High Expression Of NF-κB Research Articles

Erhuang Formula ameliorates renal damage in adenine–induced chronic renal failure rats via inhibiting inflammatory and fibrotic responses

AimsThe present study aimed to evaluate the protective effects of Erhuang Formula (EHF) and explore its pharmacological mechanisms on adenine-induced chronic renal failure (CRF). Materials and methodsThe compounds in EHF were analyzed by HPLC/MS. Adenine-induced CRF rats were administrated by EHF. The effects were evaluated by renal function examination and histology staining. Immunostaining of some proteins related cell adhesion was performedin renal tissues, including E-cadherin, β-catenin, fibronectin and laminin. The qRT-PCR was carried out determination of gene expression related inflammation and fibrosis including NF-κB, TNF-α, TGF-β1, α-SMA and osteopontin (OPN). ResultsTen compounds in EHF were identified including liquiritigenin, farnesene, vaccarin, pachymic acid, cycloastragenol, astilbin, 3,5,6,7,8,3′,4′-heptemthoxyflavone, physcion, emodin and curzerene. Abnormal renal function and histology had significant improvements by EHF treatment. The protein expression of β-catenin, fibronectin and laminin were significantly increased and the protein expression of E-cadherin significantly decreased in CRF groups. However, these protein expressions were restored to normal levels in EHF group. Furthermore, low expression of PPARγ and high expression of NF-κB, TNF-α, TGF-β1, α-SMA and OPN were substantially restored by EHF treatment in a dose-dependent manner. ConclusionsEHF ameliorated renal damage in adenine-induced CRF rats, and the mechanisms might involve in the inhibition of inflammatory and fibrotic responses and the regulation of PPARγ, NF-κB and TGF-β signaling pathways.

Chronic intermittent hypoxia induces liver fibrosis in mice with diet-induced obesity via TLR4/MyD88/MAPK/NF-kB signaling pathways

Obstructive sleep apnea (OSA) is associated with nonalcoholic fatty liver disease (NAFLD), and causes chronic intermittent hypoxia (CIH) during sleep. Inflammation is associated with the development of metabolic complications induced by CIH. Research suggests that innate immune mechanisms are involved in the pro-inflammatory pathways of liver fibrosis. The purpose of this study was to investigate whether innate immune responses induce liver fibrosis, and to evaluate mechanisms underlying hepatic inflammation related to CIH in a murine diet-induced obesity (DIO) model. Inflammatory and oxidative stress markers, TLR4, MyD88, Toll/interleukin-1-receptor-domain-containing adaptor-inducing interferon-β (TRIF), I-κB, NF-κB, p38 MAPK, c-JNK, and ERK activation, were measured in the serum and liver. As a result, α1(I)-collagen mRNA was significantly higher in DIO mice exposed to CIH than in the control groups. CIH mice exhibited liver fibrosis and significantly higher protein expression of TLR4, MyD88, phosphorylated (phospho-) I-κB, and phospho-ERK1/2 activation in the liver, and higher expression of NF-κB than that in the controls. TRIF, p38 MAPK, and JNK activation did not differ significantly between groups. We conclude that CIH in DIO mice leads to liver fibrosis via TLR4/MyD88/MAPK/NF-kB signaling pathways.

Immunohistochemical characterization of early and advanced knee osteoarthritis by NF-κB and iNOS expression.

This study was performed to determine the differences in grade of synovitis and expression of NF-κB and iNOS in knee synovial membrane between early and advanced stage of osteoarthritis (OA). Thirty synovial membrane intra-operative biopsies of patients (ten controls, ten with early and ten with advanced OA according to Kellgren-Lawrence radiological score) were immunohistochemically (NF-κB and iNOS) and hystologically (Krenn synovitis score) analyzed and correlated to WOMAC clinical score and pain duration. Krenn synovitis score of patients with radiologically early OA was significantly higher than in patients with advanced OA (p < 0.001). NF-κB expression in both synovial intima (p < 0.001) and subintima (p < 0.001) was also higher in early OA. iNOS expression in subintima was significantly higher in early than in advanced OA (p < 0.001), while in intima iNOS showed no statistical difference between groups (p = 0.07). The lymphocytic nodules, located in synovial subintima, were significantly higher in advanced OA when compared to early OA (p = 0.006) and the control group (p < 0.001). These results suggest that in early OA, there is a localized inflammation of the synovial membrane with high expression of NF-κB and iNOS. In advanced OA, number of expressed factors is reduced, with the exception of intima cells that highly express iNOS, reflecting the ongoing localized inflammatory process of lower degree. In advanced OA, the density of the resident cells is reduced and lymphocytic nodules appear, confirming the important role of adaptive immunity in later OA stage. Clinical significance of this study is better understanding possibilities of preventive measures for synovitis and OA advancement. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1990-1997, 2017.

