The return of the Scarlet Pimpernel: cobalamin in inflammation II - cobalamins can both selectively promote all three nitric oxide synthases (NOS), particularly iNOS and eNOS, and, as needed, selectively inhibit iNOS and nNOS

The up-regulation of transcobalamins [hitherto posited as indicating a central need for cobalamin (Cbl) in inflammation], whose expression, like inducible nitric oxide synthase (iNOS), is Sp1- and interferondependent, together with increased intracellular formation of glutathionylcobalamin (GSCbl), adenosylcobalamin (AdoCbl), methylcobalamin (MeCbl), may be essential for the timely promotion and later selective inhibition of iNOS and concordant regulation of endothelial and neuronal NOS (eNOS/nNOS.) Cbl may ensure controlled high output of nitric oxide (NO) and its safe deployment, because: (1) Cbl is ultimately responsible for the synthesis or availability of the NOS substrates and cofactors heme, arginine, BH(4) flavin adenine dinucleotide/flavin mononucleotide (FAD/FMN) and NADPH, via the far-reaching effects of the two Cbl coenzymes, methionine synthase (MS) and methylmalonyl CoA mutase (MCoAM) in, or on, the folate, glutathione, tricarboxylic acid (TCA) and urea cycles, oxidative phosphorylation, glycolysis and the pentose phosphate pathway. Deficiency of any of theNOS substrates and cofactors results in 'uncoupled' NOS reactions, decreasedNO production and increased or excessive O(2) (-), H(2)O(2), ONOO(-) and other reactive oxygen species (ROS), reactive nitric oxide species (RNIS) leading to pathology. (2) Cbl is also the overlooked ultimate determinant of positive glutathione status, which favours the formation of more benign NO species, s-nitrosothiols, the predominant form in which NO is safely deployed. Cbl status may consequently act as a 'back-up disc' that ensures the active status of antioxidant systems, as well as reversing and modulating the effects of nitrosylation in cell signal transduction.New evidence shows that GSCbl can significantly promote iNOS/ eNOS NO synthesis in the early stages of inflammation, thus lowering high levels of tumour necrosis factor-a that normally result in pathology, while existing evidence shows that in extreme nitrosative and oxidative stress, GSCbl can regenerate the activity of enzymes important for eventual resolution, such as glucose 6 phosphate dehydrogenase, which ensures NADPH supply, lactate dehydrogenase, and more; with human clinical case studies of OHCbl for cyanide poisoning, suggesting Cbl may regenerate aconitase and cytochrome c oxidase in the TCA cycle and oxidative phosphorylation. Thus, Cbl may simultaneously promote a strong inflammatory response and the means to resolve it.     

妊娠肝内胆汁淤积症孕妇血清和脐血清NO变化及胎盘NOS的表达

目的:通过测定妊娠肝内胆汁淤积症(ICP)患者血清、脐血清一氧化氮(NO)水平及胎盘一氧化氮合酶(NOS)的表达强度,探讨NO在ICP围产儿不良结局中的作用。方法:以ICP孕妇31例为ICP组,正常孕妇30例为对照组,采用亚硝酸盐法测定母血、脐血NO水平,免疫组化检测胎盘iNOS和eNOS表达强度,对染色强度进行图像分析。结果:ICP组脐血NO水平低于对照组(P<0.01),胎盘iNOS和eNOS表达强度均显著低于对照组(分别为P<0.001和P<0.01),两组孕妇血清NO水平无显著差异(P>0.05)。结论:ICP患者胎盘绒毛组织iNOS和eNOS表达降低,可能导致胎儿-胎盘循环阻力升高,是ICP患者发生胎儿宫内窘迫和早产的原因之一。     

Expression of NOS isoforms in internal spermatic veins of infertile men with varicocele

