TEMPOL protects human neuroblastoma SH-SY5Y cells against β-amyloid-induced cell toxicity

Amyloid-β peptide (Aβ) has been implicated in the pathogenesis of Alzheimer's disease (AD). It can cause cell death in Alzheimer's disease by evoking a cascade of oxidative damage to neurons. Antioxidant compounds may help to elucidate and develop a treatment for Alzheimer's disease. In the present study, we investigated the protective effect of TEMPOL (4-hydroxy-2,2,6,6-tetramethyl-1-piperidinyloxy), a cyclic nitroxide which is particularly effective at reducing oxidative injury, on Aβ(1-42)-induced SH-SY5Y cell toxicity. Exposure of cells to 20 μM Aβ(1-42) for 48 h caused viability loss and apoptotic increase, and pre-treatment with TEMPOL for 24 h significantly reduced the viability loss and apoptotic rate. In addition, TEMPOL inhibited Aβ(1-42)-induced superoxide anion generation and hydroxyl radical generation to a striking degree. Based on these results, it is concluded that TEMPOL effectively protects SH-SY5Y cells against β-amyloid-induced damage by suppressing the generation of reactive oxygen species especially, superoxide anion.     

Sirt1 overexpression protects murine osteoblasts against TNF-α-induced injury in vitro by suppressing the NF-κB signaling pathway

Aim:

Sirtuin 1 (Sirt1) is the class III histone/protein deacetylase that interferes with the NF-κB signaling pathway, thereby has anti-inflammatory function. This study was undertaken to investigate whether Sirt1 could protect osteoblasts against TNF-α-induced injury in vitro.

Methods:

Murine osteoblastic cell line, MC3T3-E1, was used. Overexpress of Sirt1 protein in MC3T3-E1 cells was made by transfection the cells with Sirt1-overexpressing adenovirus. The levels of mRNAs and proteins were determined with qRT-PCR and Western blotting, respectively. The activity of NF-κB was examined using NF-κB luciferase assay. The NO concentration was measured using the Griess method.

Results:

Treatment of MC3T3-E1 cells with TNF-α (2.5–10 ng/mL) suppressed Sirt1 protein expression in a concentration-dependent manner. TNF-α (5 ng/mL) resulted in an increase in apoptosis and a reduction in ALP activity in the cells. Overexpression of Sirt1 in the cells significantly attenuated TNF-α-induced injury through suppressing apoptosis, increasing ALP activity, and increasing the expression of Runx2 and osteocalcin mRNAs. Furthermore, overexpression of Sirt1 in the cells significantly suppressed TNF-α-induced NF-κB activation, followed by reducing the expression of iNOS and NO formation. Sirt1 activator resveratrol (10 μmol/L) mimicked the protection of the cells by Sirt1 overexpression against TNF-α-induced injury, which was reversed by the Sirt1 inhibitor EX-527 (5 μmol/L).

Conclusion:

Overexpression of Sirt1 protects MC3T3-E1 osteoblasts aganst TNF-α-induced cell injury in vitro, at least in part, via suppressing NF-κB signaling. Sirt1 may be a novel therapeutic target for treating rheumatoid arthritis-related bone loss.     

l-3-n-Butylphthalide attenuates β-amyloid-induced toxicity in neuroblastoma SH-SY5Y cells through regulating mitochondrion-mediated apoptosis and MAPK signaling

Alzheimer's disease (AD) is a progressive neurodegenerative disease. Amyloid-β protein (Aβ), the hallmark of AD, invokes a cascade of mitochondrial dysfunction and eventually leads to neuronal death. l-3-n-Butylphthalide (l-NBP) has shown the potent neuroprotective effects in stroke and AD animal models. The present study is to evaluate the neuroprotective effect of l-NBP on Aβ_25–35-induced neuronal injury and the possible mechanism in the human neuroblastoma SH-SY5Y cells. Our results showed that l-NBP significantly attenuated Aβ_25–35-induced cell death and reduced neuronal apoptosis. l-NBP significantly inhibited Aβ_25–35-induced mitochondrial dysfunction, including mitochondrial membrane potential reduction, and reactive oxygen species production. Furthermore, l-NBP could partially reverse the elevations of Aβ_25–35-induced active caspase-3, caspase-9, and cytochrome c expressions, and the downregulation of anti-apoptosis protein Bcl-2. Moreover, l-NBP markedly inhibited the activations of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase/stress-activated protein kinase signaling pathway. These results demonstrated that l-NBP was capable of protecting neuronal cells from Aβ_25–35-induced toxicity through a mitochondrial-dependent apoptotic pathway. Thus, l-NBP shows promising candidate of multi-target neuronal protective agent for the treatment of AD.     

