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1.
Neurosci Lett ; 676: 66-70, 2018 05 29.
Article in English | MEDLINE | ID: mdl-29626650

ABSTRACT

New psychoactive substances (NPSs), i.e., newly designed substances with chemical residues that are slightly different from those of known psychoactive substances, have been emerging since the late 2000s, and social problems related to the use of these substances are increasing globally. Two such NPSs are 4-chloro-2,5-dimethoxyamphetamine (DOC), a psychedelic substance that is structurally related to amphetamine, and AH-7921, an opioid analgesic that is used for recreational purposes and has a potency similar to that of morphine. Currently, scientific evidence for the dependence liability or toxicity of NPSs is lacking. Therefore, in this study, we performed animal behavioral tests to evaluate the dependence liability of DOC and AH-7921. The rewarding and reinforcing effects of DOC and AH-7921 were evaluated using the conditioned place preference (CPP) paradigm in mice and the self-administration (SA) procedure in rats. Both DOC and AH-7921 increased the preference for the drug-paired compartment in the CPP test at a dose of 0.3 mg/kg and increased the number of responses to the active lever in the SA test at 0.01 mg/(kg·infusion). Collectively, the data suggest that DOC and AH-7921 may have both rewarding and reinforcing effects. Further studies are needed to confirm the reinforcing effects in broader dose ranges with various schedules.


Subject(s)
Benzamides/adverse effects , DOM 2,5-Dimethoxy-4-Methylamphetamine/analogs & derivatives , Psychotropic Drugs/adverse effects , Reward , DOM 2,5-Dimethoxy-4-Methylamphetamine/adverse effects , Animals , Conditioning, Classical , Conditioning, Operant , Dose-Response Relationship, Drug , Drug-Seeking Behavior , Illicit Drugs , Male , Rats, Sprague-Dawley
2.
Cytotherapy ; 17(11): 1617-26, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26432561

ABSTRACT

BACKGROUND AIMS: The age of both the donor and the recipient has a potential influence on the efficacy of various cell therapies, but the underlying mechanisms are still being charted. We studied the effect of donor and recipient age in the context of microglia migration. METHODS: Microglia were in vitro--differentiated from bone marrow of young (3 months) and aged (12 months) mice and transplanted into young (∼ 3 months) and aged (∼ 17 months) C57BL/6 mice (n = 25) through intravenous and intranasal application routes. Recipients were not immune-suppressed or irradiated. Transplanted microglia were tracked through the use of a sex-mismatched setup or histologically with the use of cells from enhanced green fluorescent protein enhanced green fluorescent protein transgenic mice. RESULTS: No acute rejections or transplant-associated toxicity was observed. After 10 days, both intravenously and intranasally transplanted cells were detected in the brain. Transplanted cells were also found in the blood and the lymph system. The applied cells were also tracked in lungs and kidney but only after intravenous injection subjected to a "pulmonary first-pass effect." After 28 days, intravenously delivered cells were also found in the bone marrow and other organs, especially in aged recipients. Whereas in young recipients the transplanted microglia did not appear to persist, in aged brains the transplanted cells could still be identified up to 28 days after transplantation. However, when cells from aged donors were used, no signals of transplanted cells could be detected in the recipients. CONCLUSIONS: This study establishes proof of principle that in vitro--derived microglia from young but not from aged donors, intravenously or intranasally transplanted, migrate to the brain in young and aged recipients.


Subject(s)
Administration, Intranasal , Microglia/transplantation , Age Factors , Animals , Brain/cytology , Brain/physiology , Cell Differentiation , Green Fluorescent Proteins/metabolism , Green Fluorescent Proteins/pharmacokinetics , Injections, Intravenous , Male , Mice, Inbred C57BL , Mice, Transgenic , Microglia/cytology , Tissue Distribution
3.
Life Sci ; 112(1-2): 49-58, 2014 Sep 01.
Article in English | MEDLINE | ID: mdl-25058922

