Intestinal inflammation and seizure susceptibility: understanding the role of tumour necrosis factor‐α in a rat model

Objectives The aim of the study was to evaluate the correlation between colitis and susceptibility to seizures. Methods Colitis was induced in Wistar rats by a single intracolonic administration of trinitrobenzene sulfonic acid (TNBS; 20 mg in 35% ethanol). The control group were given intracolonic vehicle. One group of rats with colitis were treated with thalidomide (150 mg/kg p.o.) daily for 14 days. The other colitis group received vehicle only. On day 15, seizure susceptibility was tested by administration of pentylenetetrazole (40 mg/kg i.p.). Colonic tissue was collected for estimation of morphological score, and malondialdehyde, superoxide dismutase, catalase and glutathione peroxidase. Tumour necrosis factor (TNF)‐α levels were measured in serum and brain samples. Key findings The colitis group showed a significant increase in seizure score and reduction in onset time compared with the control group. Thalidomide was protective against seizures, resulting in decreased seizure score and significantly delaying the onset of seizures. Thalidomide also provided significant protection against TNBS‐induced colonic damage in terms of morphological and histological score and levels of lipid peroxidation, superoxide dismutase, catalase and glutathione peroxidase in colonic tissue. The level of TNF‐α in serum was also reduced significantly whereas brain TNF‐α level was reduced but not significantly. Conclusions TNBS‐induced colitis increased seizure susceptibility to a subconvulsive dose of pentylenetetrazole; the immunomodulator thalidomide was protective.     

Oxymatrine Induces Liver Injury through JNK Signalling Pathway Mediated by TNF‐α In Vivo

Oxymatrine (OMT) is a traditional Chinese medicine monomer and has been used for the treatment of chronic viral hepatitis and many other diseases. We aimed to investigate whether OMT could induce hepatotoxicity in mice and explored the preliminary mechanisms of toxic effects. Twenty‐four Institute for Cancer Research male mice were randomly divided into four groups: control group, 40, 160 and 320 mg/kg OMT‐treated group. OMT was orally administered once daily for 7 days. The OMT‐treated group exhibited an improved liver index and increase in serum alanine aminotransferase, aspartate aminotransferase and alkaline phosphatase,augmented liver histological injury, elevated levels of malondialdehyde and tumour necrosis factor alpha (TNF‐α) accompanied by the activation of caspase‐9/‐8/‐3, up‐regulated expressions of tumour necrosis factor receptor l (TNFR1), TNF receptor‐associated structure domain (TRADD) and phosphorylation of stress‐activated protein kinase/c‐jun N‐terminal protein kinases (p‐SAPK/JNK). Altogether, these results suggest that OMT at a dose of 320 mg/kg leads to liver damage and is related to the activation of JNK signalling pathway mediated by TNF‐α in the liver of mice.     

Antihypertensive methyldopa,labetalol, hydralazine,and clonidine reversed tumour necrosis factor‐α inhibited endothelial nitric oxide synthase expression in endothelial‐trophoblast cellular networks

Medications used to control hypertension in pregnancy also improve trophoblast and endothelial cellular interaction in vitro. Tumour necrosis factor‐α (TNF‐α) inhibits trophoblast and endothelial cellular interactions and simultaneously decreases endothelial nitric oxide synthase (eNOS) expression. This study investigated whether antihypertensive medications improved these cellular interactions by modulating eNOS and inducible nitric oxide synthase (iNOS) expression. Human uterine myometrial microvascular endothelial cells (UtMVECs) were pre‐incubated with (or without) low dose TNF‐α (0.5 ng/mL) or TNF‐α plus soluble fms‐like tyrosine kinase‐1 (sFlt‐1) (100 ng/mL). The endothelial cells were cultured on Matrigel. After endothelial cellular networks appeared, trophoblast derived HTR‐8/SVneo cells were co‐cultured in the presence of clinically relevant doses of methyldopa, labetalol, hydralazine or clonidine for 24 hours. Cells were retrieved from the Matrigel to extract mRNA and eNOS and iNOS expression were examined by quantitative PCR. Methyldopa, labetalol, hydralazine and clonidine reversed the inhibitory effect of TNF‐α on eNOS mRNA expression. After pre‐incubating endothelial cells with TNF‐α and sFlt‐1, all the medications except methyldopa lost their effect on eNOS mRNA expression. In the absence of TNF‐α, antihypertensive medications did not change eNOS expression. The mRNA expression of iNOS was not affected by TNF‐α or any medications. This study shows that selected antihypertensive medications used in the treatment of hypertension in pregnancy increase eNOS expression in vitro when induced by the inflammatory TNF‐α. The anti‐angiogenic molecule sFlt‐1 may antagonise the potential benefit of these medications by interfering with the NOS pathway.     

