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M Shitashige

Publications and source records attributed to M Shitashige.

18 recordsLinked to original sources
PubMed2001

Downregulation of lipopolysaccharide-induced intercellular adhesion molecule-1 expression via EP2/EP4 receptors by prostaglandin E2 in human fibroblasts.

In the present study, the effect of prostaglandin E2 (PGE2) on intercellular adhesion molecule-1 (ICAM-1) expression in human gingival fibroblasts (HGF) stimulated with lipopolysaccharides (LPS) was investigated. LPS were isolated from periodontopathic bacteria, Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) and Porphyromonas gingivalis (P. gingivalis), by the phenol-water method and Escherichia coli (E. coli) LPS was used as a control. PGE2 significantly inhibited A. actinomycetemcomitans-, P. gingivalis- and E. coli-LPS-induced ICAM-1 expression. Next, of four PGE2 receptor subtypes (EP1, EP2, EP3 and EP4), we examined which subtype(s) was involved in inhibition of LPS-elicited ICAM-1 expression by PGE2. Eleven-deoxy-PGE1, a selective EP2/EP4 agonist, and butaprost, a selective EP2 agonist, attenuated A. actinomycetemcomitans-, P. gingivalis- and E. coli-LPS-elicited ICAM-1 expression, although butaprost was less potent than PGE2 and 11-deoxy-PGE1. Sulprostone, an EP1/EP3 agonist, and ONO-AP-324, an EP3 agonist, was inert to the LPS-elicited ICAM-1 expression. Furthermore, dibutyryl cAMP, a cAMP analogue, and forskolin, an adenylate cyclase activator, downregulated A. actinomycetemcomitans-, P. gingivalis- and E. coli-LPS-elicited ICAM-1 expression in HGF. Our data suggest that PGE2 downregulates A. actinomycetemcomitans- and P. gingivalis-LPS-induced ICAM-1 expression in HGF, via EP2/EP4 receptors by cAMP-dependent signaling pathways. The cAMP-elevating agents such as EP2/EP4 receptor activators may serve to control inflammatory and immune responses in periodontal disease.

Aggregatibacter actinomycetemcomitans↗
PubMed2001

Involvement of cyclooxygenase-2 in serum-induced prostaglandin production by human oral gingival epithelial cells.

The purpose of the present study was to investigate the involvement of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) in prostaglandin (PG) production by human oral gingival epithelial (OGE) cells stimulated with proinflammatory cytokines including interleukin(IL)-1alpha, IL-1alpha and tumor necrosis factor alpha (TNFalpha), and serum. Fetal bovine serum (FBS)-stimulated OGE cells produced significant levels of PGE2, whereas IL-1alpha, IL-1beta and TNFalpha could not induce significant PGE2 production. FBS induced PGE2 production in a dose- and time-dependent manner. NS-398, a selective COX-2 inhibitor, inhibited PGE2 production by FBS-stimulated cells as completely as indomethacin, a non-selective COX-1/COX-2 inhibitor. Expression of COX-2 protein in FBS-stimulated cells was increased, compared with that in unstimulated cells, whereas COX-1 protein expression was similar both in unstimulated and in FBS-stimulated cells. COX-2 mRNA was detected in FBS-stimulated cells, but not in unstimulated cells. We suggest that COX-2 is responsible for PG production by human OGE cells stimulated with serum and that OGE cells may be involved in PG production in periodontal lesions. Selective COX-2 inhibitors, which have the advantage of reduced gastric toxicity, may provide a useful approach to treatment of periodontal disease.

Analysis of Variance↗
PubMed2001

Dissociation of Bax from a Bcl-2/Bax heterodimer triggered by phosphorylation of serine 70 of Bcl-2.

Serine 70 in the loop region of Bcl-2 is specifically phosphorylated by paclitaxel-treatment in tumor cells and BHK cells expressing Bcl-2. The phosphorylation of serine 70 of Bcl-2 (pS70-Bcl-2) peaks 24 to 48 h after paclitaxel treatment and accelerates apoptosis. Phosphorylation is effectively inhibited in the presence of actinomycin D or cycloheximide, which restore cell viability to the same level as control cells not expressing Bcl-2. These results indicate that paclitaxel-induced kinase(s) and/or its activator(s) are synthesized de novo and play an important role in paclitaxel-induced apoptosis by phosphorylating Bcl-2. In binding assays using the phosphorylation-specific antibody against pS70-Bcl-2, the induction of serine 70 phosphorylation 70 results in a loss of the binding ability of Bcl-2 to Bax, a pro-apoptotic partner, and induces subsequent cell death. When the pS70-Bcl-2 antibody was added to human breast cancer tissue, serine 70 phosphorylation was also detected, even prior to treatment with anticancer agents. Further study of breast cancers revealed 83% of tumors with high pS70-Bcl-2 expression responded to paclitaxel or docetaxel treatment, whereas 57% of those with low expression not respond. These findings suggest that pS70-Bcl-2 might be a predictive factor for prognosis and sensitivity to paclitaxel treatment for breast cancer.

