Expression Of Tumor Necrosis Factor Receptor Research Articles

Identification of tumor necrosis factor receptor 1 and its association with LPS-induced apoptosis in Onchidium reevesii

The sea slug (Onchidium reevesii) inhabits intertidal environments and face environmental pressures and disease threats. The immune system of mollusks plays a crucial role in their defense against pathogens and maintaining homeostasis. The molecular mechanisms underlying its immune defense and response to pathogens remain largely unexplored. This study aimed to isolate and identify the cDNA sequence of tumor necrosis factor receptor 1 (TNFR1) in the sea slug. The obtained cDNA sequence was named OrTNFR1. Like other members of the TNF receptor superfamily, OrTNFR1 contains a signal peptide, TNF homologous domain, and death-related domain. qPCR results showed that OrTNFR1 was highly expressed in the hepatopancreas, blood, and intestines of sea slug. This indicates that OrTNFR1 plays a role in the immune-related tissues of sea slug and is consistent with the broad expression of TNFR1 in bivalves. To investigate the inducible expression pattern and downstream effects of OrTNFR1 in response to lipopolysaccharide (LPS) stimulation, we studied the functional role of the OrTNFR1 gene by stimulating with LPS in vivo and suppressing OrTNFR1 expression using siRNA technology. Our findings demonstrate that LPS stimulation rapidly up-regulates the expression of OrTNFR1 in the hepatopancreas, blood, and intestine of the sea slug. Moreover, si-TNFR1 effectively inhibits LPS-induced TNFR1 expression and its downstream genes. Additionally, we discovered that inhibiting OrTNFR1 reduces LPS-induced cell apoptosis and the expression of apoptosis-associated protein Caspase.TNFR1 is involved in the immune defense of the sea slug, and blocking TNFR1 effectively attenuates LPS-induced cell apoptosis through the TNFR1 pathway. This finding suggests that inhibiting TNFR1 in the sea slug may serve as a potential approach to reduce cell apoptosis. As a shell-less living in harsh intertidal environments, TNFR1 likely participates in its innate immunity. Therefore, studying the TNFR1 pathway contributes to understanding the innate immune signaling pathway in the sea slug, further elucidating host defense against pathogens and apoptotic functions.

Abstract 121: Ponatinib Promotes Endothelial Cell Inflammation And Leukocyte Trafficking Via Tumor Necrosis Factor Signaling: A Potential Mechanism Of Vascular Toxicity

Background: ABL tyrosine kinase inhibitors (TKI) have dramatically improved chronic myeloid leukemia (CML) survival. Due to resistance to the first ABL-TKI imatinib (Ima), most patients are treated with newer agents, including ponatinib (Pon). Although Pon improves cancer remission, the survival benefit is mitigated by a 4-fold increase in arterial thrombosis. The mechanism of Pon-induced arterial thrombosis and the safety of Asciminib (Asc), a newly approved alternative, are not known. CML patients are older and have notably elevated serum TNFa levels. Hypothesis: Since arterial thrombotic events occur when inflamed vascular plaques rupture, we tested the hypothesis that Pon enhances endothelial cell (EC) response to TNFa, promoting EC inflammation and leukocyte trafficking. Methods and Results: In vitro: Human umbilical vein ECs (HUVECs) were treated with Pon, Ima, or Asc at concentrations observed in humans. Adhesion molecules that mediate leukocyte rolling (E- and P-selectin) and adhesion (ICAM1, VCAM1), and TNF receptors (TNFR) 1 and 2 were quantified at 24 hours by qPCR, flow cytometry (FC), and immunoblot. Pon increased TNFR2, P-selectin, VCAM, and ICAM by FC, qPCR, and immunoblot while Ima and Asc had no impact. In primary human coronary artery ECs Pon, but not Asc nor Ima, similarly increased VCAM and P-selectin mRNA. Pretreatment of HUVECs with a nonspecific TNFR inhibitor prevented Pon induction of ICAM1 and P-selectin mRNA. In vivo: Mice were treated with ABL TKIs by oral gavage to achieve drug plasma levels consistent with those in CML patients. After three days, leukocyte trafficking was quantified by intravital microscopy (IVM) in mouse mesenteric vessels. Pon, but not Asc, increased the number of rolling and adherent leukocytes, and increased serum TNFa by ELISA. Treatment with a TNFR inhibitor blocked Pon induced leukocyte trafficking by IVM. Conclusion: Pon, but not Ima nor Asc, induces expression of TNF receptors and adhesion molecules in human ECs in vitro and promotes leukocyte trafficking in vivo. Since inflamed plaques lead to acute arterial thrombosis, this mechanism could contribute to the high risk of arterial thrombosis in Pon-treated cancer patients. If so, Asc may be a safer choice as it does not induce EC inflammation.