ABSTRACT

AIMS: It has been reported that in human glioblastoma cells, phosphotase and tensin homolog (PTEN) positive cells are more prone to paclitaxel-induced apoptosis than PTEN-negative cells. We investigated whether (S)-1-(α-naphthylmethyl)-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (CKD712) enhances the therapeutic effects of paclitaxel (including effects on cellular proliferation, invasion and apoptosis) in MDA-MB-231 cells through PTEN and NF-κB activity. MAIN METHODS: Cellular proliferation, invasion and apoptosis were assessed by MTT, Western blot analysis, and TUNEL assay. KEY FINDINGS: The combination of paclitaxel and CKD712 significantly decreased cell growth, invasion and MMP-9 expression/activity compared with paclitaxel alone. CKD712 enhanced the inhibition of cell growth and invasion in response to paclitaxel in scramble siRNA-transfected, but not siPTEN-transfected cells. CKD712 significantly increased the levels of apoptosis induced by paclitaxel and this apoptosis was accompanied by reduced expression of Bcl-xL but increased activation of caspase-3. TUNEL assay further confirms that CKD712 enhanced the apoptotic effect of paclitaxel. Interestingly, over-expression of PTEN decreased phosphorylation of IκBα and NF-κB expression in the nucleus, indicating that PTEN modifies NF-κB activity in MDA-MB-231 cells. CKD712 treatment also significantly reduced expression of p-IκB and NF-κB activity in TNF-α activated cells. SIGNIFICANCE: CKD712 strongly enhances the anti-cancer effects (proliferation, invasion, and apoptosis) of paclitaxel on MDA-MB-231 cells by regulating PTEN and NF-κB activity.


Subject(s)
Antineoplastic Agents/pharmacology , Gene Expression Regulation, Neoplastic , Mammary Glands, Human/drug effects , PTEN Phosphohydrolase/genetics , Paclitaxel/pharmacology , Tetrahydroisoquinolines/pharmacology , Apoptosis/drug effects , Caspase 3/genetics , Caspase 3/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Dose-Response Relationship, Drug , Drug Synergism , Humans , I-kappa B Kinase , Mammary Glands, Human/metabolism , Mammary Glands, Human/pathology , NF-kappa B/genetics , NF-kappa B/metabolism , PTEN Phosphohydrolase/antagonists & inhibitors , PTEN Phosphohydrolase/metabolism , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Signal Transduction , Tumor Necrosis Factor-alpha/pharmacology , bcl-X Protein/genetics , bcl-X Protein/metabolism
4.
PLoS One ; 8(10): e76293, 2013.
Article in English | MEDLINE | ID: mdl-24098466

ABSTRACT

The nuclear DNA binding protein high mobility group box 1 (HMGB1) has recently been suggested to act as a late mediator of septic shock. The effect of ((S)-6,7-dihydroxy-1-(4-hydroxynaphthylmethyl)-1,2,3,4-tetrahydroisoquinoline alkaloid, also known as THI-56, in an experimental model of sepsis was investigated. THI-56 exhibited potent anti-inflammatory properties in response to LPS in RAW 264.7 cells. In particular, THI-56 significantly inhibited the expression of inducible nitric oxide synthase (iNOS) and the release of HMGB1 in activated macrophages. THI-56 activated NE-F2-regulated factor 2 (Nrf-2)/heme oxygenase 1 (HO-1). The specific knockdown of the HO-1 gene by HO-1 siRNA significantly reversed the inhibitory effects of THI-56 on iNOS expression and HMGB1 release in LPS-stimulated macrophages. Importantly, THI-56 administration protected animals from death induced by either a lethal dose of LPS or cecal ligation and puncture (CLP). Furthermore, the ALT, AST, BUN, creatinine, and HMGB1 levels in the blood were significantly increased in CLP-induced septic mice, and the administration of THI-56 reduced these levels in a concentration-dependent and zinc protoporphyrin IX (ZnPPIX)-sensitive manner. In addition, the administration of THI-56 significantly ameliorated not only lung damage but also macrophage infiltration in the livers of CLP-induced septic mice, and these effects were also abrogated in the presence of ZnPPIX. Thus, we conclude that THI-56 significantly attenuates the proinflammatory response induced by LPS and reduces organ damage in a CLP-induced sepsis model through the upregulation of Nrf-2/HO-1.


