Nuclear factor-kappaB Transcription Research Articles

The Dishevelled, EGL-10 and pleckstrin (DEP) domain-containing protein DEPDC7 binds to CARMA2 and CARMA3 proteins, and regulates NF-κB activation.

The molecular complexes containing BCL10, MALT1 and CARMA proteins (CBM complex) have been recently identified as a key component in the signal transduction pathways that regulate activation of Nuclear Factor kappaB (NF-κB) transcription factor. Herein we identified the DEP domain-containing protein DEPDC7 as cellular binding partners of CARMA2 and CARMA3 proteins. DEPDC7 displays a cytosolic distribution and its expression induces NF-κB activation. Conversely, shRNA-mediated abrogation of DEPDC7 results in impaired NF-κB activation following G protein-coupled receptors stimulation, or stimuli that require CARMA2 and CARMA3, but not CARMA1. Thus, this study identifies DEPDC7 as a CARMA interacting molecule, and provides evidence that DEPDC7 may be required to specifically convey on the CBM complex signals coming from activated G protein-coupled receptors.

Prevalence of genetic risk factors of psoriasis among the population of the Russian Federation

Goal. To assess the prevalence of polymorphisms of genes of the predisposition to psoriasis among the population of the Russian Federation. Materials and methods. The authors examined 546 psoriasis patients and 206 healthy people. The polymorphism of the following genes was assessed: genes encoding proteins of the signaling pathway of the nuclear transcription factor kappa-B - NF-κΒ (NFKBI, TRAF3IP2, TNFAIP3, REL, TYK2, TNIP1, IL-28RA) responsible for congenital immunity; genes participating in the IL-23 signaling pathway responsible for adaptive immunity (IL-23R, IL-12B); genes participating in the presentation of the antigen (ERAP1); genes responsible for skin barrier dysfunction (SERPINB8 ZNF313, ZNF816A). Peripheral blood leucocytes served as the DNA source. Polymorphisms of IL-23R, IL-28RA, SERPINB8, TRAF3IP2, TNFAIP3, REL, ZNF313, IL-12B, TNIP1, ZNF816A, ERAP1 genes were determined by the real-time PCR method; polymorphisms of NFKBI, TYK2 genes were determined by the RFLP assay (Restriction Fragment Length Polymorphism). Results. In psoriasis patients in the Russian Federation, statistically significant differences in the distribution of allele frequencies were determined for IL-23R-G/G, IL-23R-A/A, TRAF3IP2-A/A, TRAF3IP2-G/G, TNFAIP3-A/C, TNFAIP3-A/A, ZNF313-C/C, TYK2-T/T, TYK2-T/G, TNIP1-G/G, TNIP1-A/G, REL-A/A, ERAP1-G/G genotypes.

Casticin inhibits COX-2 and iNOS expression via suppression of NF-κB and MAPK signaling in lipopolysaccharide-stimulated mouse macrophages

Ethnopharmacological relevanceThe fruits of Vitex rotundifolia L. are widely used to treat inflammation of the airway in Traditional Chinese medicine. Previous studies found that casticin, isolated from Vitex rotundifolia, could induce apoptosis of tumor cells. In this study, we evaluated the anti-inflammatory effects of casticin and its underlying molecular mechanism in lipopolysaccharide (LPS)-stimulated macrophages. Materials and methodsRAW264.7 cells were pretreated with various concentrations of casticin (0.3–10μM), and then treated with LPS to induce inflammation. We assayed the levels of proinflammatory cytokines and prostaglandin E2 (PGE2) using ELISA, and examined the protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, and heme oxygenase (HO)-1 by Western blot. We also investigated the anti-inflammatory molecular mechanism by analyzing inflammatory-associated signaling pathways, including the nuclear transcription factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. ResultsWe found casticin inhibited the levels of nitric oxide and PGE2, and decreased the production of proinflammatory cytokines such as interleukin (IL)-1β, IL-6, and tumor necrosis factor α (TNF-α). In addition, iNOS and COX-2 expression levels were suppressed and casticin increased HO-1 and Nrf2 production in a concentration-dependent manner. Furthermore, casticin significantly inhibited NF-κB subunit p65 proteins in the nucleus and decreased Akt and MAPK activation. ConclusionThese results suggest that the anti-inflammatory effect of casticin is due to inhibition of proinflammatory cytokines and mediators by blocking the NF-κB, Akt, and MAPK signaling pathways.

