The host takes use of pattern recognition receptors (PRRs) to defend against pathogen invasion or cellular damage. Among microorganism-associated molecular patterns detected by host PRRs, nucleic acids derived from bacteria or viruses are tightly supervised, providing a fundamental mechanism of host defense. Pathogenic DNAs are supposed to be detected by DNA sensors that induce the activation of NFκB or TBK1-IRF3 pathway. DNA sensor cGAS is widely expressed in innate immune cells and is a key sensor of invading DNAs in several cell types. cGAS binds to DNA, followed by a conformational change that allows the synthesis of cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) from adenosine triphosphate and guanosine triphosphate. cGAMP is a strong activator of STING that can activate IRF3 and subsequent type I interferon production. Here we describe recent progresses in DNA sensors especially cGAS in the innate immune responses against pathogenic DNAs.
DNA, Bacterial ; immunology ; metabolism ; DNA, Viral ; immunology ; metabolism ; Gene Expression Regulation ; Host-Pathogen Interactions ; Humans ; Immunity, Innate ; Interferon Regulatory Factor-3 ; genetics ; immunology ; Interferon Type I ; biosynthesis ; immunology ; Membrane Proteins ; genetics ; immunology ; Models, Molecular ; NF-kappa B ; genetics ; immunology ; Nucleotides, Cyclic ; biosynthesis ; immunology ; Nucleotidyltransferases ; genetics ; immunology ; Protein Binding ; Protein-Serine-Threonine Kinases ; genetics ; immunology ; Signal Transduction

OBJECTIVE: To investigate the effect of glycyrrhizin (Gly) on human neutrophil elastase (HNE)- induced mucin (MUC) 5AC overproduction in human bronchial epithelial cells (16HBE), and the potential signaling pathway involved in this process. METHODS: The cultured cells were divided into 3 groups: a control group, cultured in serum-free DMEM medium; an HNE group, pretreated with HNE alone; and a Gly group, incubated with HNE and Gly. After stimulation with a variety of Gly concentrations, the cytotoxicity was assessed by methyl thiazolyl tetrazolium method. The mRNA expressions of p38, nuclear factor κB (NF-κB) p65, inhibitory κBα (IκBα) and MUC5AC were detected by real-time PCR. The phosphorylation levels of p38 (p-p38), NF-κB p65 (p-NF-κB p65) and IκBα (p-IκBα) were measured by Western blot while the levels of MUC5AC protein were analyzed by emzyme-linked immunosorbent assay and immunofluorescence. RESULTS: Compared with the control group, the expression levels of MUC5AC mRNA and protein in the HNE group were both significantly increased. There was a significant increase in p-p38 and p-NF-κB p65, while the production of IκBα was much lower than that in the control group. Gly significantly inhibited the increase of MUC5AC, p38 and NF-κB p65, but increased the activity of IκBα. CONCLUSION Glycyrrhizin can inhibit MUC5AC overproduction via p38-NF-κB p65/IκBα signaling pathway.
Bronchi ; cytology ; Cell Line ; Epithelial Cells ; metabolism ; Glycyrrhizic Acid ; pharmacology ; Humans ; I-kappa B Proteins ; metabolism ; Leukocyte Elastase ; metabolism ; Mucin 5AC ; biosynthesis ; NF-KappaB Inhibitor alpha ; Phosphorylation ; Real-Time Polymerase Chain Reaction ; Signal Transduction ; Transcription Factor RelA ; metabolism ; p38 Mitogen-Activated Protein Kinases ; metabolism