Although varicocele is a relatively common entity encountered in the evaluation of infertile men, the exact pathophysiology still remains unclear. Recently, as previously widely investigated in various parts of human circulatory system, nitric oxide synthase (NOS) and its product, nitric oxide (NO) have been thought to play a role in the development of varicocele and thus male infertility. In this study, we determined the concentration of NO metabolite and the expression of NOS isoforms in the internal spermatic (ISV) and superficial branch of inferior epigastric veins of infertile men with varicocele. The study included 60 infertile men with clinically unilateral or bilateral varicocele. Expression of inducible and endothelial NOS (iNOS and eNOS) isoforms were investigated in tissue arrays of internal spermatic and superficial branch of inferior epigastric veins with immunohistochemistry. NO metabolite (nitrite) levels were measured using the calorimetric method. A significantly higher expression of eNOS was observed in the varicose veins (mean score: 2.25 and 1.55, respectively; p?=?0.0001). However, statistically, there was no significant difference for expression of iNOS between varicose and control veins (p?=?0.094). The nitrite concentration and NOS expression were not found to be correlated with clinical variables (varicocele grade, maximum varicose vein diameter, and sperm concentration, motility, and morphology) (p?>?0.05). As a result, the significantly higher expression of eNOS in ISV may be responsible for the development of varicocele, although this finding is not accompanied by an increase in NO concentration. Still, the pattern of the relationship between varicocele and increased eNOS expression warrants further investigation.     

Expression of NOS isoforms in internal spermatic veins of infertile men with varicocele

Although varicocele is a relatively common entity encountered in the evaluation of infertile men, the exact pathophysiology still remains unclear. Recently, as previously widely investigated in various parts of human circulatory system, nitric oxide synthase (NOS) and its product, nitric oxide (NO) have been thought to play a role in the development of varicocele and thus male infertility. In this study, we determined the concentration of NO metabolite and the expression of NOS isoforms in the internal spermatic (ISV) and superficial branch of inferior epigastric veins of infertile men with varicocele. The study included 60 infertile men with clinically unilateral or bilateral varicocele. Expression of inducible and endothelial NOS (iNOS and eNOS) isoforms were investigated in tissue arrays of internal spermatic and superficial branch of inferior epigastric veins with immunohistochemistry. NO metabolite (nitrite) levels were measured using the calorimetric method. A significantly higher expression of eNOS was observed in the varicose veins (mean score: 2.25 and 1.55, respectively; p?=?0.0001). However, statistically, there was no significant difference for expression of iNOS between varicose and control veins (p?=?0.094). The nitrite concentration and NOS expression were not found to be correlated with clinical variables (varicocele grade, maximum varicose vein diameter, and sperm concentration, motility, and morphology) (p?>?0.05). As a result, the significantly higher expression of eNOS in ISV may be responsible for the development of varicocele, although this finding is not accompanied by an increase in NO concentration. Still, the pattern of the relationship between varicocele and increased eNOS expression warrants further investigation.     

The effect of cadmium toxicity on renal nitric oxide synthase isoenzymes

The aim of this study was to assess the cadmium (Cd) toxicity on renal nitric oxide synthase (NOS) isoenzymes. The study was carried out on 18 inbred male (Cd group: 10 and control group: 8) Wistar rats. Cd group received drinking water containing 15 mg/L Cd for 30 days; and at the end of the 30 days, plasma Cd was analysed. One kidney was snap frozen to assess the endothelial NOS (eNOS), inducible NOS (iNOS) and neuronal NOS (nNOS) expressions by Western blot analyses, and the other kidney was preserved for histopathological examination. Plasma Cd levels were significantly elevated in the Cd group. The Western blot analyses found higher levels of eNOS, iNOS and nNOS in the Cd group but only eNOS and nNOS levels were statistically significant. There was no difference in pathological assessment of the renal tissues. Cd toxicity increases NOS isoenzyme levels and may affect renal physiology.     

维生素C对镍诱导的肺泡巨噬细胞氧化损伤的保护作用

目的:研究维生素C(VC)拮抗Ni2O3的细胞氧化损伤的作用及对大鼠肺泡巨噬细胞iNOS诱导表达的影响。方法:采用体外细胞培养法测定在VC(25,50,100μmol/L)作用下,体外染镍肺泡巨噬细胞的死亡率及活力。同时观察细胞内丙二醛(MDA)、一氧化氮(nitricoxide,NO)和活性氧(ROS)的产生;超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)、诱导型一氧化氮合酶(nitricoxidesynthase,iNOS)活力的变化以及应用RT-PCR法测定iNOSmRNA的表达。结果:在体外染镍肺泡巨噬细胞中加入不同浓度的VC后可降低该细胞的死亡率并提高细胞的活力,可减少MDA、NO和活性氧的产生,降低NOS的活性,提高SOD、GSH-Px、CAT的活性,iNOSmRNA表达也较镍组降低。结论:镍可致细胞脂质过氧化,VC可通过提高抗氧化酶的活性,抑制iNOSmRNA的表达,减少活性氧及NO的产生,拮抗镍对肺泡巨噬细胞的氧化损伤。     