Molecular and genetic studies of the predisposition to the development of psoriasis among the population of the Russian Federation: a study of polymorphisms of TNFAIP3, TNIP1, TYK2 and REL genes

Goal. To study the role of polymorphic variants of genes encoding proteins of the signaling pathway of the nuclear transcription factor kappa-B, NF-kB (TNFAIP3 (rs610604), TNIP1 (rs17728338), TYK2 (rs12720356) and REL (rs702873)) in the predisposition to psoriasis among the Caucasian population of the Russian Federation. Materials and methods. The authors studied markers of the predisposition to the development of psoriasis such as TNFAIP3 (rs610604), TNIP1 (rs17728338), TYK2 (rs12720356) and REL (rs702873) using Real-Time PCR methods and allele-specific oligonucleotide probes and PFLP (Restriction Fragment Length Polymorphism) assay in whole blood samples taken from psoriatic patients (n = 301). To compare the frequency of genotypes, the authors used data obtained as a result of the genotyping of DNAs taken from healthy volunteers (n = 109). Results. The authors analyzed particular features of the distribution of polymorphic variants of TNFAIP3 (rs610604), TNIP1 (rs17728338), TYK2 (rs1272035) и REL (rs702873) genes in a sample of 301 psoriatic patients and 109 healthy volunteers from the Russian Federation. They revealed reliable differences in the frequency of registration of genotypes of these genes. The authors also determined genotypes considered as risk factors with regard to the development of psoriasis (TNFAIP3-A/C, TNIP1-A/G, TNIP1-A/A and TYK2-T/T) as well as protector genotypes (TNFAIP3-A/Ä, REL-A/A and TYK2-T/G)

Intestinal anti-inflammatory activity of Sasa quelpaertensis leaf extract by suppressing lipopolysaccharide-stimulated inflammatory mediators in intestinal epithelial Caco-2 cells co-cultured with RAW 264.7 macrophage cells.

BACKGROUND/OBJECTIVESInflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, involves chronic inflammation of the gastrointestinal tract. Previously, Sasa quelpaertensis leaves have been shown to mediate anti-inflammation and anti-cancer effects, although it remains unclear whether Sasa leaves are able to attenuate inflammation-related intestinal diseases. Therefore, the aim of this study was to investigate the anti-inflammatory effects of Sasa quelpaertensis leaf extract (SQE) using an in vitro co-culture model of the intestinal epithelial environment.MATERIALS/METHODSAn in vitro co-culture system was established that consisted of intestinal epithelial Caco-2 cells and RAW 264.7 macrophages. Treatment with lipopolysaccharide (LPS) was used to induce inflammation.RESULTSTreatment with SQE significantly suppressed the secretion of LPS-induced nitric oxide (NO), prostaglandin E2 (PGE2), IL-6, and IL-1β in co-cultured RAW 264.7 macrophages. In addition, expressions of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, and tumor necrosis factor (TNF)-α were down-regulated in response to inhibition of IκBα phosphorylation by SQE. Compared with two bioactive compounds that have previously been identified in SQE, tricin and P-coumaric acid, SQE exhibited the most effective anti-inflammatory properties.CONCLUSIONSSQE exhibited intestinal anti-inflammatory activity by inhibiting various inflammatory mediators mediated through nuclear transcription factor kappa-B (NF-kB) activation. Thus, SQE has the potential to ameliorate inflammation-related diseases, including IBD, by limiting excessive production of pro-inflammatory mediators.

