Biomedical subjects

L Boscá

Publications and source records attributed to L Boscá.

At least 19 recordsLinked to original sources

PubMed2004

Increased intrahepatic cyclooxygenase 2, matrix metalloproteinase 2, and matrix metalloproteinase 9 expression is associated with progressive liver disease in chronic hepatitis C virus infection: role of viral core and NS5A proteins.

BACKGROUND: Cyclooxygenase 2 (COX-2) and matrix metalloproteinases (MMPs) have been implicated in tissue injury and fibrogenesis in animal models but little is known regarding their role in hepatitis C virus (HCV) related liver disease in humans. AIMS: To characterise the intrahepatic expression pattern of COX-2 and MMPs in chronic HCV infection and determine whether HCV core and NS5A proteins could promote their expression in cultured hepatocyte derived cell lines. PATIENTS: Thirty two anti-HCV+ and 10 anti-HCV- patients were studied. METHODS: Western blot, reverse transcription-polymerase chain reaction (RT-PCR), enzyme immunoassay, and immunohistochemistry were used to assess the expression pattern of COX-2 and MMPs in liver biopsy samples from all patients. COX-2 gene expression and MMP-9 protein levels were also determined by immunoblot, RT-PCR, and luciferase assays in core and NS5A transfected hepatocyte derived cells. RESULTS: The intrahepatic expression level of COX-2, MMP-2, and MMP-9 was significantly higher in HCV+ than in HCV- patients, increasing with the fibrotic stage of liver disease. We further demonstrated that COX-2 mRNA, protein, and activity were induced in resting and activated core and NS5A transfectants. Both viral proteins induced transcriptional activity of the COX-2 gene promoter whereas core, but not NS5A, exerted an inducer effect on MMP-9 protein levels in cultured hepatocyte derived cells. CONCLUSIONS: Intrahepatic COX-2, MMP-2, and MMP-9 overexpression is associated with progressive hepatic fibrosis in chronic HCV infection, suggesting their pathogenic role in fibrogenesis. HCV core and NS5A proteins were able to upregulate COX-2 and MMP-9 gene expression in hepatocyte derived cells, providing a potential mechanism for hepatic fibrosis during chronic HCV infection.

Adult↗

PubMed2003

Ammonia prevents glutamate-induced but not low K(+)-induced apoptosis in cerebellar neurons in culture.

Cultured rat cerebellar granule neurons are widely used as a model system for studying neuronal apoptosis. Either low K(+) (5 mM) or low concentrations of glutamate (1-10 microM) induce apoptosis in cerebellar neurons in culture. However, the molecular mechanism(s) involved remain unclear. We show that long-term treatment with ammonia prevents glutamate-induced but not low K(+)-induced apoptosis in cerebellar neurons, as assessed by measuring DNA fragmentation and activation of caspase 3. Ammonia prevented glutamate-induced increase of intracellular calcium, depolarization of the inner mitochondrial membrane, release of cytochrome c to the cytosol, activation of caspase 3 and fragmentation of DNA. However, ammonia did not prevent low K(+)-induced activation of caspase 3 and fragmentation of DNA. These results indicate that the initial steps involved in the induction of apoptosis by low K(+) or by glutamate are different and that ammonia prevents glutamate-induced apoptosis by reducing glutamate-induced rise of intracellular Ca(2+), thus avoiding the activation of subsequent events of the apoptotic process.

Ammonia↗

PubMed2003

Terpenoids: sources, structure elucidation and therapeutic potential in inflammation.

Natural products research has lately undergone exponential growth owing to advances in isolation techniques and in synthetic methods design, as well as for the identification of a wide range of biological properties exhibited by these compounds. In the present review, general remarks on the chemical features, biosynthetic pathways, isolation and structure elucidation of terpenoids are briefly discussed. In addition to this, recent work done on anti-inflammatory terpenoids (diterpenoids, triterpenoids and sesquiterpene lactones) with special emphasis on the last new molecular targets evaluated is presented.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2002

Coexistence of translocated cytochrome c and nitrated protein in neurons of the rat cerebral cortex after oxygen and glucose deprivation.

