Activation of Mesenchymal Stem Cells by Macrophages Prompts Human Gastric Cancer Growth through NF-κB Pathway

Accumulating evidence indicate that macrophages activate mesenchymal stem cells (MSCs) to acquire pro-inflammatory phenotype. However, the role of MSCs activated by macrophages in gastric cancer remains largely unknown. In this study, we found that MSCs were activated by macrophages to produce increased levels of inflammatory cytokines. Cell colony formation and transwell migration assays revealed that supernatants from the activated MSCs could promote both gastric epithelial cell and gastric cancer cell proliferation and migration. In addition, the expression of epithelial-mesenchymal transition (EMT), angiogenesis, and stemness-related genes was increased in activated MSCs. The phosphorylated forms of NF-κB, ERK and STAT3 in gastric cells were increased by active MSCs. Inhibition of NF-κB activation by PDTC blocked the effect of activated MSCs on gastric cancer cells. Co-injection of activated MSCs with gastric cancer cells could accelerate gastric cancer growth. Moreover, human peripheral blood monocytes derived macrophages also activated MSCs to prompt gastric cancer cell proliferation and migration. Taken together, our findings suggest that MSCs activated by macrophage acquire pro-inflammatory phenotype and prompt gastric cancer growth in an NF-κB-dependent manner, which provides new evidence for the modulation of MSCs by tumor microenvironment and further insight to the role of stromal cells in gastric carcinogenesis and cancer progression.     

GEF-H1 Mediated Control of NOD1 Dependent NF-κB Activation by Shigella Effectors

Shigella flexneri has evolved the ability to modify host cell function with intracellular active effectors to overcome the intestinal barrier. The detection of these microbial effectors and the initiation of innate immune responses are critical for rapid mucosal defense activation. The guanine nucleotide exchange factor H1 (GEF-H1) mediates RhoA activation required for cell invasion by the enteroinvasive pathogen Shigella flexneri. Surprisingly, GEF-H1 is requisite for NF-κB activation in response to Shigella infection. GEF-H1 interacts with NOD1 and is required for RIP2 dependent NF-κB activation by H-Ala-D-γGlu-DAP (γTriDAP). GEF-H1 is essential for NF-κB activation by the Shigella effectors IpgB2 and OspB, which were found to signal in a NOD1 and RhoA Kinase (ROCK) dependent manner. Our results demonstrate that GEF-H1 is a critical component of cellular defenses forming an intracellular sensing system with NOD1 for the detection of microbial effectors during cell invasion by pathogens.     

Activation of α-7 Nicotinic Acetylcholine Receptor Attenuates Cardiac Inflammation Through NLRP3/Caspase-1/IL-18 Pathway

Biochemical Genetics - Activation of α-7 nicotinic acetylcholine receptor (α7nAChR) receptor might induce cardiac inflammation, cardiac remodeling, and dysfunction. In this regard, this...     

Pepsin Digest of Wheat Gliadin Fraction Increases Production of IL-1β via TLR4/MyD88/TRIF/MAPK/NF-κB Signaling Pathway and an NLRP3 Inflammasome Activation

Celiac disease (CD) is a gluten-responsive, chronic inflammatory enteropathy. IL-1 cytokine family members IL-1β and IL-18 have been associated with the inflammatory conditions in CD patients. However, the mechanisms of IL-1 molecule activation in CD have not yet been elucidated. We show in this study that peripheral blood mononuclear cells (PBMC) and monocytes from celiac patients responded to pepsin digest of wheat gliadin fraction (PDWGF) by a robust secretion of IL-1β and IL-1α and a slightly elevated production of IL-18. The analysis of the upstream mechanisms underlying PDWGF-induced IL-1β production in celiac PBMC show that PDWGF-induced de novo pro-IL-1β synthesis, followed by a caspase-1 dependent processing and the secretion of mature IL-1β. This was promoted by K+ efflux and oxidative stress, and was independent of P2X7 receptor signaling. The PDWGF-induced IL-1β release was dependent on Nod-like receptor family containing pyrin domain 3 (NLRP3) and apoptosis-associated speck like protein (ASC) as shown by stimulation of bone marrow derived dendritic cells (BMDC) from NLRP3^−/− and ASC^−/− knockout mice. Moreover, treatment of human PBMC as well as MyD88^−/− and Toll-interleukin-1 receptor domain-containing adaptor-inducing interferon-β (TRIF)^−/− BMDC illustrated that prior to the activation of caspase-1, the PDWGF-triggered signal constitutes the activation of the MyD88/TRIF/MAPK/NF-κB pathway. Moreover, our results indicate that the combined action of TLR2 and TLR4 may be required for optimal induction of IL-1β in response to PDWGF. Thus, innate immune pathways, such as TLR2/4/MyD88/TRIF/MAPK/NF-κB and an NLRP3 inflammasome activation are involved in wheat proteins signaling and may play an important role in the pathogenesis of CD.     

