Thromboxane A2 increases endothelial permeability through upregulation of interleukin-8

Thromboxane A₂ (TXA₂), a major prostanoid formed from prostaglandin H₂ by thromboxane synthase, is involved in the pathogenesis of a variety of vascular diseases. In this study, we report that TXA₂ mimetic U46619 significantly increases the endothelial permeability both in vitro and in vivo. U46619 enhanced the expression and secretion of interleukin-8 (IL-8), a major inducer of vascular permeability, in endothelial cells. Promoter analysis showed that the U46619-induced expression of IL-8 was mainly regulated by nuclear factor-κB (NF-κB). U46619 induced the activation of NF-κB through IκB kinase (IKK) activation, IκB phosphorylation and NF-κB nuclear translocation. Furthermore, the inhibition of IL-8 or blockade of the IL-8 receptor attenuated the U46619-induced endothelial cell permeability by modulating the cell-cell junctions. Overall, these results suggest that U46619 promotes vascular permeability through the production of IL-8 via NF-κB activation in endothelial cells.     

Role of nitric oxide and nuclear factor-κB in the CYP2E1 potentiation of tumor necrosis factor α hepatotoxicity in mice

Induction of CYP2E1 by pyrazole (PY) potentiated the hepatotoxicity induced by TNFα in mice. We evaluated the role of nitrosative and oxidative stress and the NF-κB activation pathway in this liver injury. The iNOS inhibitor N-(3-aminomethyl)benzylacetamindine (1400W) or the antioxidant N-acetyl-l-cysteine (NAC) prevented this liver injury. TNFα plus PY treatment triggered radical stress in the liver with increased lipid peroxidation and decreased glutathione and caused mitochondrial damage as reflected by elevated membrane swelling and cytochrome c release. The radical stress and mitochondrial damage were prevented by 1400W and NAC. TNFα plus PY treatment elevated 3-nitrotyrosine adduct formation and induced NOS2 in the liver; 1400W and NAC blocked these changes. A lower extent of liver injury and oxidative stress was found in NOS2^?/? mice treated with TNFα plus PY compared with wild-type controls. Neither 1400W nor NAC modified CYP2E1 activity or protein. Activation of JNK and p38MAPK was weaker in TNFα plus PY-treated NOS2^?/? mice and 1400W and NAC blocked the activation of JNK and p38MAPK in wild-type mice. IKKα/β protein levels were decreased by TNFα plus PY treatment, whereas IκBα and IκBβ protein levels were elevated compared with saline, PY, or TNFα alone. NF-κB DNA binding activity was increased by TNFα alone but lowered by TNFα plus PY. All these changes were blocked by 1400W and NAC. NF-κB activation products such as Bcl-2, Bcl-X_L, cFLIP_S, cFLIP_L, and Mn-SOD were reduced by TNFα plus PY and restored by 1400W or NAC. We conclude that TNFα plus CYP2E1 induces oxidative/nitrosative stress, which plays a role in the activation of JNK or p38MAPK and mitochondrial damage. These effects combine with the blunting of the NF-κB activation pathways and the synthesis of protective factors to cause liver injury.     

Hydrogen peroxide and tumour necrosis factor-α induce NF-κB-DNA binding in primary human T lymphocytes in addition to T cell lines

Reactive oxygen intermediates (ROIs), such as hydrogen peroxide (H₂O₂), have been implicated as second messengers in the activation of NF-κB by a variety of stimuli, including tumour necrosis factor-alpha (TNF-α). The aim of the present study was to examine the effects of ROIs on NF-κB activation in primary human CD3+ T lymphocytes and human peripheral blood mononuclear cells (PBMCs). For comparison purposes, Jurkat T cells (subclones JR and JE6.1) were also investigated. Cells were incubated in the presence of either H₂O₂ or TNF-α and nuclear proteins were extracted. NF-κB binding was assessed by electrophoretic mobility shift assays (EMSAs). The concentration of H₂O₂ required to activate NF-κB in human primary CD3+ T lymphocytes was as low as 1 μM. In contrast, much higher concentrations of H₂O₂ were required to activate NF-κB in PBMCs and in the JR subclone of Jurkat T cells. H₂O₂-induced NF-κB activation was not observed in the JE6.1 subclone of Jurkat T cells. NF-κB was activated by TNF-α in all four cell types tested. In PBMCs and Jurkat T cells (subclones JR and JE6.1), this activation could be inhibited by pre-treatment with the antioxidants, pyrrolidine dithiocarbamate (PDTC) and N-acetyl-l-cysteine (NAC). Our results support a role for ROIs in NF-κB-DNA binding in human primary T lymphocytes.     