Hydroxysafflor yellow A (HSYA) is a main active monomer purified from Carthamus tinctorius L. The research is to study the inhibitory effect of HSYA on the inflammatory signal transduction pathway related factors which were induced by permanent cerebral ischemia in rats. By using the successive administration at a 30 min interval of HSYA and the rats permanent focal cerebral ischemia model established by a intraluminal suture occlusion method. After cerebral artery occlusion 3, 6, 12 and 24 h, cortex was removed for the next experiments. Western blotting was used to detect the expression of p65 protein and the phospho-IkappaB-alpha (pIkappaB-alpha) in the cytoplasm and nucleus. Nuclear factor-kappaB (NF-kappaB) DNA binding activity was measured by Trans-AM transcription factor assay kits. mRNA expression of cytokines TNF-alpha, IL-1beta, IL-6 and IL-10 was measured by the RT-PCR method. The result showed that intravenous injection of HSYA (10 mg x kg(-1)) to rats after cerebral occlusion, the p65 translocation activity and the phosphorylation of IkappaB-alpha were significantly inhibited. At the same time, HSYA suppressed p65 binding activity and the transcriptional level of pro-inflammatory cytokines including TNF-alpha, IL-1beta and IL-6, and promoted the mRNA expression of anti-inflammatory cytokine IL-10. In conclusion, the anti-cerebral ischemic mechanism of HSYA may be due to its inhibition of NF-kappaB activity and the mRNA expression of cytokines in the inflammatory transduction pathway.
Animals ; Brain Ischemia ; metabolism ; Carthamus ; chemistry ; Chalcone ; analogs & derivatives ; isolation & purification ; pharmacology ; Cytokines ; biosynthesis ; genetics ; Flowers ; chemistry ; I-kappa B Proteins ; metabolism ; Interleukin-10 ; biosynthesis ; genetics ; Interleukin-1beta ; biosynthesis ; genetics ; metabolism ; Interleukin-6 ; biosynthesis ; genetics ; Male ; NF-KappaB Inhibitor alpha ; Neuroprotective Agents ; isolation & purification ; pharmacology ; Phosphorylation ; drug effects ; Plants, Medicinal ; chemistry ; Protein Transport ; Quinones ; isolation & purification ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Transcription Factor RelA ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

We demonstrated previously that Coptidis rhizoma extract (CRE) prevented S-nitroso-N-acetylpenicillamine-induced apoptotic cell death via the inhibition of mitochondrial membrane potential disruption and cytochrome c release in RINm5F (RIN) rat insulinoma cells. In this study, the preventive effects of CRE against cytokine-induced beta-cell death was assessed. Cytokines generated by immune cells infiltrating pancreatic islets are crucial mediators of beta-cell destruction in insulin-dependent diabetes mellitus. The treatment of RIN cells with IL-1beta and IFN-gamma resulted in a reduction of cell viability. CRE completely protected IL-1beta and IFN-gamma-mediated cell death in a concentration-dependent manner. Incubation with CRE induced a significant suppression of IL-1beta and IFN-gamma-induced nitric oxide (NO) production, a finding which correlated well with reduced levels of the iNOS mRNA and protein. The molecular mechanism by which CRE inhibited iNOS gene expression appeared to involve the inhibition of NF-kappa B activation. The IL-1beta and IFN-gamma-stimulated RIN cells showed increases in NF-kappa B binding activity and p65 subunit levels in nucleus, and IkappaBalpha degradation in cytosol compared to unstimulated cells. Furthermore, the protective effects of CRE were verified via the observation of reduced NO generation and iNOS expression, and normal insulin-secretion responses to glucose in IL-1beta and IFN-gamma-treated islets.
Animals ; Cell Death/drug effects ; Cell Line ; Cell Nucleus/metabolism ; Cell Survival/drug effects ; Drugs, Chinese Herbal/*pharmacology ; Gene Expression Regulation, Enzymologic/drug effects ; Glucose/pharmacology ; I-kappa B Proteins/metabolism ; Insulin/secretion ; Insulin-Secreting Cells/*cytology/*drug effects/enzymology ; Interferon-gamma/*pharmacology ; Interleukin-1beta/*pharmacology ; Male ; NF-kappa B/*metabolism ; Nitric Oxide/biosynthesis ; Nitric Oxide Synthase Type II/genetics/metabolism ; Protein Transport/drug effects ; RNA, Messenger/genetics/metabolism ; Rats ; Rats, Sprague-Dawley