Sulforaphane protects against cytokine- and streptozotocin-induced β-cell damage by suppressing the NF-κB pathway

Sulforaphane (SFN) is an indirect antioxidant that protects animal tissues from chemical or biological insults by stimulating the expression of several NF-E2-related factor-2 (Nrf2)-regulated phase 2 enzymes. Treatment of RINm5F insulinoma cells with SFN increases Nrf2 nuclear translocation and expression of phase 2 enzymes. In this study, we investigated whether the activation of Nrf2 by SFN treatment or ectopic overexpression of Nrf2 inhibited cytokine-induced β-cell damage. Treatment of RIN cells with IL-1β and IFN-γ induced β-cell damage through a NF-κB-dependent signaling pathway. Activation of Nrf2 by treatment with SFN and induction of Nrf2 overexpression by transfection with Nrf2 prevented cytokine toxicity. The mechanism by which Nrf2 activation inhibited NF-κB-dependent cell death signals appeared to involve the reduction of oxidative stress, as demonstrated by the inhibition of cytokine-induced H₂O₂ production. The protective effect of SFN was further demonstrated by the restoration of normal insulin secreting responses to glucose in cytokine-treated rat pancreatic islets. Furthermore, pretreatment with SFN blocked the development of type 1 diabetes in streptozotocin-treated mice.     

Sargachromenol protects against vascular inflammation by preventing TNF-α-induced monocyte adhesion to primary endothelial cells via inhibition of NF-κB activation

Vascular inflammation is a key factor in the pathogenesis of atherosclerosis. The purpose of this study was to investigate the protective effects of sargachromenol (SCM) against tumor necrosis factor (TNF)-α-induced vascular inflammation. SCM decreased the expression of cell adhesion molecules, including intracellular adhesion molecule-1 and vascular cell adhesion molecule-1, in TNF-α-stimulated human umbilical vein endothelial cells (HUVECs), resulted in reduced adhesion of monocytes to HUVECs. SCM also decreased the production of monocyte chemoattractant protein-1 and matrix metalloproteinase-9 in TNF-α-induced HUVECs. Additionally, SCM inhibited activation of nuclear factor kappa B (NF-κB) induced by TNF-α through preventing the degradation of inhibitor kappa B. Moreover, SCM reduced the production of reactive oxygen species in TNF-α-treated HUVECs. Overall, SCM alleviated vascular inflammation through the regulation of NF-κB activation and through its intrinsic antioxidant activity in TNF-α-induced HUVECs. These results indicate that SCM may have potential application as a therapeutic agent against vascular inflammation.     

Saikosaponin a and its epimer saikosaponin d exhibit anti-inflammatory activity by suppressing activation of NF-κB signaling pathway

Saikosaponin a (SSa) and its epimer saikosaponin d (SSd) are major triterpenoid saponin derivatives from Radix bupleuri (RB), which has been long used in Chinese traditional medicine for treatment of various inflammation-related diseases. In the present study, the anti-inflammatory activity, as well as the underlying mechanism, of SSa and SSd was investigated in lipopolysaccharide (LPS)-induced RAW264.7 cells. Our results demonstrated that both SSa and SSd significantly inhibited the expression of inducible nitric-oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-induced RAW264.7 cells, and finally resulted in the reduction of nitric oxide (NO) and prostaglandin E(2) (PGE(2)). In addition, LPS-induced production of major pro-inflammatory cytokines: the tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), was suppressed in a dose-dependent manner by the treatment of SSa or SSd in RAW264.7 cells. Further analysis revealed that both SSa and SSd could inhibit translocation of nuclear factor-κB (NF-κB) from the cytoplasm to the nucleus in the LPS-induced RAW264.7 cells. Furthermore, SSa and SSd exhibited significant anti-inflammatory activity in two different murine models of acute inflammation, carrageenan-induced paw edema in rats and acetic acid-induced vascular permeability in mice. In conclusion, SSa and SSd showed potent anti-inflammatory activity through inhibitory effects on NF-κB activation and thereby on iNOS, COX-2 and pro-inflammatory cytokines.     