Sesquiterpene Lactones Containing an α-Methylene-γ-Lactone Moiety Selectively Down-Regulate the Expression of Tumor Necrosis Factor Receptor 1 by Promoting Its Ectodomain Shedding in Human Lung Adenocarcinoma A549 Cells.

Alantolactone is a eudesmane-type sesquiterpene lactone containing an α-methylene-γ-lactone moiety. Previous studies showed that alantolactone inhibits the nuclear factor κB (NF-κB) signaling pathway by targeting the inhibitor of NF-κB (IκB) kinase. However, in the present study, we demonstrated that alantolactone selectively down-regulated the expression of tumor necrosis factor (TNF) receptor 1 (TNF-R1) in human lung adenocarcinoma A549 cells. Alantolactone did not affect the expression of three adaptor proteins recruited to TNF-R1. The down-regulation of TNF-R1 expression by alantolactone was suppressed by an inhibitor of TNF-α-converting enzyme. Alantolactone increased the soluble forms of TNF-R1 that were released into the culture medium as an ectodomain. The structure-activity relationship of eight eudesmane derivatives revealed that an α-methylene-γ-lactone moiety was needed to promote TNF-R1 ectodomain shedding. In addition, parthenolide and costunolide, two sesquiterpene lactones with an α-methylene-γ-lactone moiety, increased the amount of soluble TNF-R1. Therefore, the present results demonstrate that sesquiterpene lactones with an α-methylene-γ-lactone moiety can down-regulate the expression of TNF-R1 by promoting its ectodomain shedding in A549 cells.

Effects of moxibustion on visceral hypersensitivity and colonic inflammatory response in mice with chronic ulcerative colitis.

To observe the effects of moxibustion at "Zusanli" (ST36) on the expression levels of tumor necrosis factor (TNF)-α, TNF receptor 1 (TNF-R1), p38 mitogen-activated protein kinase (P38 MAPK), and transient receptor potential vanilloid 1 (TRPV1) in the colon tissue of mice with chronic ulcerative colitis (UC), so as to explore the underlying mechanisms of moxibustion in improving visceral hypersensitivity in chronic UC. Male C57BL/6J mice were randomly divided into normal group, normal with moxibustion (NM) group, model group, and model with moxibustion (MM) group, with 10 mice in each group. The chronic UC model was established by drinking 2.5% dextran sodium sulfate for 3 cycles. Mice in the NM and MM groups received moxibustion at ST36 for 20 min, 5 days per week with a 2-day break, for a total of 4 weeks. The disease activity index (DAI) score of each group was evaluated before and after treatment. The minimum volume threshold of abdominal wall retraction reflex (AWR) was measured to observe the intestinal sensitivity of mice. The colon length was measured. The pathological changes of colon tissue were observed by HE staining. The expression of mucin in colon goblet cells was detected by periodate Scheff staining. The intestinal fibrosis was observed by Masson staining. The number of trypsin-positive cells (i.e., mast cell) and the expression level of TNF-α in colon tissue were detected by immunofluorescence staining. The expression levels of TNF-R1, P38 MAPK and TRPV1 in colon tissue were detected by immunohistochemistry. Compared with the normal group after treatment, the model group showed increased DAI score (P<0.001), decreased AWR minimum volume threshold (P<0.01), shortened colon length (P<0.001), significant inflammatory infiltration in the colon tissue, reduced mucin secretion (P<0.01), increased collagen fiber deposition (P<0.001), and elevated expression levels of TNF-α, TNF-R1, P38 MAPK, and TRPV1 (P<0.001, P<0.01, P<0.05). Compared with the model group, the MM group showed decreased DAI score (P<0.01), increased AWR minimum volume threshold (P<0.001), elongated colon length (P<0.001), reduced inflammatory cell infiltration, improved integrity of mucosal glandular structure, enhanced mucin secretion (P<0.01), decreased collagen fiber deposition (P<0.001), decreased number of mast cells in the colon tissue (P<0.001), and decreased expression levels of TNF-α, TNF-R1, P38 MAPK, and TRPV1 (P<0.001, P<0.01, P<0.05). There were no significant differences in the above index between the NM group and the normal group. Moxibustion can reduce visceral hypersensitivity, alleviate inflammatory infiltration and fibrotic damage in the colon tissue of mice with chronic UC. These effects may be associated with the down-regulation of TNF-α, TNF-R1, P38 MAPK, and TRPV1 expression in colon.

