OBJECTIVE: To determine the effect of tetramethylpyrazine (TMP) on the expression of migration inhibitory factor (MIF) in acute spinal cord injury (ASCI) in rats. METHODS: Allen's weight-drop method was used to establish a rat model of ASCI at T10. A total of 110 adult SD rats were divided into a sham operation group (group S, n=10), a control group (group C, n=50), and a TMP group (group T, n=50). Spinal cord functionality was measured by a modified Rivilin loxotic plate degree, BBB score, and combined behavioral score (CBS) at 1, 3, 5, 7, 14 and 21 d postoperatively. The injured spinal cord tissue samples were harvested at 1, 3, 6, 12 h and 1, 3, 5, 7, 14, 21 d postoperatively (n=5 at each time point) and used to prepare continuous histological sections, in which the expression of MIF was analyzed by immunohistochemistry. RESULTS: The degree in group T measured by modified Rivlin loxotic plate test after the ASCI was significantly higher than that in group C at 7, 14, and 21 d (P<0.05). BBB score in group T was significantly higher than that in group C at 5, 7, 14, and 21 d after the ASCI (P<0.05). CBS score in group C was significantly higher than that in group T at 5, 7, 14, and 21 d after the ASCI (P<0.05). The significantly low number of MIF positive cells was shown in group T when compared with that in group C at 12 h and 1, 3, 5, 7 d after the ASCI (P<0.05). As time passed, there was negative correlation between modified Rivlin loxotic plate degree and MIF expression and also between BBB score and MIF, and there was positive correlation between CBB score and MIF expression. CONCLUSION TMP has protective effect after the ASCI, and may promote the repair of injured spinal cord tissues. TMP may decrease the MIF expression in cells after the ASCI.
Animals ; Immunohistochemistry ; Intramolecular Oxidoreductases ; metabolism ; Macrophage Migration-Inhibitory Factors ; metabolism ; Pyrazines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Injuries ; metabolism

To compare the difference in tumor immunity and autoimmunity elicited by adenovirus (Ad) encoding human or murine tyrosinase-related protein 2 (AdhTRP2 or AdmTRP2), and to find the most effective way to induce immunity by AdhTRP2 or AdmTRP2, C57BL/6 mice were immunized with AdhTRP2 or AdmTRP2 intramuscularly at different doses of 10(5), 10(6), 10(7) and 10(8) separately (10 mice for each dose). Two weeks after the immunization, in vivo CTL assay and intracellular staining (ICS) of IFN-gamma were carried out to analyze the dose-effect relationship. Tumor growth and vitiligo (as an sign of autoimmunity) were observed until 3 months after challenge with 10(5) B16F10 tumor cells. The results showed that Ad encoding AdmTrp2 induced weak tumor immune response. Similar immunization with AdhTrp-2 elicited stronger protective immunity. CTL activity and IFN-gamma-produced CD8+T cells were directly proportional to dose of AdhTrp2 or AdmTrp2. Moreover, AdhTrp2 group showed tumor rejection in 100% of challenged mice till the end of 3rd month while 60% of mice immunized with AdmTrp2 were protected against tumor. In the whole process of this experiment, no vitiligo was observed in mice immunized either with AdhTrp2 or AdmTrp2. It is concluded that anti-melanoma responses induced by genetic vaccination expressing xenoantigens breaks immune tolerance effectively and is able to elicit strong antigen-specific cytotoxic T cell response without vitiligo.
Adenoviridae/metabolism ; Antineoplastic Agents/*pharmacology ; Cell Line, Tumor ; Cytokines/metabolism ; Immune System ; Immune Tolerance ; Interferon-gamma/metabolism ; Intramolecular Oxidoreductases/*biosynthesis ; Intramolecular Oxidoreductases/*genetics ; Mice, Inbred C57BL ; T-Lymphocytes, Cytotoxic/*metabolism ; Vitiligo/*metabolism

