Expression of cyclooxygenase 1 and 2 by human gastric endothelial cells.
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BACKGROUND: Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase (COX) and cause gastric ulceration. NSAIDs also impair granulation tissue angiogenesis and healing of established gastric ulcers in humans. The mechanism whereby NSAIDs inhibit granulation tissue angiogenesis is unknown but may involve inhibition of either or both COX isoforms (COX-1 and COX-2). AIMS: To investigate COX expression by human gastric endothelial (HuGE) cells during angiogenesis in vitro. METHODS: COX-1 and COX-2 expression by HuGE cells was investigated by western blot analysis, indirect immunofluorescence, reverse transcriptase polymerase chain reaction, and measurement of prostaglandin E(2) synthesis. Plating onto basement membrane matrix and stimulation by phorbol ester were used as in vitro models of angiogenesis. RESULTS: Under normal culture conditions (30% serum), HuGE cells expressed COX-1 and low levels of COX-2. COX-2 expression was induced in HuGE cells in both angiogenesis models. Prostaglandin E(2) production and tubular structure formation by HuGE cells on basement membrane matrix was significantly inhibited by a selective COX-2 inhibitor (NS-398). CONCLUSION: Angiogenesis by HuGE cells in vitro was associated with induction of functional COX-2 expression. A selective COX-2 inhibitor significantly decreased HuGE cell angiogenesis on basement membrane matrix. Extrapolation of these data to human gastric ulcer angiogenesis in vivo suggests that selective COX-2 inhibitors could delay gastric ulcer healing to the same extent as traditional NSAIDs which are non-selective COX inhibitors. (+info)
Multiple acquisitions via horizontal transfer of a group I intron in the mitochondrial cox1 gene during evolution of the Araceae family.
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A group I intron has recently been shown to have invaded mitochondrial cox1 genes by horizontal transfer many times during the broad course of angiosperm evolution. To investigate the frequency of acquisition of this intron within a more closely related group of plants, we determined its distribution and inferred its evolutionary history among 14 genera of the monocot family Araceae. Southern blot hybridizations showed that 6 of the 14 genera contain this intron in their cox1 genes. Nucleotide sequencing showed that these six introns are highly similar in sequence (97.7%-99.4% identity) and identical in length (966 nt). Phylogenetic evidence from parsimony reconstructions of intron distribution and phylogenetic analyses of intron sequences is consistent with a largely vertical history of intron transmission in the family; the simplest scenarios posit but one intron gain and two losses. Despite this, however, striking differences in lengths of exonic co-conversion tracts, coupled with the absence of co-conversion in intron-lacking taxa, indicate that the six intron-containing Araceae probably acquired their introns by at least three and quite possibly five separate horizontal transfers. The highly similar nature of these independently acquired introns implies a closely related set of donor organisms. (+info)
Potentiation of excitotoxicity in transgenic mice overexpressing neuronal cyclooxygenase-2.
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In this study we describe the generation of a transgenic mouse model with neuronal overexpression of the human cyclooxygenase-2, h(COX)-2, to explore its role in excitotoxicity. We report that overexpression of neuronal hCOX-2 potentiates the intensity and lethality of kainic acid excitotoxicity in coincidence with potentiation of expression of the immediate early genes c-fos and zif-268. In vitro studies extended the in vivo findings and revealed that glutamate excitotoxicity is potentiated in primary cortico-hippocampal neurons derived from hCOX-2 transgenic mice, possibly through potentiation of mitochondrial impairment. This study is the first to demonstrate a cause-effect relationship between neuronal COX-2 expression and excitotoxicity. This model system will allow the systematic examination of the role of COX-2 in mechanisms of neurodegeneration that involve excitatory amino acid pathways. (+info)
Inhibition of the renin-angiotensin system upregulates cyclooxygenase-2 expression in the macula densa.
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The expression of cyclooxygenase 2 (COX-2) in the late thick ascending limb, including the macula densa, is found to be upregulated in an activated renin-angiotensin system. How this upregulation is managed is not yet known. We therefore considered the possibility that the stimulation of COX-2 expression is triggered by the activation of the renin-angiotensin system. For this purpose, we treated male Sprague-Dawley rats with the angiotensin I-converting enzyme inhibitor ramipril (10 mg/kg per day), the angiotensin II type 1 (AT(1)) receptor blocker losartan (30 mg/kg per day), and the angiotensin II type 2 (AT(2)) receptor blocker PD123319 (6 mg/kg per day) for 4 days. We determined the expression of COX-2 mRNA and protein in the renal cortex. We found that ramipril and the AT(1) receptor blocker losartan increased COX-2 mRNA and COX-2 immunoreactivity in the macula densa approximately 4-fold, whereas the AT(2) blocker PD123319 showed no effect. A low-salt diet (0.02% wt/wt) stimulated COX-2 expression in the kidney cortex <2-fold. The combination of a low-salt diet with ramipril led to a further increase of COX-2 mRNA and COX-2 immunoreactivity compared with low salt or ramipril alone. These data indicate that endogenous angiotensin II apparently inhibits COX-2 expression in the macula densa via AT(1) receptors and can therefore not account for the stimulation of COX-2 expression associated with an activated renin-angiotensin system. Because macula densa-derived prostaglandins are considered stimulators of renin secretion and renin synthesis, inhibition of macula densa COX-2 by angiotensin II could form a novel indirect negative feedback control of the renin system. (+info)
Interpreting the clinical significance of the differential inhibition of cyclooxygenase-1 and cyclooxygenase-2.
