liver and spleen, was highly upregulated by SVCV in 24 h, with a similar result in fish cells by poly(I:C) treatment. gcFKBP overexpression aggravates viral damage to cells and increases viral replication. Furthermore, SVCV engages gcFKBP5 interacting with TRAF2 (tumour necrosis factor receptor-associated factor 2) to promote host cell apoptosis for supporting viral replication. The enhanced viral replication seems not to be due to the repression of IFN and other antiviral factors as expected. For the first time, these data show the pivotal role of gcFKBP5 in the innate immune response of grass carp to SVCV infection., (Copyright © 2021. Published by Elsevier Ltd.)

Background: Abnormal expression of long noncoding RNAs (lncRNAs) has been reported in the acute stage of acute ischemic stroke (AIS). This study aimed to explore differential lncRNA expression in the subpopulations of peripheral blood mononuclear cells (PBMCs) from AIS patients and further evaluate its underlying mechanisms in stroke-induced immunosuppression., Methods: We reanalyzed lncRNA microarray data and investigated abnormally expressed lncRNAs in the subpopulations of PBMCs by magnetic cell sorting and real-time quantitative PCR. The potential mechanism of small nucleolar RNA host gene 15 (SNHG15) was explored through in vitro and in vivo approaches., Results: The stroke-induced SNHG15 acted as a checkpoint to inhibit peripheral inflammatory responses. Functional studies showed that SNHG15 promoted M2 macrophage polarization. Mechanistically, SNHG15 expression was dysregulated through the Janus kinase (JAK)-signal transducer and activator of transcription 6 (STAT6) signaling pathway. SNHG15, localized in the cytoplasm, interfered with K63-linked ubiquitination of tumor necrosis factor receptor-associated factor 2 and thereby repressed the activation of mitogen-activated protein kinase and nuclear factor kappa-B signaling pathways and prevented the production of proinflammatory cytokines. Administration of an adenovirus targeting SNHG15 improved stroke-induced immunosuppression in mice., Conclusions: This study identified SNHG15 as a negative regulator of inflammation in stroke-induced immunosuppression, suggesting it as a novel biomarker and therapeutic target in stroke-associated infection. Trial registration ClinicalTrials.gov NCT04175691. Registered November 25, 2019, https://www.clinicaltrials.gov/ct2/show/NCT04175691 ., (© 2021. The Author(s).)

Colorectal cancer (CRC) is the most common gastrointestinal cancer, with a high mortality rate but limited therapeutic targets. DIRAS family GTPase 2 ( DIRAS2 ) is a member of the Ras-related small G-protein family whose biological functions and underlying mechanism in CRC remain poorly understood. In this study, we identified the crucial roles of DIRAS2 in CRC. DIRAS2 expression was downregulated in CRC and closely correlated with poor prognosis. Functionally, DIRAS2 inhibited CRC cell proliferation and affected cell-cycle protein expression. Mechanistically, DIRAS2 blocked nuclear factor kappa light-chain enhancer of activated B-cell signaling pathways, inducing G0/G1 arrest. Moreover, DIRAS2 interacted with 26S proteasome non-ATPase regulatory subunit 2, which facilitates the degradation of DIRAS2 in a proteasome-mediated way. Together, these results demonstrate potential functions of DIRAS2 as a tumor-suppressor gene in CRC and reveal a distinct mechanism of DIRAS2 in CRC tumorigenesis, indicating its role as a potential biomarker and target for CRC therapy., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