This study is to investigate the effect of hydroxyethylpuerarin on the expression of tumor necrosis factor-alpha (TNF-alpha) and activity of nuclear factor kappa B (NF-kappaB) after middle cerebral artery occlusion (MCAO) in rats. Rats were subjected to cerebral ischemia-reperfusion injury induced by MCAO. Hydroxyethylpuerarin (10, 20, 40 mg x kg(-1), iv) was administered just 30 min before occlusion and immediately after reperfusion. After a 24 h reperfusion following 2 h of MCAO, the number of viable neurons in hippocampal CA1 region was counted by hematoxylin and eosin (HE) staining. TNF-alpha protein and its mRNA expression were examined with radioimmunoassay (RIA) and reverse transcriptasepolymerase chain reaction (RT-PCR) respectively. NF-KB activity was observed by electrophoretic mobility shift assay (EMSA), and inhibition of NF-kappaB alpha (IkappaBalpha) protein expression was evaluated by Western blotting analysis. Animals treated with hydroxyethylpuerarin had a significant increase in neuronal survival in comparison with vehicle-treated group. Hydroxyethylpuerarin significantly reduced the protein and mRNA expression of TNF-alpha following 2 h of ischemia with 24 h of reperfusion. NF-kappaB DNA binding activity and the degradation of IkappaBalpha in the cytoplasm also decreased by hydroxyethylpuerarin treatment. The protective effects of hydroxyethylpuerarin against ischemia-reperfusion injury may be mediated by decreasing the expression of TNF-alpha and the activity of NF-kappaB in rats.
Animals ; Brain ; metabolism ; pathology ; Cytoplasm ; metabolism ; DNA ; metabolism ; I-kappa B Proteins ; metabolism ; Infarction, Middle Cerebral Artery ; complications ; Isoflavones ; pharmacology ; Male ; NF-KappaB Inhibitor alpha ; NF-kappa B ; metabolism ; Neuroprotective Agents ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Wistar ; Reperfusion Injury ; etiology ; metabolism ; pathology ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics

<b>OBJECTIVEb>To investigate the effects of oxymatrine on the expression of nuclear factor kappa B (NF-kappaB) in the kidneys of rats with adriamycin-induced chronic renal fibrosis.

<b>METHODSb>Totally 120 Wistar rats were randomly assigned to normal control group, renal fibrosis model group, benazepril treatment group and oxymatrine treatment group (n=30). The rats in the normal control were injected with normal saline via the tail vein, and those in the other 3 groups with adriamycin (2 mg/kg) on the 7th day and 21st day of the experiment, respectively. Oxymatrine (100 mg/kg) or benazepril (6 mg/kg) was given by gastric perfusion after the second injection. Every 4 weeks after the second injection, 5 rats in each group were killed to evaluate the pathological changes and functional impairment of the kidney. Immunohistochemistry was used to detect the expression of NF-kappaB and inhibitory kappa B (IkappaB) in the kidney. The association of NF-kappaB expression with IkappaB expression, renal pathological changes and functional impairment were studied.

<b>RESULTSb>Oxymatrine and benazepril ameliorated renal fibrosis and functional impairment. Immunohistochemical staining revealed increased NF-kappaB expression and decreased IkappaB expression in the model group in comparison with oxymatrine and benazepril treatment groups 8 weeks after the second injection, but no significant difference was noted between the latter two groups. NF-kappaB expression in the kidneys of rats with adriamycin-induced chronic renal fibrosis showed an inverse correlation with IkappaB expression and positive correlation with pathological changes and functional impairment.

<b>CONCLUSIONb>Oxymatrine may inhibit renal fibrosis by down-regulating NF-kappaB expression, which may play a key role in protection against renal fibrosis.