TNF‐α level affects etanercept clearance: TNF‐α concentration as a new correction factor of allometric scaling to predict individual etanercept clearances in patients with ankylosing spondylitis

Etanercept (ETN) is a widely used anti‐tumour necrosis factor‐α (TNF‐α) agent, which relieves the symptoms of ankylosing spondylitis (AS) by binding to TNF‐α to inhibit its inflammation effects. In this study, the effect of TNF‐α level on ETN clearance (CL) was investigated, and the TNF‐α concentration was initially set as a correction factor for allometric scaling to improve the predictions of individual ETN CLs. Individual ETN CLs and TNF‐α concentrations in healthy volunteers and patients with AS were determined by performing ETN pharmacokinetic studies in the two cohorts. Accordingly, individual ETN CLs in both healthy volunteers and patients with AS were predicted from data of two animal species using different methods, including simple allometric scaling, scaling with a correction factor of maximum life span potential or brain weight, and scaling with a correction factor of the TNF‐α concentration. The accuracies of such predictions were evaluated by the percentage errors. Consequently, increased TNF‐α concentration was shown to improve ETN CL, by comparing both ETN CLs and TNF‐α concentrations between healthy volunteers and patients with AS. More importantly, better predictions of individual ETN CLs were achieved in patients with AS using allometric scaling with TNF‐α concentration as the correction factor. In conclusion, in vivo levels of TNF‐α can affect ETN CL, and allometric scaling corrected with the TNF‐α concentration can be used to estimate the individual CLs of anti‐TNF‐α monoclonal antibodies based on preclinical data.     

Hepatoprotective and antioxidant effects of gallic acid in paracetamol‐induced liver damage in mice

Objectives The aim of this research paper was to investigate the hepatoprotective and antioxidant effects of gallic acid in paracetamol‐induced liver damage in mice. Methods In the present study, the hepatoprotective and antioxidant effects of gallic acid were evaluated against paracetamol‐induced hepatotoxicity in mice and compared with the silymarin, a standard hepatoprotective drug. The mice received a single dose of paracetamol (900 mg/kg body weight i.p.). Gallic acid (100 mg/kg body weight i.p.) and silymarin (25 mg/kg body weight i.p.) were administered 30 min after the injection of paracetamol. After 4 h, liver marker enzymes (aspartate transaminase, alanine transaminase and alkaline phosphatase) and inflammatory mediator tumour necrosis factor‐alpha (TNF‐α) were estimated in serum, while the lipid peroxidation and antioxidant status (superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione‐S‐transferase and glutathione) were determined in liver homogenate of the control and experimental mice. Key findings Increased activities of liver marker enzymes and elevated TNF‐α and lipid peroxidation levels were observed in mice exposed to paracetamol (P < 0.05), whereas the antioxidant status was found to be depleted (P < 0.05) when compared with the control group. However gallic acid treatment (100 mg/kg body weight i.p.) significantly reverses (P < 0.05) the above changes by its antioxidant action compared to the control group as observed in the paracetamol‐challenged mice. Conclusions The results clearly demonstrate that gallic acid possesses promising hepatoprotective effects.     