Animals↗
PubMed2000

Prostaglandins E2 and I2 downregulate tumor necrosis factor alpha-induced intercellular adhesion molecule-1 expression in human oral gingival epithelial cells.

In the present study, we examined whether prostaglandin (PG) E2 and PGI2 regulated intercellular adhesion molecule-1 (ICAM-1) expression in human oral gingival epithelial cells stimulated with tumor necrosis factor alpha (TNF alpha). TNF alpha potently induced ICAM-1 expression in a dose- and time-dependent fashion. PGE2 and carbacyclin (a stable analogue of PGI2) significantly decreased ICAM-1 expression in TNF alpha-challenged oral gingival epithelial cells. Next, of the four subtypes of PGE2 receptors (EP1, EP2, EP3 and EP4), we examined which subtype(s) mediated inhibition of TNF alpha-induced ICAM-1 expression by PGE2. 11-deoxy-PGE2, an EP2/EP4 agonist, significantly suppressed TNF alpha-induced ICAM-1 expression, whereas butaprost, an EP2 agonist, sulprostone, an EP1/EP3 agonist, and ONO-AP-324, an EP3 agonist, caused no effect on it. By reverse transcriptase-polymerase chain reaction, expression of EP4 mRNA was detected in oral gingival epithelial cells. Dibutyryl cAMP, a cAMP analogue, and forskolin, a direct activator of adenylate cyclase, significantly inhibited TNF alpha-induced ICAM-1 expression in oral gingival epithelial cells. From these results, we suggest that PGE2 and PGI2 inhibit TNF alpha-elicited ICAM-1 expression by cAMP-dependent pathways via EP4 receptors and IP receptors, respectively.

Acetates↗
PubMed2000

Cyclooxygenase-2-dependent prostaglandin E2 down-regulates intercellular adhesion molecule-1 expression via EP2/EP4 receptors in interleukin-1beta-stimulated human gingival fibroblasts.

Prostaglandin E2 (PGE2), which exerts its actions via EP receptors (EP1, EP2, EP3, and EP4), is a bioactive metabolite of arachidonic acid produced by cyclooxygenase (COX)-1 and/or COX-2. We have previously demonstrated that PGE2 down-regulates intercellular adhesion molecule-1 (ICAM-1) expression in interleukin-1beta (IL-1beta)-stimulated human gingival fibroblasts (HGF). In the present study, we investigated which COX was involved in down-regulation of ICAM-1 expression by PGE2 in IL-1beta-stimulated HGF and which subtypes of EP receptors modulated the ICAM-1 expression. NS-398, a specific COX-2 inhibitor, completely inhibited PGE2 production by IL-1beta-stimulated HGF, as did indomethacin, a COX-1/COX-2 inhibitor. Northern blot analysis and immunocytochemical staining showed that mRNA and protein of COX-2 were expressed in IL-1beta-challenged HGF, but not in unstimulated HGF, and that the expression of mRNA and protein of COX-1 was similar both in unstimulated and in stimulated cells. NS-398 and indomethacin enhanced ICAM-1 expression in IL-1beta-challenged HGF. EP1, EP2, and EP4 receptor mRNA was expressed in HGF according to reverse-transcription/polymerase chain-reaction. PGE2, 11-deoxy-PGE1 (a selective EP2/EP4 agonist), and Butaprost (a selective EP2 agonist) attenuated IL-1beta-elicited ICAM-1 expression, although Butaprost was less potent than PGE2 and 11-deoxy-PGE1. AH-23848B, an EP4 antagonist, antagonized the inhibitory effect of IL-1beta-elicited ICAM-1 expression by PGE2. Sulprostone, an EP1/EP3 agonist, had no effect on IL-1beta-elicited ICAM-1 expression. Analysis of these data suggests that COX-2-derived PGE2 down-regulates ICAM-1 expression via EP2/EP4 receptors in IL-1beta-stimulated HGF.