Changes in the distribution of immunoreactive cytochrome c and protein nitration were studied in the rat cerebral cortex after oxygen and glucose deprivation by bright field, confocal and electron microscopy. In control cerebral cortex, nitrotyrosine immunoreactivity indicating protein nitration was found mostly in the neuronal nuclear region, with only a small amount distributed in the cytosol, whereas cytochrome c immunoreactivity was found at the inner membrane and in the intermembrane space of the mitochondria. During the recovery phase after oxygen and glucose deprivation, cytochrome c immunoreactivity was released from the intermembrane space of swollen mitochondria into the surrounding cytosol. The cytosol now also displayed nitrotyrosine immunoreactivity, which had diminished in the nuclear region. Both immunoreactivities were dispersed throughout the soma and processes of the cortical neurons. These changes were largely prevented by the administration of cyclosporin A, which inhibits both the mitochondrial permeability transition and the neuronal isoform of nitric oxide synthase while blocking the induction of the inducible isoform. Ischemia/reperfusion injury increases the production of nitric oxide, reactive oxygen species and intracellular factors that damage the mitochondria and liberate apoptotic factors. We suggest that translocation of cytochrome c from the mitochondria to the cytosol, which has been shown to precede the mitochondrial permeability transition, could result from peroxynitrite-mediated nitration. This phenomenon is attenuated by cyclosporin A administration, suggesting a neuroprotective role for this agent.

Animals↗

PubMed2002

Induction of apoptosis by nitric oxide in macrophages is independent of apoptotic volume decrease.

Apoptosis occurs through a sequence of specific biochemical and morphological alterations that define the progress of cell death. The changes of the mitochondrial inner membrane potential (DeltaPsi(m)), the release of cytochrome c to the cytosol, the apoptotic volume decrease (AVD) and the activation of caspases have been measured in RAW 264.7, HeLa and Jurkat T cells incubated with molecules that induce apoptosis through the mitochondrial pathway. Our data show that NO, staurosporine, etoposide and camptothecin increased DeltaPsi(m) in macrophages but not in HeLa and Jurkat cells, that exhibited a DeltaPsi(m) decrease. Moreover, the apoptosis induced by NO in macrophages, but not that promoted by staurosporine, might occur in the absence of AVD. Analysis of the sequence of apoptotic manifestations shows that DeltaPsi(m) precedes AVD and caspase activation in RAW 264.7 cells. Inhibition of AVD abrogates apoptosis in HeLa and Jurkat T cells regardless of the stimuli used. These data suggest that the changes of DeltaPsi(m) are cell-type dependent and that AVD is dispensable for apoptosis in macrophages.

Apoptosis↗

PubMed2001

Protein kinase Cepsilon is required for macrophage activation and defense against bacterial infection.

To assess directly the role of protein kinase C (PKC)epsilon in the immune system, we generated mice that carried a homozygous disruption of the PKCepsilon locus. PKCepsilon(-/-) animals appeared normal and were generally healthy, although female mice frequently developed a bacterial infection of the uterus. Macrophages from PKCepsilon(-/-) animals demonstrated a severely attenuated response to lipopolysaccharide (LPS) and interferon (IFN)gamma, characterized by a dramatic reduction in the generation of NO, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1beta. Further analysis revealed that LPS-stimulated macrophages from PKCepsilon(-/-) mice were deficient in the induction of nitric oxide synthase (NOS)-2, demonstrating a decrease in the activation of IkappaB kinase, a reduction in IkappaB degradation, and a decrease in nuclear factor (NF)kappaB nuclear translocation. After intravenous administration of Gram-negative or Gram-positive bacteria, PKCepsilon(-/-) mice demonstrated a significantly decreased period of survival. This study provides direct evidence that PKCepsilon is critically involved at an early stage of LPS-mediated signaling in activated macrophages. Furthermore, we demonstrate that in the absence of PKCepsilon, host defense against bacterial infection is severely compromised, resulting in an increased incidence of mortality.

Animals↗

PubMed2001

Contribution of cyclooxygenase 2 to liver regeneration after partial hepatectomy.

Partial hepatectomy (PH) triggers a rapid regenerative response in the remaining tissue to reinstate the organ function and the cell numbers. Among the molecules that change in the course of regeneration is an accumulation of prostaglandin E2 in the sera of rats with PH. Analysis of the cyclooxygenase (COX) isoenzymes in the remnant liver showed the preferential expression of COX-2 in hepatocytes. Cultured regenerating hepatocytes expressed significant levels of COX-2, a process that was not observed in the sham counterparts. Maximal expression of COX-2 was detected 16 h after PH with increased levels present even at 96 h. Pharmacological inhibition of COX-2 activity with NS398 shunted the up-regulation of cell proliferation after PH, which suggests a positive interaction of prostaglandins with the progression of the cell cycle. Similar results were obtained after PH of mice lacking the COX-2 gene. The expression of COX-2 in regenerating liver was concomitant with a decrease in CCAAT-enhancer binding protein (C/EBP-a) level and an increase in the expression of C/EBP-b and C/EBP-d. These results suggest a contribution of the enhanced synthesis of prostaglandins to liver regeneration observed after PH.