Subject(s)
HMGB1 Protein/metabolism , Heme Oxygenase-1/metabolism , Nitric Oxide Synthase Type II/metabolism , Sepsis/metabolism , Tetrahydroisoquinolines/pharmacology , Animals , Anti-Inflammatory Agents/administration & dosage , Anti-Inflammatory Agents/pharmacology , Cell Line , Disease Models, Animal , Dose-Response Relationship, Drug , Gene Expression Regulation/drug effects , Heme Oxygenase-1/genetics , Lipopolysaccharides/metabolism , Lipopolysaccharides/pharmacology , Macrophages/drug effects , Macrophages/metabolism , Male , Mice , NF-E2-Related Factor 2/metabolism , Nitric Oxide Synthase Type II/genetics , RNA Interference , Sepsis/drug therapy , Sepsis/etiology , Sepsis/genetics , Sepsis/mortality , Tetrahydroisoquinolines/administration & dosage
5.
Br J Pharmacol ; 168(6): 1485-96, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23088309

ABSTRACT

BACKGROUND AND PURPOSE: Given the importance of VEGF and haem oxygenase (HO)-1 in wound healing, the present study tested the hypothesis that CKD712, a synthetic tetrahydroisoquinoline alkaloid, activated VEGF production through the induction of HO-1 in human dermal fibroblasts (HDFs) and in mouse skin to stimulate wound healing. EXPERIMENTAL APPROACH: Using HDFs, the effects of CKD712 on the production of VEGF and migration were evaluated. The mechanisms responsible were investigated using various signal inhibitors and small interfering RNA techniques. The ability of CKD712 to promote wound healing was also investigated in full-thickness skin-wounded mice. KEY RESULTS: CKD712 treatment of HDFs increased VEGF production and accelerated migration, which was antagonized by anti-VEGF antibodies. Both an AMPK inhibitor (compound C) and a HO-1 activity inhibitor (SnPPIX) but not inhibitors of MAPKs, PI3K and PKC reduced the production of VEGF by CKD712. Interestingly, SnPPIX inhibited HO-1 expression but not p-AMPK, whereas compound C inhibited both p-AMPK and HO-1 induction by CKD712. Moreover, CKD712 decreased HO-1 expression without affecting the expression of p-AMPK by siHO-1 transfection, but it failed to induce HO-1 in siAMPKα1-transfected cells, suggesting that AMPK is involved in HO-1 induction by CKD712 in HDFs. Also, CKD712 shortened the time of wound closure in an SnPPIX-sensitive manner in a full-thickness skin-wounded mouse model. CONCLUSION AND IMPLICATIONS: CKD712 accelerated cutaneous wound healing, at least in part, by the production of VEGF through HO-1 induction in HDFs and mouse skin.


Subject(s)
Dermatologic Agents/therapeutic use , Heme Oxygenase-1/metabolism , Skin/drug effects , Tetrahydroisoquinolines/therapeutic use , Vascular Endothelial Growth Factor A/biosynthesis , Wound Healing/drug effects , Wounds, Penetrating/drug therapy , AMP-Activated Protein Kinases/antagonists & inhibitors , AMP-Activated Protein Kinases/genetics , AMP-Activated Protein Kinases/metabolism , Animals , Cell Movement/drug effects , Cells, Cultured , Dermatologic Agents/pharmacology , Enzyme Inhibitors/pharmacology , Heme Oxygenase-1/antagonists & inhibitors , Heme Oxygenase-1/genetics , Humans , Male , Mice , Mice, Inbred BALB C , RNA Interference , RNA, Small Interfering , Random Allocation , Signal Transduction/drug effects , Skin/cytology , Skin/injuries , Skin/metabolism , Tetrahydroisoquinolines/pharmacology , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor A/metabolism , Wounds, Penetrating/metabolism
6.
Antioxid Redox Signal ; 17(6): 878-89, 2012 Sep 15.
Article in English | MEDLINE | ID: mdl-22369644