Collagen gel protects L929 cells from TNFα-induced death by activating NF-κB

ABSTRACTPurpose: Type I collagen is one of the most abundant components of extracellular matrix. We previously illustrated that murine fibrosarcoma L929 cells grew well on type I collagen gel and escaped from TNFα-induced cell death. In this study, we investigated the mechanism underlying the protective effect of collagen gel. Material and methods: We used western blot, confocal microscopy, MTT assay and flow cytometry by introducing fluorescence staining to determine the expression levels of nuclear factor kappa B (NF-κB), inhibitory ratio and autophagy. Results: L929 cells on collagen gel showed higher expression of NF-κB in the nucleus. Inhibition of NF-κB with pyrrolidine dithiocarbamate hydrochloride (PDTC) or knockdown by NF-κB-siRNA canceled the protective effect of collagen gel on L929 cells from TNFα-induced death, suggesting for the role of NF-κB in the protection from cell death. We found a new aspect of the effect of PDTC on L929 cells cultured on collagen gel. PDTC alone without TNFα induced apoptosis in the L929 cells cultured on collagen gel but not the cells on plastic dish. The apoptosis induction of the L929 cells cultured on collagen gel with PDTC was repressed by inhibiting autophagy with chloroquine, an autophagy inhibitor, suggesting that autophagy contributes to the death induced by the treatment with PDTC. Possible underlying mechanism of this finding is discussed. Conclusion: NF-κB played an important role in protecting the L929 cells cultured on collagen gel from TNFα-induced death.

Expression of Btk and NFκB in Acute Lymphoblastic Leukemia Cells and Its Significance

To evaluate the role of Btk and NFκB in the incidence, development, prognosis and therapeutic efficacy for acute lymphoblastic leukemia(ALL) through detecting their expression in leukemia cells. Bone marrow samples from 51 ALL patients were collected, and the mononuclear cells were separated by Ficoll density gradient centrifugation. The expressions of Btk and NFκB at RNA and protein levels were detected by RT-PCR and Western blot respectively. (1)At protein level, Btk and NFκB were expressed in all newly diagnosed 51 ALL patients, among them 38 patients had higher expression level of Btk, 34 patients had high NFκB expression level. The expression of Btk and NFκB was higher in the cells from newly diagnosed ALL patients than that in the cells from patients in CR(P<0.05), and the expression of Btk and NFκB was higher in relapsed ALL patients. (2)The expression of Btk and NFκB in the ALL patients was followed-up higher expression of Btk: among the 38 newly diagnosed B-ALL cases, 27 experienced CR (71%) and 12 of which achieved CR after one course chemotherapy (one course CR) (31%). Moreover, 16 out of the 27 CR patients relapsed after a short period (less than 6 months) (59%). On the contrary, among the 13 patients with low Btk expression, 11 achieved CR (84.6%) after one course and 1 relapsed (8 months after CR) (7.6%). A similar pattern of NFκB expression was observed. Btk and NFκ B may play an important role in the incidence and progression of ALL, possibly serving as the potential therapeutic targets of ALL and the indexes for prognosis.