Animals↗

PubMed2001

Peroxisome proliferator-activated receptor-gamma-independent inhibition of macrophage activation by the non-thiazolidinedione agonist L-796,449. Comparison with the effects of 15-deoxy-delta(12,14)-prostaglandin J(2).

The effects of L-796,449 (3-chloro-4-(3-(3-phenyl-7-propylbenzofuran-6-yloxy)propylthio)phenylacetic acid; referred to henceforth as compound G), a thiazolidinedione-unrelated peroxisome proliferator activated-receptor-gamma (PPAR-gamma) agonist, on early signaling in lipopolysaccharide-activated RAW 264.7 macrophages were analyzed and compared with those elicited by 15-deoxy-Delta(12,14)-prostaglandin J(2) and the thiazolidinedione rosiglitazone. Compound G inhibited the activation of nuclear factor kappa B through the impairment of the targeting and degradation of I kappa B proteins and promoted a redistribution of I kappa B alpha and I kappa B beta in the nucleus of activated cells. Compound G inhibited I kappa B kinase (IKK) activity both in vivo and in vitro, suggesting a direct mechanism of interaction between this molecule and the IKK complex. The effect of compound G on IKK activity was independent of PPAR-gamma engagement because RAW 264.7 cells expressed negligible levels of this nuclear receptor, and rosiglitazone failed to mimic these actions. Moreover, treatment of activated macrophages with compound G enhanced the synthesis of superoxide anion, which, in combination with the NO produced under activation conditions, triggered apoptosis through the intracellular synthesis of peroxynitrite. These results suggest that compound G might contribute to the resolution of inflammation by favoring the induction of apoptosis through mechanisms independent of PPAR-gamma engagement.

Anions↗

PubMed2001

Up-regulation of TNF-alpha convertase (TACE/ADAM17) after oxygen-glucose deprivation in rat forebrain slices.

Tumour necrosis factor-alpha (TNF-alpha) is a major immunomodulatory and proinflammatory cytokine which is shed in its soluble form by a membrane-anchored zinc protease, identified as a disintegrin and metalloproteinase (ADAM) called TNF-alpha convertase (TACE; ADAM17). The role of this protease in the adult nervous system remains poorly understood. During cerebral ischemia and subsequent reperfusion, expression and release of TNF-alpha have been shown. We have investigated the expression and activity of TACE in an in vitro model of brain ischemia consisting of rat forebrain slices exposed to oxygen-glucose deprivation (OGD). OGD caused the release of TNF-alpha, an effect which was inhibited by a hydroxamate-based metalloprotease inhibitor, BB-3103, with an IC(50) of 0.1 microM, suggesting that TNF-alpha release results selectively from TACE activity. Assay of TACE enzymatic activity on a fluorescein-labelled peptide spanning the cleavage site in pro-TNF-alpha, as well as Western blot and RT-PCR analyses showed that TACE is present in control forebrain and, more interestingly, that TACE expression is increased in OGD-exposed tissue. TACE enzymatic activity from OGD-exposed slices was significantly inhibited by cycloheximide, suggesting that de novo synthesis of TACE contributes to TNF-alpha release after ischaemia. Moreover, it was also inhibited by bisindolylmaleimide I, indicating that TACE activity is regulated by PKC. These findings posed the question of what was its function therein. Among other actions, TNF-alpha has been described to be involved in the expression of inducible nitric oxide synthase (iNOS), a high-output NOS isoform associated to cellular damage, but the link between TNF-alpha release after brain ischaemia and iNOS expression in this condition has not been shown. We have now found that iNOS expression in OGD-subjected brain slices is inhibited by BB-3103 at concentrations below 1 microM, indicating that shedding of TNF-alpha by TACE plays a necessary part in the induction of this NOS isoenzyme after OGD. Taken together, these data demonstrate that (1) TACE/ADAM17 activity accounts for the majority of TNF-alpha shedding after OGD in rat forebrain slices, (2) an increase in TACE expression contributes, at least in part, to the rise in TNF-alpha after OGD and (3) iNOS expression in OGD-subjected brain slices results from TACE activity and subsequent increase in TNF-alpha levels.