ABSTRACT

AIMS: We investigated the molecular mechanism by which ethyl pyruvate (EP) induces heme oxygenase-1 (HO-1) in RAW 264.7 cells and its effect on survival rate in cecal ligation and puncture (CLP)-induced wild-type (WT) and HO-1 knockout (HO-1(-/-)) septic mice. RESULTS: EP induced HO-1 in a dose- and time-dependent manner, which was mediated through p38 mitogen-activated protein kinase (MAPK) and NF-E2-related factor 2 (Nrf2) signaling cascade in RAW 264.7 cells. EP significantly inhibited the lipopolysaccharide (LPS)-stimulated inducible nitric oxide synthase (iNOS) expression and high-mobility group box 1 (HMGB1) release in RAW 264.7 cells. The inhibitory effect of EP on LPS-stimulated iNOS expression and HMGB1 release was reversed by transfection with siHO-1RNA in RAW 264.7 cells, but EP failed to reduce them in HO-1(-/-) peritoneal macrophages treated with LPS. Moreover, treatment of cells with glutathione ethyl ester (GSH-Et), SB203580 (p38 MAPK inhibitor), siHO-1, or p38-siRNA transfection inhibited anti-inflammatory effect of EP. Interestingly, both HO-1 induction and phosphorylation of p38 by EP were reversed by GSH-Et, and antioxidant redox element-luciferase activity by EP was reversed by SB203580 in LPS-activated cells. EP increased survival and decreased serum HMGB1 in CLP-WT mice, whereas it did not increase survival or decrease circulating HMGB1 in HO-1(-/-) CLP-mice. INNOVATION AND CONCLUSION: Our work provides new insights into the understanding the molecular mechanism by showing that EP induces HO-1 through a p38 MAPK- and NRF2-dependent pathway by decreasing GSH cellular levels. We conclude that EP inhibits proinflammatory response to LPS in macrophages and increases survival in CLP-induced septic mice by upregulation of HO-1 level, in which p38 MAPK and Nrf2 play an important role.


Subject(s)
Glutathione/metabolism , Heme Oxygenase-1/metabolism , Sepsis/drug therapy , p38 Mitogen-Activated Protein Kinases/metabolism , Animals , Cell Line , HMGB1 Protein/genetics , HMGB1 Protein/metabolism , Heme Oxygenase-1/genetics , Mice , Mice, Knockout , NF-E2-Related Factor 2/genetics , NF-E2-Related Factor 2/metabolism , p38 Mitogen-Activated Protein Kinases/genetics
7.
J Ethnopharmacol ; 137(3): 1311-7, 2011 Oct 11.
Article in English | MEDLINE | ID: mdl-21843620

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: The rhizomes of Cyperus rotundus have been used as traditional folk medicine for the treatment of inflammatory diseases. However, the mechanism by which extract of rhizomes of Cyperus rotundus (ECR) elicits anti-inflammation has not been extensively investigated so far. The aim of the present study was to test whether heme oxygenase (HO)-1 induction is involved in the anti-inflammatory action of ECR. MATERIALS AND METHODS: Induction of HO-1 and inhibition of inducible nitric oxide synthase (iNOS)/NO production by ECR and its 12 constituents (3 monoterpenes, 5 sesquiterpenes, and 4 aromatic compounds) were investigated using RAW264.7 cells in vitro. In addition, anti-inflammatory action of ECR and its two active ingredients (nookkatone, valencene) were confirmed in sepsis animal model in vivo. RESULTS: ECR increased HO-1 expression in a concentration-dependent manner, which was correlated with significant inhibition of iNOS/NO production in LPS-activated RAW264.7 cells. Among 12 compounds isolated from ECR, mostly sesquiterpenes induced stronger HO-1 expression than monoterpenes in macrophage cells. Nootkatone and valencene (sesquiterpenes) significantly inhibited iNOS expression and NO production in LPS-simulated RAW264.7 cells. Inhibition of iNOS expression by nootkatone, valencene, and ECR were significantly reduced in siHO-1 RNA transfected cells. Furthermore, all three showed marked inhibition of high mobility group box-1 (HMGB1) in LPS-activated macrophages and increased survival rates in cecal ligation and puncture (CLP)-induced sepsis in mice. CONCLUSIONS: Taken together, we concluded that possible anti-inflammatory mechanism of ECR is, at least, due to HO-1 induction, in which sesquiterpenes such as nootkatone and valencene play a crucial role.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Cyperus , Heme Oxygenase-1/metabolism , Membrane Proteins/metabolism , Plant Extracts/pharmacology , Sepsis/drug therapy , Sesquiterpenes/pharmacology , Animals , Anti-Inflammatory Agents/isolation & purification , Cell Line , Cyperus/chemistry , Disease Models, Animal , Dose-Response Relationship, Drug , HMGB1 Protein/metabolism , Heme Oxygenase-1/genetics , Macrophages/drug effects , Macrophages/enzymology , Membrane Proteins/genetics , Mice , Mice, Inbred ICR , Nitric Oxide/metabolism , Nitric Oxide Synthase Type II/antagonists & inhibitors , Nitric Oxide Synthase Type II/metabolism , Plant Extracts/isolation & purification , Plants, Medicinal , Polycyclic Sesquiterpenes , RNA Interference , Rhizome , Sepsis/enzymology , Sesquiterpenes/isolation & purification , Time Factors , Transfection , Up-Regulation
8.
J Ethnopharmacol ; 133(2): 336-44, 2011 Jan 27.
Article in English | MEDLINE | ID: mdl-20937376