Nitric oxide activity in platelets of dengue haemorrhagic fever patients: the apparent paradoxical role of ADMA and l-NMMA

Dengue haemorrhagic fever (DHF) is a prevalent acute disease that occurs in patients infected by an arbovirus in tropical and subtropical regions. We have previously shown increased intraplatelet nitric oxide (NO) production in patients with dengue fever associated with reduced platelet aggregation. In this study, l-arginine transport as well as expression and activity of nitric oxide synthase (NOS) isoforms in the presence or absence of l-arginine analogues were examined in 23 DHF patients. l-arginine transport and NOS activity in platelets were increased in patients with DHF compared with controls. However, platelet endothelial NOS (eNOS) and inducible (iNOS) protein levels did not differ between healthy controls and DHF patients. Endogenous or exogenous analogues did not inhibit platelet NOS activity from DHF patients. In contrast, endogenous l-arginine analogues [N(G)-monomethyl-l-arginine (l-NMMA) and asymmetric dimethylarginine (ADMA)] inhibited NOS activity in platelets from healthy subjects. These results show the first evidence that the intraplatelet l-arginine-NO pathway is activated in DHF patients. The lack of inhibition of NO formation in vitro by all l-arginine analogues tested in DHF platelets may suggest another mechanism by which NOS activity can be regulated.     

Steroid hormones augment nitric oxide synthase activity and expression in rat uterus

Nitric oxide (NO) is synthesized in a variety of tissues, including rat uterus, from L-arginine by NO synthase (NOS), of which there are three isoforms, namely neuronal, endothelial and inducible NOS (nNOS, eNOS and iNOS, respectively). Nitric oxide is an important regulator of the biology and physiology of the organs of the reproductive system, including the uterus. Some studies have shown increased variation in NO production and NOS expression during the oestrous cycle. However, the factors that regulate NO production in the uterus remain unclear. Therefore, in the present study, we investigated the effect of sex steroids on NOS expression and activity in the ovariectomized rat uterus. Ovariectomized rats received progesterone (4 mg per rat) or 17beta-oestradiol (1 microg per rat). All rats were killed 18 h after treatment. Both progesterone and oestradiol were able to augment NOS activity. The effect of oestradiol was abolished by pre-incubation with 500 micro M aminoguanidine, an iNOS inhibitor, or by coadministration of oestradiol with 3 mg kg(-1) dexamethasone, but the effect of progesterone was not affected by these treatments. Uterine nNOS, eNOS and iNOS protein levels were assessed using Western blots. Ovariectomized rat uteri expressed iNOS and eNOS. Progesterone increased the expression of eNOS and iNOS, whereas oestradiol increased iNOS expression only. These results suggest that oestradiol and progesterone are involved in the regulation of NOS expression and activity during pregnancy and implantation in the rat.     

Differential effects of selective and non-selective NOS inhibition on renal arginine and protein metabolism during endotoxemia in rats

Background and aims: The kidney is the main endogenous producer of circulating arginine. Renal arginine disposal is directed to protein synthesis, urea production and nitric oxide synthesis. The administration of nitric oxide synthase inhibitors during sepsis may be beneficial or detrimental depending on the specificity of the inhibitor. We aimed to measure the effects of two NOS inhibitors, with different specificity, on renal arginine and protein turnover in a rat model of sepsis. Methods: Rats were subject to double hit endotoxemia and either L-NAME (non-specific), SMT (iNOS specific) or saline. Under anesthesia, vessels supplying and draining the kidney were catheterized. Systemic and intra-renal arginine and protein metabolism were measured using a primed continuous infusion of L-[2,3-(3)H]arginine and L-[2,6-(3)H]phenylalanine. Results: Non-specific NOS reduced systemic protein and arginine turnover, whereas selective iNOS inhibition did not. In the kidney, blood flow was reduced by L-NAME, but not by SMT. In conjunction with this, non-selective NOS inhibition increased renal protein breakdown, whereas selective iNOS inhibition increased renal arginine production. Conclusions: This study shows that non-selective NOS inhibition using L-NAME is detrimental for systemic and renal protein metabolism. Selective NOS inhibition stimulates renal arginine synthesis, without changing circulating arginine levels.     