TNF-expressing CD1d+ monocytes are associated with the activation of CD4- CD8- T cells in patients with Chagas cardiomyopathy.

Chagas disease cardiomyopathy is characterized by intense immune activation, with double-negative (DN) T cells as key producers of inflammatory cytokines. CD1d is an antigen-presenting molecule involved in the activation of DN T cells. We characterized CD1d+ monocytes from patients with cardiac (CARD) and indeterminate (IND) disease using flow cytometry. CARD CD1d+ monocytes exhibited higher expression of TNF, TNF-receptor, PDL-1, and Fas-L compared to those from IND. These monocytes correlated with TNF expression by DN T-cells in CARD but not in IND. CD1d+ monocytes from CARD are inflammatory and associated with DN T-cell activation, confirming that CD1d is a target for modulating inflammation in Chagas cardiomyopathy.

Altered apoptosis and proliferation in milk cells and PBMc from BLV-infected bovines with different proviral loads: Possible role of the BCL-2 family proteins, TNF-alpha, and receptors

Bovines infected by bovine leukemia virus (BLV) are characterized by presenting low proviral load (LPL) or high proviral load (HPL). It is reported that animals with HPL in peripheral blood mononuclear cells (PBMCs) present a decrease in apoptosis, an increase in viability and the proliferation rate, while animals that maintain an LPL have an intrinsic ability to control the infection, presenting an increased apoptosis rate of their PBMCs. However, there is little information on the effect of BLV on these mechanisms when the virus infects somatic milk cells (SC). This study investigates the mechanisms underlying apoptosis in milk and blood from BLV-infected animals with HPL and LPL. Relative levels of mRNA of tumor necrosis factor-α (TNF-α), TNF receptor 1 (TNF-RI), TNF receptor 2 (TNF-RII), anti-apoptotic B-cell lymphoma 2 protein (Bcl-2), and pro-apoptotic Bcl-2-like protein 4 (Bax) were measured in SC and PBMCs using quantitative reverse transcription-polymerase chain reaction (RT-qPCR) assay. A significant decrease in the expression of TNF-α in SC from HPL animals vs non-infected bovines was observed, but the infection in SC with BLV did not show a modulation on the expression of TNF receptors. A significant increase in TNF-RI expression in PBMCs from HPL bovines compared to LPL bovines was observed. No significant differences in PBMCs between HPL and LPL compared to non-infected animals concerning TNF-α, TNF-RI, and TNF-RII expression were found. There was a significant increase of both Bcl-2 and Bax in SC from LPL compared to non-infected bovines, but the Bcl-2/Bax ratio showed an anti-apoptotic profile in LPL and HPL bovines compared to non-infected ones. Reduced mRNA expression levels of Bax were determined in the PBMCs from HPL compared to LPL subjects. In contrast, BLV-infected bovines did not differ significantly in the mRNA expression of Bax compared to non-infected bovines. Our data suggest that the increased mRNA expression of Bax corresponds to the late lactation state of bovine evaluated and the exacerbated increase of mRNA expression of Bcl-2 may be one of the mechanisms for the negative apoptosis regulation in the mammary gland induced by BLV infection. These results provide new insights into the mechanism of mammary cell death in HPL and LPL BLV-infected bovine mammary gland cells during lactation.

Cannabidiol attenuates inflammatory impairment of intestinal cells expanding biomaterial-based therapeutic approaches

Cannabis-based biomaterials have the potential to deliver anti-inflammatory therapeutics specifically to desired cells, tissues, and organs, enhancing drug delivery and the effectiveness of anti-inflammatory treatment while minimizing toxicity. As a major component of Cannabis, Cannabidiol (CBD) has gained major attention in recent years because of its potential therapeutic properties, e.g., for restoring a disturbed barrier resulting from inflammatory conditions. The aim of this study was to test the hypothesis that CBD has beneficial effects under normal and inflammatory conditions in the established non-transformed intestinal epithelial cell model IPEC-J2. CBD induced a significant increase in transepithelial electrical resistance (TER) values and a decrease in the paracellular permeability of [³H]-D-Mannitol, indicating a strengthening effect on the barrier. Under inflammatory conditions induced by tumor necrosis factor alpha (TNFα), CBD stabilized the TER and mitigated the increase in paracellular permeability. Additionally, CBD prevented the barrier-disrupting effects of TNFα on the distribution and localization of sealing TJ proteins. CBD also affected the expression of TNF receptors. These findings demonstrate the potential of CBD as a component of Cannabis-based biomaterials used in the development of novel therapeutic approaches against inflammatory pathogenesis.