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: The methanol extract of Cimicifugae Rhizome has been traditionally used in various disorders including inflammation. AIM OF THE STUDY: The aim of the study is to explore whether anti-inflammatory action of 3 active compounds, two triterpenoid glycosides (cimiside E, 23-O-actylshengmanol-3-xyloside) and one furanocoumarin (isoimperatorin), isolated from Cimicifugae Rhizome is related with peroxisome proliferator-activated receptor-γ (PPAR-γ) expression in human umbilical endothelial cell line, EA.hy926 cells. MATERIALS AND METHODS: Cell viability and production of reactive oxygen species were performed. In addition, adhesion of monocyte into endothelial cells and western blot for expression of adhesion molecules and signal proteins were investigated in tumor necrosis factor-α (TNF-α)-activated cells. RESULTS: Pretreatment of test compounds significantly reduced reactive oxygen species (ROS) production and expression of vascular cell adhesion molecule-1 (VCAM-1), but not intercellular cell adhesion molecule-1 (ICAM-1). Three compounds all dose-dependently increased not only PPAR-γ expression in EA.hy926 cells but inhibited TNF-α-induced phosphorylation of Akt, extracellular-signal-regulated kinase (ERK) and protein kinase C (PKC) with different specificity. Finally, they prevented TNF-α-induced adhesion of U937 monocytic cells to EA.hy926 cells. CONCLUSIONS: The present results show that cimiside E, 23-O-actylshengmanol-3-xyloside, isoimperatorin isolated from Cimicifugae Rhizome selectively inhibits TNF-α-induced expression of VCAM-1 at least by upregulation of PPAR-γ, and signals for ERK1/2, PI3K, and PKC are involved in this effect.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Cimicifuga/chemistry , Furocoumarins/pharmacology , Saponins/pharmacology , Triterpenes/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/isolation & purification , Cell Adhesion/drug effects , Cell Line , Cell Survival/drug effects , Endothelial Cells/drug effects , Endothelial Cells/metabolism , Ethnopharmacology , Furocoumarins/isolation & purification , Humans , MAP Kinase Signaling System/drug effects , Molecular Structure , PPAR gamma/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Plants, Medicinal/chemistry , Protein Kinase C/metabolism , Reactive Oxygen Species/metabolism , Republic of Korea , Rhizome/chemistry , Saponins/isolation & purification , Signal Transduction/drug effects , Triterpenes/isolation & purification , Tumor Necrosis Factor-alpha/pharmacology , Vascular Cell Adhesion Molecule-1/metabolism
9.
J Ethnopharmacol ; 133(2): 524-30, 2011 Jan 27.
Article in English | MEDLINE | ID: mdl-20969944