Carotenoid derivatives inhibit nuclear factor kappa B activity in bone and cancer cells by targeting key thiol groups

Aberrant activation of the nuclear factor kappa B (NFkB) transcription system contributes to cancer progression, and has a harmful effect on bone health. Several major components of the NFkB pathway such as IkB Kinase (IKK) and the NFkB subunits contain cysteine residues that are critical for their activity. The interaction of electrophiles with these cysteine residues results in NFkB inhibition. Carotenoids, hydrophobic plant pigments, are devoid of electrophilic groups, and we have previously demonstrated that carotenoid derivatives, but not the native compounds activate the Nrf2 transcription system. The aim of the current study was to examine whether carotenoid derivatives inhibit NFkB, and, if so, to determine the molecular mechanism underpinning the inhibitory action. We report in the present study that a mixture of oxidized derivatives, prepared by ethanol extraction from partially oxidized lycopene preparation, inhibited NFkB reporter gene activity. In contrast, the intact carotenoid was inactive. A series of synthetic dialdehyde carotenoid derivatives inhibited reporter activity as well as several stages of the NFkB pathway in both cancer and bone cells. The activity of the carotenoid derivatives depended on the reactivity of the electrophilic groups in reactions such as Michael addition to sulfhydryl groups of proteins. Specifically, carotenoid derivatives directly interacted with two key proteins of the NFkB pathway: the IKKβ and the p65 subunit. Direct interaction with IKKβ was found in an in vitro kinase assay with a recombinant enzyme. The inhibition by carotenoid derivatives of p65 transcriptional activity was observed in a reporter gene assay performed in the presence of excess p65. This inhibition action resulted, at least in part, from direct interaction of the carotenoid derivative with p65 leading to reduced binding of the protein to DNA as evidenced by electrophoretic mobility shift assay (EMSA) experiments. Importantly, we found by using mutation in key cysteine residues of both p65 and IKK that specific thiol groups are essential for NFkB inhibition by carotenoid derivatives. In conclusion, we propose that electrophilic carotenoid derivatives contribute to cancer prevention as well as bone health maintenance via the inhibition of the NFkB transcription system. Pivotal thiol groups of both IKK and p65 play a key role in this process.

Chronotherapeutic effect of fisetin on expression of urea cycle enzymes and inflammatory markers in hyperammonaemic rats

BackgroundElevated blood ammonia leads to hyperammonaemia that affects vital central nervous system (CNS) functions. Fisetin, a naturally occurring flavonoid, exhibits therapeutic benefits, such as anti-cancer, anti-diabetic, anti-oxidant, anti-angiogenic, neuroprotective and neurotrophic effects. MethodsIn this study, the chronotherapeutic effect of fisetin on ammonium chloride (AC)-induced hyperammonaemic rats was investigated, to ascertain the time point at which the maximum drug effect is achieved. The anti-hyperammonaemic potential of fisetin (50mg/kg b.w. oral) was analysed when administered to AC treated (100mg/kg b.w. i.p.) rats at 06:00, 12:00, 18:00 and 00:00h. Amelioration of pathophysiological conditions by fisetin at different time points was measured by analysing the levels of expression of liver urea cycle enzymes (carbamoyl phosphate synthetase-I (CPS-I), ornithine transcarbamoylase (OTC) and argininosuccinate synthetase (ASS)), nuclear transcription factor kappaB (NF-κB p65), brain glutamine synthetase (GS) and inducible nitric oxide synthase (iNOS) by Western blot analysis. ResultsFisetin increased the expression of CPS-I, OTC, ASS and GS and decreased iNOS and NF-κB p65 in hyperammonaemic rats. Fisetin administration at 00:00h showed more significant effects on the expression of liver and brain markers, compared with other time points. ConclusionsFisetin could exhibit anti-hyperammonaemic effect owing to its anti-oxidant and cytoprotective influences. The temporal variation in the effect of fisetin could be due to the (i) chronopharmacological, chronopharmacokinetic properties of fisetin and (ii) modulations in the endogenous circadian rhythms of urea cycle enzymes, brain markers, redox enzymes and renal clearance during hyperammonaemia by fisetin. However, future studies in these lines are necessitated.