Ellagic Acid Prevents L-NAME-Induced Hypertension via Restoration of eNOS and p47phox Expression in Rats

The effect of ellagic acid on oxidative stress and hypertension induced by N^ω-Nitro-l-arginine methyl ester hydrochloride (L-NAME) was investigated. Male Sprague-Dawley rats were administrated with L-NAME (40 mg/kg/day) for five weeks. L-NAME induced high systolic blood pressure (SBP) and increased heart rate (HR), hindlimb vascular resistance (HVR) and oxidative stress. Concurrent treatment with ellagic acid (7.5 or 15 mg/kg) prevented these alterations. Co-treatment with ellagic acid was associated with up-regulation of endothelial nitric oxide synthase (eNOS) protein production and alleviation of oxidative stress as indicated by decreased superoxide production in the vascular tissue, reduced plasma malondialdehyde levels, reduced NADPH oxidase subunit p47^phox expression and increased plasma nitrate/nitrite levels. Our results indicate that ellagic acid attenuates hypertension by reducing NADPH oxidase subunit p47^phox expression, which prevents oxidative stress and restores NO bioavailability.     

Nutritional lipid-induced oxidative stress leads to mitochondrial dysfunction followed by necrotic death in FaO hepatocytes

ObjectiveMitochondrial dysfunction and hepatocyte cell death have been reported in fatty liver and non-alcoholic steatohepatitis. Our aim in this study was to evaluate whether direct exposure of hepatocytes to extracellular fat could facilitate such deleterious effects.MethodsFaO hepatic cells treated with fat was used as an in vitro model for steatosis. FaO hepatocytes were exposed to 0.1% triacylglycerols using commercially available lipid emulsion (LE) for various periods and studied for production of reactive oxygen species (ROS), mitochondrial function, and cell death parameters. To study the type of cell death, high-mobility group box chromosomal protein 1cellular levels, DNA fragmentation, and caspase activity were evaluated.ResultsCells incubated with LE for 6 h exhibited a marked increase in the production of intracellular ROS. Using treatments with peroxisome proliferator-activated receptor activators, mitochondrial electron-transfer chain inhibitor, and different sources of LE that did or did not contain medium-chain triacylglycerols, the mitochondria were found to be the source of ROS. LE treatment resulted in phosphorylation of adenosine monophosphate–activated protein kinase, accompanied by a decrease in adenosine triphosphate levels. Changes in intracellular ROS and energy levels were followed by cell death. FaO hepatocytes showed a significant reduction in high-mobility group box chromosomal protein-1 and little DNA fragmentation. Incubation with LE for 24 h did not change caspase-3 activity, indicating that hepatocyte death was necrotic. The antioxidant N-acetylcysteine was able to attenuate the changes in intracellular energy levels and ROS levels and to prevent cell death after exposure to LE.ConclusionThese results suggest that exposure of FaO cells to LE leads to an increase in mitochondrial ROS production and a decrease in cellular energy levels followed by necrotic cell death.     

Roles of reactive oxygen species and mitochondria in cadmium-induced injury of liver cells

The roles of reactive oxygen species (ROS) and mitochondrial damage in the cadmium (Cd)-induced injury of liver cells were studied by using N-acetyl-L-cysteine (NAC) and acetyl-L-carnitine hydrochloride (ALCAR). After exposure of experimental rats to cadmium (Cd) for 16 h, mitochondrial membrane potential (MMP), ROS production, glutathione peroxidase (GSH-Px) activity, glutathione (GSH) content, malondialdehyde (MDA) content and DNA single-strand break (DNA-SSB) were analyzed. Loss of MMP, increase of ROS production, inhibition of GSH-Px activity, elevation of GSH content, rise of MDA content and DNA-SSB level suggest the participation of ROS and mitochondrion in Cd-induced injury of liver cell. NAC pretreatment attenuated oxidative stress, reversed the decline in GSH-Px activity and reduced GSH and MDA levels significantly. However, Cd-induced loss in MMP was significantly exacerbated by NAC. For another, ALCAR did not perform as well as NAC in terms of reducing ROS production, restoring GSH-Px activity and reducing GSH content. Nevertheless, it significantly improved the recovery of MMP and reduction of MDA content. In addition, conspicuous DNA damage was observed in the samples treated with NAC or ALCAR, indicating Cd could attack DNA through other pathways. These results suggest that oxidative stress or mitochondrial impairment plays a main role in different injuries respectively.     