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: The methanol extracts of Carthamus tinctorius (MEC) have long been used in traditional medicine as anti-inflammatory agent, however, the molecular mechanism by which MEC shows anti-inflammatory action is not investigated. AIM OF THE STUDY: Induction of heme oxygenase-1 (HO-1) by many medicinal herbs has been reported excellent anti-inflammatory action. Thus, the aim of the study is to explore whether anti-inflammatory action of MEC is related with HO-1 induction in RAW 264.7 cells. MATERIALS AND METHODS: The present study was designed to investigate as to MEC induces HO-1 expression so that it reduces inflammation by suppression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression in cells activated with lipopolysaccharide (LPS). RESULTS: Expression of HO-1 protein by MEC in macrophages was increased in a concentration- and time-dependent manner. Treatment with MEC significantly inhibited upregulation of both iNOS and COX-2 in LPS-activated macrophages and consequently reduced production of NO and PGE(2), respectively. The reduced expression of iNOS and COX-2 by MEC was reversed by siHO-1 RNA transfection. In addition, NF-E2-related factor (Nrf2) was translocated from cytosol to nucleus by MEC. The binding of NF-κB as well as NF-κB luciferase activity was also significantly diminished by MEC. Finally, tumor necrosis factor (TNF)-α-mediated VCAM-1 expression in endothelial cell was significantly inhibited by MEC. CONCLUSIONS: The present results show that MEC induces HO-1 expression via Nrf2 translocation and inhibits NF-κB activity, which may be responsible for anti-inflammatory action. Therefore, we propose that anti-inflammatory action of MEC involves at least HO-1 induction.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Carthamus tinctorius , Heme Oxygenase-1/biosynthesis , Membrane Proteins/biosynthesis , Animals , Anti-Inflammatory Agents/isolation & purification , Base Sequence , Carthamus tinctorius/chemistry , Cell Line , Cyclooxygenase 2/metabolism , Dinoprostone/antagonists & inhibitors , Enzyme Induction/drug effects , Ethnopharmacology , Heme Oxygenase-1/antagonists & inhibitors , Heme Oxygenase-1/genetics , Macrophages/drug effects , Macrophages/metabolism , Medicine, Korean Traditional , Membrane Proteins/antagonists & inhibitors , Membrane Proteins/genetics , Methanol , Mice , NF-E2-Related Factor 2/metabolism , NF-kappa B/antagonists & inhibitors , Nitric Oxide/antagonists & inhibitors , Nitric Oxide Synthase Type II/antagonists & inhibitors , Plant Extracts/isolation & purification , Plant Extracts/pharmacology , Plants, Medicinal/chemistry , RNA, Small Interfering/genetics , Republic of Korea , Vascular Cell Adhesion Molecule-1/metabolism
10.
Antioxid Redox Signal ; 14(11): 2057-70, 2011 Jun.
Article in English | MEDLINE | ID: mdl-21083424

ABSTRACT

Activation of nicotinic acetylcholine receptor alpha7 subunit (α7nAChR) by nicotine leads to the improved survival rate in experimental model of sepsis. Previously, we demonstrated that heme oxygenase (HO)-1 inducers or carbon monoxide significantly increased survival of lipopolysaccharide (LPS)-induced and cecal ligation and puncture-induced septic mice by reduction of high mobility group box 1 release, a late mediator of sepsis. However, that activation of α7nAChR by nicotine provides anti-inflammatory action through HO-1 upregulation has not been elucidated. Here we show that HO-1-inducible effect by nicotine was mediated through sequential event-Ca(2+) influx, classical protein kinase C activation, and reactive oxygen species production-which activates phosphoinositol-3-kinase/Akt/Nrf-2 pathway. In addition, HO-1 is required for nicotine-mediated suppression of tumor necrosis factor-α, inducible nitric oxide synthase, and high mobility group box 1 expression induced by LPS in macrophages, as evidenced by the fact that nicotine failed to inhibit production of these mediators when HO-1 was suppressed. Importantly, nicotine-induced survival rate was reduced by inhibition of HO-1 in LPS- and cecal ligation and puncture-treated septic mice. Collectively, these data suggest that activation of α7nAChR by nicotine is critical in the regulation of anti-inflammatory process, which could be mediated through HO-1 expression. Thus, we conclude that activation of α7nAChR by nicotine provides anti-inflammatory action through HO-1 upregulation.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Heme Oxygenase-1/metabolism , Inflammation/prevention & control , Macrophages/drug effects , Nicotine/pharmacology , Nicotinic Agonists/pharmacology , Receptors, Nicotinic/metabolism , Sepsis/metabolism , Animals , Calcium Signaling/drug effects , Cell Line , Endotoxemia/genetics , Endotoxemia/metabolism , Gene Expression Regulation/drug effects , HMGB1 Protein/metabolism , Heme Oxygenase-1/antagonists & inhibitors , Heme Oxygenase-1/genetics , Humans , Inflammation/chemically induced , Kaplan-Meier Estimate , Lipopolysaccharides/toxicity , Macrophages/metabolism , Male , Mice , Mice, Inbred BALB C , NADPH Oxidases/metabolism , NF-E2-Related Factor 2/metabolism , Nitric Oxide Synthase Type II/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Protein Kinase C/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Reactive Oxygen Species/metabolism , Sepsis/genetics , Signal Transduction/drug effects , Tumor Necrosis Factor-alpha/metabolism , alpha7 Nicotinic Acetylcholine Receptor
11.
Br J Pharmacol ; 162(7): 1498-508, 2011 Apr.
Article in English | MEDLINE | ID: mdl-21091653

