Downregulation Of NF-κB Expression Research Articles

α-Synuclein Negatively Regulates Nurr1 Expression Through NF-κB-Related Mechanism.

The nuclear receptor-related 1 protein (Nurr1) is critical for the development and survival of midbrain dopamine neurons that are predominantly affected and progressively degenerated in Parkinson’s disease (PD). The expression level of Nurr1 has been proposed to be modulated by α-synuclein (α-SYN), an important pathological hallmark of PD. However, the underlying molecular mechanisms of α-SYN-Nurr1 interaction are still rarely explored. In this study, we investigated the effect and mechanism of α-SYN on the transcription level of Nurr1. Our results showed that overexpression of α-SYN (WT or A53T) reduced Nurr1 and its downstream gene expressions. α-SYN neither affected the mRNA stability nor bound with the promoter of Nurr1, but modulated the transcription activity of Nurr1 promoter region ranging from −605 bp to −418 bp, which contains the binding site of nuclear factor-kappa B (NF-κB). Moreover, overexpression of α-SYN (WT or A53T) down-regulated NF-κB expression level, thereby inhibiting the transcription factor activity of NF-κB and decreasing the binding quantity of NF-κB with Nurr1 promoter. These findings may give us new insights to better understand the molecular mechanisms underlying the α-SYN-regulated Nurr1 function, which may fascinate the investigation of dopamine neuron degeneration in PD pathogenesis.

Gliclazide attenuates acetic acid-induced colitis via the modulation of PPARγ, NF-κB and MAPK signaling pathways

Ulcerative Colitis is a universal autoimmune disease with high incidence rates worldwide. It is characterized by the existence of many other concurrent immune-associated ailments, including diabetes. The used strategies for the management of this highly costing and complicated disease face great challenges. Therefore, the urge for new medication with fewer side effects and high efficacy is growing. The peroxisome proliferator-activated receptor-gamma (PPARγ) and nuclear factor Kappa-B (NF-κB) can be considered as crucial targets for the treatment of ulcerative colitis. Several studies reported the antioxidants, anti-inflammatory, and antiapoptotic actions of gliclazide and evaluated its cardioprotective and renoprotective effects. However, its impact on ulcerative colitis has never been investigated. This study delineated the effect of gliclazide administration on ulcerative colitis induced by acetic acid in rats and the underlying molecular mechanisms. Gliclazide (10 mg/kg; p.o) prominently decreased colon tissue injury as assessed by the histopathological analysis as well as myeloperoxidase, and intercellular adhesion molecule-1 levels. Gliclazide significantly alleviated the proinflammatory mediator, IL-6, promoted the anti-inflammatory cytokine, IL-10 and, withheld oxidative stress in the injured colon tissues. The protective effect of gliclazide was mediated through the upregulation of PPARγ and downregulation of NF-κB expression. The diminution of ulcerative colitis was also accompanied by an inhibition of the elevated activity and expression of mitogen-activated protein kinases and caspase-3 as assessed by Western blot and immunohistochemistry, respectively. Our findings spotlight, for the first time, the potential of the antidiabetic agent, gliclazide, to attenuate the experimentally induced ulcerative colitis. Therefore, gliclazide might be a propitious agent for the management of ulcerative colitis in diabetic patients.

Protective Effect of Genistein on Condylar Cartilage through Downregulating NF-κB Expression in Experimentally Created Osteoarthritis Rats.

Temporomandibular joint osteoarthrosis (TMJOA) is characterised by chronic inflammatory changes, with subsequent gradual loss of joint cartilage. NF-κB is a crucial transcription factor in the course of inflammatory and immune responses, which are involved in OA pathology activated by proinflammatory cytokines. Genistein is known to have anti-inflammation and modulation of metabolic pathways through repression of the NF-κB signaling pathway in inflammatory disease. But so far, studies on the effects of genistein on TMJOA are very limited. So, the purpose of this study is to investigate the protective effect of genistein against experimentally induced condylar cartilage degradation through downregulating NF-κB expression in created osteoarthritis rats in vivo. Male SD rats were created as temporomandibular joint osteoarthritis models and administered through oral gavage with low and high dosage genistein (30 mg/kg and 180 mg/kg, respectively) daily for 4 weeks. The morphological changes of the condylar cartilage were studied with HE and Masson staining. The expressions of p65 and inflammatory cytokines (IL-1β and TNFα) were detected using immunohistochemistry and real-time PCR. The results showed that experimentally created osteoarthritis reduced the condylar cartilage thickness of rats and increased the gene expression of cytokines (IL-1β and TNFα) and positive cells of p65. Genistein treatment had positive effects on the condylar cartilage renovation, while high dose genistein treatment had more significant effects on the reversing of OA changes and reduction of the expression of p65 and inflammatory cytokines (IL-1β and TNFα). The results indicated that high dose genistein treatment had obvious therapeutic effects on condyle cartilage damages of OA rats. The mechanism may be that genistein suppresses the NF-κB expression activated by inflammatory cytokines.