(-)-Epigallocatechin-3-Gallate Protects against NO-Induced Ototoxicity through the Regulation of Caspase- 1, Caspase-3, and NF-κB Activation

Excessive nitric oxide (NO) production is toxic to the cochlea and induces hearing loss. However, the mechanism through which NO induces ototoxicity has not been completely understood. The aim of this study was to gain further insight into the mechanism mediating NO-induced toxicity in auditory HEI-OC1 cells and in ex vivo analysis. We also elucidated whether and how epigallocatechin-3-gallate (EGCG), the main component of green tea polyphenols, regulates NO-induced auditory cell damage. To investigate NO-mediated ototoxicity, S-nitroso-N-acetylpenicillamine (SNAP) was used as an NO donor. SNAP was cytotoxic, generating reactive oxygen species, releasing cytochrome c, and activating caspase-3 in auditory cells. NO-induced ototoxicity also mediated the nuclear factor (NF)-κB/caspase-1 pathway. Furthermore, SNAP destroyed the orderly arrangement of the 3 outer rows of hair cells in the basal, middle, and apical turns of the organ of Corti from the cochlea of Sprague–Dawley rats at postnatal day 2. However, EGCG counteracted this ototoxicity by suppressing the activation of caspase-3/NF-κB and preventing the destruction of hair cell arrays in the organ of Corti. These findings may lead to the development of a model for pharmacological mechanism of EGCG and potential therapies against ototoxicity.     

WAVE3-NFκB Interplay Is Essential for the Survival and Invasion of Cancer Cells

The WAVE3 cytoskeletal protein promotes cancer invasion and metastasis. We have shown that the WAVE3-mediated activation of cancer cell invasion is due, in part, to its regulation of expression and activity of key metalloproteinases (MMPs), including MMP9, which is centrally involved in invadopodia-mediated degradation of the extracellular matrix (ECM). MMP9 is also a major NFκB target gene, suggesting a potential linkage of WAVE3 to this pathway, which we sought to investigate. Mechanistically, we found that loss of WAVE3 in cancer cells leads to inhibition of NFκB signaling as a result of a decrease in the nuclear translocation of NFκB and therefore loss of activation of NFκB target genes. Conversely, overexpression of WAVE3 was sufficient to enhance NFκB activity. Both pharmacologic and genetic manipulations of NFκB effector molecules show that the biological consequence of loss of WAVE3 function in the NFκB pathway result the inhibition of invadopodia formation and ECM degradation by cancer cells, and these changes are a consequence of decreased MMP9 expression and activity. Loss of WAVE3 also sensitized cancer cells to apoptosis and cell death driven by TNFα, through the inhibition of the AKT pro-survival pathway. Our results identify a novel function of WAVE3 in NFκB signaling, where its activity is essential for the regulation of invadopodia and ECM degradation. Therefore, targeted therapeutic inhibition of WAVE3 will sensitize cancer cells to apoptosis and cell death, and suppress cancer invasion and metastasis.     

TAK1 Is Required for Survival of Mouse Fibroblasts Treated with TRAIL,and Does So by NF-κB Dependent Induction of cFLIPL

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is known as a “death ligand”—a member of the TNF superfamily that binds to receptors bearing death domains. As well as causing apoptosis of certain types of tumor cells, TRAIL can activate both NF-κB and JNK signalling pathways. To determine the role of TGF-β-Activated Kinase-1 (TAK1) in TRAIL signalling, we analyzed the effects of adding TRAIL to mouse embryonic fibroblasts (MEFs) derived from TAK1 conditional knockout mice. TAK1−/− MEFs were significantly more sensitive to killing by TRAIL than wild-type MEFs, and failed to activate NF-κB or JNK. Overexpression of IKK2-EE, a constitutive activator of NF-κB, protected TAK1−/− MEFs against TRAIL killing, suggesting that TAK1 activation of NF-κB is critical for the viability of cells treated with TRAIL. Consistent with this model, TRAIL failed to induce the survival genes cIAP2 and cFlipL in the absence of TAK1, whereas activation of NF-κB by IKK2-EE restored the levels of both proteins. Moreover, ectopic expression of cFlipL, but not cIAP2, in TAK1−/− MEFs strongly inhibited TRAIL-induced cell death. These results indicate that cells that survive TRAIL treatment may do so by activation of a TAK1–NF-κB pathway that drives expression of cFlipL, and suggest that TAK1 may be a good target for overcoming TRAIL resistance.     