ADAM Proteins↗

PubMed2001

Intracellular water motion decreases in apoptotic macrophages after caspase activation.

Triggering of the macrophage cell line RAW 264.7 with lipopolysaccharide and interferon-gamma promoted apoptosis that was prevented by inhibitors of type 2 nitric oxide synthase or caspase. Using (1)H NMR analysis, we have investigated the changes of the intracellular transverse relaxation time (T(2)) and apparent diffusion coefficient (ADC) as parameters reflecting the rotational and translational motions of water in apoptotic macrophages. T(2) values decreased significantly from 287 to 182 ms in cells treated for 18 h with NO-donors. These changes of T(2) were prevented by caspase inhibitors and were not due to mitochondrial depolarization or microtubule depolymerization. The decrease of the intracellular values of T(2) and ADC in apoptotic macrophages was observed after caspase activation, but preceded phosphatidylserine exposure and nucleosomal DNA cleavage. The changes of water motion were accompanied by an enhancement of the hydrophobic properties of the intracellular milieu, as detected by fluorescent probes. These results indicate the occurrence of an alteration in the physicochemical properties of intracellular water during the course of apoptosis.

Amino Acid Chloromethyl Ketones↗

PubMed2001

Inducible nitric oxide synthase expression in brain cortex after acute restraint stress is regulated by nuclear factor kappaB-mediated mechanisms.

The underlying mechanisms by which physical or psychological stress causes neurodegeneration are still unknown. We have demonstrated that the high-output and long-lasting synthesizing source of nitric oxide (NO), inducible NO synthase (iNOS), is expressed in brain cortex during stress and that its overexpression accounts for the neurodegenerative changes seen after 3 weeks of repeated stress. Now we have found that acute stress (restraint for 6 h) increases the activity of a calcium-independent NOS and induces the expression of iNOS in brain cortex in adult male rats. In order to elucidate the possible mechanisms involved in this induction, we studied the role of transcription nuclear factor kappaB (NF-kappaB), which is required for iNOS synthesis. We have observed that an acute restraint stress session stimulates the translocation of the NF-kappaB to the nucleus after 4 h and that the administration of the NF-kappaB inhibitor pyrrolidine dithiocarbamate [PDTC, 75 and 150 mg/kg intraperitoneally (i.p.)] at the onset of stress inhibits the stress-induced increase in iNOS expression. Since glutamate release and subsequent NMDA (N-methyl-D-aspartate) receptor activation has been recognized as an early change after exposure to stressful stimuli, and glutamate has been shown to induce iNOS in brain via a NF-kappaB-dependent mechanism, we studied the possible role of excitatory amino acids in the induction of iNOS in our model. Pretreatment with the NMDA receptor antagonist dizocilpine (MK-801, 0.1 and 0.3 mg/kg i.p.) inhibits the stress-induced NF-kappaB activation as well as the stress-induced increase in iNOS expression. Taken together, these findings indicate that excitatory amino acids and subsequent activation of NF-kappaB account for stress-induced iNOS expression in cerebral cortex, and support a possible neuroprotective role for specific inhibitors in this situation.

Active Transport, Cell Nucleus↗

PubMed2001

Expression of cyclooxygenase-2 promotes the release of matrix metalloproteinase-2 and -9 in fetal rat hepatocytes.

Treatment of primary cultures of fetal hepatocytes with proinflammatory cytokines, lipopolysaccharide (LPS), and hepatocyte growth factor promoted the expression of cyclooxygenase-2 (COX-2) and the synthesis of high amounts of prostaglandins (PGs). Under these conditions, the active forms of the matrix metalloproteinases-2 and -9 (MMPs) were released to the extracellular medium. This process was inhibited when the synthesis of PGs was suppressed pharmacologically with COX-2 inhibitors. Addition to the cell cultures of PGE(2) promoted the release of MMPs through a mechanism that involved the expression of COX-2 and the synthesis of additional PGs. Kinetic analysis of the secretion of MMPs in response to LPS and PGE(2) showed a similar time course, with a lag period of 6 hours, which suggests that PGE(2) does not act directly on the mechanism of MMP processing and release. Inhibitors of protein kinase A, p38 MAP kinase, phosphatidylinositol-3-kinase, and nuclear factor kappaB (NF-kappaB) activation impaired the release of MMPs in response to PGE(2) challenge, indicating the involvement of multiple steps in the process. The ability of fetal hepatocytes to release MMPs in response to growth factors and inflammatory stimuli constitutes a model for the study of the extracellular matrix remodeling that accompanies most liver diseases.