ABSTRACT

BACKGROUND AND PURPOSE: Recently, metformin, a well-known anti-diabetic drug, has been shown to possess anti-inflammatory activities. This study investigated the effect of metformin on the expression of pro-inflammatory cytokines including high mobility group box 1 (HMGB1) in lipopolysaccharide (LPS)-treated animals and cells. EXPERIMENTAL APPROACH: We investigated whether metformin inhibits the release of HMGB1 in LPS-treated RAW 264.7 cells and increases survival rate in endotoxaemic mice (lethal endotoxaemia was induced by an i.p. injection of LPS). This was achieved by a range of techniques including Western blotting, enzyme-linked immunosorbent assay, specific pharmacological inhibitors, knock out of α(1) -subunit of AMP-activated protein kinase (AMPK) and recombinant HMGB1. KEY RESULTS: Both pre- and post-treatment with metformin significantly improved survival of animals during lethal endotoxaemia (survival rate was monitored up to 2 weeks), decreased serum levels of tumour necrosis factor-alpha (TNF-α), interleukin-1ß, HMGB1 expression and myeloperoxidase activity in lungs. However, metformin failed to improve survival in endotoxaemic animals that had additionally been treated with recombinant HMGB1. In an in vitro study, metformin dose-dependently inhibited production of pro-inflammatory cytokines and HMGB1 release. Metformin activated AMPK by its phosphorylation. Compound C (pharmacological inhibitor of AMPK) and siAMPKα1 reversed the anti-inflammatory effect of metformin in LPS-treated cells. CONCLUSIONS AND IMPLICATIONS: Our data indicate that metformin significantly attenuates the pro-inflammatory response induced by LPS both in vivo and in vitro. Metformin improved survival in a mouse model of lethal endotoxaemia by inhibiting HMGB1 release. AMPK activation was implicated as one of the mechanisms contributing to this inhibition of HMGB1 secretion.


Subject(s)
Endotoxemia/drug therapy , HMGB1 Protein/antagonists & inhibitors , HMGB1 Protein/metabolism , Lipopolysaccharides/pharmacology , Macrophages/drug effects , Metformin/pharmacology , AMP-Activated Protein Kinases/antagonists & inhibitors , AMP-Activated Protein Kinases/metabolism , Animals , Cells, Cultured , Cytokines/antagonists & inhibitors , Cytokines/metabolism , Endotoxemia/chemically induced , Endotoxemia/immunology , Endotoxemia/metabolism , HMGB1 Protein/genetics , Interleukin-1beta/antagonists & inhibitors , Interleukin-1beta/metabolism , Macrophages/immunology , Macrophages/metabolism , Male , Mice , Mice, Inbred BALB C , Peroxidase/antagonists & inhibitors , Peroxidase/metabolism , Pyrazoles/pharmacology , Pyrimidines/pharmacology , Survival Rate , Tumor Necrosis Factor-alpha/blood , Tumor Necrosis Factor-alpha/metabolism
12.
Atherosclerosis ; 213(1): 115-21, 2010 Nov.
Article in English | MEDLINE | ID: mdl-20864106