Role and related mechanism of resolvin D1 in lung ischemia reperfusion injury in rats

Objective: To investigate the role and related mechanism of resolvin D1 (RvD1) in lung ischemia-reperfusion injury (LIRI) in rats. Methods: Forty male Sprague-Dawley rats, 7-8 weeks, weighing 220-280 g, were divided into 4 groups using a random number table method: sham operation group, lung ischemia reperfusion control group, normal saline group, and RvD1 group. The rat model of LIRI was produced by 45 min of occlusion of the left hilum of lungs followed by 150 min reperfusion. In sham group, no blocking of the left hilum of lung after thoracotomy; Normal saline 2 ml/kg and RvD1 100 μg/kg were injected respectively at 10 min of reperfusion in normal saline group and RvD1 group. Blood samples were collected from the femoral vein for determination of interleukin (IL)-6, tumor necrosis factor (TNF)-α, soluble inter-cell adhesion molecules (sICAM-1) concentrations at 150 min of reperfusion. The rats were sacrificed after collection of blood samples and then lung tissues were taken for observation of the pathological changes and for measurement of lung wet/dry weight ratio (W/D). The the contents of malondialdehyde (MDA), monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-2 and the activity of myeloperoxidase (MPO) in lung tissues were determined. The protein relative expression of nuclear factor (NF)-κB in lung tissues was detected by Western blot. Lung tissue cell apoptosis was detected with TUNEL method. Results: The plasma level of IL-6, TNF-α, sICAM-1 in normal saline group and RvD1 group were significantly higher than those in the Sham group [(110±7), (100±4) vs (72±3) ng/L, (151±8), (153±6) vs (104±5) ng/L, (2 690±133), (2 760±167) vs (1 953±125) ng/L]. Besides, NF-κB protein relative expression level of lung tissues up-regulated [(0.681±0.033), (0.664±0.024) vs (0.292±0.011)] (all P<0.05). The W/D, apoptosis index, MDA, MCP-1, MIP-2 contents and MPO activity in lung ischemia reperfusion control group, normal saline group and RvD1 group were significantly higher than those in the Sham group [(5.92±0.31), (5.85±0.24), (5.06±0.08) vs (4.14±0.10), (32.9±1.5)%, (31.9±1.3)%, (17.7±1.8)% vs (8.1±0.6)%, (72.1±2.3), (66.7±3.7), (34.0±1.4) vs (22.0±0.8) nmol/mg, (3.99±0.28), (3.86±0.25), (2.66±0.16) vs (1.47±0.17) pg/mg, (9.45±0.53), (9.68±0.62), (7.62±0.22) vs (4.70±0.41) pg/mg, (3.01±0.18), (2.92±0.19), (1.58±0.11) vs (0.98±0.07) U/g] (all P<0.05). The plasma levels of the cytokines mentioned above, the W/D, the apoptosis index, MDA, MCP-1, MIP-2 contents and MPO activity in RvD1 group were significantly lower than those in the lung ischemia reperfusion control group [(63±4) vs (110±7) ng/L, (90±8) vs (151±8) ng/L, (1 835±182) vs (2 690±133) ng/L, (5.06±0.08) vs (5.92±0.31), (17.7±1.8)% vs (32.9±1.5)%, (34.0±1.4) vs (72.1±2.3) nmol/mg, (2.66±0.16) vs (3.99±0.28) pg/mg, (7.62±0.22) vs (9.45±0.53) pg/mg, (1.58±0.11) vs (3.01±0.18) U/g]. Besides, NF-κB protein relative expression level of lung tissues down-regulated [(0.313±0.012) vs (0.681±0.033)] (all P<0.05). Inflammatory cell infiltration in LIRI groups increased significantly, while it was significantly reduced in RvD1 group. Conclusion: RvD1 can effectively alleviate the tissue damage caused by lung ischemia-reperfusion through down-regulating NF-κB expression, relieving inflammatory reaction and oxidative stress, reducing apoptosis in rats.