Oxidative stress induces inactivation of protein phosphatase 2A,promoting proinflammatory NF-κB in aged rat kidney

The molecular inflammation hypothesis of aging proposes that redox dysregulation causes an age-related activation of NF-κB and its signaling to upregulate various proinflammatory genes. In the present study, we focused on the inactive form of the protein phosphastase 2 A (PP2A). More specifically, we aimed to define the correlation between PP2A inactivation and NF-κB activation by age-related oxidative stress. Experimentations were designed to determine the effect of oxidative stress-induced PP2A inactivation on NF-κB activity, utilizing prooxidants t-BHP and AAPH, the PTP inhibitor Na₃VO₄, and the PP2A inhibitor Calyculin A and PP2A siRNA, in HEK293T cells. We also assessed the phosphorylation of PP2A catalytic subunit (PP2Ac) and the activities of PP2A and NF-κB in aged rat kidney, utilizing aging-retarding 40% calorie restriction (CR) −60% of food intake and inflammation-triggering LPS paradigms. Results revealed that an oxidative stress-induced PTK/PTP imbalance led to phosphorylation of PP2Ac, following exposures to t-BHP, AAPH, and Na₃VO₄ in HEK293T cells. Subsequently, we found that Calyculin A and PP2A siRNA activates NIK/IKK and MAPKs, leading to upregulation of NF-κB and its dependent oxidative stress. Also, the contrasting relation between PP2A inactivation and NF-κB activation was confirmed by AAPH-induced oxidative status in mice, and non-induced normal status or LPS-induced inflammatory status in aged rats while the antioxidative, anti-inflammatory, anti-aging effects of CR significantly blunted these actions. Thus, we present evidence that PP2A inactivation via PTK/PTP imbalance provoked by oxidative stress causes NF-κB activation, which contributes to the accumulation of oxidative stress in aged rat kidney.     

Forsythiaside A Exhibits Anti-inflammatory Effects in LPS-Stimulated BV2 Microglia Cells Through Activation of Nrf2/HO-1 Signaling Pathway

Inflammation and oxidative stress have been reported to play critical roles in the pathogenesis of neurodegenerative disease. Forsythiaside A, a phenylethanoside product isolated from air-dried fruits of Forsythia suspensa, has been reported to have anti-inflammatory and antioxidant effects. In this study, the anti-inflammatory effects of forsythiaside A on LPS-stimulated BV2 microglia cells and primary microglia cells were investigated. The production of inflammatory mediators TNF-α, IL-1β, NO and PGE₂ were detected in this study. NF-κB, nuclear factor-erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1) expression were detected by western blot analysis. Our results showed that forsythiaside A significantly inhibited LPS-induced inflammatory mediators TNF-α, IL-1β, NO and PGE₂ production. LPS-induced NF-κB activation was suppressed by forsythiaside A. Furthermore, forsythiaside A was found to up-regulate the expression of Nrf2 and HO-1. In conclusion, this study demonstrates that forsythiaside A inhibits LPS-induced inflammatory responses in BV2 microglia cells and primary microglia cells through inhibition of NF-κB activation and activation of Nrf2/HO-1 signaling pathway.     