ANGIOTENSIN-CONVERTING ENZYME INHIBITORS IMPROVE HEPATIC STEATOSIS BY MODULATING EXPRESSION OF TUMOUR NECROSIS FACTOR-α, INTERLEUKIN-6 AND ADIPONECTIN RECEPTOR-2 IN RATS WITH TYPE 2 DIABETES

• 1 Angiotensin‐converting enzyme inhibitors (ACEI) are hypotensive drugs that have been shown to prevent Type 2 diabetes mellitus (T2DM) in high‐risk individuals. However, in T2DM, the effects of ACEI on hepatic steatosis are not known. The aim of the present study was to examine the effects of ACEI on changes in liver histology and hepatic mRNA expression of adipokines in rats with T2DM.
• 2 Thirty‐six rats were divided into a normal control group, a T2DM group and a fosinopril‐treated group. After six weeks of treatment with 5 mg/kg per day fosinopril, an ACEI, changes in liver histology, serum fasting glucose (FG), insulin, triglyceride (TG), total cholersterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumour necrosis factor (TNF)‐α, interleukin (IL)‐6, adiponectin were evaluated, as was hepatic TNF‐α, IL‐6 and adiponectin receptor‐2 (adipoR2) mRNA expression.
• 3 The degree of hepatic steatosis and inflammation, serum FG, insulin, TG, TC, ALT, TNF‐α and IL‐6 concentrations and hepatic TNF‐α and IL‐6 mRNA expression were significantly higher in rats with T2DM than in normal controls. Serum adiponectin concentrations and hepatic adipoR2 mRNA expression in rats with T2DM were significantly lower than in normal controls. Fosinopril significantly reduced the degree of hepatic steatosis, serum FG, insulin, ALT, TNF‐α and IL‐6 concentrations and hepatic TNF‐α and IL‐6 mRNA expression. Fosinopril significantly increased serum adiponectin concentrations and hepatic adipoR2 mRNA expression.
• 4 In conclusion, the ACEI improved insulin sensitivity and hepatic steatosis in rats with T2DM by increasing circulating adiponectin and hepatic adipoR2 levels, in addition to reducing pro‐inflammatory cytokine levels in the circulation and liver.

     

Acute effect of β‐sitosterol on calcium uptake mediates anti‐inflammatory effect in murine activated neutrophils

Objectives To evaluate the effect of β‐sitosterol on ⁴⁵Ca²⁺ uptake in activated murine neutrophils, and upon myeloperoxidase and adenosine deaminase activity, and interleukin‐1β (IL‐1β) and tumour necrosis factor‐α (TNF‐α) levels, in carrageenan‐induced inflammation in the mouse air pouch model. Methods Dried Esenbeckia leiocarpa bark was macerated and extracted resulting in a crude hydroalcoholic extract (CHE) that was partitioned to obtain an alkaloid fraction. The alkaloid was then partitioned in polar and nonpolar subfractions. β‐Sitosterol was isolated from the nonpolar subfraction and identified by comparison with the literature. The effect of β‐sitosterol on ⁴⁵Ca²⁺ uptake in activated murine neutrophils, and upon myeloperoxidase and adenosine deaminase activity, IL‐1β and TNF‐α levels in carrageenan‐induced inflammation in mice were evaluated. Key findings β‐Sitosterol promoted a time‐ and dose‐dependent increase of the calcium uptake in activated neutrophils that was promptly reversed by nifedipine, BAPTA‐AM, LY294002, and colchicine. β‐Sitosterol inhibited myeloperoxidase and adenosine deaminase activity, and IL‐1β and TNF‐α levels. Conclusions β‐Sitosterol inhibited either myeloperoxidase and adenosine deaminase activity or IL‐1β and TNF‐α levels. This effect seemed to be mediated by the calcium uptake in activated neutrophils in a time‐ and concentration‐dependent manner through L‐type voltage dependent calcium channels, intracellular calcium, phosphoinositide kinase‐3, and microtubule modulation.     

