Lung preservation study by canine sequential bilateral single lung transplantation model.
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Animals↗
Biomedical subjects
Y J Jeon
Publications and source records attributed to Y J Jeon.
Transcription factor decoy for activator protein-1 (AP-1) inhibits high glucose- and angiotensin II-induced type 1 plasminogen activator inhibitor (PAI-1) gene expression in cultured human vascular smooth muscle cells.
AIMS/HYPOTHESIS: Multiple factors, including hyperglycaemia and angiotensin II (Ang II), stimulate plasminogen activator inhibitor-1 (PAI-1) gene expression in human vascular smooth muscle cells. This study tested the hypothesis that hyperglycaemia and Ang II stimulate PAI-1 gene expression through activator protein-1 (AP-1) binding sites. METHODS: We evaluated the role of AP-1 in PAI-1 gene expression in human vascular smooth muscle cells under high D-glucose and Ang II stimulation using a double-stranded cis-element AP-1 oligodeoxynucleotide (decoy ODN). RESULTS: Activator protein 1 activity was stimulated by high glucose and Ang II treatment and the AP-1 decoy ODN, but not a mismatched decoy ODN, competed for AP-1 activity. The increase in PAI-1 expression by high glucose and Ang II was significantly attenuated by the AP-1 decoy ODN (p <0.05 or p < 0.01). The increase in PAI-1 expression by high glucose and Ang II action on AP-1 sites was also confirmed by promoter analysis of PAI-1. Activator protein 1 activation in response to either high glucose or co-stimulation with high glucose and Ang II was inhibited completely by calphostin C (a PKC inhibitor) and partially by genistein (a protein tyrosine kinase inhibitor). CONCLUSION/INTERPRETATION: This study shows that high glucose and Ang II stimulate PAI-1 expression through AP-1 binding sites. Signal transduction after AP-1 activation by both high glucose and Ang II largely depends on PKC activation. These data indicate an important role for AP-1 in PAI-1 expression.
Angiotensin II↗
Activation of mitogen-activated protein kinase pathways by angelan in murine macrophages.
In our previous studies, we showed that angelan, a polysaccharide purified from Angelica gigas Nakai, specifically activated macrophages to induce cytokines including inducible nitric oxide synthase (iNOS) which has strong anti-tumor activities [Immunopharmacology, 1999; 43: 1.]. In the present study, we investigated the intracellular signal transduction pathways involved in the angelan-induced iNOS synthesis by murine macrophages. Protein tyrosine phosphorylation was induced within 5 min by angelan, and the blocking of protein tyrosine kinases (PTKs) inhibited down-stream pathways leading to iNOS production in response to angelan. Treament of RAW 264.7 cells with angelan resulted in significant activation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) and p38, while stress-activated protein kinase/c-Jun NH2 terminal kinase (SAPK/JNK) was not activated by angelan. The specific p38 inhibitor SB203580 abrogated the angelan-induced iNOS synthesis, whereas the selective mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase 1 (MEK-1) inhibitor PD98059 did not affect the iNOS induction. In conclusion, we demonstrate that PTK and p38 MAPK activation are required to transduce signals leading to iNOS expression in angelan-stimulated murine macrophages.
Animals↗
Experimental evidences and signal transduction pathways involved in the activation of NF-kappa B/Rel by angelan in murine macrophages.
