OBJECTIVETo study possible immunobiological potential of Osmunda japonica Thunb.

METHODSImmunomodulatory effects of ethanol extracts prepared from rhizomes of O. japonica and phenolic compounds isolated from the extracts were investigated under the in vitro conditions using the rat peritoneal cells (2×10(6)/mL; 24 h culture). Biosynthesis of nitric oxide (NO) was assayed by Griess reagent, production of prostaglandin E2 (PGE2) and secretion of cytokines were determined by enzyme-linked immunoabsorbent assay.

RESULTSThe extracts activated dose dependently, with the onset at 2.5-5 μmol/L concentrations, the high output NO production, and secretion of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β). Mild enhancement of NO was produced by the aldehyde-type phenolics 4-hydroxybenzaldehyde and 3,4-hydroxybenzaldehyde. In contrasts, the acetone-type phenolics 4-hydroxybenzalacetone and 3,4-hydroxybenzalacetone inhibited production of immune mediators including cytokines (TNF-α, IL-1β, IL-6), NO, and PGE2. The 3,4-hydroxybenzalacetone was more effective than 4-hydroxybenzaldehyde. The IC50s estimates ranged within the interval of 5-10 μmol/L. No signs of cytotoxicity were observed up to the 50 μmol/L concentration of the compounds.

CONCLUSIONPhenolic compounds contained in medicinal herb Osmunda japonica possess distinct immunomodulatory activity.

Animals ; Cell Survival ; drug effects ; Cells, Cultured ; Dinoprostone ; biosynthesis ; Female ; Ferns ; chemistry ; Immunologic Factors ; pharmacology ; Interferon-gamma ; pharmacology ; Lipopolysaccharides ; pharmacology ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type II ; genetics ; metabolism ; Peritoneum ; cytology ; drug effects ; Phenols ; chemistry ; isolation & purification ; pharmacology ; Plant Extracts ; chemistry ; isolation & purification ; pharmacology ; Polymyxin B ; pharmacology ; Proline ; analogs & derivatives ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Thiocarbamates ; pharmacology

Mitochondrial dysfunction and endoplasmic reticulum (ER) stress are considered the key determinants of insulin resistance. Impaired mitochondrial function in obese animals was shown to induce the ER stress response, resulting in reduced adiponectin synthesis in adipocytes. The expression of inducible nitric oxide synthase (iNOS) is increased in adipose tissues in genetic and dietary models of obesity. In this study, we examined whether activation of iNOS is responsible for palmitate-induced mitochondrial dysfunction, ER stress, and decreased adiponectin synthesis in 3T3L1 adipocytes. As expected, palmitate increased the expression levels of iNOS and ER stress response markers, and decreased mitochondrial contents. Treatment with iNOS inhibitor increased adiponectin synthesis and reversed the palmitate-induced ER stress response. However, the iNOS inhibitor did not affect the palmitate-induced decrease in mitochondrial contents. Chemicals that inhibit mitochondrial function increased iNOS expression and the ER stress response, whereas measures that increase mitochondrial biogenesis (rosiglitazone and adenoviral overexpression of nuclear respiratory factor-1) reversed them. Inhibition of mitochondrial biogenesis prevented the rosiglitazone-induced decrease in iNOS expression and increase in adiponectin synthesis. These results suggest that palmitate-induced mitochondrial dysfunction is the primary event that leads to iNOS induction, ER stress, and decreased adiponectin synthesis in cultured adipocytes.
3T3-L1 Cells ; *Adipocytes/drug effects/metabolism ; Adiponectin/biosynthesis ; Adipose Tissue/metabolism ; Animals ; Endoplasmic Reticulum Stress/drug effects ; Insulin Resistance/genetics ; Mice ; Mitochondria/drug effects/*metabolism/pathology ; Mitochondrial Turnover/drug effects/genetics ; *Nitric Oxide Synthase Type II/genetics/metabolism ; Nuclear Respiratory Factor 1 ; Obesity/genetics/metabolism ; Palmitic Acid/pharmacology ; Thiazolidinediones/pharmacology

