Activation Of Nuclear factor-kB Pathway Research Articles

The Shc protein Rai enhances T-cell survival under hypoxia.

Hypoxia occurs in physiological and pathological conditions. T cells experience hypoxia in pathological and physiological conditions as well as in lymphoid organs. Indeed, hypoxia-inducible factor 1α (HIF-1α) affects T cell survival and functions. Rai, an Shc family protein member, exerts pro-survival effects in hypoxic neuroblastoma cells. Since Rai is also expressed in T cells, we here investigated its role in hypoxic T cells. In this work, hypoxia differently affected cell survival, proapoptotic, and metabolic programs in T cells, depending upon Rai expression. By using Jurkat cells stably expressing Rai and splenocytes from Rai-/- mice, we demonstrated that Rai promotes T cell survival and affects cell metabolism under hypoxia. Upon exposure to hypoxia, Jurkat T cells expressing Rai show (a) higher HIF-1α protein levels; (b) a decreased cell death and increased Akt/extracellular-signal-regulated kinase phosphorylation; (c) a decreased expression of proapoptotic markers, including caspase activities and poly(ADP-ribose) polymerase cleavage; (d) an increased glucose and lactate metabolism; (e) an increased activation of nuclear factor-kB pathway. The opposite effects were observed in hypoxic splenocytes from Rai-/- mice. Thus, Rai plays an important role in hypoxic signaling and may be relevant in the protection of T cells against hypoxia.

Inhibition of protein kinase R protects against palmitic acid-induced inflammation, oxidative stress, and apoptosis through the JNK/NF-kB/NLRP3 pathway in cultured H9C2 cardiomyocytes.

Double-stranded RNA-dependent protein kinase (PKR) is a critical regulator of apoptosis, oxidative stress, and inflammation under hyperlipidemic and insulin resistance conditions. Saturated free fatty acids, such as palmitic acid (PA), are known inducers of apoptosis in numerous cell types. However, the underlying molecular mechanism is not fully understood. The aim of the present study was to examine the effect of PA on cultured rat H9C2 cardiac myocytes cells and to investigate the PKR mediated harmful effects of PA in vitro in cultured cardiomyocytes. PKR expression was determined by immunofluorescence and immunoblotting. Oxidative stress and apoptosis were determined by flow cytometry and assay kits. The expression of different gene markers of apoptosis, oxidative stress, and inflammation were measured by Western blot analysis and reverse transcription polymerase chain reaction. PKR expression, reactive oxygen species levels as well as apoptosis were increased in PA-treated cultured H9C2 cardiomyocytes. The harmful effects of PA were attenuated by a selective PKR inhibitor, C16. Moreover, we observed that upregulation of c-Jun N-terminal kinase (JNK), nuclear factor-kB (NF-kB) and NACHT, LRR and PYD domains-containing protein 3 (NLRP3) pathways is associated with increased expression of interleukin 6 and tumor necrosis factor-α in PA-treated cardiomyocytes and attenuation by a selective PKR inhibitor. Our study reports, for the first time, that PKR-mediated harmful effects of PA in cultured cardiomyocytes via activation of JNK, NF-kB, and NLRP3 pathways. Inhibition of PKR is one of the possible mechanistic approaches to inhibit inflammation, oxidative stress, and apoptosis in lipotoxicity-induced cardiomyocyte damage.

The role of NF-κB pathway in cancer inflammation of esophageal squamous carcinoma.

42 Background: Chronic inflammation plays an important role in tumorigenesis and tumor progression in human cancers. We aim to investigate the role of NF-kB in cancer inflammation of esophageal squamous cell carcinoma (ESCC). Methods: To generate M2-polarized macrophages, cells of human U937 monocyte cell line were treated with phorbol myristate acetate (PMA, 50 ng/ml) for 6 hours, and then cultured with PMA plus Th2 cytokines, IL-4 (20 ng/ml) and IL-13 (20 ng/ml), for another 18 hours. M2 phenotype was verified by flow cytometry and by cytokine profiling using enzyme-linked immunosorbent assay (ELISA). After co-culture with M2 macrophages, transcription nuclear factor-kB (NF-kB) activity was measured using quantitative polymerase chain reaction (Q-PCR), followed by reconfirmation with Western blot analysis for IkBα in KYSE-170 and -510 ESCC cell lines (kindly provided by Dr. Yutaka Shimada at Kyoto University, Japan). A selective inhibitor to NF-kB, Bay11-7082, was used to treat ESCC cell lines co-cultured with M2 macrophages, followed by cell proliferation, migration, invasion assays and vascular endothelial growth factor (VEGF) secretion by ELISA. The effect of Bay11-7082 (5 mg/kg) against growth of ESCC tumor was tested in xenografted tumors. Results: PMA plus Th2 cytokines treatment promoted differentiation of U937 cells into M2 macrophages. When treated with Bay11-7082, proliferation, migration, invasion and induction of VEGF expression was significantly inhibited in M2 macrophage co-cultured ESCC cells with a down-regulation of IkBα expression. Tumor growth was significantly increased in M2 macrophage co-cultured ESCC cells compared to that of the non-co-cultured controls, which was significantly retarded by treatment with Bay11-7082. Conclusions: NF-kB pathway was activated in ESCC cell lines co-cultured with M2 macrophages with an increase in cell proliferation, cell motility and angiogenic factor in vitro and tumor growth in vivo, which were significantly suppressed by a NF-kB inhibitor, Bay11-7082. These results suggest a role of M2 macrophage in promoting aggressiveness of ESCC cells, possibly through an activation of NF-kB pathway that may serve as a potential therapeutic target for ESCC.