Inflammatory cytokines tumour necrosis factor‐α and interleukin‐8 enhance airway smooth muscle contraction by increasing L‐type Ca2+ channel expression

Inflammation elevates intracellular calcium concentrations ([Ca²⁺]_i) in airway smooth muscle (ASM). The L‐type Ca²⁺ channel (L‐VDCC) plays an important role in regulating Ca²⁺ influx in ASM. However, the role of L‐VDCC in the inflammatory cytokine‐induced pathology of ASM remains unclear. In the present study, we used calcium imaging and isometric tension measurements to assess the role of L‐VDCC in agonist‐induced [Ca²⁺]_i rise and the associated contractions in mouse ASM, and we used immunoblotting to identify L‐VDCC protein expression levels in mouse ASM after exposure to tumour necrosis factor alpha (TNF‐α) or interleukin‐8 (IL‐8). Our results showed that high‐K⁺‐ or carbachol‐induced contractions of mouse ASM were significantly greater after pretreatment with TNF‐α or IL‐8 for 24 hours. Both verapamil and nifedipine, L‐VDCC inhibitors, abolished this increased contraction induced by TNF‐α or IL‐8 pretreatment. Moreover, TNF‐α treatment enhanced carbachol‐induced Ca²⁺ influx in ASM cells, and this effect was abrogated by verapamil. Additionally, immunoblotting results showed that preincubation of mouse ASM with TNF‐α or IL‐8 also enhanced L‐VDCC protein expression. On the basis of these findings, we concluded that proinflammatory cytokines, such as TNF‐α and IL‐8, increase the expression level of L‐VDCC, which in turn contributes to augmented agonist‐induced ASM contractions. This effect of inflammation on L‐VDCC expression in ASM may be associated with airway hyper‐responsiveness and involved in the development of asthma.     

Dexmedetomidine preconditioning for myocardial protection in ischaemia‐reperfusion injury in rats by downregulation of the high mobility group box 1‐toll‐like receptor 4‐nuclear factor κB signalling pathway

Pharmacological preconditioning reduces myocardial infarct size in ischaemia‐reperfusion (I‐R) injury. Dexmedetomidine, a selective α₂‐adrenoceptor agonist, has a proven cardioprotective effect when administered prior to I‐R, although the underlying mechanisms for this effect are not fully understood. We evaluated whether dexmedetomidine preconditioning could induce a myocardio‐protective effect against I‐R injury by inhibiting associated inflammatory processes through downregulation of the high mobility group box 1 (HMGB1)‐toll‐like receptor 4 (TLR4)‐nuclear factor κB (NF‐κB) signalling pathway. Seventy rats were randomly assigned to seven groups: a control and six test groups, involving I‐R for 30 and 120 minutes, respectively, in isolated rat hearts and different pretreatment protocols with dexmedetomidine (10 nmol/L) as well as yohimbine (1 μmol/L) and recombinant HMGB1 peptide (rHMGB1; 20 μg/L), alone or in combination with dexmedetomidine. Cardiac function was recorded; myocardial HMGB1, TLR4, and NF‐κB activities and levels of tumour necrosis factor‐α (TNF‐α) and interleukin‐6 (IL‐6) were measured as were lactate dehydrogenase (LDH) and creatine kinase (CK) in coronary outflow. Dexmedetomidine preconditioning significantly improved cardiac function (P<.05), downregulated the expression of HMGB1‐TLR4‐NF‐κB, reduced levels of TNF‐α and IL‐6 in isolated ventricles during I‐R injury, and significantly reduced CK and LDH levels in coronary outflow (P<.05). All of these effects were partially reversed by yohimbine (P<.05) or rHMGB1 (P<.05). Dexmedetomidine preconditioning alleviated myocardial I‐R injury in rats through inhibition of inflammatory processes associated with downregulation of the HMGB1‐TLR4‐NF‐κB signalling pathway via activation at α₂‐adrenergic receptors.     