丹蛭降糖胶囊对糖尿病大鼠肾组织NF-κB表达的影响*

Objective: To investigate the expression level of NF- κ B in renal tissue of diabetic rodent model and the regulating action of Danzhi Jiangtang Capsule (DJC) . Methods : 55 male SD rats were randomly divided into 2 groups, normal control group and experimental group. Normal control group was fed with basal diet, the experimental group were fed with high calorie diet. A single intraperitoneal injection of streptozotocin (STZ) of 35mg/kg to establish the animal model of diabetes mellitus, blood glucose was measured after 120h,when the diabetic rodent model was built, the experimental group were randomly divided into 4 groups, high does group of DJC, low does group of DJC , pioglitazone group and model group. In addition to the model group and control group, each group was kept with continuous intragastric administration of drugs for 8 weeks, high does group of DJC, low does group of DJC were treated with 1.08g/ (kg • d), 0.54 g/ (kg • d) by gavage, pioglitazone group was treated with 10mg/ (kg • d) by gavage, the blank group and the model group were treated with physiological saline 5ml/ (kg • d) by gavage. Then determined NF-κB by immunohistochemical method and detected blood glucose of rats . Results: The expression of NF-κB in model group was strongly and significantly increased compared with the normal control group （ P ＜0.05 or P ＜0.01）; after being given Danzhi Jiangtang Capsule for 8 weeks , high expression of NF-κB in high DJC group and low DJC group were significantly decreased ( P ＜0.05), much more than pioglitazone group . When the diabetic rodent model was built, the blood glucose of model group was increased significantly compared with the control group ( P ＜0.01).After the experimental group been given Danzhi Jiangtang Capsule for 2 weeks , the level of glucose was decreased significantly compared with model group ( P ＜0.01) , the blood glucose index declined steadily after 4~8 weeks（ P ＜0.01）. Conclusion : Danzhi Jiangtang Capsule can reduce blood sugar ,which delay the progression of diabetic nephropathy through inhibiting the high expression of NF- κ B in diabetic rats.

Role of immune deviation by toll-liked receptor's doping LPS in pathogenesis of allergic rhinitis

To investigate the role of TLR-NF-κB signaling pathway in pathogenesis of allergic rhinitis (AR) and the mechanism of TLR to modulate innate immunity and adaptive immunity. One hundred rats were divided into 5 groups by simple randomization, normal group(group A), modle group(group B), AR+LPS20 group(group C), AR+LPS10 group(group D), AR+LPS5 group(group E). Model of AR in B group was established by intraperitoneal injection and nasal topic delivery of ovalbumin (OVA). A group was delivered of same volume physiological saline instated of OVA, C,D,E group were interfered by nasal delivery of LPS in different concentration (including LPS 20 µg, 10 µg, 5 µg per 100 µl). Changes of nasal mucosa tissues and inflammatory cell infiltration were observed by HE staining, while neutrophil and eosinophil counted under high power microscope.Expression of IL-4, IFN-γ, and IgE in nasal mucosa tissues were measured with immunohistochemical method.Realtime-PCR and Western-blot were used to evaluate the expression level of TLR-4 and NF-κB in nasal mucosa tissues.SPSS 13.0 software was used to analyze the data. Group B was observed to have developed AR injury of nasal mucosa. Eosinophil count and the expression of IL-4, IFN-γ, and IgE were significantly higher in B group than those in A group (all P < 0.05), neutrophil count was significantly higher in C, D, E groups than that in B group (all P < 0.05). of immunohistochemical staining showed that, expression level of IFN-γ, TLR-4 and NF-κB were significantly higher than group B (all P < 0.05), while IL-4 and IgE were significantly decreased than group B (all P < 0.05) . The protein expression of TLR-4 and NF-κB was 0.888 9 ± 0.032 9 and 0.913 3 ± 0.031 1 in group C, and 0.419 2 ± 0.038 0 and 0.447 8 ± 0.033 0 in group A, 0.616 1 ± 0.025 1 and 0.748 1 ± 0.034 3 in group B, the difference was significant(all P < 0.05). TLR plays an important role of modulation between innate immunity and adaptive immunity in the pathogenesis of AR. The higher concentration of TLR doping may activate the higher expression of NF-κB then intervene the development of AR with immune deviation.

Clinical significance of nuclear factor κB and chemokine receptor CXCR4 expression in patients with diffuse large B-cell lymphoma who received rituximab-based therapy.