In our previous studies, we showed that angelan, a polysaccharide purified from Angelica gigas Nakai, activated macrophages to induce the translocation of NF-kappa B/Rel into nucleus and DNA binding to its cognate site in the promoter of iNOS gene [Immunopharmacology 43 (1999) 1; Immunopharmacology 49 (2000) 275]. In the present study, we showed that angelan induces the transcriptional activation of NF-kappa B/Rel and investigated the intracellular signal transduction pathways involved in the angelan-induced NF-kappa B/Rel activation by murine macrophages. Treatment of RAW 264.7 cells with angelan resulted in significant activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38, while stress-activated protein kinase/c-Jun NH2 terminal kinase (SAPK/JNK) was not activated by angelan. The specific p38 inhibitor SB203580 abrogated the angelan-induced NF-kappa B/Rel activation, whereas the selective MAPK/extracellular signal-regulated kinase 1 (MEK-1) inhibitor PD98059 did not affect the NF-kappa B/Rel induction. Treatment of RAW 264.7 cells with both anti-CD14 Ab and anti-CR3 Ab significantly blocked angelan-induced NF-kappa B/Rel activation. In conclusion, we demonstrate that angelan induces NF-kappa B/Rel activation through the CD14 and CR3 membrane receptor and p38 kinase that is critically involved in the signal transduction leading to NF-kappa B/Rel activation in murine macrophages.
Animals↗
Suppression of IL-8 gene expression by radicicol is mediated through the inhibition of ERK1/2 and p38 signaling and negative regulation of NF-kappaB and AP-1.
We show that radicicol, an anti-fungal agent, inhibits interleukin-8 (IL-8) production by the human monocyte line THP-1 in response to phorbol-12-myristate-13-acetate/lipopolysaccharide (PMA/LPS). IL-8 is a potent chemokine and needs for an optimal immune response--such as inflammation by activation of neutrophils. The decrease in PMA/LPS-induced IL-8 mRNA expression was demonstrated by quantitative reverse transcription-polymerase chain reaction (RT-PCR). Since the promoter in IL-8 gene contains binding motifs for NF-KB, AP-1. and NF-IL6, which appear to be important in IL-8 induction, the effects of radicicol on the activation of these transcription factors were examined. Treatment of radicicol to THP-1 cells produced a strong inhibition of NF-KB and AP-1, while NF-IL6 was not significantly affected by radicicol. Western blot analysis showed that radicicol inhibited the phosphorylation and phosphotransferase activities of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38. PD98059 and SB203580, known as a specific inhibitor of MEKI and p38 kinase, respectively, inhibited IL-8 gene expression showing that both of the kinase pathways are involved in IL-8 regulation in human monocytes. Collectively, this series of experiments indicates that radicicol inhibits IL-8 gene expression by blocking ERK1/2 and p38 signaling.
Blotting, Western↗
Molecular cloning and high-level expression of G2 protein of hantaan (HTN) virus 76-118 strain in the yeast Pichia pastoris KM71.
Hantaan viral G2 envelope gene, which is known to be one of major antigens and induce neutralizing antibodies, was cloned into expression vector pHIL-S1 which consists of AOX1 promoter, PHO1 signal sequence, HIS4 gene and other components. The recombined plasmid was transformed into methylotropic yeast, Pichia pastoris of KM71 and recombinant strains harboring multi-copy of G2 gene were selected. Expression of the cloned G2 gene was confirmed with Western blot analysis using anti-sera of guinea pig immunized with the carboxyl terminal region of G2 protein expressed in Escherichia coli. The expression of G2 gene from the recombinant strain was tightly repressed by dextrose and effectively induced by methanol, an inducer of AOX1 promoter. The highest expression level was observed from 1 day after induction and maintained at the same level for up to 4 days.
Animals↗
Paclitaxel-induced immune suppression is associated with NF-kappaB activation via conventional PKC isotypes in lipopolysaccharide-stimulated 70Z/3 pre-B lymphocyte tumor cells.