Alkaloids ; pharmacology ; Animals ; Chronic Disease ; Doxorubicin ; Fibrosis ; chemically induced ; I-kappa B Proteins ; biosynthesis ; Immunohistochemistry ; Kidney ; drug effects ; metabolism ; pathology ; Male ; NF-kappa B ; biosynthesis ; Quinolizines ; pharmacology ; Random Allocation ; Rats ; Rats, Wistar

<b>OBJECTIVEb>To explore the apoptosis induction by curcumin in androgen-dependent prostate cancer cell line LNCaP).

<b>METHODSb>After LNCaP cells were induced by 10, 25, 50, 75, 100 micromol/L curcumin respectively, the cell activity was assayed by MTT at 5, 12 and 24 hours. Flow cytometry and electronic microscopy were adopted to observe cell cycle and morphological changes of LNCaP cells at 24 hours. After 5 hours, the expression of IkappaBalpha in LNCaP cells was detected by Western blotting.

<b>RESULTSb>The growth of LNCaP cells was suppressed obviously by curcumin in dose-dependent and time-dependent manners in vitro. There were significant differences in inhibition rate among different concentrations and time groups (P < 0.05). Furthermore, curcumin could arrest the cell cycle of LNCaP cells at G2/M phase in a dose-dependent manner (P <0.01). The ratios of apoptosis were significantly higher than those of controls (P < 0. 5). Curcumin could lead to characteristic morphological changes of apoptosis in LNCaP cells after 24 hours. The expression of IkappaBalpha in LNCaP cell did not show marked changes after the exposure to different concentrations of curcumin within 5 hours.

<b>CONCLUSIONb>Curcumin can suppress the growth of LNCaP, and promotes their apoptosis.

Apoptosis ; drug effects ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Curcumin ; pharmacology ; Dose-Response Relationship, Drug ; Humans ; I-kappa B Proteins ; biosynthesis ; Male ; NF-KappaB Inhibitor alpha ; Neoplasms, Hormone-Dependent ; Prostatic Neoplasms ; metabolism ; pathology

<b>OBJECTIVEb>To investigate the curcumin-induced the expression of IkappaBalpha in androgen-dependent (LNCaP) and androgen-independent (PC3) prostate cancer cells, and to study the mechanisms of curcumin on the proliferative inhibition of prostate cancer cells.

<b>METHODSb>After LNCaP and PC3 cells were affected by 10, 25, 50, 75, 100 micromol/L curcumin respectively, the cell activity was assayed with methyl thiazolyl tetrazolium (MTT) method at 5, 12 and 24 hours; Flow cytometry was adopted to observe the cell cycle of LNCaP and PC3 cells at 24 hours. After 5 hours, the expression of IkappaBalpha in LNCaP and PC3 cells was observed with Western blotting.

<b>RESULTSb>Curcumin obviously suppressed the proliferation of LNCaP and PC3 cells in does-dependent and time-dependent manners. Curcumin could arrest the cell cycle of LNCaP and PC3 cells at G(2), M phase and then induce cell apoptosis. The expression of IkappaBalpha in LNCaP cells had no significant difference after using curcumin (F = 0.129, P > 0.05). However, the expression of IkappaBalpha in PC3 cells increased gradually with the inducement of concentration-increased curcumin (F = 31.618, P < 0.05).

<b>CONCLUSIONSb>IkappaBalpha may play a role in the curcumin inducing apoptosis of PC3 cell, while the curcumin inducing apoptosis of LNCaP cells is by antioxidation and inhibiting metabolites formation in LNCaP cells.

Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Curcumin ; pharmacology ; Dose-Response Relationship, Drug ; Humans ; I-kappa B Proteins ; biosynthesis ; Male ; NF-KappaB Inhibitor alpha ; Prostatic Neoplasms ; metabolism ; pathology

<b>AIMb>To investigate the effect of ginkgolide B on the proliferation of VSMC and the secretion of chemokines by U937 cells stimulated by oxLDL or PAF. In addition, to analyze whether the effect of oxLDL is mediated through PAF receptor.