The TNFα/TNFR2 axis mediates natural killer cell proliferation by promoting aerobic glycolysis

Natural killer (NK) cells are predominant innate lymphocytes that initiate the early immune response during infection. NK cells undergo a metabolic switch to fuel augmented proliferation and activation following infection. Tumor necrosis factor-alpha (TNFα) is a well-known inflammatory cytokine that enhances NK cell function; however, the mechanism underlying NK cell proliferation in response to TNFα is not well established. Here, we demonstrated that upon infection/inflammation, NK cells upregulate the expression of TNF receptor 2 (TNFR2), which is associated with increased proliferation, metabolic activity, and effector function. Notably, IL-18 can induce TNFR2 expression in NK cells, augmenting their sensitivity toward TNFα. Mechanistically, TNFα-TNFR2 signaling upregulates the expression of CD25 (IL-2Rα) and nutrient transporters in NK cells, leading to a metabolic switch toward aerobic glycolysis. Transcriptomic analysis revealed significantly reduced expression levels of genes involved in cellular metabolism and proliferation in NK cells from TNFR2 KO mice. Accordingly, our data affirmed that genetic ablation of TNFR2 curtails CD25 upregulation and TNFα-induced glycolysis, leading to impaired NK cell proliferation and antiviral function during MCMV infection in vivo. Collectively, our results delineate the crucial role of the TNFα-TNFR2 axis in NK cell proliferation, glycolysis, and effector function.

Parameters of TNF receptor co-expression in allergic and autoimmune processes: Differences and diagnostic significance

The authors used a method quantitative estimation density of TNFR1/TNFR2 on cells by flow cytometry with calibration particles, which allowed them to estimate the absolute number of receptors on cells regardless of the type of flow cytometer. The TNF receptor expression parameters were used to determine their association with the fact of disease and to build diagnostic models. The proposed methodological approach using a combination of flow cytometry and mathematical modeling techniques represents a promising direction for testing the diagnostic and prognostic significance of the studied biomarkers. The multifactorial regression analysis constructed on the basis of this approach made it possible to refine and supplement diagnostic schemes for determining the probability of rheumatoid arthritis and bronchial asthma in patients.

Homotherapy for heteropathy of Alzheimer's disease and anemia through reducing the expression of toll-like receptor and TNF by steamed Panax notoginseng

BackgroundOne of the effects of Steamed Panax notoginsen (SPN) is to replenish blood, which is mostly used to treat anemia in clinic. SPN has the effect of treating anemia and Alzheimer's disease (AD) in clinical and basic research. In traditional Chinese medicine, anemia and AD have the same characteristics, and their symptoms are qi and blood deficiency. MethodsFirst, data analysis was carried out through network pharmacology to predict the action targets of SPN homotherapy in the treatment of AD and anemia. Specifically, TCMSP and relevant literature were used to screen the main active ingredients of Panax notoginseng, and SuperPred was used to predict the action targets of the active ingredients. Disease targets related to AD and anemia were collected through Genecards database, and STRING and protein interaction (PPI) was used for enrichment analysis, Analyze the characteristics of the active ingredient target network on the Cytascape 3.9.0 platform, and use Metascape to enrich the gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes Pathway Enrichment (KEGG pathway). Then Drosophila was used as the AD animal model, and the effects of SPN on the climbing ability, olfactory memory and brain Aβ, with rats as anemia animal models, the improvement effect of SPN on blood routine and organ index of rats with blood deficiency induced by CTX and APH was analyzed to further explain the therapeutic effect of SPN on these two diseases. Finally, the regulatory effect of SPN on the key active target of allotherapy for AD and anemia was verified by PCR. ResultsAfter the screening, 17 active components and 92 action targets of SPN were obtained. The degree values of components and the first 15 targets are NFKB1, IL10, PIK3CA, PTGS2, SRC, ECFR, CASP3, MTOR, IL1B, ESR1, AKT1, HSP90AA1, IL6, TNF, and Toll-like receptor, it is mainly related to inflammatory response, immune regulation and antioxidation. SPN improved the climbing ability, olfactory memory ability, and Aβ42 content in the brain of Aβ flies, and significantly reduced the expression of TNF and Toll-like receptor in the brain after treatment. SPN can significantly improve the blood routine index and organ index of anemia rats, and also significantly reduce the expression of TNF and Toll-like receptor in the brain after treatment. ConclusionSPN can regulate the expression of TNF and Toll-like receptor to achieve the same treatment of AD and anemia.