TNF is a proinflammatory cytokine that is critical for the coordination of tissue homeostasis. RIPK1 and TRADD are the main participants in the transduction of TNF signaling. However, data on the cell fate-controlling functions of both molecules are quite controversial. Here, we address the functions of RIPK1 and TRADD in TNF signaling by generating RIPK1- or TRADD-deficient human cell lines. We demonstrate that RIPK1 is relevant for TNF-induced apoptosis and necroptosis in conditions with depleted IAPs. In addition, TRADD is dispensable for necroptosis but required for apoptosis. We reveal a new possible function of TRADD as a negative regulator of NIK stabilization and subsequent ripoptosome formation. Furthermore, we show that RIPK1 and TRADD do not appear to be essential for the activation of MAPK signaling. Moreover, partially repressing NF-κB activation in both RIPK1 and TRADD KO cells does not result in sensitization to TNF alone due to the absence of NIK stabilization. Importantly, we demonstrate that RIPK1 is essential for preventing TRADD from undergoing TNF-induced ubiquitination and degradation. Taken together, our findings provide further insights into the specific functions of RIPK1 and TRADD in the regulation of TNF-dependent signaling, which controls the balance between cell death and survival.

Objective To investigate the role of eight locus polymorphisms of tumor necrosis factor receptor-associated factor 2 (TRAF2) and TRAF5 gene and their interaction in the susceptibility to ankylosing spondylitis (AS) in Chinese Han population. Methods Eight single nucleotide polymorphisms (SNPs) of TRAF2 (rs3750511, rs10781522, rs17250673, rs59471504) and TRAF5 (rs6540679, rs12569232, rs4951523, rs7514863) gene were genotyped in 673 AS patients and 687 controls. Results The SNPs of TRAF2 and TRAF5 do not indicate a correlation with the susceptibility of AS in Chinese Han population. Genotype frequencies of rs3750511 were statistically significant in females between patients and controls. The allele frequencies of rs10781522 and genotype frequencies of rs3750511 were statistically significant between groups of different diseases activity. One three-locus model, TRAF2 (rs10781522, rs17250673) and TRAF5 (rs12569232), had a maximum testing accuracy of 52.67% and a maximum cross-validation consistency (10/10) that was significant at the level of P = 0.0001, after determined empirically by permutation testing. As to environmental variables, only marginal association was found between sleep quality and AS susceptibility. Conclusion TRAF2 rs3750511 polymorphism may be associated with the susceptibility and severity of AS. Besides, the interaction of TRAF2 and TRAF5 genes may be associated with AS susceptibility, but many open questions remain., (© 2021. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

CircRNAs belong to a novel class of noncoding RNAs that are generated by exons of genes by alternative mRNA splicing and involved in pathophysiological processes of ischemic stroke by regulating neuro-inflammation. A total of 982 patients were enrolled in our study for stroke recovery analysis. The aim of our study was to first explore the association between the inflammation-related circRNA polymorphism and functional outcome 3 months after ischemic stroke by using multivariate logistic regression model. Next, we further investigated the role of circRNA polymorphism in predicting stroke recurrence by using Cox proportional hazard regression model. Five circRNA polymorphisms were genotyped by using polymerase chain reaction and ligation detection reaction method. We identified circ-STAT3 (signal transducer and activator of transcription) rs2293152 GG genotype to be associated with poorer recovery 90 days after stroke (OR = 1.452, 95% CI: 1.165-4.362, p = 0.016). After adjusting for confound factors, the association for rs2293152 with 3 months outcome after IS was stronger, suggesting a mechanism that rs2293152 is an independent risk factor for stroke recovery (OR = 2.255, 95% CI: 1.034-2.038, p = 0.031). However, no other circRNA polymorphisms (circ-DLGAP4 rs41274714, circ-TRAF2 rs10870141, circ-ITCH rs10485505, rs4911154) were associated with functional outcome 3 months after stroke in any genetic models. Subgroup analysis revealed that the negative effect of rs2293152 GG genotype was greater in female and older patients, subjects with history of hypertension. Additionally, all the circRNA polymorphisms were not correlated with recurrent risk of ischemic stroke. Our results indicated that circ-STAT3 might be a novel biomarker for predicting functional outcome after stroke and an important contributor to the ischemic stroke recovery., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