Schisantherin A protects lipopolysaccharide-induced acute respiratory distress syndrome in mice through inhibiting NF-κB and MAPKs signaling pathways.

Acute respiratory distress syndrome (ARDS) is characterized by polymorphonuclear neutrophils (PMNs) adhesion, activation, sequestration and inflammatory damage to alveolar-capillary membrane. Schisantherin A, a dibenzocyclooctadiene lignan isolated from the fruit of Schisandra sphenanthera, has been reported to have anti-inflammatory properties. In the present study, we aimed to investigate the protective effects of schisantherin A on LPS-induced mouse ARDS. The pulmonary injury severity was evaluated 7 h after LPS administration and the protective effects of schisantherin A on LPS-induced mouse ARDS were assayed by enzyme-linked immunosorbent assay and Western blot. The results revealed that the wet/dry weight ratio, myeloperoxidase activity, and the number of total cells, neutrophils and macrophages in the bronchoalveolar lavage fluid (BALF) were significantly reduced by schisantherin A in a dose-dependent manner. Meanwhile, pretreatment with schisantherin A markedly ameliorated LPS-induced histopathologic changes and decreased the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in the BALF. In addition, the phosphorylation of nuclear transcription factor-kappaB (NF-κB) p65, inhibitory kappa B alpha (IκB-α), c-jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38 induced by LPS were suppressed by schisantherin A. These findings indicated that schisantherin A exerted potent anti-inflammatory properties in LPS-induced mouse ARDS, possibly through blocking the activation of NF-KB and mitogen activated protein kinases (MAPKs) signaling pathways. Therefore, schisantherin A may be a potential agent for the prophylaxis of ARDS.

Systemic injection of low-dose lipopolysaccharide fails to break down the blood-brain barrier or activate the TLR4-MyD88 pathway in neonatal rat brain.

We aimed to investigate whether peripheral low-dose lipopolysaccharide (LPS) induces the breakdown of the blood–brain barrier (BBB) and/or the activation of toll-like receptor 4 (TLR4) in the neonatal rat brain. Neonatal rats received intraperitoneal injections of low-dose LPS (0.3 mg/kg∙bw), and the BBB compromise was detected by Evans Blue extravasation and electron microscopy. Meanwhile, TLR4, adaptin myeloid differentiation factor 88 (MyD88), nuclear transcription factor kappa-B (NF-κB) p50 and tumor necrosis factor alpha (TNFα) in the neonatal rat brain were determined by quantitative real-time polymerase chain reaction (PCR) and Western Blot. Immunohistochemistry was used to determine the distribution and activation of microglia in the brain after LPS administration. It was demonstrated that Evans Blue extravasation was not observed in the brain parenchyma, and that tight junctions of cerebral endothelial cells remained intact after systemic injections of LPS in neonatal rats. Although intracerebroventricular injections of LPS activated microglia and up-regulated the expression of TLR4, MyD88, NF-κB p50 and TNFα in the neonatal rat brain, systemic LPS did not induce these responses. These findings indicate that while the neonatal rat brain responds to the direct intra-cerebral administration of LPS through robust TLR4 activation, systemic low-dose LPS does not induce the innate immune reaction or compromise the BBB in neonatal rats.

Propylene-glycol aggravates LPS-induced sepsis through production of TNF-α and IL-6.