OBJECTIVE: To investigate the effect of cardiomyopeptidin for injection on energy metabolism in isolated hearts of young rats after ischemia-reperfusion injury. METHODS: Fifty young healthy SD rats(aged 20 +/- 3 days and weighing 50-70 g) were randomly divided into 5 groups: a normal control group (NC group, n = 10 ): the isolated hearts were stable for 20 min, and then 150 min continuous perfusion; a normal + cardiomyopeptidin group (NCMP group, n = 10): the same as the normal control group, but K-H buffer solution was added with 50 mg/L cardiomyopeptidin, and 3 ischemia-reperfusion injury model groups, including a model control group (n = 10): the isolated rat hearts were perfused with K-H buffer and then arrested with cardioplegic solution; a CMP1 group (n = 10): the ST.Thomas'II cardioplegic solution was added with 100 mg/L cardiomyopeptidin; CMP2 group (n=10): K-H buffer and ST.Thomas'II cardioplegic solution was added with 50 mg/L and 100 mg/L cardiomyopeptidin respectively. The cardiac functional indexes were monitored, including heart rate, myocardial contractility and diastolic function, peak systolic and diastole myocardial velocities and coronary flow. In the 3 ischemia-reperfusion injury model groups, myocardial ultrastructure was observed through transmission electron microscopy; the creatine kinase isoenzyme (CK-MB) concentration was measured in the fluid outflow of coronary; the content of Na+-K+ ATPase, Ca2+-Mg2+ ATPase, total ATPase, superoxide dismutase (SOD), malondialdehyde (MDA), nitric oxide(NO), total nitric oxide synthase (TNOS), inducible nitric oxide synthase (iNOS) and aldosereductase were measured in the myocardium tissue; the relative expression levels of iNOS, eNOS, and Akr1b4 mRNA in the myocardial tissue were also detected by real-time fluorescence quantitative PCR. RESULTS: In the NC group, after prolonged perfusion, the cardiac function of isolated hearts had no significant change. Cardiomyopeptidin for injection had no significant effect on normal isolated hearts. Compared with the model control group, the cardiac function indexes and coronary flow in the groups treated with cardiomyopeptidin decreased much less. Cardiac myofibrillar fragmentation and mitochondrial swelling were observed in the control group, while in the CMP groups, the myocardial structure was nearly complete, and only mild mitochondria swelling and degeneration could be seen. After the reperfusion, the content of CK-MB was increased in the control group. Compared with the model control group, the CK-MB content was lower in the CMP1 and CMP2 groups. There was a slight decline in the contents of Na+-K+ ATPase, Ca2+-Mg2+ ATPase, and Total ATPase in the CMP1 and CMP2 groups, and an increase in SOD activity (P < 0.01 or P < 0.05). The concentration of NO and MDA produced after the ischemia-reperfusion injury was much lower in the CMP1 and CMP2 groups. The activity of iNOS and aldosereductase was inhibited, the expression levels of iNOS, and Akr1b4 mRNA were significantly down-regulated in the CMP1 and CMP2 groups. These changes were more prominent in the CMP2 group (P < 0.01 or P < 0.05). The eNOS mRNA levels in the CMP2 group was up-regulated (P < 0.05). CONCLUSION Cardiomyopeptidin for injection may improve the energy metabolism, improve coronary blood flow and cardiac function after the reperfusion, thus protecting immature myocardial against ischemia-reperfusion injury in young rats. Administration of it in both K-H buffer and ST.Thomas'II cardioplegic solution is better than adding it in cardioplegic solution alone. The mechanism may be associated with the inhibition the mRNA expression of iNOS and Akr1b4 in cardiomyocytes, the inhibition activity of iNOS and aldosereductase, and the decrease of NO production.
Aldehyde Reductase ; genetics ; metabolism ; Animals ; Energy Metabolism ; drug effects ; Female ; In Vitro Techniques ; Male ; Myocardial Reperfusion Injury ; metabolism ; Myocardium ; metabolism ; ultrastructure ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type II ; genetics ; metabolism ; Peptides ; pharmacology ; Random Allocation ; Rats ; Rats, Sprague-Dawley