Protumoral TSP50 Regulates Macrophage Activities and Polarization via Production of TNF-α and IL-1β, and Activation of the NF-κB Signaling Pathway

Testes-specific protease 50 (TSP50) is abnormally overexpressed in many kinds of cancers and promotes cell proliferation and migration. However, whether TSP50 can influence the tumor microenvironment, especially the function of immune cells in the microenvironment, remains largely unknown. We demonstrated that exposure to the conditioned medium from TSP50-overexpressing cells, or co-culture with TSP50-overexpressing cells, enhanced the cytokine production and phagocytic activities of macrophages, and induced M2b polarization. Further investigation showed that production of TNF-α and IL-1β was strongly induced by TSP50 in TSP50-overexpressing cells. TSP50-induced TNF-α and IL-1β were main factors that mediated the effects of TSP50-overexpressing cells on macrophages. The NF-κB pathway could be activated in macrophages upon the treatment of conditioned medium of TSP50-overexpressing cells and its activation is necessary for the observed effects on macrophages. Taken together, our results suggested that oncogenic TSP50 expressed in cells could activate surrounding macrophages and induce M2b polarization, partly through inducing TNF-α/ IL-1β secretion and subsequent NF-κB pathway activation. This implies a potential mechanism by which oncogene TSP50 regulates tumor microenvironment to support tumor development.     

IL-4Rα-Dependent Alternative Activation of Macrophages Is Not Decisive for Mycobacterium tuberculosis Pathology and Bacterial Burden in Mice

Classical activation of macrophages (caMph or M1) is crucial for host protection against Mycobacterium tuberculosis (Mtb) infection. Evidence suggests that IL-4/IL-13 alternatively activated macrophages (aaMph or M2) are exploited by Mtb to divert microbicidal functions of caMph. To define the functions of M2 macrophages during tuberculosis (TB), we infected mice deficient for IL-4 receptor α on macrophages (LysM^creIL-4Rα^-/lox) with Mtb. We show that absence of IL-4Rα on macrophages does not play a major role during infection with Mtb H37Rv, or the clinical Beijing strain HN878. This was demonstrated by similar mortality, bacterial burden, histopathology and T cell proliferation between infected wild-type (WT) and LysM^creIL-4Rα^-/lox mice. Interestingly, we observed no differences in the lung expression of inducible nitric oxide synthase (iNOS) and Arginase 1 (Arg1), well-established markers for M1/M2 macrophages among the Mtb-infected groups. Kinetic expression studies of IL-4/IL-13 activated bone marrow-derived macrophages (BMDM) infected with HN878, followed by gene set enrichment analysis, revealed that the MyD88 and IL-6, IL-10, G-CSF pathways are significantly enriched, but not the IL-4Rα driven pathway. Together, these results suggest that IL-4Rα-macrophages do not play a central role in TB disease progression.     

Morin,a Bioflavonoid Suppresses Monosodium Urate Crystal-Induced Inflammatory Immune Response in RAW 264.7 Macrophages through the Inhibition of Inflammatory Mediators,Intracellular ROS Levels and NF-κB Activation

Our previous studies had reported that morin, a bioflavanoid exhibited potent anti-inflammatory effect against adjuvant-induced arthritic rats. In this current study, we investigated the anti-inflammatory mechanism of morin against monosodium urate crystal (MSU)-induced inflammation in RAW 264.7 macrophage cells, an in vitro model for acute gouty arthritis. For comparison purpose, colchicine was used as a reference drug. We have observed that morin (100–300 μM) treatment significantly suppressed the levels of inflammatory cytokines (TNF-α, IL-1β, IL-6, MCP-1 and VEGF), inflammatory mediators (NO and PEG₂), and lysosomal enzymes (acid phosphatase, β-galactosidase, N-acetyl glucosamindase and cathepsin D) in MSU-crystals stimulated macrophage cells. The mRNA expression of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, and MCP-1), inflammatory enzymes (iNOS and COX-2), and NF-κBp65 was found downregulated in MSU crystal stimulated macrophage cells by morin treatment, however, the mRNA expression of hypoxanthine phospho ribosyl transferse (HPRT) was found to be increased. The flow cytometry analysis revealed that morin treatment decreased intracellular reactive oxygen species levels in MSU crystal stimulated macrophage cells. The western blot analysis clearly showed that morin mainly exerts its anti-inflammatory effects by inhibiting the MSU crystal-induced COX-2 and TNF-α protein expression through the inactivation of NF-κB signaling pathway in RAW 264.7 macrophage cells similar to that of BAY 11–7082 (IκB kinase inhibitor). Our results collectively suggest that morin can be a potential therapeutic agent for inflammatory disorders like acute gouty arthritis.