The role of Syk/IĸB‐α/NF‐ĸB pathway activation in the reversal effect of BAY 61‐3606, a selective Syk inhibitor,on hypotension and inflammation in a rat model of zymosan‐induced non‐septic shock

Spleen tyrosine kinase (Syk), a non‐receptor tyrosine kinase, plays an important role in allergic diseases and inflammation. Syk triggers several intracellular signalling cascades including Toll‐like receptor signalling to activate inflammatory responses following fungal infection but the role of this enzyme in zymosan (ZYM)‐induced non‐septic shock and its impacts on hypotension and inflammation in rats is not well understood. This study was conducted to determine the effects of Syk inhibition on ZYM‐induced alterations in the expression and/or activities of Syk, inhibitor ?B (I?B)‐α, and nuclear factor‐?B (NF‐?B) p65. We also examined the effect of Syk inhibition on inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)‐2, and tumour necrosis factor (TNF)‐α, and activity of myeloperoxidase (MPO) that contribute to hypotension and inflammation. Administration of ZYM (500 mg/kg, ip) to male Wistar rats decreased blood pressure and increased heart rate. These changes were associated with increased expression and/or activities of Syk, NF‐κB p65, iNOS and COX‐2 and decreased expression of IκB‐α with enhanced levels of nitrite, nitrotyrosine, 6‐keto‐PGF_1α, and TNF‐α and activity of MPO in renal, cardiac and vascular tissues. ZYM administration also elevated serum and tissue nitrite levels. The selective Syk inhibitor BAY 61‐3606 (3 mg/kg, ip) given 1 hour after ZYM injection reversed all of these changes induced by ZYM. These results suggest that Syk/I?B‐α/NF‐?B pathway activation contributes to hypotension and inflammation caused by the production of vasodilator and proinflammatory mediators in the zymosan‐induced non‐septic shock model.     

Bis‐(3‐hydroxyphenyl) diselenide inhibits LPS‐stimulated iNOS and COX‐2 expression in RAW 264.7 macrophage cells through the NF‐kB inactivation

Objectives Previously, we reported that diaryl diselenide compounds have strong inhibitory effects on lipopolysaccharide (LPS)‐induced nitric oxide (NO) production in macrophages. In this study, we investigated the molecular mechanisms underlying NO suppression and prostaglandin E₂ (PGE₂) production by diaryl diselenide compounds, bis‐(2‐hydroxyphenyl) diselenide (DSE‐A), bis‐(3‐hydroxyphenyl) diselenide (DSE‐B), bis‐(4‐hydroxyphenyl) diselenide (DSE‐C), dipyridyl diselenide (DSE‐D) and diphenyl diselenide (DSE‐E). Methods The effect of these compounds on NO suppression and PGE₂ production was investigated in RAW 264.7 macrophages. Key findings Our data indicate that of the above, DSE‐B most potently inhibits NO and PGE₂ production, and that it also significantly reduces the releases of tumour necrosis factor (TNF)‐α, interleukin(IL)‐1β and IL‐6. Consistent with these observations, DSE‐B also reduced the protein levels of inducible NO synthase (iNOS) and cyclooxygenase‐2 (COX‐2), and the mRNA levels of iNOS, COX‐2, TNF‐α, IL‐1β and IL‐6. Furthermore, DSE‐B inhibited LPS‐induced nuclear factor‐κB (NF‐κB) activation, which was associated with the prevention of the inhibitor κB‐α (IκB‐α) degradation and a subsequent reduction in nuclear p65 protein levels. Conclusions Taken together, our data suggest that the anti‐inflammatory properties of DSE‐B are due to reduction in the expression of iNOS, COX‐2, TNF‐α, IL‐1β and IL‐6 through the down‐regulation of NF‐κB binding activity.     