MHC class II expression is controlled mainly at transcriptional level by class II transactivator (CIITA), which is a non-DNA binding coactivator and serves as a master control factor for MHC class II genes expression. Here, we describe the function of a novel splice-isoform of CIITA, DC-expressed caspase inhibitory isoform of CIITA (or DC-CASPIC), and we show that the expression of DCCASPIC in DC is upregulated upon lipopolysaccharides (LPS) induction. DC-CASPIC localizes to mitochondria, and protein-protein interaction study demonstrates that DC-CASPIC interacts with caspases and inhibits its activity in DC. Consistently, DC-CASPIC suppresses caspases-induced degradation of nitric oxide synthase-2 (NOS2) and subsequently promotes the synthesis of nitric oxide (NO). NO is an essential regulatory molecule that modulates the capability of DC in stimulating T cell proliferation/activation in vitro; hence, overexpression of DC-CASPIC in DC enhances this stimulation. Collectively, our findings reveal that DC-CASPIC is a key molecule that regulates caspases activity and NO synthesis in DC.
Alternative Splicing ; Amino Acid Sequence ; Animals ; Base Sequence ; CARD Signaling Adaptor Proteins ; genetics ; metabolism ; Cell Line ; Dendritic Cells ; drug effects ; immunology ; metabolism ; Humans ; In Vitro Techniques ; Lipopolysaccharides ; pharmacology ; Lymphocyte Activation ; Mice ; Mice, Inbred C57BL ; Mitochondria ; metabolism ; Molecular Sequence Data ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type II ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; Protein Isoforms ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; T-Lymphocytes ; immunology ; metabolism ; Trans-Activators ; genetics ; metabolism ; Up-Regulation ; drug effects

OBJECTIVETo evaluate the effect of clearance of superoxide anion by catechin on the expression of nitrogen monoxidum (NO) and endothelial nitricoxide synthase (eNOS) and apoptosis in endothelial progenitor cells (EPCs) induced by angiotensin II (Ang II).

METHODSThe marrow endothelial progenitor cells of Sprague-Dawley rats were isolated and assigned to control (no treatment), Ang II treatment and Ang II + catechin treatment groups. After 48 hrs of culture, the concentration of O2*- in the supernate was measured by the NBT method, and NO concentration in the supernate was measured by the nitrate reductase method; the apoptosis rate of EPCs was detected by the TUNEL method; the mRNA expression of eNOS was detected by RT-PCR; the protein expression of eNOS was detected by Western blot analysis.

RESULTSAng II of 10-6 mol/L was determined as the suitable concentration for cell induction by the MTT test. Catechin of 400 mg/L was determined as an advisable intervention dosage. The apoptosis rate of EPCs in the control, the Ang II and the Ang II+catechin treatment groups were 2.48+/-0.12%, 54.18+/-0.77% and 16.87+/-0.35%, respectively, and there were significant differences among the three groups (P<0.01). The O2*- concentration in the Ang II and the Ang II+catechin treatment groups (81.7+/- 3.6 and 62.3+/- 2.2 U/L respectively) was significantly higher than that in the control group (33.7+/- 2.8 U/L) (P<0.01). An increased NO concentration was also found in the Ang II (189. 8+/- 9.0 micromol/L) and the Ang II+catechin treatment groups (276.4+/- 10.1 micromol/L) compared with that in the control group (105.8+/- 9.8 micromol/L) (P<0.01). There were significant differences in the concentrations of O2*- and NO between the Ang II and the Ang II+catechin treatment groups (P<0.05). The mRNA (P<0.05) and protein expression (P<0.01) of eNOS in the Ang II and the Ang II+catechin treatment groups increased significantly compared with those in the control group. The Ang II+catechin treatment group showed increased eNOS protein expression compared with the Ang II group (P<0.05).

CONCLUSIONSAng II may induce the generation of O2*-, inactivate NO and increase gene and protein expression of eNOS in EPCs. Catechin might decrease the apoptosis of EPCs through the effective clearance of O2*-and the reduction of NO inactivation and of eNOS protein uncoupling.