Downregulation of nuclear factor-kappa B (NF-κB) pathway by silibinin in human monocytes challenged with Paracoccidioides brasiliensis

AimsSilibinin is the major active component of silymarin, a polyphenolic plant flavonoid that has anti-inflammatory effects. The modulatory effect of silibinin on monocyte function against Paracoccidioides brasiliensis (Pb18) has not yet been demonstrated. The present study investigated whether the effect of silibinin on nuclear factor-kappa B (NF-κB) pathways may affect the production of tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), transforming growth factor beta (TGF-β1), prostaglandin E₂ (PGE₂), nitric oxide (NO) and fungicidal activity of human monocytes challenged in vitro with Pb18.Main methodsPeripheral blood monocytes from healthy individuals were treated with silibinin and challenged with Pb18 for 18 h. TNF-α, IL-10, TGF-β1 and PGE₂ expression were determined by immunoenzymatic assay (ELISA) and NO release was determined by the accumulation of nitrite in culture supernatants. Fungicidal activity of monocytes was analyzed after treatment with interferon-gamma plus silibinin and challenge with Pb18. NF-κB activation in cultured monocytes was evaluated by flow cytometry and ELISA.Key findingsSilibinin partially inhibited p65NF-κB activation as the number of cells expressing this factor was reduced and the concentration of nuclear p65NF-κB was low, compared to untreated controls. The addition of silibinin also resulted in suppression of TNF-α, IL-10, TGF-β1, PGE₂ and NO production but did not affect the fungicidal activity of monocytes against Pb18.SignificanceSilibinin exerts anti-inflammatory and anti-fibrotic effects on CD14± human monocytes challenged by Pb18 by partial inhibition of p65NF-κB activation.     

Oxidative stress-induced expression of HSP70 contributes to the inhibitory effect of 15d-PGJ2 on inducible prostaglandin pathway in chondrocytes

The inhibitory effect of 15-deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂) on proinflammatory gene expression has been extensively documented and frequently ascribed to its ability to prevent NF-κB pathway activation. We and others have previously demonstrated that it was frequently independent of the peroxisome proliferator activated receptor (PPAR)γ activation. Here, we provide evidence that induction of intracellular heat shock protein (HSP)70 by oxidative stress is an additional regulatory loop supporting the anti-inflammatory effect of 15d-PGJ₂ in chondrocytes. Using real-time quantitative PCR and Western blotting, we showed that 15d-PGJ₂ stimulated HSP70, but not HSP27 expression while increasing oxidative stress as measured by spectrofluorimetry and confocal spectral imaging. Using N-acetylcysteine (NAC) as an antioxidant, we demonstrated further that oxidative stress was thoroughly responsible for the increased expression of HSP70. Finally, using an HSP70 antisense strategy, we showed that the inhibitory effect of 15d-PGJ₂ on IL-1-induced activation of the NF-κB pathway, COX-2 and mPGES-1 expression, and PGE₂ synthesis was partly supported by HSP70. These data provide a new anti-inflammatory mechanism to support the PPARγ-independent effect of 15d-PGJ₂ in chondrocyte and suggest a possible feedback regulatory loop between oxidative stress and inflammation via intracellular HSP70 up-regulation. This cross talk is consistent with 15d-PGJ₂ as a putative negative regulator of the inflammatory reaction.     

MAPKs and NF-κB differentially regulate cytokine expression in the diaphragm in response to resistive breathing: the role of oxidative stress

Inspiratory resistive breathing (IRB) induces cytokine expression in the diaphragm. The mechanism of this cytokine induction remains elusive. The roles of MAPKs and NF-κB and the impact of oxidative stress in IRB-induced cytokine upregulation in the diaphragm were studied. Wistar rats were subjected to IRB (50% of maximal inspiratory pressure) via a two-way nonrebreathing valve for 1, 3, or 6 h. Additional groups of rats subjected to IRB for 6 h were randomly assigned to receive either solvent or N-acetyl-cysteine (NAC) or inhibitors of NF-κB (BAY-11-7082), ERK1/2 (PD98059), and P38 MAPK (SB203580) to study the effect of oxidative stress, NF-κB, and MAPKs in IRB-induced cytokine upregulation in the diaphragm. Quietly breathing animals served as controls. IRB upregulated cytokine (IL-6, TNF-α, IL-10, IL-2, IL-1β) protein levels in the diaphragm and resulted in increased activation of MAPKs (P38, ERK1/2) and NF-κB. Inhibition of NF-κB and ERK1/2 blunted the upregulation of all cytokines except that of IL-6, which was further increased. P38 inhibition attenuated all cytokine (including IL-6) upregulation. Both P38 and ERK1/2 inhibition decreased NF-κB/p65 subunit phosphorylation. NAC pretreatment blunted IRB-induced cytokine upregulation in the diaphragm and resulted in decreased ERK1/2, P38, and NF-κB/p65 phosphorylation. In conclusion, IRB-induced cytokine upregulation in the diaphragm is under the regulatory control of MAPKs and NF-κB. IL-6 is regulated differently from all other cytokines through a P38-dependent and NF-κB independent pathway. Oxidative stress is a stimulus for IRB-induced cytokine upregulation in the diaphragm.     