This article has been retracted: please see Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). This article has been retracted at the request of the Editor-in-Chief. Concern was raised about the reliability of the Western blot results in Figs. 1E, 2G and 6C, which appear to have the same eyebrow shaped phenotype as many other publications tabulated here (https://docs.google.com/spreadsheets/d/149EjFXVxpwkBXYJOnOHb6RhAqT4a2llhj9LM60MBffM/edit#gid=0). Concerns were also raised over the provenance of the flow cytometry plots in Fig. 8A. The journal requested the corresponding author comment on these concerns and provide the raw data. However the authors were not able to satisfactorily fulfil this request and therefore the Editor-in-Chief decided to retract the article., (Copyright © 2021. Published by Elsevier Inc.)

Context: The role of immune-related genes (IRGs) in thyroid cancer dedifferentiation and accompanying immune exhaustion remains largely unexplored., Objective: To construct a significant IRG-based signature indicative of dedifferentiation and immune exhaustion in thyroid cancer., Design and Settings: One exploratory cohort and 2 validation cohorts were used to identify stably dysregulated IRGs in dedifferentiated thyroid cancer (DDTC) and to obtain independent risk factors for dedifferentiation. The IRGs formed a gene signature, whose predictive value was tested by the receiver operating characteristic curve. Correlations between the signature and differentiation-related genes, immune checkpoints, and prognosis were analyzed. Gene set enrichment analyses were performed to identify related signaling pathways., Results: Four IRGs (PRKCQ, PLAUR, PSMD2, and BMP7) were found to be repeatedly dysregulated in DDTC, and they formed an IRG-based signature with a satisfactory predictive value for thyroid cancer dedifferentiation. Correlation analyses revealed that immune checkpoints were closely related to the 4 IRGs and the IRG-based signature, which was significantly associated with the histological subtype (P = 0.026), lymph node metastasis (P = 0.001), and BRAFV600E mutation (P < 0.001). The downregulated expression of PRKCQ shortened the disease-free survival for patients with thyroid cancer. Furthermore, we identified several signaling pathways inherently associated with the IRG-based signature., Conclusions: This study suggests that IRGs participate in the dedifferentiation and immune exhaustion process of thyroid cancer and are potential biomarkers for DDTC., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Glutathione peroxidase (GPx) is a selenocysteine-containing peroxidase enzyme that defends mammalian cells against oxidative stress, but the role of GPx signaling is poorly characterized. Here, we show that GPx type 1 (GPx1) plays a key regulatory role in the apoptosis signaling pathway. The absence of GPx1 augmented TNF-α-induced apoptosis in various RIPK3-negative cancer cells by markedly elevating the level of cytosolic H 2 O 2 , which is derived from mitochondria. At the molecular level, the absence of GPx1 led to the strengthened sequential activation of sustained JNK and caspase-8 expression. Two signaling mechanisms are involved in the GPx1-dependent regulation of the apoptosis pathway: (1) GPx1 regulates the level of cytosolic H 2 O 2 that oxidizes the redox protein thioredoxin 1, blocking ASK1 activation, and (2) GPx1 interacts with TRAF2 and interferes with the formation of the active ASK1 complex. Inducible knockdown of GPx1 expression impaired the tumorigenic growth of MDA-MB-231 cells (>70% reduction, P = 0.0034) implanted in mice by promoting apoptosis in vivo. Overall, this study reveals the apoptosis-related signaling function of a GPx family enzyme highly conserved in aerobic organisms.