Wogonin inhibits H2O2-induced angiogenesis via suppressing PI3K/Akt/NF-κB signaling pathway

Wogonin, a natural monoflavonoid extracted from Scutellariae radix, has been reported for its ability of inhibiting tumor angiogenesis. In this study, we assessed the effect of wogonin on angiogenesis induced by low level of H₂O₂ (10 μM) in human umbilical vein endothelial cells (HUVECs). Wogonin suppressed H₂O₂-induced migration and tube formation of HUVECs as well as microvessel sprouting from rat aortic rings in vitro. Meanwhile, wogonin suppressed vessel growth in chicken chorioallantoic membrane (CAM) model in vivo. Mechanistic studies showed that wogonin suppressed H₂O₂-activated PI3K/Akt pathway and reduced the expression of vascular endothelial growth factor (VEGF) up-regulated by H₂O₂ in both protein and mRNA levels. In addition, wogonin also inhibited nuclear translocation of NF-κB, and decreased the binding ability of NF-κB with exogenous consensus DNA oligonucleotide. Then we further investigated the effect of wogonin on over-activated PI3K/Akt pathway by insulin-like growth factor-1 (IGF-1) and H₂O₂. We found that wogonin suppressed phosphorylation of Akt, up-regulation of VEGF and angiogenesis in vitro which was further induced by IGF-1 and H₂O₂. Moreover, in NF-κB overexpressed HUVECs, wogonin could also reduce the expression of VEGF and inhibited the migration and tube formation. Taken together, these results suggested that wogonin was potential in inhibiting H₂O₂-induced angiogenesis in vitro and in vivo via suppressing PI3K/Akt pathway and NF-κB signaling.     

α-Mangostin protects lipopolysaccharide-stimulated nucleus pulposus cells against NLRP3 inflammasome-mediated apoptosis via the NF-κB pathway

Intervertebral disc degeneration (IDD) is a common and chronic inflammatory disorder. α-Mangostin exhibits a novel biological function against inflammation in various inflammatory diseases. Here, we aimed to explore the role of α-mangostin in IDD using an in vitro cell model. Human nucleus pulposus cells (NPCs) were exposed to lipopolysaccharide (LPS) to induce inflammatory injury. Cell viability of NPCs was determined by CCK-8 assay. ELISA was performed to examine the production of interleukin (IL)-1β and IL-18. Apoptotic cell death in NPCs was detected by TUNEL staining. The expression levels of apoptotic-associated proteins were detected by western blotting. Nuclear factor-kappa B (NF-κB) activation was examined by determining the expression levels of p-p65, p65, and nuclear p65. Results showed that treatment with α-mangostin improved the viability of LPS-treated NPCs. α-Mangostin treatment also inhibited the LPS-induced increase in expression levels of NLRP3, ASC and pro-caspase-1, as well as the production of IL-1β and IL-18 in NPCs. Moreover, treatment with α-mangostin or NLRP3 inhibitor (MCC950) significantly decreased apoptotic cell death in NPCs, as compared with treatment with LPS. In addition, the expression levels of cleaved caspase-3 and Bax were decreased, while Bcl-2 expression was increased in α-mangostin- or MCC950-treated NPCs. Treatment with α-mangostin also suppressed LPS-induced increase of p-p65/p65 and nuclear p65 levels. Moreover, inhibition of NF-κB by PDTC aggravated the inhibitory effects of α-mangostin on NLRP3 inflammasome activation and apoptosis in LPS-induced NPCs. These findings suggested that α-mangostin exerted a protective effect on NLRP3 inflammasome-mediated apoptosis in LPS-induced NPCs through regulating NF-κB signaling.     

