NF-κB P65 DNA Binding Activity Research Articles

Antiinflammatory effects of the cyclopentenone isoprostane 15-A 2-IsoP in human gestational tissues

Proinflammatory prostaglandins and cytokines are involved in the initiation of human labor and delivery. Although cyclopentenone prostaglandins regulate the formation of these prolabor mediators via nuclear factor-κB (NF-κB) and/or peroxisome proliferator-activated receptor-γ, recent evidence suggests that they do not exist in vivo. Cyclopentenone isoprostanes (IsoPs), which are highly reactive structural isomers of bioactive cyclopentenone prostaglandins, do exist physiologically and have been shown to inhibit the inflammatory response in macrophages. Therefore the aim of this study was to determine the effect of the synthetic cyclopentenone IosP 15-A 2-IsoP on the expression of prolabor mediators in human gestational tissues. Human placenta and gestational membranes ( n = 5) were incubated in the absence or presence of 12.5, 25, and 50 μM 15-A 2-IsoP with 10 μg/ml lipopolysaccharide (LPS). Treatment of placenta and fetal membranes with 15-A 2-IsoP caused a dose-dependent decrease in LPS-stimulated release of the cytokines IL-1β, IL-6, IL-8, and TNF-α and the prostaglandins PGE 2 and PGF 2α. NF-κB p65 DNA binding activity was significantly inhibited by treatment with 50 μM 15-A 2-IsoP. Collectively, these data suggest that 15-A 2-IsoP exhibits antiinflammatory properties via antagonism of NF-κB activity. Cyclopentenone IsoPs may serve as negative feedback regulators of the inflammatory response in human gestational tissues.

Sulfasalazine and BAY 11-7082 Interfere with the Nuclear Factor-κB and IκB Kinase Pathway to Regulate the Release of Proinflammatory Cytokines from Human Adipose Tissue and Skeletal Muscle in Vitro

There is much evidence to indicate a role for adipocytokines in insulin resistance and/or type 2 diabetes mellitus. In experimental models, oral salicylates, through their ability to interfere with the nuclear factor-kappa B (NF-kappa B) transcription pathway, have been demonstrated to reverse insulin resistance. The aim of this study was to investigate whether NF-kappa B regulates the release of adipocytokines in human adipose tissue and skeletal muscle. Human sc adipose tissue and skeletal muscle (obtained from normal pregnant women) were incubated in the absence (control) or presence of two NF-kappa B inhibitors sulfasalazine (1.25, 2.5, and 5 mm) and BAY 11-7082 (25, 50, and 100 microm). After an 18-h incubation, the tissues were collected, and NF-kappa B p65 DNA-binding activity and I kappa B kinase (IKK-beta) and insulin receptor-beta protein expression were assessed by ELISA and Western blotting, respectively. The incubation medium was collected, and the release of TNF-alpha, IL-6, IL-8, resistin, adiponectin, and leptin was quantified by ELISA. Treatment of adipose tissue and skeletal muscle with sulfasalazine and BAY 11-7082 significantly inhibited the release of IL-6, IL-8, and TNF-alpha; NF-kappa B p65 DNA-binding activity; and IKK-beta protein expression (P < 0.05, by Newman-Keuls test). There was no effect of sulfasalazine and BAY 11-7082 on resistin, adiponectin, or leptin release. Both sulfasalazine and BAY 11-7082 increased the adipose tissue and skeletal muscle expression of insulin receptor-beta. The data presented in this study demonstrate that the IKK-beta/NF-kappa B transcription pathway is a key regulator of IL-6, IL-8, and TNF-alpha release from adipose tissue and skeletal muscle. Control of the IKK-beta/NF-kappa B pathway may therefore provide an alternative therapeutic strategy for regulating aberrant cytokine release and thereby alleviating insulin resistance in type 2 diabetes mellitus.

Effect of nuclear factor-kappa B inhibitors and peroxisome proliferator-activated receptor-gamma ligands on PTHrP release from human fetal membranes

Parathyroid hormone-related protein (PTHrP) has been implicated in many processes during normal and pathological pregnancies. In the human fetal membranes, PTHrP exhibits cytokine-like actions. We have recently shown that inhibitors of the nuclear factor-kappa B (NF-κB) and activators of the peroxisome proliferator-activated receptor (PPAR)-γ signalling pathways down-regulate cytokine release from human gestational tissues. Therefore, the aim of this study was to determine whether NF-κB and PPAR-γ also regulate PTHrP release from human fetal membranes. Human amnion and choriodecidua explants were incubated in the absence (control) or presence of two known NF-κB inhibitors (1, 5 and 10 mM sulphasalazine (SASP) or 5, 10 and 15 mM N-acetyl-cysteine (NAC)), and two PPAR-γ ligands (15 and 30 μM 15-deoxy-Δ 12,14-PGJ 2 (15d-PGJ 2) or 15 and 30 μM troglitazone), under basal conditions. After 18 h incubation, the tissues were collected and NF-κB p65 DNA binding activity in nuclear extracts was assessed by ELISA, and the incubation medium was collected and the release of PTHrP was quantified by RIA. Treatment of amnion and choriodecidual tissues with SASP concentrations greater than 5 mM, 15 mM NAC, 30 μM 15d-PGJ 2 and 30 μM troglitazone significantly reduced the release of PTHrP ( p<0.05). This study demonstrates that PTHrP release from human fetal membranes is regulated by inhibitors of NF-κB, and ligands of PPAR-γ.