Analysis of Variance↗
PubMed2000

Cyclooxygenase-2-dependent prostaglandin production by peripheral blood monocytes stimulated with lipopolysaccharides isolated from periodontopathogenic bacteria.

BACKGROUND: Prostaglandin E2 (PGE2) plays important roles in the pathogenesis of periodontal disease. Recent studies have revealed the existence of 2 isozymes of cyclooxygenase (COX), called COX-1 and COX-2. The purpose of the present study was to investigate the contribution of COX-1 and COX-2 to PGE2 production by human peripheral blood monocytes that are stimulated with lipopolysaccharides (LPS) from periodontopathogenic bacteria. METHODS: LPS were isolated from Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) and Porphyromonas gingivalis (P. gingivalis) by the phenol-water method. Peripheral blood monocytes were stimulated with LPS for the indicated periods, and the levels of PGE2 or interleukin (IL)-1 beta in the culture media were measured by enzyme-linked immunosorbent assay. Expression of COX-1 and -2 proteins was studied by immunocytochemical staining, and COX-2 mRNA expression was examined by Northern blot analysis. RESULTS: Peripheral blood monocytes stimulated with A. actinomycetemcomitans- or P. gingivalis-LPS produced PGE2 in a time- and dose-dependent manner. Indomethacin, a non-selective COX-1/COX-2 inhibitor, and NS-398, a specific COX-2 inhibitor, completely inhibited PGE2 production. Immunocytochemical staining of COX-1 and COX-2 proteins showed that expression of COX-2 protein was increased in monocytes that were stimulated with A. actinomycetemcomitans- or P. gingivalis-LPS, compared with that in unstimulated monocytes, whereas expression of COX-1 protein was not altered. Northern blot analysis showed that monocytes stimulated with A. actinomycetemcomitans- or P. gingivalis-LPS expressed COX-2 mRNA, while COX-2 mRNA was not detectable in unstimulated cells. Treatment of A. actinomycetemcomitans-LPS-stimulated monocytes with NS-398 induced a significant increase of IL-1 beta production to the same extent as treatment with indomethacin. CONCLUSIONS: These results suggest that COX-2 is induced in monocytes stimulated with LPS derived from A. actinomycetemcomitans and P. gingivalis and that the COX-2 is primarily responsible for PGE2 production. COX-2 may be pivotal in PGE2 production in periodontal lesions and may be involved in inflammatory responses.

Aggregatibacter actinomycetemcomitans↗
PubMed1999

Interleukin-4 and interferon-gamma inhibit prostaglandin production by interleukin-1beta-stimulated human periodontal ligament fibroblasts.

The purpose of the present study was to investigate the involvement of cyclooxygease-1 (COX-1) and cyclooxygenase-2 (COX-2) in prostaglandin (PG) production by human periodontal ligament (PDL) fibroblasts stimulated with a proinflammatory cytokine, inerleukin-1beta (IL-1beta), and to examine the effect of interleukin-4 (IL-4), a Th2 cytokine, and interferon-gamma (IFN-gamma), a Th1 cytokine, on PG production by the cells. IL-1beta-stimulated PDL fibroblasts produced prostaglandin E2 (PGE2) in a time-dependent manner. Indomethacin, a non-selective COX-1/COX-2 inhibitor, and NS-398, a selective COX-2 inhibitor, completely inhibited PGE2 production by IL-1beta-stimulated cells. Northern blot analysis showed that COX-2 mRNA was detected in IL-1beta-stimulated PDL cells, although not detected in unstimulated cells, while expression of COX-1 mRNA was in the same extent in both the cells. Dexamethasone inhibited COX-2 mRNA expression, COX activity and PGE2 production in IL-1beta-stimulated cells. IL-4 and IFN-gamma suppressed PGE2 production by IL-1beta-stimulated PDL fibroblasts, but COX activity enhanced by IL-1beta treatment was significantly inhibited by IL-4, not by IFN-gamma. Northern blot analysis showed that IL-4 depressed COX-2 mRNA expression with no effect on COX-1 mRNA expression. On the other hand, IFN-gamma had no effect on expression of COX-1 and -2 mRNA. These data suggest that COX-2 is primarily responsible for PGE2 production by IL-1beta-stimulated human PDL fibroblasts and that IL-4 inhibited PGE2 production by IL-1beta-stimulated PDL fibroblasts through down-regulation of COX-2 expression, while IFN-gamma suppressed the PGE2 production with no effect on COX-2 expression.