POTENTIAL ROLE OF REDOX SENSITIVE SERINE KINASE PATHWAYS IN THE PATHOGENESIS OF INSULIN RESISTANCE IN RHEUMATOID ARTHRITIS

Objectives: Insulin resistance and oxidative stress have been reported in rheumatoid arthritis (RA). However, the molecular basis of insulin resistance in RA is not clearly understood. Recent studies have documented the role of redox sensitive serine kinase pathways in insulin resistance. The aim of the study is to investigate insulin resistance, oxidative stress andlymphocyte redox sensitive - nuclear factor - kB (NF-kB), c-Jun N-terminal kinase (JNK) and p38 mitogen activated protein kinase (p38MAPK) pathways in RA subjects. Methods: Twenty newly diagnosed RA patients and 20 healthy volunteers in the age group of 30-50 years were included in this study. Overnight fasting blood was collected from the subjects, red blood cells, lymphocytes and plasma were separated using Ficoll hypaque. Blood oxidative stress parameters, fasting glucose and insulin were estimated. Redox sensitive serine kinase pathways in lymphocytes also studied. Results: The fasting insulin and HOMA-IR were significantly (p<0.001) higher in RA subjects compared to control subjects. Lymphocyte and erythrocyte reduced glutathione, erythrocyte catalase activity and plasma total antioxidant capacity were significantly (p<0.001) lower in RA subjects. Plasma MDA levels were significantly (p<0.001) higher in RA subjects. Western blotting analysis shows activation of NF-kB pathway in the lymphocytes of RA subjects. There was no significant change in the lymphocyte p38MAPK and JNK pathways between RA and control subjects. Conclusion: Further studies which explores the role of redox sensitive serine kinase pathways on insulin action in target tissues of insulin will help in the identification of novel therapeutic target for insulin resistance and its complications in RA.

Thrombin Inhibits NF-κb and RhoA Pathways in Cytokine-Stimulated Vascular Endothelial Cells When EPCR Is Occupied by Protein C.

We recently demonstrated that the occupancy of endothelial protein C receptor (EPCR) by protein C switches the PAR-1-dependent signaling specificity of thrombin from a permeability-enhancing to a barrier-protective response in endothelial cells. To determine whether the occupancy of EPCR by protein C renders thrombin a protective enzyme, thus up-regulating the expression of signaling molecules in the antiinflammatory pathways, we investigated the effects of thrombin and thrombin receptor agonist peptides (TRAP) on TNF-a-stimulated HUVECs in the absence and presence of the catalytically inactive protein C-S195A by monitoring the expression of cell surface adhesion molecules (VCAM- 1, ICAM-1 and E-selectin), adhesion of neutrophils to cytokine-stimulated endothelial cells, regulation of the Rho family of small GTPases and the activation of nuclear factorkB (NF-kB) pathway. Analysis of the results indicates that both thrombin and TRAP initiate proinflammatory responses in endothelial cells, thus neither thrombin nor TRAP influenced the proinflammatory effects of TNF-a in the absence of the protein C mutant. Interestingly, however, the occupancy of EPCR by the protein C mutant switched the PAR-1-dependent signaling specificity of thrombin and TRAP, thus leading to inhibition of the expression of all three adhesion molecules as well as the binding of neutrophils to TNFa-stimulated endothelial cells. Furthermore, similar to activated protein C, both thrombin and TRAP activated Rac1 and inhibited the activation of RhoA and NF-kB pathways in response to TNF-a in cells pretreated with protein C-S195A. Based on these results we conclude that when EPCR is bound by its natural ligand protein C, the cleavage of PAR-1 by thrombin initiates antiinflammatory responses in vascular endothelial cells.

Abnormal growth factor/cytokine network in gastric cancer.

Gastric cancer cells express a broad spectrum of the growth factor/cytokine receptor systems that organize the complex interaction between cancer cells and stromal cells in tumor microenvironment, which confers cell growth, apoptosis, morphogenesis, angiogenesis, progression and metastasis. However, these abnormal growth factor/cytokine networks differ in the two histological types of gastric cancer. Importantly, activation of nuclear factor-kB pathway by Helicobacter pylori infection may act as a key player for induction of growth factor/cytokine networks in gastritis and pathogenesis of gastric cancer. Better understanding of these events will no doubt provide new approaches for biomarkers of diagnosis and effective therapeutic targeting of gastric cancer.