SIRT3 protects cardiomyocytes from oxidative stress-mediated cell death by activating NF-κB

Oxidative stress-mediated cell death in cardiomyocytes reportedly plays an important role in many cardiac pathologies. Our previous report demonstrated that mitochondrial SIRT3 plays an essential role in mediating cell survival in cardiac myocytes, and that resveratrol protects cardiomyocytes from oxidative stress-induced apoptosis by activating SIRT3. However, the exact mechanism by which SIRT3 prevents oxidative stress remains unknown. Here, we show that exposure of H9c2 cells to 50 μM H₂O₂ for 6 h caused a significant increase in cell death and the down-regulation of SIRT3. Reactive oxygen species (ROS)-mediated NF-κB activation was involved in this SIRT3 down-regulation. The SIRT3 activator, resveratrol, which is considered an important antioxidant, protected against H₂O₂-induced cell death, whereas the SIRT inhibitor, nicotinamide, enhanced cell death. Moreover, resveratrol negatively regulated H₂O₂-induced NF-κB activation, whereas nicotinamide enhanced H₂O₂-induced NF-κB activation. We also found that SOD2, Bcl-2 and Bax, the downstream genes of NF-κB, were involved in this pathological process. These results suggest that SIRT3 protects cardiomyocytes exposed to oxidative stress from apoptosis via a mechanism that may involve the NF-κB pathway.     

Fibrinopeptide A induces C-reactive protein expression through the ROS-ERK1/2/p38-NF-κB signal pathway in the human umbilical vascular endothelial cells

Atherosclerosis is a chronic inflammatory disease of the arterial wall. Inflammation causes endothelial injury and dysfunction, which is an initial step of atherosclerosis. Fibrinopeptide A (FPA) is a biomarker of the activation of the coagulation system, and a high concentration of FPA in the blood occurs in patients with ischemic cardiocerebrovascular diseases. The present research observed that FPA stimulated the generation of C-reactive protein (CRP), IL-1β, and IL-6 in human umbilical vascular endothelial cells (HUVECs); and anti-IL-1 β and anti-IL-6 neutralizing antibodies did not alter FPA-induced CRP expression in HUVECs. The subchronic administration of FPA into rats increased the plasma FPA and CRP levels. Further studies showed that FPA stimulated superoxide anion generation, activated ERK1/2 and p38, promoted nuclear factor κB (NF-κB) nuclear translocation, and raised the NF-κB level in the nuclei of HUVECs. Antioxidant N-acetylcysteine (NAC), complex II inhibitor thenoyltrifluoroacetone (TTFA), and NADPH oxidase inhibitor diphenyleneiodonium (DPI) inhibited FPA-stimulated generation of superoxide anion, and NAC reduced FPA-induced expressions of the phosphorylated ERK1/2 and p38. NAC, TTFA, DPI, inhibitors of ERK1/2, p38, and NF-κB all downregulated FPA-induced CRP expression. These results indicate that FPA induces CRP expression in HUVECs via the ROS-ERK1/2/p38-NF-κB signal pathway. Moreover, this is the first report that FPA produces a proinflammatory effect on the vascular endothelial cells.     