Cells, Cultured↗
PubMed1999

Prostaglandin E2 downregulates interferon-gamma-induced intercellular adhesion molecule-1 expression via EP2 receptors in human gingival fibroblasts.

In the present study, the effect of prostaglandin E2 (PGE2) on intercellular adhesion molecule-1 (ICAM-1) expression in interferon-gamma (IFN-gamma)-stimulated human gingival fibroblasts (HGF) was investigated. Addition of PGE2 to HGF inhibited ICAM-1 expression elicited by IFN-gamma. As PGE2 elevated intercellular cyclic AMP (cAMP) levels in HGF in a dose-dependent fashion, the effect of dibutyryl cAMP and 8-bromo-cAMP, cAMP analogues, on IFN-gamma-elicited ICAM-1 expression was examined. Both the agents downregulated ICAM-1 expression in IFN-gamma-stimulated HGF. Next, we examined which subtype(s) of the four PGE2 receptor subtypes (EP1, EP2, EP3 and EP4) modulated the ICAM-1 expression elicited by IFN-gamma, using subtype-specific agonists or antagonists. An EP2/EP4 agonist, 11-deoxy-PGE1, attenuated IFN-gamma-elicited ICAM-1 expression in a concentration-dependent manner. A specific EP4 antagonist, AH-23848B, showed no effect on inhibition of IFN-gamma-elicited ICAM-1 expression by PGE2 and 11-deoxy-PGE1. Butaprost, an EP2-selective agonist, mimicked inhibition of IFN-gamma-elicited ICAM-1 expression by 11-deoxy-PGE1. An EP3 agonist, ONO-AP-324, was inert with respect to IFN-gamma-elicited ICAM-1 expression. Sulprostone, an EP1/EP3 agonist, showed stimulatory effect on ICAM-1 expression elicited by IFN-gamma. From these results, we suggest that PGE2 downregulates IFN-gamma-induced ICAM-1 expression in HGF, primarily via EP2 receptors by cAMP-dependent signaling pathways.

Antineoplastic Agents↗
PubMed1999

Existence of endogenous inhibitor(s) of prostaglandin endoperoxide H synthase activities in murine NIH3T3 fibroblasts.

The production of prostaglandins (PGs) is regulated by several processes, i.e. arachidonic acid release, prostaglandin endoperoxide H synthase (PGHS) activity and its induction. In the present study, we investigated the possibility that inhibitor(s) of PGHS activities exist endogenously and regulate PG production. NIH3T3 cell, a cloned murine fibroblast, expressed PGHS-1 under unstimulated conditions and induced PGHS-2 protein under stimulation by serum. When PGHS activity was measured by individual intact cell assay, there was heterogeneity in PGHS activities in individual cells after serum treatment. We isolated and cultured many cloned cells which showed different PGHS activities after serum treatment. In these cells, however, there was no difference in the expression of PGHS-1 and PGHS-2 mRNA. The protein level of PGHS-1 and PGHS-2 was also not different in these cloned cells. The lysate obtained from the cells that showed lower PGHS activity after the serum treatment suppressed PGHS-1 activities of sheep seminal vesicle and PGHS-2 activity in purified ovine placenta. The suppression by this lysate was stronger than that by the lysate obtained from the cells which showed higher PGHS activity after the serum treatment. The production of the inhibitor(s) was up-regulated by the serum treatment and abolished by actinomycin D and cycloheximide during the serum treatment. From these results, we suggest that there is some endogenous inhibitor(s) of PGHS-1 and -2 activities in NIH3T3 cells, and that the inhibitor(s) is induced by serum.

3T3 Cells↗
PubMed1999

Prostaglandin E2 receptors of the EP2 and EP4 subtypes downregulate tumor necrosis factor alpha-induced intercellular adhesion molecule-1 expression in human gingival fibroblasts.