Paclitaxel, a potent antitumor agent, has been shown to be lipopolysaccharide (LPS) mimetic in mice, stimulating signaling pathways and gene expression indistinguishably from LPS. In the present study, we showed the intracellular signaling pathway of paclitaxel-induced nuclear factor-kappaB (NF-kappaB) activation and its suppressive effect on LPS-induced signaling in murine 70Z/3 pre-B cells. Stimulation of 70Z/3 cells with LPS for 30 min caused activation of NF-kappaB in the nuclei by detection of DNA-protein binding specific to NF-kappaB. Similarly, paclitaxel also produced a marked and dose-related NF-kappaB activation. However, pretreatment of cells with 10 microM paclitaxel for 18 h resulted in complete inhibition of LPS-mediated NF-kappaB activation. Interestingly, the activity of IkappaB kinase (IKK-beta), which plays an essential role in NF-kappaB activation through IkappaB phosphorylation, was largely enhanced in paclitaxel-treated cells, detected as IkappaBalpha phosphorylation. Because protein kinase C (PKC) is implicated in the activation of NF-kappaB via IKK-beta, the effect of paclitaxel on PKC activation was also measured. It was shown that NF-kappaB nuclear translocation and DNA binding in response to paclitaxel was completely blocked by the conventional PKC inhibitor, Gö 6976. Moreover, immunoblotting analysis with paclitaxel-treated cell extract demonstrated that the conventional PKC isotype PKC-alpha was found to be involved in the regulation of paclitaxel-induced NF-kappaB activation, as determined by electrophoretic mobility shift of PKC. Therefore, these data suggest that paclitaxel may activate IKK-beta via conventional PKC isotypes, resulting in NF-kappaB activation and, finally, desensitization of LPS-inducible signaling pathway in 70Z/3 pre-B cells.
Animals↗
Polychlorinated biphenyls activate caspase-3-like death protease in vitro but not in vivo.
We prove here that serum albumin inhibits apoptosis induced by polychlorinated biphenyls (PCBs), confirming that serum albumin binds to PCB, and that the albumin-PCB complexes inhibit apoptosis in HL-60 cells. We found that PCB (50 microM) increased the activity of caspase-3-like protease when HL-60 cells, as well as splenocytes, were cultured in "serum-free medium." Benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-fmk) inhibited apoptosis in cells cultured in the serum-free medium containing 50 microM PCB. To elucidate whether or not PCBs induce apoptosis in vivo, we examined apoptosis of splenocytes by administering PCB to ICR mice (100, 500, 1000 mg x kg(-1) x d(-1)) for 5 d and characterizing splenocytes. Interestingly, splenocytes treated with PCB did not show any changes characteristic of apoptosis. These results demonstrate that PCB activates the caspase-3-like death protease in vitro in serum-free medium, but does not induce apoptosis of splenocytes in vivo, suggesting that blood serum may mask the apoptosis induced by PCB.
Animals↗
New immunosuppressive activity of dykellic acid.
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Animals↗
Prodigiosin blocks T cell activation by inhibiting interleukin-2Ralpha expression and delays progression of autoimmune diabetes and collagen-induced arthritis.
Prodigiosin (PDG) was previously reported to be a T cell-specific immunosuppressant. Here we describe the mechanism of action of PDG in T cells and the effect of PDG on autoimmune diseases. PDG selectively suppresses concanavalin A (Con A)-induced T cell proliferation, but has little effect on lipopolysaccharide-induced proliferation of B cells and nitric oxide production of macrophages. Although PDG does not block interleukin (IL)-2 production, it efficiently inhibits interleukin-2 receptor alpha-chain (IL-2Ralpha) expression, and this results in a disruption of the IL-2/IL-2R signaling pathway, on which a great part of the regulation of T cell activation depends. PDG blocks T cell differentiation into effector helper T cells secreting interferon-gamma and IL-4 as well as into effector cytotoxic T lymphocytes expressing perforin, which is at least in part resulting from inhibition of the IL-2/IL-2R signaling. PDG indirectly blocks signal transducer and activator of transcription activation by inhibiting cytokine signalings in Con A-activated T cells, although it does not inhibit the activation of nuclear factor-kappaB, nuclear factor of activated T cells, and activator protein-1. As direct evidence of immunosuppression in vivo, we show that PDG markedly reduced blood glucose levels and cellular infiltration into the pancreatic islets in nonobese diabetic mice, and that it also delays the onset of collagen-induced arthritis in DBA/1 mice. In conclusion, our results demonstrate that PDG has a unique mode of action, namely, that it blocks T cell activation by inhibiting primarily IL-2Ralpha expression in the IL-2/IL-2R signaling, and show that this compound represents a promising immunosuppressant candidate for the treatment of autoimmune diseases.