Propylene glycol (1,2-propanediol, PG) is a commonly used solvent for oral, intravenous, as well as topical pharmaceutical preparations. While PG is generally considered to be safe, it has been known that large intravenous doses given over a short period of time can be toxic. To evaluate the effect of PG in sepsis induced by the bacterial endotoxin lipopolysaccharide (LPS). Balb/c mice were treated with LPS (1 mg/kg b.w., i.p.) with or without PG (5 g/kg b.w. i.v.). The survival rate and the production of inflammatory cytokines were measured. In RAW264.7 mouse macrophages encoding NF-kB-luc reporter gene, the nuclear transcription factor kappa-B (NF-kB) activation was measured. We found that intravenous PG increased the mortality rate in sepsis induced by the bacterial endotoxin lipopolysaccharide (LPS) in mice. In accordance with that, PG enhanced LPS-induced production of inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in vivo. PG also increased the LPS-induced macrophage activation in vitro as detected by measuring NF-kB activation. Our results indicate that drugs containing high doses of PG can pose a risk when administered to patients suffering from or prone to Gram negative bacterial infection.

Rosiglitazone inhibits chlorpyrifos-induced apoptosis via modulation of the oxidative stress and inflammatory response in SH-SY5Y cells

Oxidative stress can lead to expression of inflammatory transcription factors, which are important regulatory elements in the induction of inflammatory responses. One of the transcription factors, nuclear transcription factor kappa-B (NF-κB) plays a significant role in the inflammation regulatory process. Inflammatory cell death has been implicated in neuronal cell death in some neurodegenerative disorders such as Parkinson's disease (PD). In this study, we investigated the molecular mechanisms underlying apoptosis initiated by chlorpyrifos (CPF)-mediated oxidative stress. Based on the cytotoxic mechanism of CPF, we examined the neuroprotective effects of rosiglitazone (RGZ), a peroxisome proliferator-activated receptor gamma (PPAR-γ) agonist, against CPF-induced neuronal cell death. The treatment of SH-SY5Y cells with CPF induced oxidative stress. In addition, CPF activated the p38, JNK and ERK mitogen-activated protein kinases (MAPKs), and induced increases in the inflammatory genes such as COX-2 and TNF-α. CPF also induced nuclear translocation of NF-κB and inhibitors of NF-κB abolished the CPF-induced COX-2 expression. Pretreatment with RGZ significantly reduced ROS generation and enhanced HO-1 expression in CPF-exposed cells. RGZ blocked the activation of both p38 and JNK signaling, while ERK activation was strengthened. RGZ also attenuated CPF-induced cell death through the reduction of NF-κB-mediated proinflammatory factors. Results from this study suggest that RGZ may exert an anti-apoptotic effect against CPF-induced cytotoxicity by attenuation of oxidative stress as well as inhibition of the inflammatory cascade via inactivation of signaling by p38 and JNK, and NF-κB.

Gene structure, cDNA characterization and RNAi-based functional analysis of a myeloid differentiation factor 88 homolog in Tenebrio molitor larvae exposed to Staphylococcus aureus infection

Myeloid differentiation factor 88 (MyD88), an intracellular adaptor protein involved in Toll/Toll-like receptor (TLR) signal processing, triggers activation of nuclear factor-kappaB (NF-κB) transcription factors. In the present study, we analyzed the gene structure and biological function of MyD88 in a coleopteran insect, Tenebrio molitor (TmMyD88). The TmMyD88 gene was 1380bp in length and consisted of five exons and four introns. The 5′-flanking sequence revealed several putative transcription factor binding sites, such as STAT-4, AP-1, cJun, cfos, NF-1 and many heat shock factor binding elements. The cDNA contained a typical death domain, a conservative Toll-like interleukin-1 receptor (TIR) domain, and a C-terminal extension (CTE). The TmMyD88 TIR domain showed three significantly conserved motifs for interacting with the TIR domain of TLRs. TmMyD88 was grouped within the invertebrate cluster of the phylogenetic tree and shared 75% sequence identity with the TIR domain of Tribolium castaneum MyD88. Homology modeling of the TmMyD88 TIR domain revealed five parallel β-strands surrounded by five α-helices that adopted loop conformations to function as an adaptor. TmMyD88 expression was upregulated 7.3- and 4.79-fold after 12 and 6h, respectively, of challenge with Staphylococcus aureus and fungal β-1,3 glucan. Silencing of the TmMyD88 transcript by RNA interference led to reduced resistance of the host to infection by S. aureus. These results indicate that TmMyD88 is required for survival against Staphylococcus infection.