Prostaglandin E2 (PGE2) exerts its biological actions via EP receptors, which are divided into 4 subtypes of EP1, EP2, EP3 and EP4. In the present study, we investigated whether PGE2 regulated intercellular adhesion molecule-1 (ICAM-1) expression in human gingival fibroblasts (HGF) stimulated with tumor necrosis factor-alpha (TNF alpha) and if so, which subtype(s) of PGE2 receptors was involved. Exogenous addition of PGE2 to HGF inhibited ICAM-1 expression elicited by TNF alpha in a concentration-dependent manner. Treatment of HGF with indomethacin, a cyclo-oxygenase inhibitor, had no effect on TNF alpha-elicited ICAM-1 expression, although indomethacin completely inhibited PGE2 production enhanced by TNF alpha. Next, we examined which subtype(s) of the 4 EP receptors modulated the ICAM-1 expression elicited by TNF alpha, using subtype-specific agonists and antagonists. 11-deoxy-PGE1, a selective EP2/EP4 agonist, inhibited TNF alpha-elicited ICAM-1 expression as potently as PGE2, while butaprost, a selective EP2 agonist, was somewhat less effective than PGE2. AH23848B, an EP4 antagonist, antagonized the inhibitory effect of TNF alpha-elicited ICAM-1 expression by PGE2. Sulprostone, an EP1/EP3 agonist, and ONO-AP-324, an EP3 agonist, were inert to TNF alpha-elicited ICAM-1 expression. As EP2 and EP4 receptors are linked to elevation of intracellular cyclic AMP (cAMP), the effect of dibutyryl cAMP and 8-bromo-cAMP, cAMP analogs, on TNF alpha-elicited ICAM-1 expression was examined. Both the agents downregulated ICAM-1 expression in TNF alpha-stimulated HGF. From these data, we suggest that PGE2 downregulates TNF alpha-induced ICAM-1 expression in HGF, via EP2 and EP4 receptors by cAMP-dependent signaling pathways, which may result in control of inflammatory and immunological responses in periodontal disease.

Analysis of Variance↗
PubMed1999

Involvement of cyclooxygenase-2 in interleukin-1alpha-induced prostaglandin production by human periodontal ligament cells.

BACKGROUND: Human periodontal ligament (PDL) cells produce prostaglandin (PG) E2 in response to proinflammatory cytokines. However, the mechanism of PGE2 production is not well understood. The purpose of the present study was to investigate the involvement of cyclooxygenase (COX)-1 and COX-2 in PGE2 production by PDL cells stimulated with a proinflammatory cytokine, interleukin-1alpha (IL-1alpha), and to examine the regulation of PGE2 production by cell-cell interaction of human gingival keratinocytes and PDL cells. METHODS: The levels of PGE2 in the culture media of PDL cells stimulated with IL-1alpha or culture media of human gingival keratinocytes were determined by an enzyme-linked immunosorbent assay. Expression of COX-1 and -2 mRNA and protein was studied by Northern blot analysis and Western blot analysis, respectively. RESULTS: IL-1alpha-stimulated PDL cells produced PGE2 in a time-dependent manner. Indomethacin, a non-selective COX-1/COX-2 inhibitor, and NS-398, a selective COX-2 inhibitor, completely inhibited PGE2 production by the IL-1alpha-stimulated cells. COX-2 mRNA was detected after IL-1alpha stimulation, although it was not detected in unstimulated cells. There was no difference in expression of COX-1 mRNA between unstimulated cells and IL-la-stimulated cells. Expression of COX-2 protein in IL-1alpha-stimulated cells was increased, compared with that in unstimulated cells, whereas COX-1 protein expression was almost the same in both the cells. Treatment of IL-1alpha-stimulated PDL cells with dexamethasone, known to inhibit COX-2 expression, prevented PGE2 production and COX-2 mRNA expression. Addition of the culture media of human gingival keratinocytes to PDL cells increased PGE2 production. The PGE2 production was depressed by treatment of the cells with IL-1 receptor antagonist and anti- IL-1alpha antibody, not with anti-IL-1beta antibody. The PGE2 production was also inhibited by treatment with NS-398 and dexamethasone. CONCLUSIONS: We suggest that PDL cells stimulated with IL-1alpha produce PGE2 through de novo synthesis of COX-2 and that the cell interaction of gingival keratinocytes and PDL cells controls COX-2 expression and PGE2 production via IL-1alpha or 1alpha IL-la-like factor(s). Selective COX-2 inhibitors, which have the advantage of reduced gastric toxicity, may provide a useful approach to treatment of periodontal disease.