TNF Receptor Associated Factor 2 (TRAF2) is a trimeric protein that belongs to the TNF receptor associated factor family (TRAFs). The TRAF2 oligomeric state is crucial for receptor binding and for its interaction with other proteins involved in the TNFR signaling. The monomer-trimer equilibrium of a C- terminal domain truncated form of TRAF2 (TRAF2-C), plays also a relevant role in binding the membrane, causing inward vesiculation. In this study, we have investigated the conformational dynamics of TRAF2-C through circular dichroism, fluorescence, and dynamic light scattering, performing temperature-dependent measurements. The data indicate that the protein retains its oligomeric state and most of its secondary structure, while displaying a significative increase in the heterogeneity of the tyrosines signal, increasing the temperature from ≈15 to ≈35 °C. The peculiar crowding of tyrosine residues (12 out of 18) at the three subunit interfaces and the strong dependence on the trimer concentration indicate that such conformational changes mainly involve the contact areas between each pair of monomers, affecting the oligomeric state. Molecular dynamic simulations in this temperature range suggest that the interfaces heterogeneity is an intrinsic property of the trimer that arises from the continuous, asymmetric approaching and distancing of its subunits. Such dynamics affect the results of molecular docking on the external protein surface using receptor peptides, indicating that the TRAF2-receptor interaction in the solution might not involve three subunits at the same time, as suggested by the static analysis obtainable from the crystal structure. These findings shed new light on the role that the TRAF2 oligomeric state might have in regulating the protein binding activity in vivo.

Abstract: Thyroid cancer is a common endocrine malignancy; however, surgery remains its primary treatment option. A novel targeted drug for the development and application of targeted therapy in thyroid cancer treatment remain underexplored.We obtained RNA sequence data of thyroid cancer from The Cancer Genome Atlas database and identified differentially expressed genes (DEGs). Then, we constructed co-expression network with DEGs and combined it with differentially methylation analysis to screen the key genes in thyroid cancer. PockDrug-Server, an online tool, was applied to predict the druggability of the key genes. Finally, we constructed protein-protein interaction (PPI) network to observe potential targeted drugs for thyroid cancer.We identified 3 genes correlated with altered DNA methylation level and oncogenesis of thyroid cancer. According to the druggable analysis and PPI network, we predicted TRAF2 and NCK-interacting protein kinase (TNIK) sever as the drug targeted for thyroid cancer. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis indicated that genes in protein-protein interaction network of TNIK enriched in mitogen-activated protein kinase signaling pathway. For drug repositioning, we identified a targeted drug of genes in PPI network.Our study provides a bioinformatics method for screening drug targets and provides a theoretical basis for thyroid cancer targeted therapy., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc.)

Tumour necrosis factor (TNF) receptor associated factor (TRAF) family members share a common domain architecture, but play non-redundant physiological roles in cell signalling. At the N terminus, most TRAFs have a RING domain, followed by a series of Zinc finger (ZF) domains. The RING domain of TRAF6 dimerizes, and the RING homodimer together with the first ZF assembles ubiquitin chains that form a platform which facilitates activation of downstream kinases. The RING dimer interface is conserved amongst TRAF proteins, suggesting that functional heterodimers could be possible. Here we report the structure of the TRAF5-TRAF6 RING heterodimer, which accounts for the stability of the heterodimer as well as its ability to assemble ubiquitin chains. We also show that the RING domain of TRAF6 heterodimerizes with TRAF3 and TRAF2, and demonstrate that the linker helix and first ZF of TRAF2 can cooperate with TRAF6 to promote chain assembly. Collectively our results suggest that TRAF RING homo- and hetero-dimers have the potential to bridge interaction of nearby TRAF trimers and modulate TRAF-mediated signalling., Competing Interests: Declaration of interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