Angiotensin II ; pharmacology ; Animals ; Apoptosis ; drug effects ; Catechin ; pharmacology ; Cell Survival ; drug effects ; Endothelial Cells ; drug effects ; metabolism ; Female ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type III ; analysis ; genetics ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Stem Cells ; drug effects ; metabolism ; Superoxides ; metabolism

The effect of extracellular beta-(1-->3), (1-->6)-glucan, produced by Paenibacillus polymyxa JB115, on nitric oxide (NO) production in RAW264.7 macrophages was investigated. beta-glucan induced the production of NO by RAW264.7 macrophages in a concentration- and time-dependent manner. Moreover, beta-glucan stimulation increased the mRNA expression of iNOS, COX-2 and IL-6 in RAW264.7 macrophages in a concentration-dependent manner.
Animals ; Bacillus/*metabolism ; Cell Line ; Cyclooxygenase 2/biosynthesis/genetics ; Interleukin-6/biosynthesis/genetics ; Lipopolysaccharides/pharmacology ; Macrophages/*drug effects/enzymology/immunology ; Mice ; Nitric Oxide/*biosynthesis/immunology ; Nitric Oxide Synthase Type II/biosynthesis/genetics/metabolism ; RNA, Messenger/biosynthesis/genetics ; Reverse Transcriptase Polymerase Chain Reaction ; beta-Glucans/metabolism/*pharmacology

The purpose of this study was to identify the effect of sildenafil citrate on IL-1 beta induced nitric oxide (NO) synthesis and iNOS expression in human synovial sarcoma SW982 cells. IL-1 beta stimulated the cells to generate NO in both dose- and time-dependent manners. The IL-1 beta -induced NO synthesis was inhibited by guanylate cyclase (GC) inhibitor, LY83583. When the cells were treated with 8-bromo-cGMP, a hydrolyzable analog of cGMP, NO synthesis was increased upto 5-fold without IL-1 beta treatment suggesting that cGMP is an essential component for increasing the NO synthesis. Synoviocytes and chondrocytes contain strong cGMP phosphodiesterase (PDE) activity, which has biochemical features of PDE5. When SW982 cells were pretreated with sildenafil citrate (Viagra), a PDE5 specific inhibitor, sildenafil citrate significantly inhibited IL-1 beta -induced NO synthesis and iNOS expressions. From this result, we noticed that PDE5 activity is required for IL-1 beta -induced NO synthesis and iNOS expressions in human synovial sarcoma cells, and sildenafil citrate may be able to suppress an inflammatory reaction of synovium through inhibition of NO synthesis and iNOS expression by cytokines.
Anti-Inflammatory Agents/immunology/pharmacology ; Cell Line, Tumor ; Cyclic GMP/analogs & derivatives/immunology/metabolism ; Cyclic Nucleotide Phosphodiesterases, Type 2/antagonists & inhibitors/metabolism ; Humans ; Interleukin-1beta/*metabolism ; Male ; Nitric Oxide/*biosynthesis/genetics/immunology ; Nitric Oxide Synthase Type II/*biosynthesis/genetics/immunology ; Phosphodiesterase Inhibitors/immunology/*pharmacology ; Piperazines/immunology/*pharmacology ; Purines/immunology/pharmacology ; Signal Transduction/drug effects/genetics/immunology ; Sulfones/immunology/*pharmacology ; Synovial Membrane/enzymology/immunology

OBJECTIVETo observe the effect of IPS-B2 on mouse peritoneal macrophages and the transcription of IL-1beta, IL-6, TNF-alpha and iNOS.

METHODELISA method and Griess method were used to detect the effect of mouse peritoneal macrophages produce cytokines IL-1beta, IL-6, TNF-alpha and cytotoxic effectors NO. The transcription of IL-1beta, IL-6, TNF-alpha and iNOS was detected by real-time RT-PCR method.

RESULTIPS-B2 could not promote mouse peritoneal macrophage production, but it could significantly improve the IL-1beta, IL-6, TNF-alpha content in mouse peritoneal macrophages culture supernatant, and increase the gene expression of IL-1beta, IL-6, TNF-alpha and iNOS.