Lonicera japonica THUNB. protects 6-hydroxydopamine-induced neurotoxicity by inhibiting activation of MAPKs, PI3K/Akt, and NF-κB in SH-SY5Y cells

In this study, we investigated the neuroprotective effects of Lonicera japonica THUNB. extract (LJ) on 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in SH-SY5Y cells. We found that LJ significantly increased cell viability decrease, lactate dehydrogenase release (LDH), morphological changes, nuclear condensation, fragmentation, and reactive oxygen species (ROS) production induced by 6-OHDA in SH-SY5Y cells. The cytoprotection afforded by pretreatment with LJ was associated with increases of the glutathione (GSH) level, superoxide dismutase (SOD) activity, and catalase (CAT) activity in 6-OHDA-induced SH-SY5Y cells. In addition, LJ strikingly inhibited 6-OHDA-induced mitochondrial dysfunctions including reduction of mitochondria membrane potential (MMP) and activation of cleaved poly-ADP-ribose polymerase (PARP), cleaved caspase-3, cleaved caspase-9, increased Bax, as well as decreased Bcl-2 and Bcl-xL. Additionally, LJ dramatically attenuated 6-OHDA-induced phosphorylation of c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase 1/2 (ERK 1/2), and phosphoinositide 3-kinase (PI3K)/Akt. Meanwhile, LJ counteracted nuclear factor-κB (NF-κB) activation by blocking its translocation to the nucleus. These findings suggest that LJ has a potent anti-parkinsonism; this effect was mediated, at least in part, by inhibition of neurotoxicity, apoptotic cascade events, and oxidative stress via activation of MAPKs, PI3K/Akt, and NF-κB.     

Akt/GSK3β signaling is involved in fipronil-induced apoptotic cell death of human neuroblastoma SH-SY5Y cells

Fipronil (FPN) is a phenylpyrazole insecticide acted on insect gamma-aminobutyric acid (GABA) receptors. Although action of FPN is restricted on insect neuronal or muscular transmitter system, a few studies have assessed the effects of this neurotoxicant on neuronal cell death. To determine the mechanisms underlying FPN-induced neuronal cell death, we investigated whether reactive oxygen species (ROS) plays a role in FPN-induced apoptosis, using an in vitro model of human dopaminergic SH-SY5Y cells. FPN was cytotoxic to these cells and its cytotoxicity showed a concentration-dependent manner. Additionally, FPN treatment significantly decreased the tyrosine hydroxylase (TH) expression without change of glutamic acid decarboxylase 65 (GAD65) expression. FPN-induced dopaminergic cell death involved in increase of ROS generation since pretreatment with N-acetyl cysteine (NAC), an anti-oxidant, reduced cell death. After FPN treatment, dopamine (DA) levels decreased significantly in both cell and culture media, and oxidative effects of DA were blocked by NAC pretreatment. We showed that cell death in response to FPN was due to apoptosis since FPN increased cytochrome c release into the cytosol and activated caspase-3. It also led to nuclear accumulation of p53 and reduced the level of Bcl-2 protein in a concentration-dependent manner. Additionally, FPN altered the level of Akt/glycogen synthase kinase-3 (GSK3β) phosphorylation. FPN reduced the Akt phosphorylation on Ser473, and in parallel with the inactivation of Akt, phosphorylation of GSK3β on Ser9 which inactivates GSK3β, decreased after treatment with FPN. Furthermore, inhibition of the GSK3β signal protected the cell against FPN-induced cell death. These results suggest that regulation of GSK3β activity may control the apoptosis induced by FPN-induced oxidative stress associated with neuronal cell death.     

Ghrelin protects H9c2 cells from hydrogen peroxide-induced apoptosis through NF-κB and mitochondria-mediated signaling