Animals↗

PubMed2000

Contribution of cyclopentenone prostaglandins to the resolution of inflammation through the potentiation of apoptosis in activated macrophages.

Activation of the macrophage cell line RAW 264.7 with LPS and IFN-gamma induces apoptosis through the synthesis of high concentrations of NO due to the expression of NO synthase-2. In addition to NO, activated macrophages release other molecules involved in the inflammatory response, such as reactive oxygen intermediates and PGs. Treatment of macrophages with cyclopentenone PGs, which are synthesized late in the inflammatory onset, exerted a negative regulation on cell activation by impairing the expression of genes involved in host defense, among them NO synthase-2. However, despite the attenuation of NO synthesis, the percentage of apoptotic cells increased with respect to activated cells in the absence of cyclopentenone PGs. Analysis of the mechanisms by which these PGs enhanced apoptosis suggested a potentiation of superoxide anion synthesis that reacted with NO, leading to the formation of higher concentrations of peroxynitrite, a more reactive and proapoptotic molecule than the precursors. The effect of the cyclopentenone 15-deoxy-Delta(12,14)-PGJ(2) on superoxide synthesis was dependent on p38 mitogen-activated protein kinase activity, but was independent of the interaction with peroxisomal proliferator-activated receptor gamma. The potentiation of apoptosis induced by cyclopentenone PGs involved an increase in the release of cytochrome c from the mitochondria to the cytosol and in the nitration of this protein. These results suggest a role for cyclopentenone PGs in the resolution of inflammation by inducing apoptosis of activated cells.

Adjuvants, Immunologic↗

PubMed2000

Mechanisms of the neuroprotective effect of aspirin after oxygen and glucose deprivation in rat forebrain slices.

Acetylsalicylic acid (ASA, Aspirin) is an anti-inflammatory drug with a wide spectrum of pharmacological activities and multiple sites of action. Apart from its preventive actions against stroke due to its antithrombotic properties, recent data in the literature suggest that high concentrations of ASA also exert direct neuroprotective effects. We have used an in vitro model of brain ischaemia using rat forebrain slices deprived of oxygen and glucose to test ASA neuroprotective properties. We have found that ASA inhibits neuronal damage at concentrations lower than those previously reported (0.1-0.5 mM), and that these effects correlate with the inhibition of excitatory amino acid release, of NF-kappaB translocation to the nucleus and iNOS expression caused by ASA. All of these three mechanisms may mediate the neuroprotective effects of this drug. Our results also show that the effects of ASA are independent of COX inhibition. Taken together, our present findings show that ASA is neuroprotective in an in vitro model of brain ischaemia at doses close to those recommended for its antithrombotic effects.

Animals↗

PubMed2000

Anti-inflammatory action of type I interferons deduced from mice expressing interferon beta.

Type I interferons (IFN) are widely used for the therapeutic treatment of viral infections, tumor growth and various chronic diseases such as multiple sclerosis. Antagonism between type I IFNs and IFN-gamma has been described in cells of the immune system, in particular in the activation of macrophages. To study the systemic effects of type I IFNs we used transgenic mice carrying a human IFN-beta (hIFN-beta) gene under the control of the rat insulin I promoter. These animals expressed high levels of hIFN-beta in beta-pancreatic cells, and the ability of the macrophages to respond to pro-inflammatory stimuli was analyzed. Transgenic mice exhibited an increased extravasation of cells to the peritoneal cavity after eliciting with thioglycollate broth. The expression of the inducible form of nitric oxide synthase and cyclooxygenase-2, two enzymes involved in inflammation, was impaired in transgenic animals challenged with lipopolysaccharide and IFN-gamma. Analysis of the mechanisms leading to this attenuated inflammatory response showed a decrease in the serum levels of TNF-alpha and an inhibition of the activation of the transcription factor NF-KB in various tissues. These results indicate that systemic administration of IFN-beta might influence the response to pro-inflammatory stimuli, in particular through the antagonism of IFN-gamma signaling.

Animals↗

PubMed2000

Implication of glutamate in the expression of inducible nitric oxide synthase after oxygen and glucose deprivation in rat forebrain slices.