To investigate the effect of a microsomal prostaglandin E synthase-1 (mPGES-1) inhibitor MK886 on cell cycle of the human acute myeloid leukemia HL-60 cells. HL-60 cells were treated with different concentration of MK886 (10, 25, 50 µmol/L) for 24 h. Flow cytometry, Western blot and ELISA were used to measure cell cycle, cyclin D1, mPGES-1, PGE(2), Akt, P-Akt and C-MYC. The results indicated that after treated with MK886, the percentage of HL-60 cells decreased in G(0)/G(1) phase and increased in S phase, and expressions of mPGES-1, cyclin D1, P-Akt and C-MYC and synthesis of PGE(2) decreased significantly. It is concluded that MK886 can arrest HL-60 cells in G(0)/G(1) phase, the mechanism of which is possibly associated to inhibition of mPGES-1 expression, reduction of PGE(2) synthesis, suppression of Akt phosphorylation and C-MYC expression, down-regulation of cyclin D1 expression.
Cell Cycle ; drug effects ; HL-60 Cells ; Humans ; Indoles ; pharmacology ; Intramolecular Oxidoreductases ; antagonists & inhibitors ; Leukemia ; metabolism ; pathology ; Prostaglandin-E Synthases

Antigens, CD34 ; metabolism ; Biomarkers ; metabolism ; Cell Differentiation ; physiology ; Hair Follicle ; cytology ; Humans ; Intramolecular Oxidoreductases ; metabolism ; Keratinocytes ; metabolism ; Melanins ; metabolism ; Melanocytes ; metabolism ; PAX3 Transcription Factor ; metabolism ; Stem Cells ; metabolism

OBJECTIVE: To investigate the expression of MIF, NF-κB p65 and IL-1β in the tissue of nasal polyps and normal inferior turbinate, to analyze their relevance, and to explore their role in nasal polyps. METHOD: The infiltrating results of EOS and others inflammatory cells in 48 cases diagnosed as nasal polyps (nasal polyps group) were detected by HE staining, and the expression of MIF, NF-κB p65 and IL-1β were investigated by immunohistochemistry. Twenty-one patients who were performed septoplasty orthotics were included as the control group; the VAS and Lund-Kennedy score were used to evaluate the degree of nasal polyps in patients and the correlation analysis was conducted between the disease severity and the expression levels of this three factors. RESULT: (1) The infiltrating results of EOS and the expression level of MIF, NF-κB p65, IL-1β in nasal polyps group are obviously higher than these in the control group (P < 0.05); Spearman correlation analysis shows that MIF, NF-κb p65 and IL-1β are positively correlated with each other (r = 0.74, 0.66, 0.60, P < 0.05); the nuclear activation rate of NF-κB p65 is positively correlated with MIF, IL-1β (r = 0.67, 0.63, P < 0.05); the infiltration degree of EOS is positively correlated with MIF, IL-1β (r = 0.49, 0.55, P < 0.05), but has no correlation with the NF-κB p65 and its nuclear activation rate. (2) The VAS grade of the nasal polyps group is 8.24 ± 1.72 and the nasal endoscopic examination grade is 8.63 ± 3.81. Spearman correlation analysis shows that the VAS grade is positively correlated with the level of MIF (r = 0.71, P < 0.05), but had no correlation with NF-κB p65, its nuclear activation rate and IL-1β. The nasal endoscopic examination grade is positively correlated with MIF and the nuclear activation rate of NF-κB p65 (r = 0.79, 0.73, P < 0.05), but has no correlation with the level of NF-κB p65 and IL-1β (P > 0.05). CONCLUSION MIF, NF-κB p65 and IL-1β may promote the development of the nasal polyps, and there may exist the IL-1β--NF-κB--MIF approach in nasal polyps; MIF and NF-κB may participate in maintaining physiological function of inferior turbinate and have relations with the lightest sustained inflammation of inferior turbinate. The MIF and NF-κB p65 nuclear activation rate can be used as a standard of the nasal polyp severity and the judgement prognosis.
Humans ; Inflammation ; metabolism ; Interleukin-1beta ; metabolism ; Intramolecular Oxidoreductases ; metabolism ; Macrophage Migration-Inhibitory Factors ; metabolism ; Nasal Polyps ; metabolism ; Transcription Factor RelA ; metabolism