Background/AimsThis study investigated the expression of nuclear factor κB (NF-κB) and the chemokine receptor (CXCR4) in patients with diffuse large B-cell lymphoma (DLBCL) who received rituximab-based therapy.MethodsSeventy patients with DLBCL and treated with rituximab-CHOP (R-CHOP) were included, and immunohistochemistry was performed to determine the expression of NF-κB (IκB kinase α, p50, and p100/p52) and CXCR4. To classify DLBCL cases as germinal center B-cell-like (GCB) and non-GCB, additional immunohistochemical expression of CD10, bcl-6, or MUM1 was used in this study. The expression was divided into two groups according to the intensity score (negative, 0 or 1+; positive, 2+ or 3+).ResultsThe median age of the patients was 66 years (range, 17 to 87), and 58.6% were male. Twenty-seven patients (38.6%) had stage III or IV disease at diagnosis. Twenty-three patients (32.9%) were categorized as high or high-intermediate risk according to their International Prognostic Indexs (IPIs). The overall incidence of bone marrow involvement was 5.7%. Rates of positive NF-κB and CXCR4 expression were 84.2% and 88.6%, respectively. High NF-κB expression was associated with CXCR4 expression (p = 0.002), and 56 patients (80.0%) showed coexpression. However, the expression of NF-κB or CXCR4 was not associated with overall survival and EFS. On multivariate analysis that included age, gender, performance status, stage, and the IPI, no significant association between the grade of NF-κB or CXCR4 expression and survival was observed.ConclusionsThe current study suggests that the tissue expression of NF-κB and CXCR4 may not be an independent prognostic marker in DLBCL patients treated with R-CHOP.

Tumor refractoriness to endostatin anti-angiogenesis is associated with the recruitment of CD11b+Gr1+ myeloid cells and inflammatory cytokines

A major challenge in developing antiangiogenic therapies is tumor intrinsic refractoriness and the emergence of treatment-induced resistance. Recently, such resistance is considered to be associated with inflammatory changes in the tumor microenvironment. However, no information has been acquired about the effect of endostatin on tumor microenvironment in this field. We established two tumor models refractory to endostatin treatment and sought to determine the role of inflammatory changes in the development of tumor refractoriness to antiangiogenic therapy. Three xenograft tumor murine models were treated with low-dose endostatin or high-dose endostatin for 10 days. The effect of endostatin on tumor growth was observed, and tumors refractory to endostatin treatment were defined. Flow cytometry were carried out to assess the presence of CD11b+Gr1+ myeloid cells in the peripheral blood and in the tumor. Inflammatory cytokine levels in peripheral blood were measured using the enzyme-linked immunosorbent assay. The expression of NF-κB, versican and hypoxia-inducible factor-1α in the tumor was evaluated using immunohistochemistry. LLC and B16F1 tumors were defined as animal models of refractoriness to endostatin treatment. CD11b+Gr1+ myeloid cells were inherently recruited into the peripheral blood and the tumor microenvironment in the LLC tumor-bearing mice, and levels of serum G-CSF and TNF-α were increased along with the progression of tumor growth. In the B16F1 tumor-bearing mice, CD11b+Gr1+ myeloid cells were acquiredly recruited by endostatin into the peripheral blood and the tumor microenvironment. Additionally, high levels of G-CSF and TNF-α in serum and high expression of NF-κB, versican and hypoxia-inducible factor-1α in tumor tissue were found in B16F1 tumor-bearing mice after endostatin administration. A tumor can grow inherently or acquiredly with refractoriness to endostatin treatment in vivo. Recruitment of CD11b+Gr1+ myeloid cells and inflammatory cytokines may play an important role in the development of tumor refractoriness to endostatin anti-angiogenesis.

PI3K-Akt/PKB signaling pathway in neutrophils and mononuclear cells exposed to N-nitrosodimethylamine

Neutrophils (PMN) play diverse regulatory and effector functions in the immune system through the release of reactive nitrogen species, including nitric oxide (NO). The enzyme responsible for NO synthesis in PMN is inducible nitric oxide synthase (iNOS) that is regulated by various signaling pathways, e.g. PI3K-Akt/PKB, and transcription factors. N-Nitrosodimethylamine (NDMA), a xenobiotic widespread in the human environment, affects immune cells. The study objective here was to examine the role of the PI3K-Akt/PKB pathway in induction of NO synthesis (with involvement of iNOS) in human PMN, as well as in autologous mononuclear cells (PBMC), exposed to NDMA. Isolated cells were incubated for 2 h with a sub-lethal dose of NDMA and then the expression of several select proteins in the cell cytoplasmic and nuclear fractions were determined by Western blot analyses. The results indicated that NDMA enhanced expression of iNOS, phospho-PI3K, and phospho-IκBα in the cytoplasmic fraction of the PMN and PBMC. The nuclear fraction of these cells also had a higher NF-κB expression. Moreover, in PMN, NDMA caused an increased expression of phospho-Akt (T308), phospho-Akt (S473), and phospho-IKKαβ in the cytoplasm, and c-Jun and FosB in the nuclear fraction. Blocking of PI3K caused a decrease in expression of all these proteins in NDMA-exposed PMN. However, inhibition of PI3K led to a drop in expression of iNOS, phospho-PI3K, and phospho-IκBα in the cytoplasm, and in NF-κB in the nuclear fraction, of PBMC. The results of these studies indicated to us that NDMA activates the PI3K-Akt/PKB pathway in human PMN and that this, in turn, contributes to the activation of transcription factors NF-κB, c-Jun, and FosB involved in NO production (through modulation of iNOS expression).