AphaMax®, an Aphanizomenon Flos-Aquae Aqueous Extract,Exerts Intestinal Protective Effects in Experimental Colitis in Rats

Background: Aphanizomenon flos-aquae (AFA) is a unicellular cyanobacterium considered to be a “superfood” for its complete nutritional profile and beneficial properties. We investigated possible beneficial effects of an AFA extract, commercialized as AphaMax^®, containing concentrated amount of phycocyanins and phytochrome, in 2,4 dinitrobenzensulfonic acid(DNBS)-induced colitis in rats. Methods: Effects of preventive oral treatment of AphaMax^® (20, 50 or 100 mg/kg/day) in colitic rats were assessed and then macroscopic and microscopic analyses were performed to evaluate the inflammation degree. Myeloperoxidase (MPO) activity and NF-κB, pro-inflammatory citockines, cycloxygenase-2 (COX-2), and inducible NOS (iNOS) levels of expression were determined, as Reactive Oxygen Species (ROS) and nitrite levels. Results: AphaMax^® treatment attenuated the severity of colitis ameliorating clinical signs. AphaMax^® reduced the histological colonic damage and decreased MPO activity, NF-κB activation, as well as iNOS and COX-2 expression. AphaMax^® treatment improved the altered immune response associated with colonic inflammation reducing IL-1β, IL-6 expression. Lastly, AphaMax^® reduced oxidative stress, decreasing ROS and nitrite levels. Conclusions: Preventive treatment with AphaMax^® attenuates the severity of the inflammation in DNBS colitis rats involving decrease of the NF-kB activation, reduction of iNOS and COX-2 expression, and inhibition of oxidative stress. Due its anti-inflammatory and antioxidant proprieties AphaMax^® could be a good candidate as a complementary drug in inflammatory bowel disease (IBD) treatment.     

The Nutraceutical Antihypertensive Action of C-Phycocyanin in Chronic Kidney Disease Is Related to the Prevention of Endothelial Dysfunction

C-phycocyanin (CPC) is an antihypertensive that is not still wholly pharmacologically described. The aim of this study was to evaluate whether CPC counteracts endothelial dysfunction as an antihypertensive mechanism in rats with 5/6 nephrectomy (NFx) as a chronic kidney disease (CKD) model. Twenty-four male Wistar rats were divided into four groups: sham control, sham-treated with CPC (100 mg/Kg/d), NFx, and NFx treated with CPC. Blood pressure was measured each week, and renal function evaluated at the end of the treatment. Afterward, animals were euthanized, and their thoracic aortas were analyzed for endothelium functional test, oxidative stress, and NO production. 5/6 Nephrectomy caused hypertension increasing lipid peroxidation and ROS production, overexpression of inducible nitric oxide synthase (iNOS), reduction in the first-line antioxidant enzymes activities, and reduced-glutathione (GSH) with a down-expression of eNOS. The vasomotor response reduced endothelium-dependent vasodilation in aorta segments exposed to acetylcholine and sodium nitroprusside. However, the treatment with CPC prevented hypertension by reducing oxidative stress, NO system disturbance, and endothelial dysfunction. The CPC treatment did not prevent CKD-caused disturbance in the antioxidant enzymes activities. Therefore, CPC exhibited an antihypertensive activity while avoiding endothelial dysfunction.     

Protective role of oligonol from oxidative stress-induced inflammation in C6 glial cell

BACKGROUND/OBJECTIVES

Natural products or active components with a protective effect against oxidative stress have attracted significant attention for prevention and treatment of degenerative disease. Oligonol is a low molecular weight polyphenol containing catechin-type monomers and oligomers derived from Litchi chinensis Sonn. We investigated the protective effect and its related mechanism of oligonol against oxidative stress.

MATERIALS/METHODS

Oxidative stress in C6 glial cells was induced by hydrogen peroxide (H₂O₂) and the protective effects of oligonol on cell viability, nitric oxide (NO) and reactive oxygen species (ROS) synthesis, and mRNA expression related to oxidative stress were determined.