Nitric oxide synthesis by inducible nitric oxide synthase (iNOS) has been postulated to contribute to ischemia-reperfusion neurotoxicity. The expression of this enzyme has been demonstrated in cells present in the postischemic brain. The mechanisms of iNOS expression after cerebral ischemia are a subject of current research. We therefore decided to investigate whether glutamate, which is released in ischemia and is implicated in neurotoxicity, might be involved in the mechanisms by which oxygen and glucose deprivation (OGD) leads to the expression of iNOS in rat forebrain slices. In this model, we have shown previously that 20 min of OGD causes the expression of iNOS. We have now found that the NMDA receptor antagonist MK-801 blocks the expression of iNOS, suggesting that the activation of the NMDA subtype of glutamate receptor is implicated in the mechanisms that lead to the expression of this isoform. Moreover, we have found that glutamate alone could trigger the induction process, as shown by the appearance of a Ca(2+)-independent NOS activity and by the detection of iNOS mRNA and protein in slices exposed to glutamate. Glutamate-dependent iNOS expression was concentration-dependent and was blocked by EGTA and by the inhibitors of nuclear factor kappaB (NF-kappaB) activation pyrrolidine dithiocarbamate and MG132. In addition, glutamate induced NF-kappaB translocation to the nucleus, an effect that was inhibited by MG132. Taken together, our data suggest that activation of NMDA receptors by glutamate released in ischemia is involved in the expression of iNOS in rat forebrain slices via a Ca(2+)-dependent activation of the transcription factor NF-kappaB. To our knowledge, this is the first report showing an implication of excitatory amino acids in the expression of iNOS caused by ischemia.

Animals↗

PubMed2000

Chronic stress induces the expression of inducible nitric oxide synthase in rat brain cortex.

Long-term exposure to stress has detrimental effects on several brain functions in many species, including humans, and leads to neurodegenerative changes. However, the underlying neural mechanisms by which stress causes neurodegeneration are still unknown. We have investigated the role of endogenously released nitric oxide (NO) in this phenomenon and the possible induction of the inducible NO synthase (iNOS) isoform. In adult male rats, stress (immobilization for 6 h during 21 days) increases the activity of a calcium-independent NO synthase and induces the expression of iNOS in cortical neurons as seen by immunohistochemical and western blot analysis. Three weeks of repeated immobilization increases immunoreactivity for nitrotyrosine, a nitration product of peroxynitrite. Repeated stress causes accumulation of the NO metabolites NO2+ NO3- (NOx-) accumulation in cortex, and these changes occur in parallel with lactate dehydrogenase (LDH) release and impairment of glutamate uptake in synaptosomes. Administration of the selective iNOS inhibitor aminoguanidine (400 mg/kg i.p. daily from days 7 to 21 of stress) prevents NOx- accumulation in cortex, LDH release, and impairment of glutamate uptake in synaptosomes. Taken together, these findings indicate that a sustained overproduction of NO via iNOS expression may be responsible, at least in part, for some of the neurodegenerative changes caused by stress and support a possible neuroprotective role for specific iNOS inhibitors in this situation.

Animals↗

PubMed2000

Regulation of cyclooxygenase 2 expression in hepatocytes by CCAAT/enhancer-binding proteins.

BACKGROUND & AIMS: Expression of cyclooxygenase (COX)-2 in response to lipopolysaccharide (LPS) challenge has been analyzed in cultured fetal, neonatal, and adult hepatocytes and in hepatoma cell lines. METHODS: To study the mechanisms of LPS-dependent expression of COX-2 in these cells, the activity of the COX-2 promoter and the levels of CCAAT/enhancer-binding proteins (C/EBPs) were determined. RESULTS: COX-2 was induced in fetal hepatocytes, but this response declined rapidly after birth. This loss of inducibility of COX-2 paralleled the expression of C/EBP-alpha in neonatal hepatocytes. Transfection of fetal and adult hepatocytes with sequences corresponding to the 5'-flanking region of the rat COX-2 gene confirmed the absence of promoter activity in adult hepatocytes. Moreover, transient expression of C/EBP-alpha, but not C/EBP-delta, in the hepatoma cell line AT3F cells abolished the COX-2 promoter activity. Prolonged culture of adult hepatocytes restored the induction of COX-2 after complete disappearance of C/EBP-alpha. CONCLUSIONS: These results suggest that the presence of high levels of C/EBP-alpha is involved in the impairment of COX-2 expression in adult hepatocytes challenged with proinflammatory stimuli.

Age Factors↗