NF‐κBP65 promotes invasion and metastasis of oesophageal squamous cell cancer by regulating matrix metalloproteinase‐9 and epithelial‐to‐mesenchymal transition

NF-κB has been recognized as one of the factors responsible for the development of cancer; however, the mechanism by which high expression of NF-κB contributes to the progression of human oesophageal squamous cell cancer (ESCC) is not fully understood. In our investigations, NF-κBP65 was overexpressed in human ESCC tissues, especially in ESCC tissues with deep invasion and lymph node metastasis. Suppression of NF-κBP65 by siRNA decreased the invasion and proliferation ability of EC9706 cells in vitro. Furthermore, siRNA-mediated NF-κBP65 knock-down could lead to the downregulation of MMP-9, a metastasis-related gene. Reduced E-cadherin is a hallmark of invasive carcinomas that have acquired epithelial-mesenchymal transition (EMT) phenotypes and Vimentin is another molecule that is used widely as a marker of the EMT. We found upregulation of E-cadherin expression and downregulation of Vimentin was induced by NF-κBP65 siRNA, which suggests that NF-κBP65 siRNA could inhibit the invasion and proliferation ability of ECSS through attenuating the expression of MMP-9 and EMT. Thus, ESCC NF-κBP65 could be a useful target for cancer prevention and therapy.

Abstract 1298: The role of nuclear factor kappa B in survival of human papillomavirus-positive head and neck cancer cell lines.

Abstract Nuclear factor kappa B (NFκB) has been implicated in the development of head and neck squamous cell carcinoma (HNSCC) from premalignant changes to tumor progression, and in treatment resistance and recurrence. However, the roles of NFκB in the sensitivity to chemoradiation of human papillomavirus (HPV)-associated HNSCC have not been well defined. Previously we reported that high expression of NFκB detected by immunohistochemistry in pretreated HPV+ oropharyngeal cancer patients was significantly associated with poor progression free survival (AACR 2012 Abstract #4489). Therefore, we hypothesized that NFκB could play a role in pro-survival in HPV+ head and neck cancer cells. We first examined the sensitivity of five HPV+ head and neck cancer cell lines (HPV16: UDSCC2, 93-VU-147T, UM-SCC47, UPCI:SCC90, HPV18: SCC1483) to radiation and cisplatin, Consistent with current models, when we knocked down expression of the HPV16E6 oncogene, SCC47 and SCC1483 cells with restored expression of p53 displayed a significant reduction in S phase entry as revealed by cell cycle analysis (SCC47: scrambled control 14.08% ± 0.55% versus siRNA-E6 6.39% ± 2.40, p=0.004; SCC1483: scrambled control 13.39% ± 1.28% versus 7.49% ± 0.83, p=0.002). In addition, all five cell lines displayed reduced sensitivity to cisplatin compared with cells treated with scrambled control siRNA. The basal level of activated NFκB in HPV+ cell lines was correlated with the sensitivity to radiation and cisplatin as assessed by colony formation and cell growth assays. Suppression of NFκBp65 by siRNA resulted in marked increases in apoptosis associated with reduction of Bcl-2 in both SCC47 and SCC1483 cells. This pattern was more pronounced with increasing doses of the proteasome inhibitor bortezomib (32.5 to 130 nM for 48 to 72 hr). Furthermore, SCC47 and SCC1483 cells treated with siRNA NFκBp65 displayed a significantly greater level of apoptosis induced by radiation or cisplatin compared to cells treated with non-targeted control siRNA, indicating a pro-survival effect for NFκB. Further studies are warranted to delineate the effects of HPV oncogene interactions with NFκB signaling pathways on chemoradiation sensitivity. (This study was supported by the V Foundation). Citation Format: Hongzheng Zhang, Zhongliang Hu, Ning Jiang, Rossella Marullo, Paul W. Doetsch, William D. Hunt, Georgia Z. Chen, Dong M. Shin. The role of nuclear factor kappa B in survival of human papillomavirus-positive head and neck cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1298. doi:10.1158/1538-7445.AM2013-1298

NFkB activation drives mesenchymal transformation and susceptibility to calcification in aortic valve endothelial cells

Treatment of calcific aortic valve disease (CAVD) remains difficult due to limited understanding of the initial pathological activators. The early inflammation stage is known to stimulate an endothelial to mesenchymal transition (EndMT) of valve endothelial cells (VEC), but the role of the transformed cells in later disease is unknown. Characterization of EndMT VEC revealed high expression of NFκB, α-SMA, Snai1, MMP-9, and extracellular matrix remodeling capabilities. Transfection of VEC with NFκB over-expression and inhibition plasmids showed that NFκB is sufficient and necessary to induce EndMT. Further, when cultured in an osteogenic environment, VEC+NFκB had increased α-SMA (50.4+/− 3.1 fold), osteocalcin (16.1+/−1.8 fold) runx-2 (14.7+/−.8 fold), and calcium deposition compared to controls. In diseased human valves, cells with activated NFkB localized near calcific nodules and co-expressed CD31 and α-SMA, suggesting EndMT origin. Together, these results suggest that transformed VEC are key players in aortic valve calcification and NFκB-inhibition is an attractive valve specific intervention strategy in the early stages of CAVD. Funding: NIH 1R01HL110328–01

Expression of inflammatory factors on the vessel wall of patients with lower extremity arterial occlusive disease

Objective To analyze the expression of nuclear factor-κB (NF-κB) and platelet derived growth factor (PDGF) on the intima and media by using quantitative analysis,and investigate the changes in the expression patterns of NF-κB and PDGF on the vessel wall of patients with lower extremity arterial occlusive disease.Methods Nineteen patients with lower extremity arterial occlusive disease served as study group,and 3 patients with other diseases as control group,respectively.The expression of NF-κB and PDGF on the intima and media of the resected arteries was detected by using real-time polymerase chain reaction (RT-qPCR).Results The mRNA expressions of NF-κB (35.42 ±5.69),PDGF-A (34.38 ±5.80) and PDGF-B (33.95 ± 5.92) on the vessel wall of study group was significantly higher than that in control group (P ＜0.05).There was a positive linear correlation between NF-κB and PDGF-A (r=0.926) or between NF-κB and PDGF-B (r=0.925,P＜0.05).Conclusion There was higher expression of NF-κB and PDGF on the atherosclerotic vesssel walls of patients with lower extremity arterial occlusive disease,and a positive linear correlation between NF-κB and PDGF. Key words: Lower extremity arterial occlusive disease; Nuclear factor-κB; Platelet-derived growth factor

The biological and clinical impact of inhibition of NF‐κB‐initiated apoptosis in diffuse large B cell lymphoma (DLBCL)

NF-κB is frequently over-expressed in a variety of non-Hodgkin's lymphomas (NHLs) and has been implicated in lymphomagenesis; however, its role in diffuse large B cell lymphoma (DLBCL) as a prognostic biomarker has not been fully elucidated. Therefore, we investigated the role of NF-κB and its association with clinicopathological features in a tissue microarray cohort of 230 DLBCL patient samples. We then elucidated the role of NF-κB inhibition on cell viability and apoptosis in vitro, using DLBCL cell lines. Using immunohistochemistry, NF-κB was detected in 25.6% (52/203) DLBCL tumours, was associated with activated B cell (ABC) phenotype (p = 0.0054), Epstein-Barr virus (EBV; p = 0.0080) and over-expression of the anti-apoptotic marker XIAP (p = 0.0013). DLBCL cases with nuclear expression of NF-κB showed a significantly poorer overall survival as compared to those without NF-κB expression (p = 0.0236). In a multivariate analysis using a Cox proportional hazard model for IPI and NF-κB expression, the relative risk was 2.97 for high NF-κB expression (95% CI 1.27-6.94; p = 0.0113) and 7.55 for the high-IPI group (95% CI 3.34-18.35; p < 0.0001). In vitro, Bay 11-7085 inhibited constitutively active NF-κB expression in a dose-dependent manner and inhibition of NF-κB also down-regulated expression of the downstream target gene products Bcl-2, Bcl-XL (BCL2L1), XIAP and Survivin, leading to apoptosis via activation of the mitochondrial apoptotic pathway. NF-κB over-expression was found to be an independent prognostic marker for poor survival in DLBCL. Altogether, these results suggest that NF-κB may be a useful prognostic biomarker and a potential target for therapeutic intervention in DLBCL.