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The International Consensus Meeting on the Mode of Action of COX-2 Inhibition (ICMMAC) brought together 17 international experts in arthritis, gastroenterology and pharmacology on 5 6 December 1997. The meeting was convened to provide a definition of COX-2 specificity and to consider the clinical relevance of COX-2-specific agents. These compounds are a new class of drugs that specifically inhibit the enzyme COX-2 while having no effect on COX-1 across the whole therapeutic dose range. The objectives of the meeting were to review the currently available data regarding the roles and biology of COX-1 and COX-2, and to foster a consensus definition on COX-2 specificity. At the present time, no guidelines exist for the in vitro and in vivo assessment of COX specificity, and it was felt that consensus discussion might clarify some of these issues. The meeting also reviewed recent clinical data on COX-2-specific inhibitors. The following article reflects discussion at this meeting and provides a consensus definition of COX-2-specific inhibitors. (+info)
Regulation of cyclooxygenase-2 by interferon gamma and transforming growth factor alpha in normal human epidermal keratinocytes and squamous carcinoma cells. Role of mitogen-activated protein kinases.
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Treatment of normal human epidermal keratinocytes (NHEK) with interferon-gamma (IFN-gamma) causes a 9-fold increase in the level of cyclooxygenase-2 (COX-2) mRNA expression. Nuclear run-off assays indicate that this induction is at least partly due to increased transcription. Activation of the epidermal growth factor receptor (EGFR) signaling pathway due to the enhanced transforming growth factor alpha (TGFalpha) expression plays an important role in the induction of COX-2 by IFN-gamma. This is supported by the ability of TGFalpha to rapidly induce COX-2 and the inhibition of the IFN-gamma-mediated COX-2 mRNA induction by an EGFR antibody and EGFR-selective kinase inhibitors. Deletion and mutation analysis indicates the importance of the proximal cAMP-response element/ATF site in the transcriptional control of this gene by TGFalpha. The increase in COX-2 mRNA by TGFalpha requires activation of both the extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) pathways. Inhibition of p38 MAPK decreases the stability of COX-2 mRNA, while inhibition of MAPK/ERK kinase (MEK) does not. These results suggest that the p38 MAPK signaling pathway controls COX-2 at the level of mRNA stability, while the ERK signaling pathway regulates COX-2 at the level of transcription. In contrast to NHEK, IFN-gamma and TGFalpha are not very effective in inducing TGFalpha or COX-2 expression in several squamous carcinoma cell lines, indicating alterations in both IFN-gamma and TGFalpha response pathways. (+info)
Expression of cyclo-oxygenase in human endometrium during the implantation period.
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Prostaglandins (PG) are known to be involved in the process of human implantation. In several animal species treatment with prostaglandin synthesis inhibitors will prevent implantation. Cyclo-oxygenase (COX) is the rate-limiting enzyme in the synthesis of PGs and exists in two isoforms, COX-1 and COX-2. Defective implantation in COX-2-deficient mice has been demonstrated recently. In the present study, the expression of COX-1 and COX-2 was studied during the implantation period in healthy fertile women before and after treatment with the antiprogesterone, mifepristone. The study consisted of one control cycle and one treatment cycle. The subjects served as their own control. During the treatment cycle the subjects received 200 mg of mifepristone 2 days after the luteinizing hormone (LH) surge. Endometrial biopsies were obtained in the mid-luteal phase (LH + 6 to LH + 8) in both cycles. Using polyclonal antibodies against COX-1 and COX-2, immunostaining for COX-1 was found mainly in the glandular and luminal epithelium and for COX-2 mainly in the luminal epithelium and the perivascular cells. After treatment with mifepristone, expression of COX-1 in glandular epithelium and COX-2 in luminal epithelium significantly decreased whilst the immunostaining for COX-2 in the perivascular cells remained strong. This study shows the expression of both COX-1 and COX-2 during the implantation period and also indicates that treatment with mifepristone in early luteal phase impairs glandular epithelial function and endometrial receptivity. (+info)
Nitric oxide donors stimulate prostaglandin F(2alpha) and inhibit thromboxane B(2) production in the human cervix during the first trimester of pregnancy.
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Nitric oxide (NO) donors are capable of ripening the human cervix during pregnancy. The purpose of this study was to examine how NO donors may be involved in this process. Cervical biopsies were obtained from pregnant women randomized to receive isosorbide mononitrate (n = 10) or no treatment (n = 10) prior to suction termination. Enzyme-linked immunosorbent assays (ELISA) were performed on culture supernatant for interleukin (IL)-1, IL-6, IL-8, IL-10, IL-15, tumour necrosis factor-alpha, monocyte chemoattractant protein-1 and prostaglandin metabolites. Immunohistochemistry was performed to localize these cytokines, cyclooxygenase (COX)-1, COX-2 and prostaglandin dehydrogenase in cervical tissue and reverse transcription-polymerase chain reaction (RT-PCR) to identify COX-1 and COX-2 expression. Biopsies treated with the NO donor isosorbide mononitrate (IMN) produced significantly greater amounts of prostaglandin F(2alpha) in culture and lower amounts of thromboxane B(2) than controls (572.8 versus 34.9 pg/ml, P < 0.05; 53.3 pg/ml versus 530.9 pg/ml, P < 0.01 respectively). The release of other prostaglandins and of cytokines was not affected by treatment with NO. Inflammatory mediators were localized to cervical tissue and COX-1 and COX-2 expression was confirmed by RT-PCR. In conclusion, the mechanism of NO donor-induced cervical ripening during pregnancy may be mediated in part via increased prostaglandin F(2alpha) synthesis. (+info)