Animals↗
2-Amino-3-methylimidazo[4,5-f]quinoline inhibits nitric oxide production in lipopolysaccharide-stimulated RAW 264.7 cells by blocking p38 kinase activation.
We show that 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), a heterocyclic amine, significantly inhibits nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated macrophages. The decrease in NO production was found to correlate well with a decrease in inducible NO synthase (iNOS) mRNA expression as demonstrated by Northern blot analysis. Treatment of RAW 264.7 cells with IQ selectively inhibited the activation of NF-kappaB/Rel, an important transcription factor of iNOS gene expression, while neither AP-1 nor Oct was affected by IQ. Since iNOS transcription has been shown recently to be under the control of the p38 kinase signaling cascade, we assessed the effect of IQ on p38 kinase activation. Treatment of RAW 264.7 with IQ inhibited LPS-stimulated p38 kinase phosphorylation in a dose-related manner. IQ also inhibited the p38 kinase activity. Collectively, this series of experiments indicates that IQ inhibits LPS-induced expression of iNOS gene in RAW 264.7 cells. Based on our findings, the most likely mechanism that can account for this biological effect involves the negative regulation of NF-kappaB/Rel and p38 kinase pathway.
Animals↗
Dexamethasone inhibits IL-1 beta gene expression in LPS-stimulated RAW 264.7 cells by blocking NF-kappa B/Rel and AP-1 activation.
In the present study, the mechanism by which dexamethasone (DEX) inhibited IL-1beta gene expression in bacterial lipopolysaccharide (LPS)-activated RAW 264.7 cells was investigated. The decrease in LPS-induced IL-1beta mRNA expression was demonstrated by quantitative reverse transcription polymerase chain reaction (RT-PCR). Since the promoter in IL-1beta gene contains binding motifs for NF-kappaB/Rel, AP-1, NF-IL6, and CREB/ATF, which appear to be important in LPS-mediated IL-1beta induction, the effects of DEX on the activation of these transcription factors were examined. Treatment of DEX to RAW 264.7 cells induced a dose-related inhibition of NF-kappaB/Rel and AP-1 in chloramphenicol acetyltransferase activity, while neither NF-IL6 nor CREB/ATF activation was affected by DEX. Treatment of RAW 264.7 cells with DEX inhibited DNA binding of NF-kappaB/Rel and AP-1 proteins to their cognate DNA sites as measured by electrophoretic mobility shift assay (EMSA). DEX treatment caused a significant reduction in nuclear c-rel, p65, and p50 protein contents, and these decreases were paralleled by the accumulation of cytoplasmic c-rel, p65, and p50. DEX treatment of RAW 264.7 cells did not inhibit the nuclear translocation of c-jun and c-fos. We found that the inhibition of IL-1beta production by DEX is not related to p38, which is important in the IL-1beta induction. These results suggest that DEX may inhibit IL-1beta gene expression by a mechanism involving the blocking of LPS-induced NF-kappaB/Rel and AP-1 activation.
Animals↗
A new SUMO-1-specific protease, SUSP1, that is highly expressed in reproductive organs.
A full-length cDNA encoding a SUMO-1-specific protease, named SUSP1, was identified and cloned for the first time from the human brain. Nucleotide sequence analysis of the cDNA containing an open reading frame of 3336 base pairs revealed that the protease consists of 1112 amino acids with a calculated molecular mass of 126,116 Da. Like yeast Ulp1, SUSP1 is a cysteine protease containing the well conserved His/Asp/Cys catalytic triad. SUSP1 expressed in Escherichia coli cells efficiently released SUMO-1 from SUMO-1. beta-galactosidase fusion but not from other ubiquitin-like protein fusions, including Smt3.beta-galactosidase, suggesting its role in the generation of matured SUMO-1 specifically from its precursors. Interestingly, reproductive organs, such as testis, ovary, and prostate, contained much higher amounts of SUSP1 mRNA than colon and peripheral blood leukocyte, whereas other tissues, such as heart and spleen, had little or none. In addition, confocal microscopy using green fluorescent protein.SUSP1 fusion showed that SUSP1 is exclusively localized to the cytoplasm of NIH3T3 and HeLa cells. These results suggest that SUSP1 may play a role in the regulation of SUMO-1-mediated cellular processes particularly related to reproduction.