PURPOSE: In addition to cyclooxygenase-2 (COX-2) which is related to prostaglandin E2 synthesis, other enzymes such as cytosolic phospholipase A2 (cPLA2), microsomal prostaglandin E2 synthase-1 (mPGES-1), and 15-prostaglandin dehydrogenase (15-PGDH) have been suggested to be related to carcinogenesis of colorectal cancer (CRC). The aim of this study was to investigate the roles of cPLA2, COX-2, mPGES-1, and 15-PGDH in tumor progression. MATERIALS AND METHODS: cPLA2, COX-2, mPGES-1, 15-PGDH, and vascular endothelial growth factor (VEGF) expressions were immunohistochemically examined in 89 CRC, and their expressions were compared with each other or clinicopathologic parameters as well as VEGF as tumor progression parameters. RESULTS: cPLA2 was expressed in 54.5%, COX-2 in 80.5%, mPGES-1 in 96.4%, 15-PGDH in 46.1%, and VEGF in 65.9%. The expression of cPLA2 correlated with VEGF expression. COX-2 expression was correlated with the depth of invasion, tumor stage, cPLA2, and VEGF expressions. Moreover, VEGF revealed the highest expression in the tissues positive for both cPLA2 and COX-2. Furthermore, 15-PGDH expression was inversely correlated with VEGF expression. CONCLUSION: The present study demonstrates that cPLA2 and mPGES-1, in addition to COX-2, are constitutively overexpressed, and that 15-PGDH might be attenuated in colorectal cancer. Furthermore, cPLA2 and 15-PGDH as well as COX-2 could have an important role in tumor progression.
Aged ; Colorectal Neoplasms/*enzymology ; Cyclooxygenase 2/*metabolism ; Female ; *Gene Expression Regulation, Enzymologic ; Group IV Phospholipases A2/*metabolism ; Humans ; Hydroxyprostaglandin Dehydrogenases/*metabolism ; Immunohistochemistry ; Intramolecular Oxidoreductases/metabolism ; Male ; Middle Aged

OBJECTIVETo investigate the effect of lipopolysaccharide of Porphyromonas gingivalis (Pg-LPS) on the bio-thythetic pathway of prostaglandin E2 (PGE2) and its difference from lipopolysaccharide of Escherichia coli (Ec-LPS).

METHODSPurified Pg-LPS and Ec-LPS were used to stimulate a human monocytic cell strain THP-1. PGE2 concentration was determined by an enzyme immunoassay kit. The release of tritium labeled arachidonic acid (AA) was detected by a liquid scintillation counter. Reverse transcription polymerase chain reaction and western blot were used to analyse the expression of cytosolic phospholipase A2 (cPLA2) enzyme, cyclooxygenase-2 (COX-2), and microsomal prostaglandin E synthase-1 (mPGES-1).

RESULTSThe effect of Pg-LPS on induction of PGE2 and release of AA was significantly weaker than that of Ec-LPS (P < 0.05).Increased secretion of PGE2 was observed after stimulation with Pg-LPS for 6 h, which peak at 24 h at (221.40 +/- 29.46) ng/L; or with Ec-LPS for 1-48 h, at (161.80 +/- 17.31) approximately (379.80 +/- 37.35) ng/L. The highest levels of COX-2 and mPGES-1 were shown after 16 h treatment by Pg-LPS, or after 8 h and 16 h by Ec-LPS respectively.cPLA2 inhibitor AACOCF3 could lower the level of LPS-induced release of AA, while it did not influence the production of PGE2. COX-2 inhibitor NS-398 could remarkably reduce the concentration of PGE2.

CONCLUSIONSPg-LPS showed delayed and weaker effect on PGE2 biosynthetic pathway than Ec-LPS. Pg-LPS-induced PGE2 synthesis was mainly due to enhanced expression of COX-2 and mPGES-1, whereas cPLA2 played an insignificant role.

Cell Line ; Cyclooxygenase 2 ; metabolism ; Dinoprostone ; biosynthesis ; Humans ; Intramolecular Oxidoreductases ; metabolism ; Lipopolysaccharides ; pharmacology ; Monocytes ; drug effects ; metabolism ; Porphyromonas gingivalis ; Prostaglandin-E Synthases