Oxidative stress is a major mechanism underlying the pathogenesis of cardiovascular disease. Herein we investigate the protective effects of ghrelin in H₂O₂-induced apoptosis of H9c2 cells, as well as the possible molecular mechanisms involved. To study apoptosis, the cells were assessed by morphologic examination, MTS assay, Annexin V–propidium iodide dual staining and TUNEL analysis. Intracellular reactive oxygen species (ROS) production and mitochondrial membrane potential were also measured. To investigate the underlying molecular mechanisms, the expression of Bcl-2, Bax, active caspase-9 and NF-κB were assessed by Western blotting, and caspase-3 activity was determined by a colorimetric activity assay kit. After stimulation with H₂O₂ for 18 h, H9c2 cells viability decreased significantly; a large fraction of cells underwent apoptosis. We observed a dose-dependent rescue of H9c2 cells from H₂O₂-induced apoptosis in the presence of different ghrelin concentrations. Preincubation with ghrelin also restored the ROS and mitochondrial membrane potential levels that had been altered by H₂O₂ treatment. Moreover, ghrelin decreased H₂O₂-induced Bax production and caspase-9 activation, and increased Bcl-2 levels. NF-κB phosphorylation was also significantly inhibited by ghrelin in H₂O₂-treated cells. Caspase-3 activation was suppressed by ghrelin in H₂O₂-treated H9c2 cells in a dose-dependent manner. In summary, ghrelin protects H9c2 cells from oxidative stress-induced apoptosis through downregulation of Bax expression, caspase-9 activation and NF-κB phosphorylation, and upregulation of Bcl-2 expression. Caspase-3 activation was also reduced in a dose-dependent manner. These data suggest that ghrelin might protect against cardiovascular disease by protecting the mitochondria.     

Tetrandrine down-regulates ERK/NF-κB signaling and inhibits activation of mesangial cells

Objectives

Tetrandrine (TET), a bisbenzylisoquinoline alkaloid isolated from Stephania tetrandra S. Moore of the Menispermaceae, possesses anti-inflammatory activity. We examined the effect of tetrandrine on interleukin-1β (IL-1β)-provoked inflammatory response in mesangial cells.

Materials and methods

Primary rat mesangial cells (PRMCs) were treated with IL-1β to induce inflammation to resemble glomerulonephritis. Cell viability, morphology and NO production were evaluated. Western blotting was applied for expression of matrix metalloproteinase-9 (MMP-9), inducible NO synthase (iNOS), extracellular signal-regulated kinase (ERK) and NF-κB-related molecules. Electrophoretic mobility shift assay was performed to examine the DNA-binding activity of NF-κB.

Results

TET, at concentrations up to 10 μg/ml, had no significant effect on viability of PRMCs. At non-toxic concentrations, TET inhibited expression of phosphorylated ERK as well as phosphorylated IKK, enhanced degradation of IκBα and reduced the DNA-binding activity of NF-κB in IL-1β-primed PRMCs, suggesting an inhibitory effect on ERK/NF-κB signaling. TET attenuated the IL-1β-provoked expression of iNOS and release of NO. Moreover, both the protein expression and gelatinase activity of MMP-9, but not MMP-2, were markedly suppressed by TET.

Significance

TET down-regulated ERK/NF-κB signaling and inhibited the expression of inflammatory mediators NO and MMP-9. Since these mediators appear to activate mesangial cells, TET may play an important role in prevention of glomerulonephritis.     

Caffeine suppresses lipopolysaccharide-stimulated BV2 microglial cells by suppressing Akt-mediated NF-κB activation and ERK phosphorylation

Since the anti-inflammatory effect of caffeine is unclear in microglial cells, we performed whether caffeine attenuates the expression of pro-inflammatory mediators in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. Caffeine substantially suppressed the LPS-induced pro-inflammatory mediators nitric oxide (NO), prostaglandin E₂ (PGE₂) and tumor necrosis factor-α (TNF-α) in BV2 microglial cells. These effects resulted from the inhibition of their regulatory genes inducible NO synthase (iNOS), cycloxygenase-2 (COX-2) and TNF-α. In addition, caffeine significantly decreased LPS-induced DNA-binding activity of nuclear factor-κB (NF-κB) by suppressing the nuclear translocation of p50 and p65 subunits. A specific NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), attenuated the LPS-induced expression of iNOS, COX-2 and TNF-α genes. In addition, we elucidated that inhibition of Akt phosphorylation plays a crucial role in caffeine-mediated NF-κB regulation in LPS-stimulated BV2 microglial cells. Caffeine also attenuated the LPS-induced phosphorylation of extracellular signal-regulated kinase (ERK) and a specific inhibitor of ERK, PD98059, subsequently downregulated the expression of the pro-inflammatory genes iNOS, COX-2 and TNF-α. Taken together, our data indicate that caffeine suppresses the generation of pro-inflammatory mediators, such as NO, PGE₂ and TNF-α as well as their regulatory genes in LPS-stimulated BV2 microglial cells by inhibiting Akt-dependent NF-κB activation and the ERK signaling pathway.     