Analysis of Variance↗
PubMed1998

Different substrate utilization between prostaglandin endoperoxide H synthase-1 and -2 in NIH3T3 fibroblasts.

Recent studies suggested that prostaglandin endoperoxide H synthase- and prostaglandin endoperoxide H synthase-2 (PGHS-1 and PGHS-2) utilize different pools of arachidonic acid for synthesizing prostanoids. Using cultured murine NIH3T3 fibroblasts, we investigated the mechanism for the different utilization of arachidonic acid between PGHS-1 and -2. Histofluorescence staining for PGHS activity in intact cells demonstrated that quiescent 3T3 cells expressed only PGHS-1 activity and serum-activated 3T3 cells pretreated with aspirin expressed only PGHS-2 activity. Endogenous arachidonic acid released by calcium ionophore A23187 was not converted by PGHS-1 but exclusively converted by PGHS-2. In the cell free system, the kinetics of PGHS-1 were not so much different from those of PGHS-2. However, in intact cells, arachidonic acid at concentrations lower than 2.5 microM was converted by PGHS-2 alone but not by PGHS-1. Our findings indicated that this small amount of arachidonic acid as released by some stimuli is converted exclusively by PGHS-2. Furthermore, treating the PGHS-2-expressing cells with sodium selenite or ebselen, reducing agents of intracellular peroxides, only decreased PGHS-2 activity. We speculate that only PGHS-2 has been activated by intracellular peroxides and subsequently, it can convert the arachidonic acid released endogenously.

3T3 Cells↗
PubMed1997

Cationic amino acid transporter-2 mRNA induction by tumor necrosis factor-alpha in vascular endothelial cells.

Nitric oxide (NO) synthesis may be coupled to the activity of the cellular L-arginine transporter, namely the cationic amino acid transporter. The present study examined tumor necrosis factor (TNF)-alpha-induced alterations in the gene expression of the cationic amino acid transporter (CAT) and NO production in human umbilical vein endothelial cells. In quiescent endothelial cells, CAT-1 mRNA expression, determined by reverse transcription-polymerase chain reaction, was dominant to that of CAT-2. TNF-alpha (10 ng/ml for 1-24 h) induced a time-dependent increase in CAT-2 but not CAT-1 expression. Moreover, TNF-alpha (1-30 ng/ml) treatment for 6 h induced a concentration-dependent increase in CAT-2 mRNA expression. The upregulation of CAT-2 expression by TNF-alpha was associated with enhanced nitrite accumulation in the culture medium (70% increase compared with vehicle-treated cells at 24 h). Thus, induction of the cationic amino acid transporter may constitute one mechanism for the TNF-alpha-induced NO production in human umbilical vein endothelial cells.

Amino Acid Transport Systems, Basic↗
PubMed1996

Prostaglandin production via induction of cyclooxygenase-2 by human gingival fibroblasts stimulated with lipopolysaccharides.

The purpose of the present study was to investigate the involvement of cyclooxygenase-1(COX-1) and cyclooxygenase-2 (COX-2) in PGE2 production by human gingival fibroblasts stimulated with lipopolysaccharides (LPS) from periodondopathogenic bacteria. LPS were isolated from Porphyromonas gingivalis (P. gingivalis), Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) and Eschericia coli (E coli) by the phenol-water procedure. The three LPS preparations produced PCE2 up to 48 h in a time-dependent manner in human gingival fibroblasts. P. gingivalis-LPS was the most potent stimulator of PGE2 production and, to a lesser extent, A actinomycetemcomitans- and E coli-LPS. Treatment of the cells with indomethacin, a non selective COX-1/COX-2 inhibitor and NS-398, a selective COX-2 inhibitor, completely depressed PGE2 production. Treatment of dexamethasone, known to inhibit COX-2 expression, also significantly prevented PGE2 production. Immunohistochemical staining of COX-2 protein demonstrated that expression of COX-2 protein was increased at 24 h after P gingivalis-LPS stimulation, while expression of COX-1 protein was not affected by P. gingivalis-LPS. In order to investigate the regulation of PGE2 production. P. gingivalis-LPS-stimulated cells were treated with herbimycin A and genistein, both inhibitors of tyrosine kinases. Both the inhibitors significantly inhibited PGE2 production. Herbimycin A treatment depressed expression of COX-2 protein. These data suggest that human gingival fibroblasts stimulated with LPS from periodontopathogenic bacteria mainly produce PGE2 not by COX-1, but by COX-2, induction of which may be regulated by tyrosine kinase and that the produced PGE2 may be involved in the pathogenesis of periodontal diseases.