<b>METHODSb>Using 3H-Tdr incorporation assay, the proliferation of VSMC was measured. The protein and mRNA level of MCP-1 and IL-8 in U937 cells were determined by RT-PCR and ELISA. Using Western blotting the p65 and IkappaB was quantified. The binding of oxLDL to U937 cell was measured by a radio-ligand binding assay of 3H-PAF.

<b>RESULTSb>Ginkgolide B inhibited, in dose-dependent manner, the proliferation of VSMC and the secretion of chemokines by U937 cells stimulated by oxLDL, and inhibited the oxLDL-induced p65 activation and depletion of IKappaB. oxLDL inhibited PAF binding to U937 cells.

<b>CONCLUSIONb>Ginkgolide B, as a PAF antagonist, possesses the effect of inhibiting the proliferation of VSMC and the secretion of chemokines by U937 cells stimulated by oxLDL in vitro. The effect of oxLDL is, at least in part, mediated through PAF receptor.

Animals ; Aorta, Thoracic ; cytology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Chemokine CCL2 ; biosynthesis ; genetics ; Diterpenes ; isolation & purification ; pharmacology ; Dose-Response Relationship, Drug ; Ginkgo biloba ; chemistry ; Ginkgolides ; Humans ; I-kappa B Proteins ; metabolism ; Interleukin-8 ; biosynthesis ; genetics ; Lactones ; isolation & purification ; pharmacology ; Lipoproteins, LDL ; pharmacology ; Male ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; drug effects ; metabolism ; Plants, Medicinal ; chemistry ; Platelet Activating Factor ; antagonists & inhibitors ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Wistar ; Synaptotagmin I ; metabolism ; U937 Cells

<b>OBJECTIVEb>To investigate the effect of homocysteine (HCY) on activation of nuclear factor (NF-kappaB) and inhibitory factor IkappaB-alpha in human monocyte cell line THP-1, as well as its association with macrophage inflammatory protein (MIP-1alpha) upregulation.

<b>METHODSb>THP-1 monocytes were incubated with HCY, with and without NF-kappaB inhibitor pyrolidine dithiocarbamate (PDTC) pretreatment. Northern blot analysis and flow cytometry were used to detect MIP-1alpha mRNA and protein respectively. The nuclear protein NF-kappaB P65 subunit and the inhibitory protein IkappaB-alpha were analyzed by Western blotting.

<b>RESULTSb>Compared with controls, HCY, at a concentration of 0.1 mmol/L, was able to enhance the expression of MIP-1alpha mRNA (up to 3.69-fold) and protein (1.16-fold) in THP-1 monocytes, as well as enhance NF-kappaB P65 transcription to nuclear proteins. These actions were significantly suppressed after pretreatment with 100 micromol/L PDTC for 30 minutes before HCY incubation; whereas incubation of THP-1 monocytes with PDTC only had no effect on both the expression of MIP-1alpha and nuclear transcription of NF-kappaB P65. Moreover, the level of IkappaB-alpha protein in THP-1 monocytes decreased after a 30-minute incubation with HCY, which gradually increased after 120 minutes.

<b>CONCLUSIONSb>Homocysteine at a pathologic concentration stimulates MIP-1alpha expression in THP-1 monocytes, probably via NF-kappaB activation. Such activation may be caused by enhanced phosphorylation and degradation of the inhibitor protein IkappaB-alpha.

Cell Line, Tumor ; Chemokine CCL3 ; Chemokine CCL4 ; Homocysteine ; pharmacology ; Humans ; I-kappa B Proteins ; metabolism ; Leukemia, Monocytic, Acute ; metabolism ; pathology ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Monocytes ; metabolism ; NF-KappaB Inhibitor alpha ; NF-kappa B ; antagonists & inhibitors ; Phosphorylation ; Proline ; analogs & derivatives ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics ; Thiocarbamates ; pharmacology ; Transcription Factor RelA ; biosynthesis ; genetics ; Transcription, Genetic