TRAF2 and TRAF3 genes of tumor necrosis factor receptor (TNF-R)-associated factor (TRAF) family are involved in diverse cell signaling, and function as both tumor suppressor gene and oncogene. Alterations of TRAF2 and TRAF3 in colon cancer (CC) along with their regional difference and microsatellite instability (MSI) are largely unknown. In the present study, we analyzed TRAF2 and TRAF3 frameshift mutations in 168 sporadic CCs (100 high MSI (MSI-H) and 68 microsatellite-stable (MSS) CCs). We identified TRAF2 and TRAF3 frameshift mutations in 4 (4%) and 3 CCs (3%) with MSI-H, respectively, but none in 68 cases of MSS CCs. Of the 168 CCs, we analyzed the mutations in multi-regions for 39 CCs (16 MSI-H and 23 MSS CCs), and discovered that 12.5% (2/16) and 6.3% (1/16) of MSI-H CCs exhibited regional difference in TRAF2 and TRAF3 mutations, respectively. In the multi-region samples of 23 MSS CCs, neither TRAF2 nor TRAF3 frameshift mutation was found. In 40% of CCs, both TRAF2 and TRAF3 expressions were increased compared to normal colon cells. Our data indicate that TRAF2 and TRAF3 frameshift mutations and their regional difference as well as altered expressions are present in MSI-H CCs, which could contribute to MSI-H cancer development., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Moon, Son, Choi, Yoo and Lee.)

Chronic lymphocytic leukemia (CLL)/Small lymphocytic lymphoma (SLL) is a heterogeneous disease consisting of at least two separate subtypes, based on the mutation status of the immunoglobulin heavy chain variable gene (IGHV) sequence. Exposure to antigens seems to play a role in malignant transformation and in the selection and expansion of more aggressive CLL clones. Furthermore, a biased usage of particular IGHV gene subgroups and the existence of stereotyped B-cell receptors (BCRs) are distinctive characteristics of human CLL. We have previously described that Traf2 DN/ BCL2 double-transgenic (tg, +/+ ) mice develop CLL/SLL with high incidence with aging. In this model, TNF-Receptor Associated Factor (TRAF)-2 deficiency cooperates with B cell lymphoma (BCL)-2 in promoting CLL/SLL in mice by specifically enforcing marginal zone (MZ) B cell differentiation and rendering B cells independent of BAFF for survival. In this report, we have performed the sequencing of the IGHV-D-J rearrangements of B cell clones from the Traf2 DN/ BCL2 -tg +/+ mice with CLL/SLL. The results indicate that these mice develop oligoclonal and monoclonal B cell expansions. Allotransplantation of the oligoclonal populations into immunodeficient mice resulted in the preferential expansion of one of the parental clones. The analysis of the IGHV sequences indicated that 15% were mutated (M) and 85% unmutated (UM). Furthermore, while the Traf2 DN/ BCL2 -tg -/- (wild-type), -/+ ( BCL2 single-tg) and +/- ( Traf2DN DN single-tg) littermates showed the expression of various IGHV gene subgroups, the CLL/SLL expanded clones from the Traf2 DN/ BCL2 -tg +/+ (double-transgenic) mice showed a more restricted IGHV gene subgroup usage and an overrepresentation of particular IGHV genes. In addition, the HCDR3-encoded protein sequence indicates the existence of stereotyped immunoglobulin (Ig) in the BCRs and strong similarities with BCR recognizing autoantigens and pathogen-associated antigens. Altogether, these results highlight the remarkable similarities between the CLL/SLL developed by the Traf2 DN/ BCL2 -tg +/+ mice and its human counterpart., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Perez-Chacon and Zapata.)

Intervertebral disc degeneration (IVDD) is a common cause of low back pain. This study is aimed at investigating the role of microRNAs (miRNAs) in regulating human nucleus pulposus (NP) cell injury induced by tumor necrosis factor- (TNF-) α in IVDD. In this study, we induced NP cells with 20 ng/mL TNF- α in vitro, which promoted the obvious apoptosis of NP cells and the activation of nuclear transcription factor (NF)- κ B. In contrast, using the specific NF- κ B inhibitor BAY 11-7082 to treat cells greatly impaired the activation of NF- κ B and increased the sensitivity of NP cells to TNF- α -induced apoptosis. Moreover, both TNF- α and BAY 11-7082 treatments were associated with marked miRNA dysregulation, with miR-502 being upregulated by TNF- α treatment and downregulated by BAY 11-7082 treatment, respectively. And the overexpression of miR-502 enhanced NF- κ B activation and suppressed apoptosis of human NP cells induced by TNF- α , whereas the opposite was observed following miR-502 inhibition. Last, through bioinformatic analyses and luciferase reporter gene experiments, we identified TRAF2, an important activator of NF- κ B, as a miR-502 target gene. Similarly, siRNA-mediated knockdown of the TRAF2 expression also suppressed TNF- α -induced apoptosis and enhanced NF- κ B activation. Our findings provide evidence indicating that miR-502 is a key regulator of apoptosis of human NP cells induced by TNF- α by targeting TRAF2 and activating NF- κ B., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2021 Zhao Guo et al.)