Amino Acid Sequence↗
Suppression of IL-2 and IL-4 gene expression by nodularin through the reduced NF-AT binding activity.
Nodularin is a cyclic peptide produced by cyanobacteria. In the present study, the inhibitory effect of nodularin on T lymphocyte functions was demonstrated. Direct addition of nodularin to B6C3F1 mouse splenocyte cultures produced a concentration-dependent inhibition of the lymphoproliferative response to concanavalin A stimulation. Nodularin inhibited PMA plus ionomycin (Io)-induced IL-2 mRNA expression in murine splenocytes and thymocytes as determined by quantitative/competitive RT-PCR. To further characterize the mechanism for the transcriptional regulation of IL-2, the binding activity of transcription factors, NF-AT, AP-1, NF-kappaB, and Oct, was evaluated by electrophoretic mobility shift assays in mouse splenocytes. Nodularin reduced the NF-AT binding activity in PMA/Io-induced splenocytes, but no significant effect was observed on AP-1, NF-kappaB, or Oct binding activity. Nodularin also inhibited IL-4 mRNA expression in PMA/Io-stimulated murine splenocytes. These results suggest that T lymphocyte is a possible cellular target of nodularin, and the inhibitory effect of nodularin on T-cell specific transcription factor NF-AT induces T-cell dysfunction, which leads to a diminution in IL-2 and IL-4 gene transcription.
Animals↗
Effect of extracellular cations on the chemotherapeutic efficacy of anticancer drugs.
Cancer development and the efficiency of chemotherapy relies on the patients calcium-related pathological status such as hyper- or hypocalcemia. In the present study, we investigated the effect of extracellular cations such as calcium and magnesium on the therapeutic efficacy of antitumor drugs. The analytic parameters used were cellular drug uptake/excretion and the chemosensitivity of the human breast cancer cell lines, MCF7 and MCF7/ADR. Both calcium and magnesium ions decreased the membrane permeability of cancer cells, which was determined by cell size analysis. These divalent ions also lowered the drug uptake and the cytoplasmic levels of rhodamine 123 and adriamycin, suggesting that they might interfere with the diffusion of these drugs by modifying the physical properties of the cytoplasmic membrane. The acute cytotoxicity of adriamycin after a short period of incubation correlated with changes in its cytoplasmic level. Our results indicate that these extracellular cations might play an important role in the therapeutic activities of anticancer drugs in cancer patients. These results also provide insight a new aspect of chemotherapy, because they suggest that the therapeutic doses of anti-cancer drugs should be modified in cancer-bearing patients presenting with abnormal blood calcium levels.
Antineoplastic Agents↗
Differential activation of murine macrophages by angelan and LPS.
In our previous studies, we showed that angelan, a polysaccharide purified from Angelica gigas Nakai, is a potent LPS-mimetic in murine macrophages [Jeon, Y.J., Han, S.B., Ahn, K.S., Kim, H.M., 1999. Activation of NF-kB/Rel in angelan-stimulated macrophages. Immunopharmacology 43, 1-9]. Angelan stimulates murine macrophage to produce cytokines including iNOS and activate NF-kappaB/Rel. In the present study, we investigated the role of CD14 and complement receptor type 3 (CR3) in mediating NO production and NF-kappaB/Rel activation induced by angelan and LPS. Three major differences between angelan and LPS were observed. First, angelan does not require serum proteins for NO response and NF-kappaB/Rel activation, while the activation by LPS requires serum proteins. Second, blocking of either CD14 or CR3 decreased angelan-induced NO response, while LPS-mediated NO production was inhibited by anti-CD14 mAb only. Third, angelan induced strong NF-kappaB/Rel and slight AP-1 DNA binding, whereas LPS potently activated both NF-kappaB/Rel and AP-1. Both angelan and LPS degraded IkappaB proteins and subsequently induced the mobilization of NF-kappaB/Rel proteins (p65, c-rel and p50) into nucleus. This suggests that macrophages display a common signaling machinery leading to the NF-kappaB/Rel activation in response to different stimulants. In conclusion, angelan and LPS use the membrane receptor CD14 and CR3 differentially for signaling NF-kappaB/Rel activation and NO production.