Crocin protects against cardiotoxicity induced by doxorubicin through TLR-2/NF-κB signal pathway in vivo and vitro

Doxorubicin (DOX) is widely used to treat multiple of tumors, but its clinical trials are allied with some serious adverse events mainly cardiac functional abnormalities. So the objective of our investigation is to identify the cardioprotective action of crocin (CRO), a natural compound derived from saffron, against DOX-induced cardiotoxicity. CRO was injected intraperitoneally (i.p.) to rats for six consecutive days and DOX (i.p.) was administered on the fourth day. H9c2 cells were treated with DOX for 24 h after being pre-treated by CRO for 2 h. CRO reduced tachycardia and J-point elevation, decreased the levels of serum creatine kinase, lactate dehydrogenase, glutamic-oxalacetic transaminase and glutamic-pyruvic transaminase. CRO exerted positive effect on DOX-induced ROS production and changes of oxidative stress biomarkers. CRO significantly decreased intracellular Ca²⁺ concentration and increased mitochondria membrane potential in H9c2 cells. CRO also resisted the DOX-induced high expression of tumor necrosis factor-α and interleukin-6, inhibited apoptosis and improved the abnormal expression levels of Bcl-2, Bax and Caspase-3 proteins. CRO obviously restrained DOX-mediated high expression of toll-like receptor-2 (TLR-2) and nuclear factor kappa-B (NF-κB) in ventricular tissue. In brief, CRO distinctly restrained DOX-mediated cardiotoxicity by inhibiting oxidative stress, inflammation, apoptotic and redressing cardiomyocyte calcium dyshomeostasis and mitochondria damage. These cardioprotective effects may be related closely with the TLR2/NF-κB pathway.     

Oligo-peptide I-C-F-6 inhibits hepatic stellate cell activation and ameliorates CCl4-induced liver fibrosis by suppressing NF-κB signaling and Wnt/β-catenin signaling

Oligo-peptide I-C-F-6 is a Carapax trionycis extract component that has an effect on hepatic fibrosis, however, its mechanism of action is still unclear. This study investigated whether oligo-peptide I-C-F-6 could inhibit liver fibrosis by suppressing NF-κB and Wnt/β-catenin signaling, which are important in liver fibrosis. HSC-T6 cells were treated with oligo-peptide I-C-F-6, and rats were divided randomly into five groups: control (saline), CCl₄, CCl₄ plus oligo-peptide I-C-F-6 (0.12 and 0.24 mg/kg), and CCl₄ plus colchicine (0.11 mg/kg). Here, we demonstrated that oligo-peptide I-C-F-6 ameliorated liver injury, inflammation, and hepatic fibrogenesis induced by CCl₄. Oligo-peptide I-C-F-6 also inhibited the activation of hepatic stellate cells (HSCs) in vivo and in vitro, as evaluated by the expression of transforming growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA), which is a specific marker of HSC activation. Moreover, oligo-peptide I-C-F-6 significantly reduced the expression and distribution of β-catenin, P-AKT, phospho (P)-GSK-3β, nuclear factor κB (NF-κB) P65, phospho-P65, and IκB kinase α/β (IKK-α/β) levels; additionally, IκB-α level was elevated both in vivo and in vitro. Together, these results indicate that oligo-peptide I-C-F-6 has hepatoprotective and anti-fibrotic effects in animal models of liver fibrosis, the mechanism of which may be related to modulating NF-κB and Wnt/β-catenin signaling.     