Reactive oxygen species mediate IL-8 expression in Down syndrome candidate region-1-overexpressed cells

Reactive oxygen species (ROS) have been considered to mediate inflammation in Down syndrome (DS). The present study is purposed to examine the mechanism of increased ROS levels and inflammatory cytokine IL-8 expression in Down syndrome candidate region-1 (DSCR1)-transfected cells, by determining ROS levels, IL-8 expression, NF-κB activation, and SOD1 levels in human embryonic kidney (HEK) 293 cells. The cells were treated with an antioxidant N-acetyl cysteine (NAC) or a calcium chelator BAPTA and stimulated with or without IL-1β. As a result, basal levels of ROS, IL-8, and NF-κB-DNA binding activity were higher, and basal SOD1 levels were higher in DSCR1-transfected cells than pcDNA-transfected cells. BAPTA and NAC inhibited increase in ROS (intracellular and mitochondrial levels) in DSCR-1-transfected cells without treatment of IL-1β. DSCR1 transfection-induced changes were increased by treatment with IL-1β, which was suppressed by NAC and BAPTA. Transfection of SOD1 inhibited ROS levels in DSCR1-transfected cells. In conclusion, ROS activate NF-κB and IL-8 induction in DSCR1-transfected cells in a calcium-dependent manner, which is augmented by IL-1β since IL-1β increases calcium and ROS levels in the cells. Reducing ROS levels by treatment of antioxidants may be beneficial for preventing DS-associated inflammation by suppressing cytokine expression.     

Electrolytically generated acid functional water inhibits NF-κB activity by attenuating nuclear-cytoplasmic shuttling of p65 and p50 subunits

Background: Human β-defensin 2 (hBD2) gene expression is dependent on nuclear factor kappa B (NF-κB) activity. We have previously demonstrated that electrolytically generated acid functional water (FW) induces the expression of hBD2 in the human oral squamous cell carcinoma (OSCC) cell line Ca9-22. However, the induction was not dependent on NF-κB activity; in fact, FW inhibited NF-κB activity. Therefore, we hypothesized that FW might reduce spontaneous interleukin 8 (IL-8) secretion by Ca9-22 cells, which is heavily dependent on NF-κB activity. This study aimed at demonstrating the inhibitory effect of FW on NF-κB activity. Methods: Ca9-22 cells were incubated with FW, and spontaneous IL-8 secretion was observed by enzyme-linked immunosorbent assay. Luciferase assay was performed using the 5′-untranslated region of the IL-8 gene. The steps of NF-κB activation blocked by FW were evaluated by localization of the NF-κB subunits p65 and p50 by immunofluorescence staining. Western blotting was further performed to confirm the changes in NF-κB subunit localization. Results: The Ca9-22 cells spontaneously secreted IL-8, which was rapidly and drastically inhibited by FW treatment. The luciferase assay demonstrated the inhibitory action of FW, which was diminished by deletion of the NF-κB binding site from this construct. FW treatment altered the distribution of both the p65 and p50 subunits. P65, which was localized in the nucleus during the resting state, moved to the cytoplasm after FW treatment, whereas, p50, localized in the cytoplasm during the resting state, moved to the nucleus subsequent to FW treatment. Conclusions: The results from this study indicate that FW might inhibit spontaneous IL-8 secretion by redistribution of the NF-κB subunits within the cells.     

Immunohistochemical localization of a H2S-generating enzyme and its potential molecular target in hepatopancreas of the freshwater crayfish Procambarus clarkii

ABSTRACT

There is increasing evidence showing that the gasotransmitter hydrogen sulfide (H₂S) and NF-κB-mediated signaling may play an important role in regulating immune response. However, essentially nothing is known about them in crustaceans, especially the red swamp crayfish (Procambarus clarkii). In this work, we used immunohistochemical method to investigate, for the first time, the presence and distribution of a H₂S biosynthetic enzyme (cystathionine β-synthase, CBS) and a member of NF-κB family (RelA/p65) in crayfish hepatopancreas. Results indicated that (1) CBS-like and NF-κB p65-like immunopositive cells coexisted in hepatopancreas; (2) NF-κB p65 was diffusely localized and highly expressed in the cytoplasm of R- and B-cells, but CBS was observed mainly in connective tissue, especially in sinusoids; (3) and normally, at mean optical density (MOD) level, their expression was about 0.0032 ± 0.0028 and 0.0072 ± 0.0039, and proportion (%) of positive cells was 14.37 ± 5.76 and 23.5 ± 14.3, respectively.     