ABSTRACT

Phosphotase and tensin homolog deleted on chromosome 10 (PTEN) is a potent negative regulator of PI3K/Akt pathway. Here, we tried to elucidate the role of PTEN in the regulation of endothelial adhesion molecules, vascular cell adhesion molecule (VCAM)-1 and intracellular adhesion molecule (ICAM)-1, induced by TNF-α in human endothelial cells (ECs). Transfection with PTEN overexpressing vector resulted in the significant decrease in phosphorylation of Akt in TNF-α-treated ECs. PTEN strongly inhibited VCAM-1 but not ICAM-1, however this inhibitory effect was reversed by co-transfection with constitutively active-Akt (CA-Akt-HA) in TNF-α-stimulated ECs. Additionally, silencing of PTEN with specific siRNA showed significant increase of phosphor-Akt compared with TNF-α alone treated ECs. siPTEN significantly upregulated VCAM-1 but was indifferent to ICAM-1 in TNF-α-treated cells. Further, chromatin immunoprecipitation (ChIP) assay showed that PTEN targets GATA-6 but not IRF-1 binding to VCAM-1 promoter. In addition, GATA-6 is associated with glycogen synthesis kinase-3beta (GSK-3ß) which is in turn regulated by PTEN-dependent Akt activity. Finally, PTEN significantly prevented monocyte adhesion to TNF-α-induced ECs probably through VCAM-1 regulation. It is concluded that PTEN selectively inhibits expression of VCAM-1 but not ICAM-1 through modulation of PI3K/Akt/GSK-3ß/GATA-6 signaling cascade in TNF-α-treated ECs.


Subject(s)
GATA6 Transcription Factor/metabolism , Gene Expression Regulation , Glycogen Synthase Kinase 3/metabolism , Intercellular Adhesion Molecule-1/metabolism , PTEN Phosphohydrolase/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Tumor Necrosis Factor-alpha/metabolism , Vascular Cell Adhesion Molecule-1/metabolism , Cells, Cultured , Glycogen Synthase Kinase 3 beta , Humans , Phosphorylation , RNA, Small Interfering/metabolism , Signal Transduction
13.
Shock ; 34(6): 608-14, 2010 Dec.
Article in English | MEDLINE | ID: mdl-20442692

ABSTRACT

Reduction of high-mobility group box 1 (HMGB1) and NO levels may be important therapeutic strategy for treatment of sepsis. Recently, we found that carbon monoxide (CO) can reduce HMGB1 levels in septic animal models. Here, we tried to elucidate the molecular machinery of how CO inhibits HMGB1 release in toll-like receptor (TLR)-activated macrophages. Carbon monoxide-releasing molecule 2 (CORM-2) specifically inhibited the expression of iNOS (NO), but not of cyclooxygenase 2 (COX-2) (PGE2) in RAW 264.7 cells activated either by peptidoglycan (TLR-2 agonist), polyinosinic-polycytidylic acid (TLR-3 agonist), or LPS (TLR-4 agonist); this inhibition seemed to be mediated via the JAK2/STAT1 pathway. Treatment with neutralizing antibody to IFN-ß, a JAK2 inhibitor (AG490), or a STAT1 inhibitor (fludarabine) selectively inhibited iNOS, but not COX-2 in this system. Moreover, deletion of STAT1 by siRNA also showed preferential inhibition of iNOS but not COX-2 in LPS-treated cells. Carbon monoxide-releasing molecule 2 reduced IFN-ß production and phosphorylation of JAK2 and STAT1 in LPS-activated RAW264.7 cells. Carbon monoxide-releasing molecule 2 failed to inhibit iNOS and HMGB1 levels in the presence of recombinant IFN-ß and NO donor (NOC-18), respectively. Finally, plasma levels of HMGB1 and iNOS protein expression in lung tissues of cecal ligation and puncture-induced septic mice were decreased in the presence of CORM-2. Taken together, it is concluded that CO selectively inhibits iNOS over COX-2, at least through IFNß/JAK2/STAT1 signals, and this regulation plays an important role in the CORM-2-mediated inhibitory effect on HMGB1 release in macrophages.


Subject(s)
Carbon Monoxide/pharmacology , Cyclooxygenase 2/metabolism , HMGB1 Protein/metabolism , Interferon-beta/metabolism , Janus Kinase 2/metabolism , Nitric Oxide Synthase Type II/metabolism , Nitric Oxide/metabolism , Organometallic Compounds/pharmacology , Animals , Blotting, Western , Carbon Monoxide/chemistry , Cell Line , Cell Survival/drug effects , Macrophages/drug effects , Macrophages/metabolism , Male , Mice , Mice, Inbred BALB C , Organometallic Compounds/chemistry , Peptidoglycan/pharmacology , Poly I-C/pharmacology , Signal Transduction/drug effects , Toll-Like Receptors/agonists , Toll-Like Receptors/metabolism
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