Effects of Modified Glucosamine on the Chondrogenic Potential of Circulating Stem Cells under Experimental Inflammation.

Glucosamine (GlcN) is a glycosaminoglycan (GAGs) constituent in connective tissues. It is naturally produced by our body or consumed from diets. In the last decade, in vitro and in vivo trials have demonstrated that the administration of GlcN or its derivates has a protective effect on cartilage when the balance between catabolic and anabolic processes is disrupted and cells are no longer able to fully compensate for the loss of collagen and proteoglycans. To date, these benefits are still controversial because the mechanism of action of GlcN is not yet well clarified. In this study, we have characterized the biological activities of an amino acid (AA) derivate of GlcN, called DCF001, in the growth and chondrogenic induction of circulating multipotent stem cells (CMCs) after priming with tumor necrosis factor-alpha (TNFα), a pleiotropic cytokine commonly expressed in chronic inflammatory joint diseases. In the present work, stem cells were isolated from the human peripheral blood of healthy donors. After priming with TNFα (10 ng/mL) for 3 h, cultures were treated for 24 h with DCF001 (1 μg/mL) dissolved in a proliferative (PM) or chondrogenic (CM) medium. Cell proliferation was analyzed using a Corning® Cell Counter and trypan blue exclusion technique. To evaluate the potentialities of DCF001 in counteracting the inflammatory response to TNFα, we measured the amount of extracellular ATP (eATP) and the expression of adenosine-generating enzymes CD39/CD73, TNFα receptors, and NF-κB inhibitor IκBα using flow cytometry. Finally, total RNA was extracted to perform a gene expression study of some chondrogenic differentiation markers (COL2A1, RUNX2, and MMP13). Our analysis has shed light on the ability of DCF001 to (a) regulate the expression of CD39, CD73, and TNF receptors; (b) modulate eATP under differentiative induction; (c) enhance the inhibitory activity of IκBα, reducing its phosphorylation after TNFα stimulation; and (d) preserve the chondrogenic potentialities of stem cells. Although preliminary, these results suggest that DCF001 could be a valuable supplement for ameliorating the outcome of cartilage repair interventions, enhancing the efficacy of endogenous stem cells under inflammatory stimuli.

Autoimmune Valvular Carditis Requires Endothelial Cell TNFR1 Expression.

Inflammation is a key driver of cardiovascular pathology, and many systemic autoimmune/rheumatic diseases are accompanied by increased cardiac risk. In the K/B.g7 mouse model of coexisting systemic autoantibody-mediated arthritis and valvular carditis, valve inflammation depends on macrophage production of TNF (tumor necrosis factor) and IL-6 (interleukin-6). Here, we sought to determine if other canonical inflammatory pathways participate and to determine whether TNF signaling through TNFR1 (tumor necrosis factor receptor 1) on endothelial cells is required for valvular carditis. We first asked if type 1, 2, or 3 inflammatory cytokine systems (typified by IFNγ, IL-4, and IL-17, respectively) were critical for valvular carditis in K/B.g7 mice, using a combination of in vivo monoclonal antibody blockade and targeted genetic ablation studies. To define the key cellular targets of TNF, we conditionally deleted its main proinflammatory receptor, TNFR1, in endothelial cells. We analyzed how the absence of endothelial cell TNFR1 affected valve inflammation, lymphangiogenesis, and the expression of proinflammatory genes and molecules. We found that typical type 1, 2, and 3 inflammatory cytokine systems were not required for valvular carditis, apart from a known initial requirement of IL-4 for autoantibody production. Despite expression of TNFR1 on a wide variety of cell types in the cardiac valve, deleting TNFR1 specifically on endothelial cells protected K/B.g7 mice from valvular carditis. This protection was accompanied by reduced expression of VCAM-1 (vascular cell adhesion molecule), fewer valve-infiltrating macrophages, reduced pathogenic lymphangiogenesis, and diminished proinflammatory gene expression. TNF and IL-6 are the main cytokines driving valvular carditis in K/B.g7 mice. The interaction of TNF with TNFR1 specifically on endothelial cells promotes cardiovascular pathology in the setting of systemic autoimmune/rheumatic disease, suggesting that therapeutic targeting of the TNF:TNFR1 interaction could be beneficial in this clinical context.