Mitofusin 2 deficiency leads to oxidative stress that contributes to insulin resistance in rat skeletal muscle cells

Mitofusin 2 (Mfn2) is a dynamin-like protein anchored in the outer mitochondrial membrane that plays a crucial role in ensuring optimal mitochondrial morphological homeostasis. It has been shown that reduced expression of Mfn2 is associated with insulin resistance, but the mechanism is still unclear. We investigated whether Mfn2 deficiency leads to impaired insulin sensitivity via elevated oxidative stress. L6 skeletal muscle cells were treated with palmitate and Mfn2 expression was repressed by transfection with antisense Mfn2. Levels of antioxidant enzymes, reactive oxygen species (ROS), the phosphorylation of c-Jun N-terminal Kinase (JNK) and nuclear factor-κB (NF-κB) and the mitochondrial membrane potential (Δψ_m) were measured. The results showed palmitate-induced insulin resistance of skeletal muscle cells was accompanied by Mfn2 repression. Meanwhile, the cells had decreased Δψ_m and activity of antioxidant enzymes which could increase production of ROS, phosphorylation of JNK and NF-κB. When Mfn2 was up-regulated in palmitate-treated cells, oxidative stress and insulin resistance was alleviated. Furthermore, knock-down of Mfn2 in control cells enhanced oxidative stress. Mfn2 deficiency led to increased superoxide concentration and activation of JNK as well as NF-κB associated with insulin signaling. In conclusion, Mfn2 is a potent repressor for oxidative stress and regulation of Mfn2 expression may prove to be a potential method to circumvent insulin resistance.     

Differences in TLR9-dependent inhibitory effects of H2O2-induced IL-8 secretion and NF-kappa B/I kappa B-alpha system activation by genomic DNA from five Lactobacillus species

Lactic acid bacteria (LAB) show anti-inflammatory effects, and their genomic DNA was identified as one of the anti-inflammatory components. Despite the differences in anti-inflammatory effects between live LAB dependent not only on genus but also species, this effect has not been compared at the genomic DNA level. We compared the anti-inflammatory effects of the genomic DNA from five Lactobacillus species—Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus gasseri, Lactobacillus plantarum, and Lactobacillus reuteri—using Caco-2 cells. To evaluate anti-inflammatory effects, decreases in H₂O₂-induced IL-8 secretion and inhibition of H₂O₂-induced NF-κB/IκB-α system activation were examined. All LAB genomic DNAs dose-dependently decreased H₂O₂-induced IL-8 secretion and inhibited H₂O₂-induced NF-κB/IκB-α system activation. Comparison of these effects between Lactobacillus species showed that the anti-inflammatory effects of L. acidophilus genomic DNA are lower than those of the other species. Furthermore, suppression of Toll-like receptor 9 (TLR9), a specific receptor of bacterial DNA, expression by RNAi abolished the decrease of H₂O₂-induced IL-8 secretion and inhibition of H₂O₂-induced NF-κB/IκB-α system activation by LAB genomic DNA. Our results demonstrated that the anti-inflammatory effects of genomic DNA differ between Lactobacillus species and TLR9 is one of the major pathways responsible for the anti-inflammatory effect of LAB genomic DNA.     

Curcumin protects against acute liver damage in the rat by inhibiting NF-κB,proinflammatory cytokines production and oxidative stress