Osteoarthritis (OA) is a chronic joint disease, which occurs in the elderly. The regulatory mechanisms of circRNAs were involved in the occurrence and development of various diseases. However, the potential regulatory network of circRNA in OA remains further research and clarification. The expression of circ&#95;0114876 was increased in OA tissues and inhibition of circ&#95;0114876 could induce cell viability and suppress inflammation as well as inhibit cell apoptosis in IL-1β induced CHON-001 cells. Circ&#95;0114876 regulated TRAF2 expression via sponging miR-671 in CHON-001 cells. Down-regulated miR-671 expression could reverse the effects of low circ&#95;0114876 expression on cell progression and inflammation in IL-1β induced CHON-001 cells. Overexpression of TRAF2 could weaken the promotion effects of high miR-671 expression on cell progression and inflammation in IL-1β induced CHON-001 cells. Circ&#95;0114876 targeted miR-671 to regulate cell progression and inflammation via modulating TRAF2 expression in IL-1β induced CHON-001 cells, and played an important regulatory mechanism in IL-1β-induced chondrocyte injury, providing a novel diagnostics and therapeutics in OA.

Astrocytes play pivotal roles in the brain and they become reactive under stress conditions. Here, we carried out, for the first time, an integrative meta-analysis of genome-wide expression profiling of astrocytes from human and mouse exposed to different stressful stimuli (hypoxia, infections by virus and bacteria, cytokines, ethanol, among others). We identified common differentially expressed genes and pathways in human and murine astrocytes. Our results showed that astrocytes induce expression of genes associated with stress response and immune system regulation when they are exposed to stressful stimuli, whereas genes related to neurogenesis are found as downregulated. Several of the identified genes showed to be important hubs in the protein-protein interaction analysis (TRAF2, CDC37 and PAX6). This work demonstrates that despite astrocytes are highly heterogeneous and complex, there are common gene expression signatures that can be triggered under distinct detrimental stimuli, which opens an opportunity for exploring other possible markers of reactivity., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Inhibition of phosphodiesterase 4 (PDE4) protects against neuronal apoptosis induced by cerebral ischemia. However, the exact mechanisms responsible for the protection of PDE4 inhibition have not been completely clarified. Roflumilast (Roflu) is an FDA-approved PDE4 inhibitor for the treatment of chronic obstructive pulmonary disease. The potential protective role of Roflu against ischemic stroke-associated neuronal injury remains unexplored. In this study, we investigated the effect and mechanism of Roflu against ischemic stroke using in vitro oxygen-glucose deprivation reperfusion (OGD/R) and in vivo rat middle cerebral artery occlusion (MCAO) models. We demonstrated that Roflu significantly reduced the apoptosis of HT-22 cells exposed to OGD/R, enhanced the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf-2), and reduced oxidative stress. Treatment with Roflu increased the phosphorylation of protein kinase B (Akt) and glycogen synthase kinase 3β (GSK3β) but decreased the level of phosphorylated inositol requiring enzyme 1α (IRE1α). Interestingly, constitutively active GSK3β (S9A) mutation abolished the effects of Roflu on oxidative stress and IRE1α phosphorylation. Moreover, Roflu decreased the binding of IRE1α to tumor necrosis factor receptor-associated factor 2 (TRAF2) and attenuated the phosphorylation of c-Jun N-terminal kinase (JNK). We also found that PDE4B knockdown reduced the phosphorylation of both IRE1α and JNK, while overexpression of PDE4B antagonized the role of PDE4B knockdown on the activation of IRE1α and JNK. Besides, the inhibition of PDE4 by Roflu produced similar effects in primary cultured neurons. Finally, Roflu ameliorated MCAO-induced cerebral injury by decreasing infarct volume, restoring neurological score, and reducing the phosphorylation of IRE1α and JNK. Collectively, these data suggest that Roflu protects neurons from cerebral ischemia reperfusion-mediated injury via the activation of GSK3β/Nrf-2 signaling and suppression of the IRE1α/TRAF2/JNK pathway. Roflu has the potential as a protective drug for the treatment of cerebral ischemia., (Copyright © 2020 Elsevier Inc. All rights reserved.)