Cobalt oxide nanoparticles induced oxidative stress linked to activation of TNF‐α/caspase‐8/p38‐MAPK signaling in human leukemia cells

The purpose of this study was to determine the intracellular signaling transduction pathways involved in oxidative stress induced by nanoparticles in cancer cells. Activation of reactive oxygen species (ROS) has some therapeutic benefits in arresting the growth of cancer cells. Cobalt oxide nanoparticles (CoO NPs) are an interesting compound for oxidative cancer therapy. Our results showed that CoO NPs elicited a significant (P <0.05) amount of ROS in cancer cells. Co‐treatment with N‐aceyltine cystine (an inhibitor of ROS) had a protective role in cancer cell death induced by CoO NPs. In cultured cells, the elevated level of tumor necrosis factor‐alpha (TNF‐α) was noted after CoO NPs treatment. This TNF‐α persuaded activation of caspase‐8 followed by phosphorylation of p38 mitogen‐activated protein kinase and induced cell death. This study showed that CoO NPs induced oxidative stress and activated the signaling pathway of TNF‐α‐Caspase‐8‐p38‐Caspase‐3 to cancer cells. Copyright © 2015 John Wiley & Sons, Ltd.     

Cyanidin‐3‐glucoside inhibits inflammatory activities in human fibroblast‐like synoviocytes and in mice with collagen‐induced arthritis

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by joint tissue inflammation. Cyanidin‐3‐glucoside (C3G) is a major component in the flavonoid family and has shown anti‐inflammatory, anti‐oxidant and anti‐tumour activity. In this study, we investigated the effects of C3G on lipopolysaccharides (LPS)‐induced inflammation on human rheumatoid fibroblast‐like synoviocytes (FLS) and on collagen‐induced arthritis (CIA) mice model. We treated FLS with C3G followed by LPS induction, the expressions of tumour necrosis factor alpha (TNF‐α), interleukin 1 beta (IL‐1β) and IL‐6 and the activation of nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) and mitogen‐activated protein kinase (MAPK) signalling pathway were analyzed. CIA was induced in mice and the arthritic mice were treated with C3G for 3 weeks. The disease severity was compared between control and C3G treated mice. The serum levels of TNF‐α, IL‐1β and IL‐6 were analyzed by ELISA. C3G inhibited LPS‐induced TNF‐α, IL‐1β and IL‐6 expression in FLS. Moreover, C3G inhibited LPS‐induced p65 production and IκBa, p38, ERK and JNK phosphorylation. Administration of C3G significantly attenuated disease in mice with CIA and decreased the serum level of TNF‐α, IL‐1β and IL‐6. C3G inhibited LPS‐induced inflammation in human FLS by inhibiting activation of NF‐κB and MAPK signalling pathway. C3G exhibited therapeutic effects in mice with CIA.     

Investigations of Free Anthraquinones from Rhubarb Against α‐Naphthylisothiocyanate‐induced Cholestatic Liver Injury in Rats

Abstract: The protective effects of anthraquinones from Rhubarb, a Chinese herbal medicine, consisting of the root and rhizome of Rheum palmatum L., R. tanguticum Maxim. Ex Balf., or R. officinale Baill. (family Polygonaceae) were investigated and compared in rats with liver injury induced by α‐naphthylisothiocyanate. α‐Naphthylisothiocyanate was given intragastrically in rats, liver injury with cholestasis developed within 36 hrs, as indicated by characteristic serum levels of glutamate‐pyruvate transaminase, glutamic oxaloacetic transaminase, total bilirubin, direct bilirubin, alkaline phosphatase, γ‐glutamyltransferase and total bile acid. The intragastrical administration of rhein, aloe‐emodin and physione to α‐naphthylisothiocyanate‐treated rats reduced significantly the serum level of both glutamate‐pyruvate transaminase, glutamic oxaloacetic transaminase and the serum total bilirubin, direct bilirubin, alkaline phosphatase, γ‐glutamyltransferase and total bile acid. For all hepatic biochemical markers and cholestasis index, rhein was most efficient. By comparison, the administration of emodin and chrysophanol did not reduce the serum levels of hepatic enzymes glutamate‐pyruvate transaminase and glutamic oxaloacetic transaminase but decreased the levels of serum total bilirubin, direct bilirubin, alkaline phosphatase, γ‐glutamyltransferase, and total bile acid, showing their partial protective effects on cholestatic liver injury. The liver in α‐naphthylisothiocyanate‐treated rats showed cholangiolitic hepatitis characterized by intrahepatic cholestasis, necrosis of hepatocytes and biliary epithelial cells and bile obstruction. The concurrent intragastrical administration of rhein reduced the severity of all morphological alteration, especially the neutrophil infiltration and sinusoid congestion. Rhein, aloe‐emodin, and physione all exhibited protective effects on hepatocytes and cholangiocytes against α‐naphthylisothiocyanate‐induced damage, whereas emodin and chrystophanol provided partial protection.     