Flavonoid components in Scutellaria baicalensis inhibit nicotine-induced proliferation, metastasis and lung cancer-associated inflammation in vitro

The objective of the present study was to investigate the therapeutic efficacy of flavonoid components in Scutellariabaicalensis on proliferation, metastasis and lung cancer-associated inflammation during nicotine induction in the A549 and H1299 lung cancer cell lines. After experimental period, augmentation of proliferation was observed, accompanied by marked decrease in apoptotic cells in nicotine-induced lung cancer cells; additionally, nicotine-exposed cells exhibited increased invasive and migratory abilities based on invasion and wound-healing assay. Flavones in Scutellaria, baicalin, baicalein and wogonin significantly counteracted the above deleterious changes. Moreover, assessment of tumor apoptotic and metastatic factors on mRNA levels by quantitative PCR and protein levels by western blotting revealed that these phytochemical treatments effectively negated nicotine-induced upregulated expression of bcl-2, bcl-2/bax ratio, caspase-3, matrix metalloproteinase (MMP)-2 and MMP-9 as well as downregulated expression of bax. Further analysis of inflammatory markers such as tumor necrosis factor (TNF)-α and interleukin (IL)-6 in cell culture supernatant and mRNA and protein expression of nuclear transcription factor-kappaB (NF-κB) and I kappa B-alpha (IκB-α) was carried out to substantiate the anti-inflammatory effect of flavones in Scutellaria in nicotine-exposed lung cancer cells. The therapeutic effects observed in the present study are attributed to the potent potential against proliferation, metastasis and inflammatory microenvironment by flavonoid components in Scutellaria in nicotine-induced lung cancer cells.

D-ribosylation induces cognitive impairment through RAGE-dependent astrocytic inflammation

Non-enzymatic glycation of proteins by reducing saccharides for instance D-glucose is an important post-translational modification regulating protein function. Already two centuries ago, D-glucose (Glc) was identified in the urine of diabetic patients. Recently, abnormally high level of D-ribose (Rib) in the urine of type 2 diabetics has been discovered, which is highly active in protein glycation, resulting in the production of advanced glycation end products (AGEs). Accumulation of AGEs leads to altered cellular function, for example AGE accumulation in the nervous system impairs cognitive ability, yet the mechanisms mediating this process for Rib are unknown. Here we found that treatment with Rib accelerated AGE formation in U251 and U87MG astrocytoma cells and in mouse brain, inducing upregulation of receptor for AGEs (RAGE). Astrocytoma cells with elevated levels of RAGE displayed enhanced activity of the proinflammatory nuclear transcription factor kappaB and increased expression of tumor necrosis factor alpha and glial fibrillary acidic protein. Moreover, injection of Rib induced astrocyte activation in mouse hippocampus and impaired spatial learning and memory abilities. These results indicate that mouse spatial cognitive impairment caused by Rib-derived AGEs is correlated with activation of an astrocyte-mediated, RAGE-dependent inflammatory response. This study may provide insights into the mechanism of Rib-involved cognitive impairments and diabetic encephalopathy.

Effects of plant-based Korean food extracts on lipopolysaccharide-stimulated production of inflammatory mediators in vitro.