It has been shown that sphingosine-1-phosphate (S1P) is elevated in patients with pulmonary arterial hypertension (PAH) and promotes the proliferation of pulmonary artery smooth muscle cells (PASMCs). Meanwhile, S1P has been found to induce the activation of autophagy in several types of human diseases including cancers. However, it is still unclear whether activation of autophagy mediates S1P-induced PASMCs proliferation, and detailed mechanisms responsible for these processes are indefinite. The aims of this study are to address these issues. S1P dose- and time-dependently reduced the expression of E-cadherin/CDH1 and stimulated PASMCs proliferation; this was accompanied with the elevation of TNF receptor-associated factor 2 (TRAF2), up-regulation and ubiquitination of BECN1 and the activation of autophagy. Prior silencing TRAF2 or BECN1 using siRNA or pre-incubation of cells with autophagy inhibitor chloroquine phosphate (CQ) suppressed S1P-induced autophagy activation and subsequent CDH1 degradation and further PASMCs proliferation. Taken together, our study indicates that S1P promotes the activation of autophagy by accelerating TRAF2-mediated BECN1 up-regulation and ubiquitination, which in turn results in CDH1 reduction and contributes to PASMCs proliferation., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The BCR recognizes foreign Ags to initiate humoral immunity that needs isotype-switched Abs generated via class switch recombination (CSR); however, stimulating the BCR in the absence of costimulation (e.g., CD40) does not induce CSR; thus, it remains elusive whether and how the BCR induces CSR mechanistically. Autoreactive B cells can maintain anergy via unresponsiveness of their BCRs to self-antigens. However, it remains unknown what molecule(s) restrict BCR signaling strength for licensing BCR-induced CSR and whether deficiency of such molecule(s) disrupts autoreactive B cell anergy and causes B cell-mediated diseases by modulating BCR signaling. In this study, we employ mouse models to show that the BCR's capacity to induce CSR is restrained by B cell-intrinsic checkpoints TRAF3 and TRAF2, whose deletion in B cells enables the BCR to induce CSR in the absence of costimulation. TRAF3 deficiency permits BCR-induced CSR by elevating BCR-proximal signaling intensity. Furthermore, NF-κB2 is required for BCR-induced CSR in TRAF3-deficient B cells but not for CD40-induced or LPS-induced CSR, suggesting that TRAF3 restricts NF-κB2 activation to specifically limit the BCR's ability to induce CSR. TRAF3 deficiency also disrupts autoreactive B cell anergy by elevating calcium influx in response to BCR stimulation, leading to lymphoid organ disorders and autoimmune manifestations. We showed that TRAF3 deficiency-associated autoimmune phenotypes can be rectified by limiting BCR repertoires or attenuating BCR signaling strength. Thus, our studies highlight the importance of TRAF3-mediated restraint on BCR signaling strength for controlling CSR, B cell homeostasis, and B cell-mediated disorders., (Copyright © 2020 by The American Association of Immunologists, Inc.)