Effects of lactoferrin on X-ray-induced intestinal injury in Balb/C mice

Intestinal injury is common after radiotherapy. We aim to investigate the effects of lactoferrin (Lf) on X-ray-induced intestinal injury in Balb/C mice. In assessing animal survival, a total of 40 animals were assigned randomly to 8Gy group, 2mg Lf+8Gy group, 4mg Lf+8Gy group, and 6mg Lf+8Gy group. Mice were administered with Lf intraperitoneally and exposed to single whole-body X-ray irradiation. Lf administration lasted for 3 days. Survival rate was compared among groups. For the observation of intestinal injury, a total of 60 animals were divided randomly into control group, 5Gy group, 2mg Lf+5Gy group, and 4mg Lf+5Gy group. Lf was administered once a day. Five mice in each group were randomly sacrificed at days 1, 3, and 9 after irradiation. Fasting blood was used to determine serum levels of pro-inflammatory cytokines IL-6 and TNF-α, and anti-inflammatorycytokine IL-10. Intestinal tissues were collected to examine histological changes and determine the protein expression levels of NF-κB, IKKα and IKKβ. Mean survival time was 4.30±1.34, 4.20±0.71, 5.75±2.44 and 6.70±2.54 days in four groups, respectively, with significantly longer duration in 6mg Lf+8Gy group than in the 8Gy group. Survival rate was significantly higher in 4mg Lf+8Gy group and 6mg Lf+8Gy group, compared with the 8Gy group. For intestinal histology, the radiation-induced injury was considerably improved in the 2mg Lf+5Gy and 4mg Lf+5Gy groups. Villus length and its ratio to crypt depth significantly increased in the two Lf intervention groups. Compared with 5Gy group, serum levels of IL-6 and TNF-α significantly decreased in the two Lf intervention groups at days 3 and 9. Furthermore, Lf significantly reduced the radiation-induced expression of IKKα/β and NF-κB at day 3 and/or day 9. Lf extended the survival time of radiated mice and improved intestinal injury by decreasing inflammatory cytokines and downregulating NF-κB expression.

BST-104, a Water Extract of Lonicera japonica, Has a Gastroprotective Effect via Antioxidant and Anti-Inflammatory Activities.

The gastroprotective effects of BST-104 (a water extract of Lonicera japonica) and the mechanisms involved were investigated in murine models of gastritis and peptic ulcer. The gastroprotective effects of BST-104 and its active components were evaluated in rat models of HCl/ethanol-induced gastritis and acetic acid-induced gastric ulcer. After orally administering BST-104, chlorogenic acid, rebamipide (positive control), or vehicle to each animal model, gastric lesion sizes, gastric mucus statuses, proinflammatory cytokine levels, and oxidative stress were measured. Superoxide dismutase (SOD), catalase, and malondialdehyde (MDA) levels and oxidized/reduced glutathione (GSH) ratios in gastric mucosal tissues were measured to evaluate oxidative stress. To clarify the action mechanism of BST-104, we investigated nuclear factor (NF)-κB pathway involvement by real-time polymerase chain reaction (PCR). In the acetic acid-induced ulcer model, oral administration of BST-104 at 50, 100, or 200 mg/kg significantly reduced gastric lesions by 38%, 43%, and 55%, respectively, compared with vehicle controls. BST-104 significantly increased gastric mucus contents and this was accompanied by higher levels of hexosamine, sialic acid, and prostaglandin E2 in gastric mucus. Furthermore, BST-104 treatment increased antioxidant activities, as evidenced by higher levels of catalase, SOD, and oxidized/reduced GSH and lower MDA levels. In addition, BST-104 significantly suppressed proinflammatory cytokine (tumor necrosis factor-α, interleukin [IL]-6, and IL-1β) increases, and real-time PCR showed that BST-104 significantly downregulated NF-κB expression. In summary, BST-104 and its active component, chlorogenic acid, were found to have gastroprotective effects by virtue of their antioxidant and anti-inflammatory properties through downregulation of NF-κB expression.

Pioglitazone ameliorates glomerular NLRP3 inflammasome activation in apolipoprotein E knockout mice with diabetes mellitus

ObjectiveThe NLRP3 inflammasome plays an important role in the pathogenesis of inflammation in diabetic nephropathy (DN). Pioglitazone (PIO) has been found to exert an anti-inflammatory effect in patients with diabetes mellitus, but it is still unclear whether PIO exhibits a similar effect in DN. We aimed to explore the effect and underlying mechanism of PIO on DN, as well as investigate if NLRP3 is a pharmacologic target of PIO.MethodsWe divided 48 apolipoprotein E (apoE) (-/-) mice into 4 groups: apoE (-/-), apoE (-/-) with PIO, diabetic apoE (-/-), and diabetic apoE (-/-) with PIO. Wild type male C57BL/6 mice were used as controls (n = 8 per group). After 8 weeks of PIO treatment, we examined the baseline characteristics and metabolic parameters of each group, and we used enzyme-linked immunosorbent assay (ELISA), western blot, and immunohistochemical staining to evaluate the expression levels of advanced glycation end products (AGEs), receptor for advanced glycation end products (RAGE), NLRP3, nuclear factor—kappa B (NF-κB), caspase-1, interleukin (IL)-18, and IL-1β in each group.ResultsCompared to the diabetic apoE (-/-) group, PIO treatment decreased blood glucose, cholesterol, serum blood urea nitrogen (BUN), and creatinine levels. It also depressed the glomerular mesangial expansion. PIO down-regulated expression of AGEs, RAGE, and NF-κB, all of which further depressed NLRP3, caspase-1, IL-18, and IL-1β levels.ConclusionPioglitazone can ameliorate diabetic renal damage, and this effect is related to the inhibition of renal AGE/RAGE axis activation and the down-regulation of NF-κB expression. These effects lead to a decline in NLRP3 levels and downstream secretion of inflammatory cytokines.