Effects of Age and Suntan on the Expression of Second Messenger Signaling Pathways of Necroptosis in Skin Cells during Facelifting Surgery.

We studied the effects of age and suntan on the expression of necroptosis signaling molecules (RIPK1, RIPK3, and MLKL kinases) and first TNF receptor (TNFR1) in isolated skin cells from women undergoing facelift surgery. In women above 50 years, the expression of the TNFR1, kinases RIPK1, RIPK3, and MLKL, the phosphorylated forms of these kinases was significantly (p<0.05) increased in comparison with the corresponding parameters in women under 30 years. The expression of all necroptosis proteins and TNFR1 in women with suntan was significantly (p<0.05) higher than in those without tan. Cells from the surgical material were incubated with TNFα to determine the level of induced necroptosis. In women aged >50 years and women with suntan, the expression of phosphorylated forms of kinases was significantly increased, which attested to necroptosis activation. This study allowed identifying the targets on skin cells for prevention of necrosis and inflammation after facelift surgery.

TNFR2 as a Potential Biomarker for Early Detection and Progression of CKD

The inflammatory pathway driven by TNF-α, through its receptors TNFR1 and TNFR2, is a common feature in the pathogenesis of chronic kidney disease (CKD), regardless of the initial disease cause. Evidence correlates the chronic inflammatory status with decreased renal function. Our aim was to evaluate the potential of TNF receptors as biomarkers for CKD diagnosis and staging, as well as their association with the progression of renal lesions, in rat models of early and moderate CKD. We analyzed the circulating levels of inflammatory molecules-tumor necrosis factor-alpha (TNF-α), tumor necrosis factor receptor 1 (TNFR1) and 2 (TNFR2) and tissue inhibitor of metalloproteinase-1 (TIMP-1)-and studied their associations with TNFR1 and TNFR2 renal expression, glomerular and tubulointerstitial lesions, and with biomarkers of renal (dys)function. An increase in all inflammatory markers was observed in moderate CKD, as compared to controls, but only circulating levels of both TNFR1 and TNFR2 were significantly increased in the early disease; TNFR2 serum levels were negatively correlated with eGFR. However, only TNFR2 renal expression increased with CKD severity and showed correlations with the score of mild and advanced tubular lesions. Our findings suggest that renal TNFR2 plays a role in CKD development, and has potential to be used as a biomarker for the early detection and progression of the disease. Still, the potential value of this biomarker in disease progression warrants further investigation.

The Influence of Severity and Disease Duration on TNF Receptors’ Redistribution in Asthma and Rheumatoid Arthritis

One of the mechanisms of cellular dysfunction during the chronization of immune-system-mediated inflammatory diseases is a change in the profile of expression and co-expression of receptors on cells. The aim of this study was to compare patterns of redistribution of TNF receptors (TNFRs) among patients with different durations of rheumatoid arthritis (RA) or asthma. Subgroup analysis was performed on RA (n = 41) and asthma (n = 22) patients with disease duration<10 years and >10 years and on 30 comparable healthy individuals. The co-expression profile of TNFR1 and TNFR2 was assessed in T cells, B cells, monocytes, regulatory T cells, T-helper subsets, and cytotoxic T-lymphocyte subsets. Percentages of cells with different co-expression combinations and receptor density per cell were estimated. Longer disease duration was significantly associated with a redistribution of receptors in immunocompetent cell subsets with an increase in the expression of TNFR1 in asthma but did not correlate with significant unidirectional changes in receptor expression in RA. In asthma, a higher proportion of cells with a certain type of TNF receptor (as compared with the healthy group) was correlated with a simultaneous greater density of this receptor type. In RA, an inverse correlation was observed (compensatory lower receptor density). Mechanisms of long-term changes in the expression of TNF receptors differ significantly between the diseases of autoimmune and allergic etiology. The formation of irreversible morphostructural alterations was strongly correlated with changes in the expression of TNFR1 in asthma and with changes in the expression of TNFR2 in RA.