BACKGROUND/OBJECTIVESThe traditional Korean diet is plant-based and rich in antioxidants. Previous studies have investigated the potential health benefits of individual nutrients of Korean foods. However, the cumulative effects of a Korean diet on inflammation remain poorly understood. Therefore, the aim of this study was to investigate the anti-inflammatory effects of a plant-based Korean diet.MATERIALS/METHODSUsing data from the Fifth Korean National Health and Nutrition Examination Survey, 75 individual plant food items were selected which represent over 1% of the total diet intake of the Korean diet. These items were classified into ten different food groups, and the vegetable (Veg) and fruit (Fruit) groups were studied based on their high antioxidant capacity. For comparison, a mixture of all ten groups (Mix) was prepared. To produce a model of inflammation with which to test these Veg, Fruit, and Mix plant-based Korean food extracts (PKE), RAW264.7 macrophages were treated with lipopolysaccharide (LPS).RESULTSLevels of nitric oxide (NO) and prostaglandin E2 (PGE2), as well as protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were found to be lower following PKE treatment. Furthermore, PKE treatment was found to suppress tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) via the nuclear transcription factor kappa-B (NF-κB) signaling pathway. Overall, the Mix group exhibited the greatest anti-inflammatory effects compared with Veg and Fruit PKE group.CONCLUSIONSInhibition of LPS-induced pro-inflammatory mediators by the PKE tested was found to involve an inhibition of NF-kB activation. Moreover, PKE tested have the potential to ameliorate various inflammation-related diseases by limiting the excessive production of pro-inflammatory mediators.

Association of Acute Interstitial Nephritis with Carnivora, a Venus Flytrap Extract, in a 30-Year-Old Man with Hodgkin's Lymphoma.

Acute interstitial nephritis (AIN) is a common cause of acute kidney injury and has been associated with a variety of medications. This is the case of 30-year-old man with Hodgkin's lymphoma who on routine labs before chemotherapy was found to have acute nonoliguric renal failure. A kidney biopsy was performed and confirmed the diagnosis of acute interstitial nephritis. The patient had taken several medications including a higher dose of Carnivora, a Venus flytrap extract, composed of numerous amino acids. The medication was discontinued and kidney function improved towards the patient's baseline indicating that this may be the possible cause of his AIN. Proximal tubular cell uptake of amino acids increasing transcription of nuclear factor-kappaB is a proposed mechanism of AIN from this compound.

Wedelolactone inhibits LPS-induced pro-inflammation via NF-kappaB Pathway in RAW 264.7 cells

BackgroundWedelolactone (WEL), a major coumestan ingredient in Wedelia chinensis, has been used to treat septic shock, hepatitis and venom poisoning in traditional Chinese medicines. The objective of the study was to elucidate the anti-inflammatory effects and mechanism of WEL with a cellular model of lipopolysaccharide (LPS)-induced RAW 264.7 cells.ResultsTo study the role of WEL in pro-inflammation, we measured key inflammation mediators and end products including nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and tumor necrosis factor-α (TNF-α) by using the Griess method, enzyme linked immunosorbent assay (ELISA) and Western blotting. Nuclear factor-kappaB (NF-κB) transcription activity was detected by luciferase reporter assay. The important pro-inflammatory transcription factors, NF-κB p65 and inhibitory kappaB alpha (IκB-α); and mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 MAPK (p38) were analyzed by Western blotting. Our study showed that WEL (0.1, 1, 10 μM) significantly inhibited the protein expression levels of iNOS and COX-2 in LPS-stimulated cells, as well as the downstream products, including NO, PGE2 and TNF-α. Moreover, WEL also inhibited LPS-induced NF-κB p65 activation via the degradation and phosphorylation of IκB-α and subsequent translocation of the NF-κB p65 subunit to the nucleus.ConclusionsOur results revealed that WEL has a potential to be a novel anti-inflammatory agent targeting on the NF-κB signaling pathway.

Anti-inflammatory effect of sophoraflavanone G isolated from Sophora flavescens in lipopolysaccharide-stimulated mouse macrophages