Rosmarinic Acid Protects against Inflammation and Cardiomyocyte Apoptosis during Myocardial Ischemia/Reperfusion Injury by Activating Peroxisome Proliferator-Activated Receptor Gamma.

The cardiac ischemia-reperfusion (I/R) injury greatly influences the therapeutic effect and remains an urgent challenge in clinical therapy. Polypharmacology opens a new therapeutic opportunity to design drugs with a specific target for improving the efficacy. In this study, we first forecasted that Rosmarinic acid (RosA) could be used for the treatment of cardiovascular disease using text mining, chemometric and chemogenomic methods. Consistent with the effect of the positive drug (pioglitazone, PIO), we subsequently validated that RosA pretreatment could restore the decreased cardiac hemodynamic parameters (LVDP, ± dp/dtmin, ± dp/dtmax and CF), decreased the infarct size and the cardiomyocyte apoptosis in a rat model of cardiac I/R injury. Furthermore, RosA pre-treatment inhibited the levels of inflammatory cytokines (IL-6, TNF-α and CRP), up-regulated PPARγ expression and down-regulated NF-κB expression in myocardial tissue isolated from the rat model of I/R-induced myocardial injury. In addition, the effects of RosA were reversed by co-treatment with PPAR-γ inhibitor GW9662 and T0070907, respectively. These data suggest that RosA attenuates cardiac injury through activating PPARγ and down-regulating NF-κB-mediated signaling pathway, which inhibiting inflammation and cardiomyocyte apoptosis in a rat model of cardiac I/R injury.

Total Flavonoids Extracted from Oxytropis falcata Bunge Improve Insulin Resistance through Regulation on the IKKβ/NF-κB Inflammatory Pathway.

Background. Insulin resistance (IR) is the main etiology of type 2 diabetes mellitus (T2DM). It has been known that total flavonoid extracts can markedly improve the hypoglycemic symptoms caused by IR. Nevertheless, the relevant molecular mechanism remains unclarified. Aim. This study aimed to investigate the antihyperglycemic effects and mechanism of the total flavonoid extract from Oxytropis falcata Bunge. Methods. STZ-induced T2DM rats (n = 35) were divided into 5 groups: model, low-, medium-, and high-dose total flavonoids, and pioglitazone groups. Ten healthy rats were used as controls. The serum insulin and inflammatory cytokines (MCP-1, TNF-α, and IL-6) level was measured by ELISA. The concentration of IRS-1, p-IRS-1, PKB p-PKB, PI3Kp85, and p-PI3K in skeletal muscles was determined by Western blot. The mRNA level of GLUT4, IκB, and NF-κB in skeletal muscle was detected by qRT-PCR. Results. The treatment of medium- and high-dose total flavonoids significantly reduced the FPG and P2hPG and enhanced insulin level in T2DM rats (P < 0.05). When compared with controls, the serum level of MCP-1, TNF-α, IL-6, IRS-1, and p-IRS-1 was significantly increased in T2DM rats, but the level of PKB, p-PKB, PI3Kp85, and p-PI3K expression was reduced (P < 0.05). The GLUT4 and IκB mRNA expression were significantly decreased, and NF-κB mRNA level was increased (P < 0.05). The treatment of low-, medium-, or high-dose total flavonoids markedly reversed the changes above (P < 0.05). Conclusion. Our study has confirmed the therapeutic effects of total flavonoids from Oxytropis falcata Bunge on IR. The flavonoids might reduce the production of inflammatory cytokines through downregulation of NF-κB expression in inflammatory pathway and regulate the IRS-1-PI3-K-PKB/Akt insulin pathway and thereby increased the GLUT4 expression.

Novel amino acid derivatives bearing thieno[2,3-d]pyrimidine moiety down regulate NF-κB in γ-irradiation mediated rat liver injury

A series of different amino acid-bearing thieno[2,3-d]pyrimidine moiety was synthesized via green chemistry aspects incorporating water as a solvent to give 2a–f, which were then acidified to attain the target compounds 3–9. Moreover, a tricyclic imidazothienopyrimidine of the glycine derivative (3) was synthesized to give (10). The title compounds were characterized by FT-IR, Mass, 13C-1HNMR spectroscopy and microanalysis. All the obtained amino acid-derivatives were screened for their post-irradiation protective efficacy in young rats. γ-Irradiation exposure was employed to induce oxidative stress. Radiation triggered significant alterations in the hematologic and blood - biochemical parameters. Further, a significant deterioration of hepatic antioxidant status was observed. Furthermore, a significant elevation of Nuclear Factor-kappa B (NF-κB) protein expression and level, which induced elevation in Cyclooxygenase-2 (COX-2) activity and cytochrome P450 2E1 (CYP2E1) gene expression were detected in hepatic tissues. Additionally, elevated levels of Tumor Necrosis Factor–alpha (TNF-α), and Interleukin-6 (IL-6) were perceived in serum of γ-irradiated young rats. However, most of the newly synthesized derivatives showed significant protective effects against injuries induced by γ-irradiation exposure, via ameliorating the altered hematopoietic system and activity of different biochemical parameters in blood. The results indicate that the antioxidant and anti-inflammatory mechanism of these compounds could be attributed to the significant down-regulation NF-κB protein expression in hepatic tissues. Subsequently, NF-κB could regulate TNF-α and IL-6 levels, COX-2 activity and CYP2E1 gene expression. Methionine derivative 8 was found to be the most active one. ConclusionThese new amino acid-derivatives showed promising outcomes as curative agents against γ-irradiation induced oxidative stress and physiological disturbance in different organs of young animals. Beyond, they may represent a novel selective class for treating liver injury induced by γ-irradiation in young rats, via down-regulation of NF-κB expression. Further investigations are warranted prior to the clinical use of these new compounds.

Effects of andrographolide on intrahepatic cholestasis induced by alpha-naphthylisothiocyanate in rats

Cholestasis is a cardinal manifestation of liver diseases but effective therapeutic approaches are limited. Therefore, alternative therapy for treating and preventing cholestatic liver diseases is necessary. Andrographolide, a promising anticancer drug derived from the medicinal plant Andrographis paniculata, has diverse pharmacological properties and multi-spectrum therapeutic applications. However, it is unknown whether andrographolide has a hepatoprotective effect on intrahepatic cholestasis. The aims of this study were to investigate the protective effect and possible mechanisms of andrographolide in a rat model of acute intrahepatic cholestasis induced by alpha-naphthylisothiocyanate (ANIT). Andrographolide was administered intragastrically for four consecutive days, with a single intraperitoneal injection of ANIT on the second day. Liver injury was evaluated biochemically and histologically together with hepatic gene and protein expression analysis. Rats pretreated with andrographolide prior to ANIT injection demonstrated lower levels of serum alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, gamma-glutamyltransferase, as well as bilirubin and bile acids as compared to rats treated with ANIT alone. Andrographolide also decreased the incidence and extent of periductular fibrosis and bile duct proliferation. Analysis of protein expression in livers from andrographolide-treated cholestatic rats revealed markedly decreased expression of alpha-smooth muscle actin and nuclear factor kappa-B (NF-κB). In conclusion, andrographolide has a potent protective property against ANIT-induced cholestatic liver injury. The mechanisms that underlie this protective effect are mediated through down-regulation of NF-κB expression and inhibition of hepatic stellate cell activation. These findings suggest that andrographolide could be a promising therapeutic option in prevention and slowing down the progression of cholestatic liver diseases.

Neuroprotective Effect of Hydrogen-Rich Saline in Global Cerebral Ischemia/Reperfusion Rats: Up-Regulated Tregs and Down-Regulated miR-21, miR-210 and NF-κB Expression.

Recently, it has been suggested that molecular hydrogen (H2) can selectively reduce the levels of hydroxyl radicals (.OH), and ameliorate oxidative and inflammatory injuries to organs in global cerebral ischemia reperfusion models. Global cerebral ischemia/reperfusion (I/R) can induce a sudden activation of inflammatory cytokines and later influence the systemic immunoreactivity which may contribute to a worse outcome. Regulatory T cells (Tregs) are involved in several pathological aspects of cerebral I/R. In addition, miRNA took part in the processes of cellular response to hypoxia. Since the expression of a specific set of miRNA called “hypoxamirs” is upregulated by hypoxia. Therefore, the aim of this study was to analyze the effect of HRS on I/R inducing cerebral damage, Tregs, and specific miRNA. Our results showed that rats undergone global cerebral I/R and treated with HRS have milder injury than I/R animals without HRS treatment. miR-210 expression in the hippocampus of the I/R group at 6, 24 and 96 h after reperfusion was significantly increased at each time point, while its expression in the group treated with HRS was significantly decreased. In addition, Tregs number in group I/R was decreased at each time points, while its number in the group treated with HRS was increased at 24 and 96 h after reperfusion. We focus on the relationship among Tregs, TGF-β1, TNF-α and NF-κB at 24 h, and we found that there is a high correlation among them. Therefore, our results indicated that the brain resuscitation mechanism in the HRS-treated rats may be related with the effect of upregulating the number of Treg cells.

HN protein of Newcastle disease virus sensitizes HeLa cells to TNF-α-induced apoptosis by downregulating NF-κB expression.

Hemagglutinin neuraminidase (HN) is a membrane protein of Newcastle disease virus (NDV) with the ability to induce apoptosis in many transformed cell lines. TNF-α is a multi-factorial protein that regulates cell survival, differentiation and apoptosis. In a previous study, we reported that HN protein induces apoptosis by downregulating NF-κB expression. Further, we speculated that downregulation of NF-κB expression might sensitize HeLa cells to TNF-α-mediated apoptosis. Therefore, the present study was undertaken to investigate if HN protein could sensitize HeLa cells to TNF-α and to examine the apoptotic potential of the HN protein and TNF-α in combination. The results revealed that the pro-apoptotic effects were more pronounced with the combination of HN and TNF-α than with HN or TNF-α alone, which indicates that the HN protein indeed sensitized the HeLa cells to TNF-α-induced cell death. The results of the study provide a mechanistic insight into the apoptotic action of HN protein along with TNF-α, which could be valuable in treating tumor types that are naturally resistant to TNF-α.

Plumbagin exerts protective effects in nucleus pulposus cells by attenuating hydrogen peroxide-induced oxidative stress, inflammation and apoptosis through NF-κB and Nrf-2.

Plumbagin, one of the constituents responsible for the various biological activities of Plumbagozeylanica has been demonstrated to possess antioxidant activity, which may inhibit lipid peroxidation in a dose- and time-dependent manner. In the present study, we aimed to examine the protective effects of plumbagin as well as the underlying mechansim through which plumbagin attenuates hydrogen peroxide(H2O2)-induced oxidative stress in nucleus pulposus cells(NPCs). For this purpose, the NPCs were incubated with fresh medium containing H2O2 (200µM) at 37˚C in a humidified 5%CO2 atmosphere for 6h, and cultured with various concentrations of plumbagin (0, 0.5, 1, 2, 5, 10 and 20µM). Treatment with plumbagin significantly increased the viability of the H2O2-exposed NPCs in a dose‑dependent manner. Moreover, plumbagin significantly reduced the generation of reactive oxygen species, lipid peroxidation, as well as the levels of tumor necrosis factor-α(TNF-α), interleukin(IL)-1β and IL-6 in the H2O2‑exposed NPCs. Glutathione(GSH) content, as well as the activity of catalase(CAT), superoxide dismutase(SOD) and glutathione peroxdiase(GSH-Px) were increased. We found that the administration of plumbagin significantly inhibited the activity of caspase-9 and-3, and downregulated NF-κB expression and upregulated Nrf-2 expression in the H2O2-exposed NPCs. Taken together, these findings suggest that plumbagin exerts neuroprotective effects in NPCs by attenuating H2O2‑induced oxidative stress, inflammation and apoptosis through mediating the expression of NF-κB and Nrf-2.

Nobiletin promotes antioxidant and anti-inflammatory responses and elicits protection against ischemic stroke in vivo

BackgroundPost-ischemic oxidative stress and inflammation play pivotal roles in the pathogenesis of ischemic stroke and may represent therapeutic targets. Nobiletin (NOB) has been reported to elicit a variety of biological effects through its anti-oxidant and anti-inflammatory properties. Our previous study has demonstrated the beneficial effect of NOB in ischemic stroke, but the underlying mechanisms remain poorly defined. We therefore further investigated the role of NOB in cerebral ischemia and its potential mechanisms. MethodsAdult male Sprague-Dawley rats were randomly assigned to five groups: Sham (sham-operated+0.05% Tween-80), permanent middle cerebral artery occlusion (pMCAO+0.9% saline), Vehicle (pMCAO+0.05% Tween-80), NOB-L (pMCAO+NOB 10mg/kg) and NOB-H (pMCAO+NOB 25mg/kg) groups. Rats were pre-administered intraperitoneally once daily for 3 days prior to ischemia and then received once again immediately after surgery. Neurological deficit, brain edema and infarct volume were evaluated at 24h after stroke. Immunohistochemistry, western blot and RT-qPCR were used to detect the expression of Nrf2, HO-1, SOD1, NF-κB and MMP-9. SOD1, GSH and MDA were measured by spectrophotometer. ResultsCompared with Vehicle group, neurological deficits and brain edema were relieved in NOB-H group, infarct volume was lessened in both NOB-L and NOB-H groups (P<0.05). NOB significantly increased the expression of Nrf2, HO-1, SOD1 and GSH, while decreased the levels of NF-κB, MMP-9 and MDA (P<0.05). ConclusionNOB may have a neuroprotective effect on cerebral ischemia, and this protection may be through upregulating Nrf2, HO-1 and downregulating NF-κB expression.