Aggregatibacter actinomycetemcomitans↗
PubMed1995

Reduction of ubiquinone in membrane lipids by rat liver cytosol and its involvement in the cellular defence system against lipid peroxidation.

Rat liver homogenates reduced ubiquinone (UQ)-10 to ubiquinol (UQH2)-10 in the presence of NADPH rather than NADH. This NADPH-dependent UQ reductase (NADPH-UQ reductase) activity that was not inhibited by antimycin A and rotenone, was located mainly in the cytosol fraction and its activity accounted for 68% of that of the homogenates. Furthermore, the NADPH-UQ reductase from rat liver cytosol efficiently reduced both UQ-10 incorporated into egg yolk lecithin liposomes, and native UQ-9 residing in rat microsomes, to the respective UQH2 form in the presence of NADPH. The gross redox ratios of UQH2-9/(UQ-9 + UQH2-9) in individual tissues of rat correlated positively with the log of their respective cytosolic NADPH-UQ reductase activities, while the redox ratios in every intracellular fraction from liver were at about the same level, irrespective of NADPH-UQ reductase activities in the respective fractions. The combined addition of rat liver cytosol and NADPH inhibited to a great extent 2,2'-azobis(2,4-dimethyl-valeronitrile)-induced lipid peroxidation of UQ-10-fortified lecithin liposomes and completely inhibited such peroxidation in the liposomes in which UQH2-10 replaced UQ-10. The NADPH-UQ reductase activity was clearly separated from DT-diaphorase (EC 1.6.99.2) activity by means of Cibacron Blue-immobilized Bio-Gel A-5m chromatography. In conclusion, the NADPH-UQ reductase in cytosol, which is a novel enzyme to our knowledge, was presumed to be responsible for maintaining the steady-state redox levels of intracellular UQ and thereby to act as an endogenous antioxidant in protecting intracellular membranes from lipid peroxidation that is inevitably induced in aerobic metabolism.

Animals↗
PubMed1995

Oxidative modification of low density lipoprotein by diesel exhaust particles.

Oxidized low density lipoprotein (LDL) has a variety of hazardous influences on biological systems. Oxidative modification of LDL by diesel exhaust particles (DEP) was studied in vitro to assess its in vivo health effects. DEP suspensions (1, 10 and 100 micrograms/ml) were incubated for 1 h with LDL (1 mg protein/ml) at 37 degrees C. Conjugated diene formation and negative charges in LDL were increased by DEP-treatment in a dose-dependent manner. When native LDL and DEP-treated LDL (DEP-LDL) were incubated for 18 h with macrophage, J774A.1 cell at 37 degrees C, significantly more DEP-LDL was taken up into cells than native LDL. Accumulation of cholesterol ester in cells incubated with DEP-LDL was 4 to 8 times higher than that with native LDL while there was no significant difference between them in free cholesterol content. Incubation (18 h) of J774A.1 with DEP-LDL caused an increase in leakage of lactate dehydrogenase from cells in a DEP-concentration dependent manner, but the incubation with native LDL or DEP per se did not increase the leakage except at the highest concentration of DEP. These results suggest that DEP oxidatively modified LDL giving it cytotoxic, inflammatory and atherogenic properties characteristic of so-called oxidized LDL; these initial modifications of LDL may be one of the underlying mechanisms for diseases associated with DEP.

Animals↗
PubMed1992

A novel ubiquinone reductase activity in rat cytosol.

Ubiquinone (UQ) reductase activity which reduces UQ to ubiquinol (UQH2) in rat tissues was roughly proportional to the UQH2/total UQ ratio in respective tissues. The highest activity was found in the liver, showing the highest UQH2/total UQ ratio. A greater part of liver UQ reductase activity was located in the cytosol. Within a week, the liver UQ reductase activity decreased by 80% even at -20 degrees C. The DT-diaphorase activity was stable. UQ reductase required NADPH as the hydrogen donor and was not inhibited by a less than 1 microM concentration of dicoumarol. There was no stimulation of UQ reductase in the presence of bovine serum albumin nor in Triton X-100. Yet, both stimulated DT-diaphorase. As a result, UQ reductase appeared to be a novel NADPH-UQ oxidoreductase and responsible for the UQ redox state in liver.

Animals↗