Acute-Phase Proteins↗
Paclitaxel causes mouse splenic lymphocytes to a state hyporesponsive to lipopolysaccharide stimulation.
Multiple immune system actions have been ascribed to paclitaxel (taxol), a novel anticancer drug, including the capacity to induce macrophage antitumor cytotoxic molecule production. In the present studies, we demonstrated that paclitaxel produced a selective inhibition of lipopolysaccharide (LPS)-induced B cell proliferation. Similarly, in vitro polyclonal antibody-forming cell responses also were found to be inhibited by paclitaxel. Conversely, paclitaxel exhibited no inhibitory effects on concanavalin A (Con A)-induced T cell proliferation. To study the pathway leading to paclitaxel-induced immunosuppression, we analyzed Raf-1/ERK and JNK/p38 MAPK pathways, both of which have been reported to be involved in LPS signaling. Our results indicate that taxol treatment inhibits Raf-1 kinase activation while having no effect on ERK activation suggesting that ERK activation is distinct from upstream Raf-1 kinase in taxol-induced immunomodulation. Furthermore, paclitaxel pretreatment caused down-regulation of stress-activated MAPKs, JNK and p38 MAPK in lipopolysaccharide (LPS)-stimulated mouse splenic lymphocytes, demonstrating that spleen cells are induced to a state hyporesponsive to LPS stimulation by pre-exposing them to paclitaxel. Taken together, these results suggest that down-regulation of JNK/p38 MAP kinase may contribute to paclitaxel-induced immunosuppression in mouse splenic lymphocytes.
Animals↗
Radicicol suppresses expression of inducible nitric-oxide synthase by blocking p38 kinase and nuclear factor-kappaB/Rel in lipopolysaccharide-stimulated macrophages.
We show that radicicol, a fungal antibiotic, produces a marked inhibition of p38 kinase, nuclear factor-kappaB/Rel (NF-kappaB/Rel), and inducible nitric-oxide synthase (iNOS) transcription by the macrophage line RAW 264.7 in response to lipopolysaccharide (LPS). Treatment of RAW 264.7 with radicicol inhibited LPS-stimulated p38 kinase phosphorylation in a dose-related manner. iNOS transcription, which is regulated in part by the NF-kappaB/Rel family of transcription factors, has been shown to be under the control of the p38 kinase signaling cascade. Our data also show that the p38 kinase pathway is specifically involved in LPS-induced NF-kappaB/Rel activation and iNOS expression because NF-kappaB/Rel DNA binding and iNOS mRNA production in the presence of a specific inhibitor of p38 kinase, SB203580, were dramatically diminished. In contrast, PD98059, a specific inhibitor of mitogen-activated protein kinase/extracellular signal-regulated protein kinase kinase 1 had no effect on NF-kappaB/Rel activation and iNOS expression. LPS-induced loss of inhibitory proteins IkappaB-alpha and IkappaB-beta and translocation of p65, c-Rel, and p50 was inhibited by radicicol. Collectively, this series of experiments indicates that radicicol inhibits iNOS gene expression by blocking p38 kinase signaling. Due to the critical role that NO release plays in mediating inflammatory responses, the inhibitory effects of radicicol on iNOS suggest that this potent antifungal agent may represent a useful anti-inflammatory agent.
Animals↗