Curcumin, an anti-inflammatory and antioxidant compound, was evaluated for its ability to suppress acute carbon tetrachloride-induced liver damage. Acute hepatotoxicity was induced by oral administration of CCl₄ (4 g/kg, p.o.). Curcumin treatment (200 mg/kg, p.o.) was given before and 2 h after CCl₄ administration. Indicators of necrosis (alanine aminotransferase) and cholestasis (γ-glutamyl transpeptidase and bilirubins) resulted in significant increases after CCl₄ intoxication, but these effects were prevented by curcumin treatment. As an indicator of oxidative stress, GSH was oxidized and the GSH/GSSG ratio decreased significantly by CCl₄, but was preserved within normal values by curcumin. In addition to its antioxidants properties, curcumin is capable of preventing NF-κB activation and therefore to prevent the secretion of proinflammatory cytokines. Therefore, in this study we determined the concentrations of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) mRNA, and NF-κB activation. CCl₄-administered rats depicted significant increases in TNF-α, IL-1β, and IL-6 production, while curcumin remarkably suppressed these mediators of inflammation in liver damage. These results were confirmed by measuring TNF-α, and IL-1β protein production using Western Blot analysis. Accordingly, these proteins were increased by CCl₄ and this effect was abolished by curcumin. Administration of CCl₄ induced the translocation of NF-κB to the nucleus; CCl₄ induced NF-κB DNA binding activity was blocked by curcumin treatment. These findings suggest that curcumin prevents acute liver damage by at least two mechanisms: acting as an antioxidant and by inhibiting NF-κB activation and thus production of proinflammatory cytokines.     

Pellino-1 Protects Periodontal Ligament Stem Cells Against H<Subscript>2</Subscript>O<Subscript>2</Subscript>-Induced Apoptosis via Activation of NF-κB Signaling

To determine the protective effects of Pellino-1 against H₂O₂-induced apoptosis in periodontal ligament stem cells (PDLSC). We demonstrated that H₂O₂ decreases PDLSC viability by 40 and 50% with the concentrations of 400 and 500 μM, respectively, with an observed downregulation of Pellino-1 mRNA and protein; we further concluded that overexpression of Pellino-1 significantly lowers 8-hydroxy-2′-deoxyguanosine levels by 10% and upregulates superoxide dismutase 1, glutathione peroxidase levels, and catalase mRNA levels by 200, 40, and 250%, respectively. More importantly, we found that overexpression of Pellino-1 inhibited H₂O₂-induced cellular apoptosis through the activation of the NF-κB signaling pathway. Pellino-1 may be critically important for cell survival in the presence of oxidative elements; activation of the NF-κB signaling cascade was required for the overexpression of Pellino-1 to protect the cells from H₂O₂-induced apoptosis.     

Ataxia telangiectasia mutated inhibits oxidative stress-induced apoptosis by regulating heme oxygenase-1 expression

Ataxia telangiectasia (AT) is caused by mutational inactivation of the ataxia telangiectasia mutated (Atm) gene, which is involved in DNA repair. Increased oxidative stress has been shown in human AT cells and neuronal tissues of Atm-deficient mice. Heme oxygenase-1 (HO-1) is an inducible antioxidant enzyme and protects cells against oxidative stress. The purpose of this study is to determine whether ATM induces antioxidant enzyme HO-1 and protects cells from oxidative stress-mediated apoptosis by driving the activation of PKC-δ and NF-κB, by increasing cell viability, and by downregulating DNA fragmentation and apoptotic indicators (apoptosis-inducing factor and cleaved caspase-3). AT fibroblasts stably transfected with human full-length ATM cDNA (YZ5 cells) or the empty vector (MOCK cells) were treated with H₂O₂ as a source of reactive oxygen species (ROS). As a result, transfection with ATM inhibited ROS-induced cell death and DNA fragmentation in MOCK cells. Transfection with ATM induced expression of HO-1 which was mediated by PKC-δ and NF-κB in H₂O₂-treated MOCK cells. ZnPP, an HO-1 inhibitor, and transfection with HO-1 siRNA increased ROS levels and apoptosis, whereas hemin, an HO-1 activator, reduced ROS levels and apoptosis in H₂O₂-treated YZ5 cells. Rottlerin, a PKC-δ inhibitor, inhibited NF-κB activation and HO-1 expression in H₂O₂-treated YZ5 cells. MOCK cells showed increased cell death, DNA fragmentation, and apoptotic indicators compared to YZ5 cells exposed to H₂O₂. In addition, transfection with p65 siRNA increased ROS levels and DNA fragmentation, but decreased HO-1 protein levels in H₂O₂-treated YZ5 cells. In conclusion, ATM induces HO-1 expression via activation of PKC-δ and NF-κB and inhibits oxidative stress-induced apoptosis. A loss of HO-1 induction may explain why AT patients are vulnerable to oxidative stress.