Down-regulation of NF-κB expression by n-3 fatty acid-rich linseed oil is modulated by PPARγ activation, eicosanoid cascade and secretion of cytokines by macrophages in rats fed partially hydrogenated vegetable fat.

The industrially produced partially hydrogenated vegetable fat (PHVF) contains trans fatty acid mostly comprising of elaidic acid (18:1 ∆9t). PHVF is used as a cooking medium in Southeast Asian countries. The purpose of this study is to evaluate the effects of dietary PHVF on inflammatory mediators and possible ameliorative effects of n-3 fatty acid (α-linolenic acid, ALA)-rich linseed oil (LSO) on the inflammatory mediators. Male Wistar weaning rats were fed AIN-93-purified diet supplemented with one of the following lipids for 60days, groundnut oil (GNO, 10wt%), PHVF (10wt%), LSO (10wt%), PHVF blended with LSO at 2.5, 5.0 and 7.5wt% levels. The final fat level in the diet was maintained at 10wt%. The macrophages from rats fed PHVF showed higher levels of total cholesterol and free cholesterol as compared to those from rats fed GNO and LSO. Macrophages from rats fed PHVF down-regulated the expression of PPARγ and up-regulated the expressions of cytosolic phospholipase A2, cyclooxygenase-2, 5-lipoxygenase and nuclear factor-kappa B p65. The macrophages from rats fed PHVF secreted higher levels of pro-inflammatory eicosanoids and cytokines. The rats fed PHVF blended with LSO at incremental amounts showed a significant reduction in the expressions of pro-inflammatory markers in dose-dependent manner. Detrimental effects of dietary PHVF in enhancing pro-inflammatory agents in rats could be significantly reduced by providing ALA (n-3 PUFA)-rich LSO.

Anthocyanins in Rubus fruits and antioxidant and anti-inflammatory activities in RAW 264.7 cells

Antioxidant and anti-inflammatory activities of ethanol extracts (E) and anthocyanin fractions (AF) from the 3 Rubus fruits blackberries, Korean raspberries, and black raspberries (BR) produced in Korea were determined. BR contained the highest levels of total polyphenols, flavonoids, anthocyanins, and the highest antioxidant activity. E and AF protected RAW 264.7 cells against H2O2-induced cytotoxicity. AF at 20 µg/mL significantly (p<0.05) decreased intracellular ROS levels (39.1-43.5%) and increased SOD (7.1-35.9%) and GPx (7.5-48.6%) activities in H2O2-treated RAW 264.7 cells, compared with controls. AF also increased Nrf2 protein expression. All AF significantly (p<0.05) reduced H2O2-induced DNA oxidation, compared with controls. AF suppressed NO (32.4-42.9%) and PGE2 (17.4-49.0%) production in LPS-stimulated RAW 264.7 cells along with a decrease in LPS-induced iNOS and COX-2 protein expressions. AF significantly (p<0.05) down-regulated NF-κB expression, compared with controls, via inhibition of I-κBα degradation in LPS-stimulated RAW 264.7 cells.

Ganoderic acid C1 isolated from the anti-asthma formula, ASHMI™ suppresses TNF-α production by mouse macrophages and peripheral blood mononuclear cells from asthma patients