Effect of different types of immunosuppressive therapy on the parameters of TNF receptor expression in patients with rheumatoid arthritis

Background. The balance of TNF receptor expression on immune cells is a key factor determining cytokine-induced activation of proapoptotic or proliferative signaling pathways. As a result, the changes in cytokine level and in expression of its receptors may be one of the mechanisms that regulate the level of systemic and local inflammation in rheumatoid arthritis (RA) and determine the degree of therapy effectiveness. The aim. To study the effect of rheumatoid arthritis therapy on the change in the patterns of TNF receptors expression in terms of co-expression and the number of receptors on the main subpopulations of immunocompetent cells.Materials and methods. A comparative analysis of the profiles of TNF receptors type 1 and 2 (TNFR1/2) co-expression was carried out in patients with RA (n = 16) before and after having inpatient effective therapy and in comparison with a group of healthy individuals (n = 21). We compared the number of receptors and the proportion of cells expressing the corresponding receptor using flow cytometry and studied the subpopulations of regulatory T cells, T cells, B cells, and monocytes. Results. In patients with RA, there is a significant redistribution of TNFR1 and TNFR2 expression on immunocompetent cells, while the intensity of changes is associated not only with disease severity indicators, but also with the therapy received. The key adaptive mechanism of the TNF system in long-term treatment refractory course of RA is a change in the proportion of double-positive TNFR1+TNFR2+ cells, while the effectiveness of therapy and clinical indicators of the disease severity are associated with individual variability in the parameters of type 2 receptors expression. Conclusions. The data obtained confirm the existence of a relationship between an imbalance in the expression of type 1 and type 2 TNF receptors on immunocompetent cells and the effectiveness of response to therapy. The identified patterns of typical changes in TNFR1/2 co-expression in RA can be used as potential therapeutic targets and predictive factors for the effectiveness of therapy.

TNFα/TNFR1 signal induces excessive senescence of decidua stromal cells in recurrent pregnancy loss

Defects in decidual response are associated with adverse pregnancy outcomes which includes recurrent pregnancy loss (RPL). It is reported that cellular senescence happens during decidualization and pro-senescent decidual response in the luteal phase endometrium is related to RPL. However, the underlying mechanisms of how excessive decidual senescence takes place in RPL decidua cells remain largely unexplored. The senescent phenotype of RPL decidua and tumor necrosis factor receptor 1(TNFR1) expression were analyzed by using our previously published single-cell sequencing dataset of decidua cells from 6 RPL and 5 matched normal decidua, which were further verified by PCR and WB in decidual tissues. Effects of TNFα on the decidual stromal cells (DSCs) senescence and underlying molecular pathways were analyzed using the in vitro decidualization model of human endometrial stromal cells (HESCs). We showed that decidual stroma cells from RPL patients exhibited transcriptomic features of cellular senescence by analysis of single-cell datasets. The TNFα level and TNFR1 expression were increased in RPL decidua tissues. Furthermore, in vitro cell model demonstrated that increased TNFα induced excessive senescence during decidualization and TNFR1/p53/p16 pathway mediates TNFα-induced stromal senescence. In addition, we also found that the expression of IGFBP1 was regulated by TNFα-TNFR1 interaction during decidualization. Taken together, the present findings suggest that the increased secretion of TNFα induced stromal cell excessive senescence in RPL decidua, which is mediated via TNFR1, and thus provide a possible therapeutic target for the treatment of RPL.

A TNFR1–UBCH10 axis drives lung squamous cell carcinoma dedifferentiation and metastasis through a cell-autonomous signaling loop

Tumor necrosis factor receptor 1 (TNFR1), encoded by TNFRSF1A, is a critical transducer of inflammatory pathways, but its physiological role in human cancer is not completely understood. Here, we observed high expression of TNFR1 in many human lung squamous cell carcinoma (SCCs) samples and in spontaneous lung SCCs derived from kinase-dead Ikkα knock-in (KA/KA) mice. Knocking out Tnfrf1a in KA/KA mice blocked lung SCC formation. When injected via tail vein, KALLU+ lung SCC cells that highly expressed TNFR1/TNF, Sox2, c-Myc, Twist1, Bcl2, and UBCH10, generated dedifferentiated spindle cell carcinomas with epithelial–mesenchymal transition markers in mouse lungs. In contrast, KALLU+ cells with silenced TNFR1 and KALLU- cells that expressed low levels of TNFR1 generated well-differentiated lung SCCs and were less tumorigenic and metastatic. We identified a downstream effector of TNFR1: oncogenic UBCH10, an E2 ubiquitin-conjugating enzyme with targets including Twist1, c-Myc, and Sox2, which enhanced SCC cell dedifferentiation. Furthermore, Tg-K5.TNFR1;KA/KA mice, which expressed transgenic TNFR1 in keratin 5-positve epithelial cells, developed more poorly differentiated and metastatic lung SCCs than those found in KA/KA mice. These findings demonstrate that an overexpressed TNFR1–UBCH10 axis advances lung carcinogenesis and metastasis through a dedifferentiation mechanism. Constituents in this pathway may contribute to the development of differentiation-related therapies for lung SCC.