RESULTS

Treatment with oligonol inhibited NO and ROS formation under cellular oxidative stress in C6 glial cells. In addition, it recovered cell viability in a dose dependent-manner. Treatment with oligonol also resulted in down-regulated mRNA expression related to oxidative stress, nuclear factor kappa-B (NF-κB) p65, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS), compared with the control group treated with H₂O₂. In particular, expression of NF-κB p65, COX-2, and iNOS was effectively reduced to the normal level by treatment with 10 µg/mL and 25 µg/mL of oligonol.

CONCLUSIONS

These results indicate that oligonol has protective activity against oxidative stress-induced inflammation. Oligonol might be a promising agent for treatment of degenerative diseases through inhibition of ROS formation and NF-κB pathway gene expression.     

Effect of thiol-containing molecule cysteamine on the induction of inducible nitric oxide synthase in hepatocytes

BACKGROUND: Cysteamine, which is a known antioxidant and anti-inflammatory agent, is believed to be a key regulator of essential metabolic pathways in organisms. Cysteamine has beneficial effects in liver damaged by a variety of insults. During liver injury, inducible nitric oxide synthase (iNOS) is induced by lipopolysaccharide or proinflammatory cytokines, leading to excessive nitric oxide (NO) production. Accumulated evidence indicates that NO is an important factor associated with hepatic dysfunction. We examined whether cysteamine influences the induction of iNOS in hepatocytes. METHODS: Primary cultured rat hepatocytes were treated with interleukin (IL)-1beta in the presence and absence of cysteamine. NO production, iNOS induction, and iNOS signal were analyzed. RESULTS: IL-1beta stimulated the inhibitory protein kappaB (IkappaB)/nuclear factor kappaB (NFkappaB) pathway, resulting in the activation of NFkappaB (nuclear translocation and DNA binding), which was followed by the induction of iNOS and NO production. The addition of IL-1beta and cysteamine (1-4 mmol/L) markedly inhibited NO production, with a maximal effect at 4 mmol/L (80%-90% inhibition). Cysteamine also decreased the levels of iNOS protein and mRNA. Transfection experiments revealed that cysteamine decreased the transactivation activity of the iNOS promoter. An electrophoretic mobility shift assay demonstrated that cysteamine inhibited the activation of NFkappaB. Furthermore, cysteamine decreased the mRNA levels of the NFkappaB subunit p65 but increased those of the inhibitory protein IkappaB. CONCLUSIONS: These findings suggest that cysteamine inhibits iNOS induction at the step of NFkappaB activation. Further study is necessary to define the molecular basis of this effect of cysteamine on the regulation of NFkappaB and its pharmacologic implications.     

Elental® amino acid component has protective effects on primary cultured hepatocytes and a rat model of acute liver injury

Amino acids can exert protective effects on the liver either when administered as a medication or following an operation. In this study, we examined the protective effects of amino acids on the liver using in vitro and in vivo models by studying their influence on the induction of inducible nitric oxide synthase (iNOS) and nitric oxide production as a liver injury marker in cultured hepatocytes and liver-protective effects in d-galactosamine and lipopolysaccharide (GalN/LPS)-treated rats, respectively. Primary cultured rat hepatocytes were treated with interleukin (IL)-1β in the presence or absence of Elental® amino acid component (EleAA; 17 amino acids). Rats were pretreated with either EleAA or a diet containing selected amino acids followed by GalN/LPS injection. Survival rate and mRNA expression were analyzed. EleAA inhibited iNOS induction through reduction of mRNA synthesis and stability in cultured hepatocytes, indicating prevention of liver injury, but did not show a liver-protective effect in GalN/LPS rats. Among EleAA, Lys, Trp, His, and Arg (4AA) markedly decreased nitric oxide production and inhibited nuclear factor-κB (NF-κB) activation. In GalN/LPS rats, 4AA (3% of each amino acid in diet) increased survival rate by 50% and decreased mRNA expression of iNOS, tumor necrosis factor-α, and cytokine-induced neutrophil chemoattractant-1 in the liver. 4AA reduced NF-κB activation induced by GalN/LPS. 4AA inhibited the expression of inflammatory mediators, in part through inhibition of NF-κB activation in cultured hepatocytes and GalN/LPS-treated rats. The results suggest that EleAA has therapeutic potential for organ injuries including liver.