Hepatoprotective effects of syringin on fulminant hepatic failure induced by D‐galactosamine and lipopolysaccharide in mice

The prognosis for fulminant hepatic failure (FHF) still remains extremely poor with a high mortality and, therefore, better treatments are urgently needed. Syringin, a main active substance isolated from Eleutherococcus senticosus, has been reported to exhibit immunomodulatory and anti‐inflammatory properties. In this study, we investigated the effects and underlying mechanisms of syringin on lipopolysaccharide (LPS) and D‐galactosamine (D‐GalN)‐induced FHF in mice. Mice were administered syringin (10, 30 and 100 mg kg^–1, respectively) intraperitoneally (i.p) 30 min before LPS/D‐GalN then mortality and liver injury were evaluated subsequently. We found that syringin dose‐dependently attenuated LPS/D‐GalN‐induced FHF, as indicated by reduced mortality, inhibited aminotransferase and malondialdehyde (MDA) content, an increased glutathione (GSH) concentration and alleviated pathological liver injury. In addition, syringin inhibited LPS/D‐GalN‐induced hepatic caspase‐3 activation and hepatocellular apoptosis, myeloperoxidase (MPO) activity and intercellular adhesion molecule‐1 (ICAM‐1) expression, as well as hepatic tissues tumor necrosis factor‐alpha (TNF‐α) production and NF‐κB activation in a dose‐dependent manner. These experimental data indicate that syringin might alleviate the FHF induced by LPS/D‐GalN through inhibiting NF‐κB activation to reduce TNF‐α production. Copyright © 2013 John Wiley & Sons, Ltd.     

Suppression of the TNF‐alpha level is mediated by Gan‐Lu‐Yin (traditional Chinese medicine) in human oral cancer cells through the NF‐kappa B,AKT, and ERK‐dependent pathways

Oral cancer is one of the major causes of deaths in the male population of Taiwan. Gan‐Lu‐Yin (GLY) is used for an adjuvant treatment of Traditional Chinese Medicine in clinical patients. In this study, we investigated the molecular mechanisms in oral cancer cell lines after exposure to GLY. The cytometric bead‐based array (CBA) method was used for the examining and analyzing of tumor necrosis factor‐alpha (TNF‐α) secretion level. TNF‐α mRNA expression was determined by real‐time PCR analysis. Nuclear factor kappa B (NF‐κB) activity and other relative proteins were determined by NF‐κB promoter assay, Western blotting, electrophoretic mobility shift assay (EMSA), and immuno‐staining analyses. GLY decreased the secretion of TNF–α from the oral cancer CAL 27 cells. Furthermore, 2000 μg/mL of GLY significantly suppressed TNF‐α mRNA expression of CAL 27 cells in a time‐dependent manner. GLY reduced the levels of proteins, including nuclear NF‐κB (p65 and p50), p‐IKK (ser176), p‐IκB, p‐AKT, p‐ERK, and nuclear Egr‐1 in a time and dose‐dependent manner. GLY also suppressed the NF‐κB activity and translocation in CAL 27 cells. We suggest that GLY might promote the cure of oral cancer through decreasing the level of TNF‐α cytokine, and these actions were mediated partially through the NF‐κB, AKT, and ERK‐dependent pathways in vitro. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1196–1205, 2016.