Extracellular sulfatase-2 is overexpressed in rheumatoid arthritis and mediates the TNF-α-induced inflammatory activation of synovial fibroblasts.

Extracellular sulfatase-2 (Sulf-2) influences receptor-ligand binding and subsequent signaling by chemokines and growth factors, yet Sulf-2 remains unexplored in inflammatory cytokine signaling in the context of rheumatoid arthritis (RA). In the present study, we characterized Sulf-2 expression in RA and investigated its potential role in TNF-α-induced synovial inflammation using primary human RA synovial fibroblasts (RASFs). Sulf-2 expression was significantly higher in serum and synovial tissues from patients with RA and in synovium and serum from hTNFtg mice. RNA sequencing analysis of TNF-α-stimulated RASFs showed that Sulf-2 siRNA modulated ~2500 genes compared to scrambled siRNA. Ingenuity Pathway Analysis of RNA sequencing data identified Sulf-2 as a primary target in fibroblasts and macrophages in RA. Western blot, ELISA, and qRT‒PCR analyses confirmed that Sulf-2 knockdown reduced the TNF-α-induced expression of ICAM1, VCAM1, CAD11, PDPN, CCL5, CX3CL1, CXCL10, and CXCL11. Signaling studies identified the protein kinase C-delta (PKCδ) and c-Jun N-terminal kinase (JNK) pathways as key in the TNF-α-mediated induction of proteins related to cellular adhesion and invasion. Knockdown of Sulf-2 abrogated TNF-α-induced RASF proliferation. Sulf-2 knockdown with siRNA and inhibition by OKN-007 suppressed the TNF-α-induced phosphorylation of PKCδ and JNK, thereby suppressing the nuclear translocation and DNA binding activity of the transcription factors AP-1 and NF-κBp65 in human RASFs. Interestingly, Sulf-2 expression positively correlated with the expression of TNF receptor 1, and coimmunoprecipitation assays demonstrated the binding of these two proteins, suggesting they exhibit crosstalk in TNF-α signaling. This study identified a novel role of Sulf-2 in TNF-α signaling and the activation of RA synoviocytes, providing the rationale for evaluating the therapeutic targeting of Sulf-2 in preclinical models of RA.

Cucurbitacin B Down-Regulates TNF Receptor 1 Expression and Inhibits the TNF-α-Dependent Nuclear Factor κB Signaling Pathway in Human Lung Adenocarcinoma A549 Cells.

Pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), induce the expression of intracellular adhesion molecule-1 (ICAM-1) by activating the nuclear factor κB (NF-κB) signaling pathway. In the present study, we found that cucurbitacin B decreased the expression of ICAM-1 in human lung adenocarcinoma A549 cells stimulated with TNF-α or interleukin-1α. We further investigated the mechanisms by which cucurbitacin B down-regulates TNF-α-induced ICAM-1 expression. Cucurbitacin B inhibited the nuclear translocation of the NF-κB subunit RelA and the phosphorylation of IκBα in A549 cells stimulated with TNF-α. Cucurbitacin B selectively down-regulated the expression of TNF receptor 1 (TNF-R1) without affecting three adaptor proteins (i.e., TRADD, RIPK1, and TRAF2). The TNF-α-converting enzyme inhibitor suppressed the down-regulation of TNF-R1 expression by cucurbitacin B. Glutathione, N-acetyl-L-cysteine, and, to a lesser extent, L-cysteine attenuated the inhibitory effects of cucurbitacin B on the TNF-α-induced expression of ICAM-1, suggesting that an α,β-unsaturated carbonyl moiety is essential for anti-inflammatory activity. The present results revealed that cucurbitacin B down-regulated the expression of TNF-R1 at the initial step in the TNF-α-dependent NF-κB signaling pathway.