CONCLUSIONIPS-B2 can enhance the ability of peritoneal macrophages to excrete bioactive substances and promote the transcription of bioactive substances to antitumor.

Agaricales ; chemistry ; Animals ; Gene Expression Regulation ; drug effects ; Interleukin-1beta ; biosynthesis ; genetics ; Interleukin-6 ; biosynthesis ; genetics ; Macrophages, Peritoneal ; drug effects ; metabolism ; Mice ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type II ; metabolism ; Polymerase Chain Reaction ; Polysaccharides ; pharmacology ; RNA, Messenger ; biosynthesis ; Transcription, Genetic ; drug effects ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics

To explore the mechanism of action of butylphalide (NBP) against the injury following oxygen glucose deprivation/reoxygenation (OGD/R) in rat cortical neurons, neurons of Wistar newborn rats were prepared by filtering through a mesh, centrifugation and trypsogen digestion. A simple, stable and reliable in vitro model of OGD/R of neurons was established. We studied the activation, the nuclear translocation of NF-kappaB p65 and the mRNA expression of iNOS affected by NBP in each group neuron by RT-PCR. NBP is proved to be able to add cellular vigor and decrease LDH release. The mRNA expression of iNOS in neurons after OGD 4 h/R 8 h decreased when treated with NBP. There is statistical difference between each concentration of NBP that it adds cellular vigor, decreases LDH release and expression of iNOS in neurons after OGD 4 h/R 8 h. There is also statistical difference between NBP (100 micromol x L(-1)) and PDTC (100 micromol x L(-1)). It is proved that NBP can protect neurons, block upregulation of iNOS mRNA, and restrain activation of NF-kappaB in neurons.
Animals ; Animals, Newborn ; Antioxidants ; pharmacology ; Benzofurans ; pharmacology ; Cell Hypoxia ; Cerebral Cortex ; cytology ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Female ; Glucose ; deficiency ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Male ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Nitric Oxide Synthase Type II ; biosynthesis ; genetics ; Pyrrolidines ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Wistar ; Thiocarbamates ; pharmacology ; Transcription Factor RelA ; metabolism

OBJECTIVETo observe the influence of hypoxia-inducible factor 1 alpha (HIF-1alpha) on angiogenesis in prostate carcinoma in vivo and to investigate its molecular mechanism.

METHODSLNCaP/HIF-1alpha and LNCaP cells were cultured, the level of PSA in the supernatant of the culture medium detected by ELISA assay before and after the transfection, and the cellular cycle measured by flow cytometry. Nude mouse models of subcutaneous tumor were established with LNCaP/HIF-1alpha and LNCaP cells, the tumor growth observed, and tumor specimens collected for immunohistochemical staining.

RESULTSCompared with the LNCaP cells, LNCaP/HIF-1alpha cells showed an obviously decreased PSA level (t = 8.243, P < 0.05) and enhanced proliferous activity. The tumorigenesis rate increased and the tumorigenesis time advanced in the LNCaP/HIF-1alpha group of the nude mice. Immunohistochemistry displayed higher expressions of VEGF, iNOS and Ang-2 in the LNCaP/HIF-1alpha than in the LNCaP group.

CONCLUSIONThe over-expression of HIF-1alpha can up-regulate VEGF and iNOS involved in angiogenesis in vivo and contribute to the invasive potency of LNCaP cells. HIF-1alpha may have no influence on Ang-2 either in vitro or in vivo, while the expression of Ang-2 is regulated by other factors in vivo.

Animals ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Flow Cytometry ; Gene Expression Regulation, Neoplastic ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; physiology ; Immunohistochemistry ; Male ; Mice ; Mice, Nude ; Neoplasms, Experimental ; blood supply ; metabolism ; pathology ; Neovascularization, Pathologic ; physiopathology ; Nitric Oxide Synthase Type II ; biosynthesis ; Prostatic Neoplasms ; blood supply ; genetics ; pathology ; Transfection ; Transplantation, Heterologous ; Tumor Burden ; Vascular Endothelial Growth Factor A ; biosynthesis