Evaluation of nuclear factor κB and chemokine receptor CXCR4 co‐expression in patients with prostate cancer in the Radiation Therapy Oncology Group (RTOG) 8610

To determine the frequency of nuclear factor κB (NFκB) and the chemokine receptor CXCR4 co-expression in prostate cancer specimens from men with locally advanced disease. Paraffin-embedded samples from patients enrolled on the Radiation Therapy Oncology Group (RTOG) 8610 trial underwent immunohistochemical staining for NFκB and CXCR4. The amount of NFκB and CXCR4 was scored by a 'blinded' pathologist for the percentage of cells stained (0-100%) and staining intensity (0-3 +). Cox proportional hazard models were used for overall survival and disease-free survival to examine if NFκB and/or CXCR4 expression were associated with patient outcomes with and without adjustment for covariates. Available material and successful staining allowed NFκB and CXCR4 status to be determined for 55 and 63 patients, respectively. Both NFκB and CXCR4 status were available for 51 patients. Of these, 53% were 2/3 + for cytoplasmic NFκB staining and 56% were 2/3 + for CXCR4. In all, 18 of the 51 patients were 2/3 + for both NFκB and CXCR4 (P = 0.129). Ten of 11 patients with 3 + NFκB had 2/3 + CXCR4 (P= 0.004). In this small study, neither NFκB nor CXCR4 were associated with prostate cancer outcomes. High NFκB expression is associated with CXCR4 expression and they are co-expressed in about one third of patients with clinically localized prostate cancer. Larger studies to accurately determine the frequency of co-expression and prognostic utility of NFκB and CXCR4 alone and in combination are warranted.

Nuclear factor-kappa B pathway and response in a phase II trial of bortezomib and docetaxel in patients with recurrent and/or metastatic head and neck squamous cell carcinoma

BackgroundOur previous study has shown that nuclear factor-kappa B (NF-κB)-signaling pathway was associated with a higher rate of recurrence in head and neck squamous cell carcinoma (HNSCC). The combination of bortezomib, an NF-κB inhibitor by inhibition of proteasomes, plus docetaxel was assessed for efficacy and toxicity. Materials and methodsPatients with recurrent and/or metastatic HNSCC were enrolled on a phase II bortezomib/docetaxel trial (bortezomib 1.6 mg/m2 and docetaxel 40 mg/m2 on days 1 and 8 of a 21-day cycle). Response was assessed using RECIST. Tissue specimens were evaluated for the presence of human papillomavirus (HPV) and expression of NF-κB-associated genes. ResultsTwenty-one of 25 enrolled patients were assessable for response; one partial response (PR, 5%), 10 stable disease (SD, 48%) and 10 progressive disease (PD, 48%). Patients with PR/SD had significantly longer survival compared with patients with PD and the regimen was well tolerated. Only one of 20 tumors was positive for HPV. Patients with PD had higher expression of NF-κB and epidermal growth factor receptor-associated genes in their tumors by gene expression analysis. ConclusionFurther understanding of treatment resistance and interactions between bortezomib and docetaxel may provide novel approaches in managing HNSCC.

The changes of nuclear factor-κB expression in perihematoma region and brain edema of experimental in tracerebral hemorrhage rats after local mild to moderate hypothermia

Objective To investigate the effects of local mild to moderate hypothermia on brain edema and nuclear factor-κB (NF-κB) expression in the perihematoma region of experimental intracerebral hemorrhage rats.Methods A total of 144 Wistar rats were randomly divided into a normothermia group and a mild to moderate hypothermia group.The two groups each comprised control,6 h intracerebral hemorrhage,24 h intracerebral hemorrhage,48 h intracerebral hemorrhage,72 h intracerebral hemorrhage,and 1 week intracerebral hemorrhage subgroups,with 12 rats in each subgroup.Rat models of intracerebral hemorrhage were established by injection of autologous blood into the rat candate nucleus.Rats in the mild to moderate hypothermia group were subject to local mild to moderate hypothermia for 4 hours immediately after the injection,with the intracerebral temperature maintained at (33.0±0.5)℃,while those in the normothermia were maintained with a body temperature of 37℃.The changes of brain water content were measured and the expression of NF-κB in perihematoma region of intracerebral hemorrhage was observed with immunohistochemistry and Western blot methods. Results The brain edema and the NF-κB expression peaked at 48 h after intracerebral hemorrhage.There was obvious difference in terms of brain edema between the normothermia group and the mild to moderate hypothermia group (P＜0.05).The NF-κB expression reduced significandy in the mild to moderate hypothermia group compared with the normothermia group at the 24th,48th and 72nd hours,as well as 1 week after intracerebral hemorrhage (P＜0.05). Conclusions The severity of brain edema was associated with high expression of NF-κB.Treatment with local mild to moderate hypothermia can significantly inhibit overexpression of NF-κB and relieve brain edema following intracerebral hemorrhage. Key words: Intracerebral hemorrhage; Brain edema; Mild to moderate hypothermia; Nuclear factor-κB

Over-expression of the A3 adenosine receptor in tumor and PBMC of hepatocellular carcinoma patients: A specific therapeutic target

14085 Background: The Gi protein associated A3 adenosine receptor (A3AR) is highly expressed in colon carcinoma tissue and is reflected in peripheral blood mononuclear cells (PBMC) of the patients. Drug candidates with high affinity to the receptor possess anti-cancer effect when administered orally to colon carcinoma bearing animals. The receptor was suggested as a potent therapeutic target and predictive biological marker. Since there is no adequate therapy for Hepatocellular Carcinoma (HCC) we looked at the presence of A3AR in tumor and PBMC of HCC patients and tested the efficacy of the A3AR agonist, CF102 in an experimental rat HCC model. Methods: mRNA and protein A3AR expression level were examined in paraffin embedded slides and PBMC from HCC patients utilizing RT-PCR and Western blot (WB) analysis, respectively. Rat HCC model was established by inoculating N1S1 cells to the animals’ liver. Oral treatment with CF102 was initiated 5 days after tumor inoculation. Signaling proteins in tumor lesions and PBMC from the animals was tested by WB analysis. Results: A3AR over-expression was found in tumor and PBMC of HCC patients. Receptor over-expression was attributed to high expression of NF-κB, known to act as a transcription factor of A3AR. In the HCC bearing rats, A3AR was over-expressed in the tumor tissue and PBMC. Upon treatment with CF102 A3AR expression was down-regulated followed by inhibition of tumor size (60%±18.4%, p&lt;0.05). Mechanistically, CF102 de-regulated the NF-κB and Wnt signaling pathways. Conclusions: A3AR which is up- regulated in HCC tumors may be suggested as a new target to treat this malignancy and as a biological marker to follow up tumor response to the drug. No significant financial relationships to disclose.

Phosphoprotein levels, MAPK activities and NFκB expression are affected by fisetin

Flavonoids, polyphenolic phytochemicals, are ubiquitous in plants and are commonly present in the human diet. They may exert diverse beneficial effects, including antioxidant and anticarcinogenic activities. The present study was designed to evaluate three biomolecules that play important roles in the apoptotic process: mitogen-activated protein kinases, protein phosphatases and NFκB, using HL60 cells treated with fisetin as an experimental model. Our results demonstrated that cells treated with fisetin presented high expression of NFκB, activation of MAPK p38 and an increase of phosphoprotein levels; inhibition of enzymes involved in redox status maintenance were also observed. Our findings reinforce the hypothesis that fisetin is likely to exert beneficial and/or toxic actions on cells not through its potential as antioxidant but rather through its modulation of protein kinase and phosphatase signaling cascades. Additionally, our results also indicate that the cellular effects of fisetin will ultimately depend on the cell type and on the extent to which they associate with the cells, either by interactions at the membrane or by uptake into the cytosol.