Hydrogen sulfide inhibits cigarette smoke-induced inflammation and injury in alveolar epithelial cells by suppressing PHD2/HIF-1α/MAPK signaling pathway

Chronic obstructive pulmonary fibrosis (COPD) is a chronic and fatal lung disease with few treatment options. Sodium hydrosulfide (NaHS), a donor of hydrogen sulfide (H₂S), was found to alleviate cigarette smoke (CS)-induced emphysema in mice, however, the underlying mechanisms have not yet been clarified. In this study, we investigated its effects on COPD in a CS-induced mouse model in vivo and in cigarette smoke extract (CSE)-stimulated alveolar epithelial A549 cells in vitro. The results showed that NaHS not only relieved emphysema, but also improved pulmonary function in CS-exposed mice. NaHS significantly increased the expressions of tight junction proteins (i.e., ZO-1, Occludin and claudin-1), and reduced apoptosis and secretion of pro-inflammatory cytokines (i.e., TNF-α, IL-6 and IL-1β) in CS-exposed mouse lungs and CSE-incubated A549 cells, indicating H₂S inhibits CS-induced inflammation, injury and apoptosis in alveolar epithelial cells. NaHS also upregulated prolyl hydroxylase (PHD)2, and suppressed hypoxia-inducible factor (HIF)-1α expression in vivo and in vitro, suggesting H₂S inhibits CS-induced activation of PHD2/HIF-1α axis. Moreover, NaHS inhibited CS-induced phosphorylation of ERK, JNK and p38 MAPK in vivo and in vitro, and treatment with their inhibitors reversed CSE-induced ZO-1 expression and inflammation in A549 cells. These results suggest that NaHS may prevent emphysema via the suppression of PHD2/HIF-1α/MAPK signaling pathway, and subsequently inhibition of inflammation, epithelial cell injury and apoptosis, and may be a novel strategy for the treatment of COPD.     

Aflatoxin B1 induces microglia cells apoptosis mediated by oxidative stress through NF-κB signaling pathway in mice spinal cords

Many studies have shown that aflatoxin B1 (AFB1) can cause cytotoxicity in numerous cells and organs induced by oxidative stress. However, the toxic effects and related mechanism of AFB1 on the microglia cells in the spinal cords have not been studied yet. Our results showed that AFB1 significantly reduced the number of microglia cells, increased the oxidants (malonaldehyde and hydrogen peroxide) but decreased the anti-oxidants (superoxide dismutase and total antioxidant capacity) in a dose dependent manner in mice spinal cords. In addition, AFB1 significantly increased the oxidative stress, promoted apoptosis and cell cycle arrest in G2-M phase, and activated NF-κB phosphorylation in BV2 microglia cells. However, the addition of active oxygen scavenger N-acetylcysteine can significantly reduce the ROS production, improve cell cycle arrest, reduce apoptosis, and the expression of phosphorylated NF-κB in BV2 microglia cells. These results indicate that AFB1 induces microglia cells apoptosis through oxidative stress by activating NF-κB signaling pathway.     

The immunostimulating activity of quercetin 3-O-xyloside in murine macrophages via activation of the ASK1/MAPK/NF-κB signaling pathway

Quercetin is a natural plant flavonoid that has been reported to possess a wide range of beneficial health effects, including anti-cancer and anti-inflammatory activities. Glycosylation of natural flavonoids with various sugar moieties can affect their physical, chemical, and biological properties. In this study, quercetin 3-O-xyloside (Quer-xyl) was enzymatically synthesized, and the immunomodulatory activities of quercetin and Quer-xyl were evaluated and compared. The results showed that Quer-xyl more effectively induced the secretion of TNF-α and IL-6 than quercetin by 2.5 and 1.5-fold, respectively. Quer-xyl dose-dependently induced the inducible nitric oxide synthase (iNOS) expression and increased the production of nitric oxide (NO) 1.3-fold more than quercetin. Quer-xyl also increased the phosphorylation of ASK1 and MAPKs (JNK and p38). Treatment with NQDI-1 (an inhibitor of ASK1) significantly attenuated the Quer-xyl-induced up-regulation of TNF-α secretion. The activation and subsequent nuclear translocation of NF-κB were substantially enhanced upon treatment with Quer-xyl (2.5–20 μM), while NQDI-1 treatment blocked the nuclear translocation of NF-κB. These results demonstrated that Quer-xyl can enhance the early innate immunity more effectively than quercetin by activating macrophages to secrete TNF-α and IL-6 through up-regulation of the redox-dependent ASK1/MAPK/NF-κB signaling pathway, suggesting for the first time that Quer-xyl may represent a new immunostimulator.