Asthma is a heterogeneous airway inflammatory disease, which is associated with Th2 cytokine-driven inflammation and non-Th2, TNF-α mediated inflammation. Unlike Th2 mediated inflammation, TNF-α mediated asthma inflammation is generally insensitive to inhaled corticosteroids (ICS). ASHMITM, aqueous extract of three medicinal herbs—Ganoderma lucidum (G. lucidum), Sophora flavescens Ait (S. flavescens) and Glycyrrhiza uralensis Fischer (G. uralensis), showed a high safety profile and was clinically beneficial in asthma patients. It also suppresses both Th2 and TNF-α associated inflammation in murine asthma models. We previously determined that G. uralensis flavonoids are the key active compounds responsible for ASHMITM suppression of Th2 mediated inflammation. Until now, there are limited studies on anti-TNF-α compounds presented in ASHMITM. The objective of this study was to isolate and identify TNF-α inhibitory compounds in ASHMITM. Here we report that G. lucidum, but not the other two herbal extracts, S. flavescens or G. uralensis inhibited TNF-α production by murine macrophages; and that the methylene chloride (MC)–triterpenoid-enriched fraction, but not the polysaccharide-enriched fraction, contained the inhibitory compounds. Of the 15 triterpenoids isolated from the MC fraction, only ganoderic acid C1 (GAC1) significantly reduced TNF-α production by murine macrophages (RAW 264.7 cells) and peripheral blood mononuclear cells (PBMCs) from asthma patients. Inhibition was associated with down-regulation of NF-κB expression, and partial suppression of MAPK and AP-1 signaling pathways. Ganoderic acid C1 may have potential for treating TNF-α mediated inflammation in asthma and other inflammatory diseases.

Protective Effects of Turbinaria ornata and Padina pavonia against Azoxymethane-Induced Colon Carcinogenesis through Modulation of PPAR Gamma, NF-κB and Oxidative Stress.

The aim of this study was to investigate the antiproliferative and protective effects of the brown seaweeds, Turbinaria ornata and Padina pavonia, against azoxymethane (AOM)-induced colon carcinogenesis in mice. Both algal extracts showed anti-proliferative effects on the human carcinoma cell line HCT-116 in vitro, with T. ornata demonstrating a more potent effect. Male albino Swiss mice received intraperitoneal injections of AOM (10 mg/kg) once a week for two consecutive weeks and 100 mg/kg of either T. ornata or P. pavonia extracts. AOM-induced mice exhibited alterations in the histological structure of the colon, elevated lipid peroxidation and nitric oxide, declined glutathione content and reduced activity of superoxide dismutase and glutathione peroxidase. In addition, AOM induced downregulation of peroxisome proliferator activated receptor gamma (PPARγ) and p53 mRNA expression, with concomitant upregulation of nuclear factor-kappa B (NF-κB) in colon tissue. Administration of either algal extract markedly alleviated the recorded alterations. In conclusion, the current study suggests that T. ornata and P. pavonia, through their antioxidant and anti-inflammatory effects, are able to attenuate colon inflammation by downregulating NF-κB expression. Furthermore, the protective effects of both algae against AOM-initiated carcinogenesis were attributed, at least in part, to their ability to upregulate colonic PPARγ and p53 expression.

Upregulation of AIOLOS induces apoptosis and enhances etoposide chemosensitivity in Jurkat leukemia cells.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplastic disorder of immature hematopoietic precursors committed to T-cell lineage. T-ALL accounts for ~15% of pediatric ALL cases and is prone to early relapse. With new and improved treatment protocols, the prognosis of T-ALL has improved particularly in children; however, the outcome of relapsed T-ALL cases remains poor. The AIOLOS gene is necessary to control lymphocyte differentiation and may be a potential target of T-ALL therapy. In the present study, Jurkat cells were divided into three groups: untransfected (UT) control, lentiviral vector control (Lenti-Mock) and AIOLOS-overexpressing (Lenti-AIOLOS) groups. Lenti-AIOLOS Jurkat cells were constructed by lentiviral transduction; cell cycle analysis, apoptosis and cytotoxicity assays were then performed to evaluate the effects of AIOLOS on cell cycle distribution, apoptosis and cell chemosensitivity to etoposide of Jurkat cells in vitro. Moreover, the expression levels of genes associated with apoptosis and cell cycle were investigated by quantitative reverse transcription-polymerase chain reaction. Results showed that the percentage of Jurkat cells in the G0/G1 phase increased from 71.5 (UT) to 85.4% (Lenti-AIOLOS; P<0.05), yet the percentage of cells in the S-phase decreased from 15.1 (UT) to 11.6% (Lenti‑AIOLOS; P<0.05). The percentage of total apoptotic cells was significantly increased in the AIOLOS-transfected Jurkat cells (21.93%) compared with this percentage in the Lenti-Mock (13.35%) or the UT group (13.30%; P<0.05). Consistent with these results, AIOLOS overexpression induced P21 and P27 upregulation and CCND3 and SKP2 downregulation. Furthermore, AIOLOS overexpression synergistically increased the cytotoxic effects of etoposide and downregulated NF-κB expression. Our findings revealed that lentivirus-mediated AIOLOS overexpression in Jurkat cells induced cell apoptosis, arrested the cell cycle at the G0/G1 phase, and synergistically increased the sensitivity of Jurkat cells to etoposide by inhibiting NF-κB activity.