Sophoraflavanone G (SG; 5,7,D, 2',4'-tetrahydroxy-8-lavandulylflavanone) has been isolated from Sophora flavescens and found to be effective against bacteria and to decrease cyclooxygenase (COX)-2 expression in RAW 264.7 macrophage. However, the anti-inflammatory mechanisms of SG are not well understood. RAW 264.7 cells were pretreated with various concentrations of SG (2.5-20 μM) and inflammatory responses were induced with lipopolysaccharide. Using enzyme-linked immunosorbent assay, the levels of pro-inflammatory cytokines and prostaglandin E2 (PGE2) were determined. Western blot was used to examine the protein expression of inducible nitric oxide synthase (iNOS), COX-2, and heme oxygenase-1 (HO-1). To investigate the molecular mechanism, we analyzed inflammatory-associated signaling pathways, including nuclear transcription factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK). SG inhibited the levels of nitric oxide and PGE2 and decreased the production of pro-inflammatory cytokines, such as interleukin (IL)-1β, IL-6, and tumor necrosis factor α. The expression of iNOS and COX-2 was also suppressed. However, SG increased HO-1 production in a concentration-dependent manner and significantly decreased MAPK activation and inhibited NF-κB subunit p65 proteins to translocate into the nucleus. These results suggest that SG has an anti-inflammatory effect, inhibiting pro-inflammatory cytokines and mediators production via interruption of the NF-κB and MAPK signaling pathways.

A Functional Insertion/Deletion Polymorphism in the Promoter of PDCD6IP Is Associated with the Susceptibility of Hepatocellular Carcinoma in a Chinese Population

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. Apart from environmental factors such as hepatitis B virus (HBV) or hepatitis C virus, alcohol abuse, and exposure to dietary aflatoxin, genetic factors are also involved in the pathogenesis of HCC. By analyzing 390 HCC cases and 431 healthy controls in a Chinese population, we used a candidate gene approach to evaluate the association between a 15-bp insertion/deletion (indel) polymorphism (rs28381975) in the promoter region of the programmed cell death 6 interacting protein (PDCD6IP) gene and HCC susceptibility. Logistic regression analysis demonstrated that subjects carrying ins/del or ins/ins genotypes had significantly increased risk for HCC than individuals carrying del/del genotypes (adjusted odds ratio=1.39, 95% confidence interval=1.01-1.91, p=0.033]. Carrying the 15-bp insertion allele was associated with a 1.26-fold risk for HCC (95% CI=1.04-1.54, p=0.018). Moreover, significant differences were observed within HCC patients concerning genotypic frequencies of rs28381975 after stratifying by tumor stages and HBV infection. Computational modeling suggests that rs28381975 could disrupt the binding patterns of c-rel, a key subunit of nuclear factor-kappaB transcription factor. Further luciferase-based transient transfection assays revealed that rs28381975 can affect the promoter activity of PDCD6IP, indicating its possible functional significance. Taken together, our data suggest that common genetic variations in PDCD6IP may influence HCC risk, possibly through promoter activity-mediated regulation. Replication of our studies in other populations and further functional analysis will strengthen our understanding of this association.

Phloretin and phlorizin promote lipolysis and inhibit inflammation in mouse 3T3‐L1 cells and in macrophage‐adipocyte co‐cultures

Previous studies found that phloretin (PT) and phlorizin (PZ) could inhibit glucose transport, with PT being a better inhibitor of lipid peroxidation. This study aimed to evaluate the antiobesity effects of PT and PZ in 3T3-L1 cells and if they can modulate the relationship between adipocytes and macrophages. Differentiated 3T3-L1 cells were treated with PT or PZ. Subsequently, transcription factors of adipogenesis and lipolysis proteins were measured. In addition, RAW 264.7 macrophages treated with PT or PZ were cultured in differentiated media from 3T3-L1 cells to analyze inflammatory mediators and signaling pathways. PT significantly enhanced glycerol release and inhibited the adipogenesis-related transcription factors. PT also promoted phosphorylation of AMP-activated protein kinase and increased activity of adipose triglyceride lipase and hormone-sensitive lipase. PT suppressed the nuclear transcription factor kappa-B and mitogen-activated protein kinase pathways when RAW 264.7 cells were cultured in differentiated media from 3T3-L1 cells. PZ improved lipolysis and inhibited the macrophage inflammatory response less effectively than PT. This study suggests that PT is more effective than PZ at increasing lipolysis in adipocytes. In addition, PT also suppresses inflammatory response in macrophage that is stimulated by differentiated media from 3T3-L1 cells.