Objective To investigate the effect of 15-Deoxy-△(12,14)-prostaglandin J2 (15 d-PGJ2) on the expression of macrophage migration inhibitory factor (MIF) and its underlying mechanism in J774A.1. Methods The murine monocyte/macrophage cell line J774A.1 were divided into six groups:lipopolysaccharide (LPS) group,incubated with 1 μg/ml LPS for 1 h;normal control group,incubated with PBS for 1 h;negative control group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h;15 d-PGJ2 group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h followed by 1 μg/ml LPS for 1 h;GW9662 group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h following GW9662 10 μmol/L for 1 h,and then incubated with 1 μg/ml LPS for 1 h;and Vehicle group,control of GW9662,GW9662 was replaced by its solvent DMSO. The expression of MIF was detected via immunofluorescence and agarose gel electrophoresis. RT-qPCR and Western blotting were used to test whether 15 d-PGJ2 could regulate mRNA and protein expression of MIF in J774A.1 upon LPS challenge. The effect of peroxisome proliferator-activated receptor-γ (PPAR-γ) antagonist GW9662 on the regulation of MIF by 15 d-PGJ2 was observed. The effects of 15 d-PGJ2 on the nuclear translocation of PPAR-γ upon LPS challenge were detected via high content screening analysis. Results MIF DNA and protein expressions were detected in J774A.1. MIF mRNA expression was up-regulated (1.75±0.09,P=0.037) when challenged with LPS and 15 d-PGJ2 inhibited its upregulation (0.84±0.08,P=0.026) in J774A.1. The protein level was consistent with the mRNA level. PPAR-γ antagonist GW9662 reversed the effect of 15 d-PGJ2 (mRNA,1.48±0.06,P=0.016;protein,1.28). Furthermore,nuclear translocation of PPAR-γ was regulated by 15 d-PGJ2 in J774A.1 upon LPS challenge(1.39±0.02 vs. 1.01±0.03,P=0.003). Conclusion 15 d-PGJ2 may down-regulate the MIF expression in J774A.1 in a PPAR-γ-dependent manner.
Anilides ; pharmacology ; Animals ; Cell Line ; Intramolecular Oxidoreductases ; metabolism ; Lipopolysaccharides ; Macrophage Migration-Inhibitory Factors ; metabolism ; Mice ; Monocytes ; drug effects ; PPAR gamma ; antagonists & inhibitors ; Prostaglandin D2 ; analogs & derivatives ; pharmacology

OBJECTIVETo investigate the gene expression of MAPEG in the cortex of concanavalin A (Con A)-induced mouse immune inflammatory model and the effect of cyclosporine A (Cs A).

METHODSMale Balb/c mouse immune inflammation model was developed by intravenous injection of Con A (20 mg/kg). Cs A (150 mg/kg) was intravenously infected prior to Con A administration. The MAPEG expressions were determined by RT-PCR.

RESULTmGST1, mGST3, LTC(4)S, FLAP and mPGES-1 were detected by RT-PCR but not mGST2. Eight hours after Con A treatment, mGST1 level was up-regulated to 1.2 approximately 1.5 folds of control with or without Cs A treatment. mGST3ìLTC(4)S, FLAP and mPGES-1 mRNA levels were not influenced by Con A administration.

CONCLUSIONImmune mechanism may be not involved in mGST1 up-regulation in this model and Con A does not alter arachidonic acid metabolism in cortex.

5-Lipoxygenase-Activating Proteins ; Animals ; Brain ; metabolism ; Carrier Proteins ; genetics ; metabolism ; Concanavalin A ; toxicity ; Cyclosporine ; pharmacology ; Eicosanoids ; metabolism ; Glutathione ; metabolism ; Glutathione Transferase ; genetics ; metabolism ; Intramolecular Oxidoreductases ; genetics ; metabolism ; Male ; Membrane Proteins ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; Prostaglandin-E Synthases

Glycyrrhizin (GR), triterpenoid saponin composed of one glycyrrhetinic acid (GA) and two glucuronic acids, is a main constituent of the hydrophilic fraction of licorice (Glycyrrhiza glabra) extracts and is known to have a wide range of pharmacological actions. In this study, we investigated the mechanism of GR effect on melanogenesis in B16 murine melanoma cells. The cellular levels of tyrosinase mRNA, protein, enzyme activities and melanin contents were increased by GR in a dose dependent manner. Expression of tyrosinase-related protein-2 (TRP-2) mRNA was also increased by GR, however, no significant change was observed on TRP-1. No cytotoxicity was observed at the effective concentration range of GR. GA showed no effect on melanogenesis at the equivalent nontoxic concentrations, indicating that glycoside structure is important in the stimulatory effect of GR on melanogenesis. These results indicate that GR-induced stimulation of melanogenesis is likely to occur through the transcriptional activation.
Animal ; Blotting, Western ; Glycyrrhetinic Acid/pharmacology ; Glycyrrhizic Acid/*pharmacology ; Intramolecular Oxidoreductases/genetics/metabolism ; Melanins/*biosynthesis ; Melanoma, Experimental/enzymology/*metabolism ; Mice ; Monophenol Monooxygenase/genetics/metabolism ; Proteins/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction