NF-κB Mediates Tumor Necrosis Factor α-Induced Expression of Optineurin, a Negative Regulator of NF-κB

Optineurin is a ubiquitously expressed multifunctional cytoplasmic protein encoded by OPTN gene. The expression of optineurin is induced by various cytokines. Here we have investigated the molecular mechanisms which regulate optineurin gene expression and the relationship between optineurin and nuclear factor κB (NF-κB). We cloned and characterized human optineurin promoter. Optineurin promoter was activated upon treatment of HeLa and A549 cells with tumor necrosis factor α (TNFα). Mutation of a putative NF-κB-binding site present in the core promoter resulted in loss of basal as well as TNFα-induced activity. Overexpression of p65 subunit of NF-κB activated this promoter through NF-κB site. Oligonucleotides corresponding to this putative NF-κB-binding site showed binding to NF-κB. TNFα-induced optineurin promoter activity was inhibited by expression of inhibitor of NF-κB (IκBα) super-repressor. Blocking of NF-κB activation resulted in inhibition of TNFα-induced optineurin gene expression. Overexpressed optineurin partly inhibited TNFα-induced NF-κB activation in Hela cells. Downregulation of optineurin by shRNA resulted in an increase in TNFα-induced as well as basal NF-κB activity. These results show that optineurin promoter activity and gene expression are regulated by NF-κB pathway in response to TNFα. In addition these results suggest that there is a negative feedback loop in which TNFα-induced NF-κB activity mediates expression of optineurin, which itself functions as a negative regulator of NF-κB.     

NF-κB p100 limits TNF-induced bone resorption in mice by a TRAF3-dependent mechanism

TNF and RANKL mediate bone destruction in common bone diseases, including osteoarthritis and RA. They activate NF-κB canonical signaling directly in osteoclast precursors (OCPs) to induce osteoclast formation in vitro. However, unlike RANKL, TNF does not activate the alternative NF-κB pathway efficiently to process the IκB protein NF-κB p100 to NF-κB p52, nor does it appear to induce osteoclast formation in vivo in the absence of RANKL. Here, we show that TNF limits RANKL- and TNF-induced osteoclast formation in vitro and in vivo by increasing NF-κB p100 protein accumulation in OCPs. In contrast, TNF induced robust osteoclast formation in vivo in mice lacking RANKL or RANK when the mice also lacked NF-κB p100, and TNF-Tg mice lacking NF-κB p100 had more severe joint erosion and inflammation than did TNF-Tg littermates. TNF, but not RANKL, increased OCP expression of TNF receptor–associated factor 3 (TRAF3), an adapter protein that regulates NF-κB p100 levels in B cells. TRAF3 siRNA prevented TNF-induced NF-κB p100 accumulation and inhibition of osteoclastogenesis. These findings suggest that upregulation of TRAF3 or NF-κB p100 expression or inhibition of NF-κB p100 degradation in OCPs could limit bone destruction and inflammation-induced bone loss in common bone diseases.     

Lansoprazole, a Proton Pump Inhibitor, Suppresses Production of Tumor Necrosis Factor-α and Interleukin-1β Induced by Lipopolysaccharide and Helicobacter Pylori Bacterial Components in Human Monocytic Cells via Inhibition of Activation of Nuclear Factor-κB and Extracellular Signal-Regulated Kinase

Pathogenic bacterial components play critical roles in initiation of gastrointestinal inflammation via activation of intracellular signaling pathways which induce proinflammatory cytokines such as tumor necrosis factor (TNF)-α and interleukin (IL)-1β. Lansoprazole (LANSO), a proton pump inhibitor, has been widely used for the treatment of peptic ulcers and reflux esophagitis due to its potent acid-suppressive effect. It has also been reported to have anti-inflammatory effects. In this study we investigated the effects of LANSO on the production of TNF-α and IL-1β induced by lipopolysaccharide (LPS) and Helicobacter pylori water-soluble extract (HpWE) in the human monocytic cell line (THP-1). LANSO (100 µM) significantly reduced mRNA expression and production of TNF-α and IL-1β by THP-1 cells stimulated by LPS and HpWE. LANSO inhibited phosphorylation and degradation of inhibitory factor κB-α (IκB-α) and phosphorylation of extracellular signal-regulated kinase (ERK) induced by LPS and HpWE in THP-1 cells. These findings suggest that LANSO exerts anti-inflammatory effects by suppressing induction of TNF-α and IL-1β via inhibition of nuclear factor (NF)-κB and ERK activation.     

High-sensitivity CRP discriminates HNF1A-MODY from other subtypes of diabetes

OBJECTIVE

Maturity-onset diabetes of the young (MODY) as a result of mutations in hepatocyte nuclear factor 1-α (HNF1A) is often misdiagnosed as type 1 diabetes or type 2 diabetes. Recent work has shown that high-sensitivity C-reactive protein (hs-CRP) levels are lower in HNF1A-MODY than type 1 diabetes, type 2 diabetes, or glucokinase (GCK)-MODY. We aim to replicate these findings in larger numbers and other MODY subtypes.

RESEARCH DESIGN AND METHODS

hs-CRP levels were assessed in 750 patients (220 HNF1A, 245 GCK, 54 HNF4-α [HNF4A], 21 HNF1-β (HNF1B), 53 type 1 diabetes, and 157 type 2 diabetes).

RESULTS

hs-CRP was lower in HNF1A-MODY (median [IQR] 0.3 [0.1–0.6] mg/L) than type 2 diabetes (1.40 [0.60–3.45] mg/L; P < 0.001) and type 1 diabetes (1.10 [0.50–1.85] mg/L; P < 0.001), HNF4A-MODY (1.45 [0.46–2.88] mg/L; P < 0.001), GCK-MODY (0.60 [0.30–1.80] mg/L; P < 0.001), and HNF1B-MODY (0.60 [0.10–2.8] mg/L; P = 0.07). hs-CRP discriminated HNF1A-MODY from type 2 diabetes with hs-CRP <0.75 mg/L showing 79% sensitivity and 70% specificity (receiver operating characteristic area under the curve = 0.84).

CONCLUSIONS

hs-CRP levels are lower in HNF1A-MODY than other forms of diabetes and may be used as a biomarker to select patients for diagnostic HNF1A genetic testing.Maturity-onset diabetes of the young (MODY) is a rare monogenic form of diabetes and is often misdiagnosed as type 1 diabetes or type 2 diabetes (1,2). A correct genetic diagnosis of MODY is important for predicting the clinical course of disease, risk to relatives, and optimizing treatment. Therefore, cheap and novel biomarkers that help identify these patients are desirable.The CRP gene has hepatocyte nuclear factor 1-α (HNF1A) binding sites in its promoter, and common variants in and around HNF1A are associated with circulating high-sensitivity C-reactive protein (hs-CRP) levels (3,4). Recently, Owen et al. (5) demonstrated that hs-CRP levels were reduced in 31 HNF1A-MODY patients compared with type 1 diabetes, type 2 diabetes, glucokinase (GCK) MODY, and normal control subjects, making it potentially a useful clinical test.We aim to replicate this initial study in a larger cohort of patients and assess whether the reduction in hs-CRP is also seen in patients with mutations of other genes encoding hepatic nuclear factors (hepatocyte nuclear factor 4-α [HNF4A] and hepatocyte nuclear factor 4-β [HNF1B]).     

IκBα Overexpression Delays Tumor Formation in v-rel Transgenic Mice

We have previously shown that transgenic mice expressing the oncoprotein v-Rel under the control of a T cell–specific promoter develop T cell lymphomas. Tumor formation was correlated with the presence of p50/v-Rel and v-Rel/v-Rel nuclear κB-binding activity. Since experimental evidence has led to the suggestion of a potential tumor suppressor activity for IκBα, we have studied the role of IκBα in the transforming activity of v-Rel by overexpressing IκBα in v-rel transgenic mice. Overexpression of IκBα in v-rel transgenic mice resulted in an extended survival, and the development of cutaneous T cell lymphomas of CD8⁺CD4⁻ phenotype. These phenotypic alterations were associated with a dramatic reduction of p50/v-Rel, but not v-Rel/v-Rel nuclear DNA binding activity and an increased expression of the intercellular adhesion molecule 1. Our results indicate that v-Rel homodimers are active in transformation and that the capacity of v-Rel–containing complexes to escape the inhibitory effect of IκBα may be a key element in its transforming capability.     

Rcan1 negatively regulates FcεRI-mediated signaling and mast cell function

Aggregation of the high affinity IgE receptor (FcεRI) activates a cascade of signaling events leading to mast cell activation. Subsequently, inhibitory signals are engaged for turning off activating signals. We identified that regulator of calcineurin (Rcan) 1 serves as a negative regulator for turning off FcεRI-mediated mast cell activation. FcεRI-induced Rcan1 expression was identified by suppression subtractive hybridization and verified by real-time quantitative polymerase chain reaction and Western blotting. Deficiency of Rcan1 led to increased calcineurin activity, increased nuclear factor of activated T cells and nuclear factor κB activation, increased cytokine production, and enhanced immunoglobulin E–mediated late-phase cutaneous reactions. Forced expression of Rcan1 in wild-type or Rcan1-deficient mast cells reduced FcεRI-mediated cytokine production. Rcan1 deficiency also led to increased FcεRI-mediated mast cell degranulation and enhanced passive cutaneous anaphylaxis. Analysis of the Rcan1 promoter identified a functional Egr1 binding site. Biochemical and genetic evidence suggested that Egr1 controls Rcan1 expression. Our results identified Rcan1 as a novel inhibitory signal in FcεRI-induced mast cell activation and established a new link of Egr1 and Rcan1 in FcεRI signaling.