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Biomedical subjects

M Lucas

Publications and source records attributed to M Lucas.

At least 19 recordsLinked to original sources

Protein kinase C involvement in apoptosis.

1. Conflicting observations on the involvement of PKC in apoptosis point to a great variability depending on cell type, agent or condition causing apoptosis, phase of cell cycle and intracellular signaling pathway. 2. Inhibition by PKC of store-operated calcium entry mechanisms, which are sensitive to the oncoprotein bcl-2, should block the activation of calcium-dependent enzymes triggering the apoptotic cell death. 3. Activation of phosphatases by ceramide and inhibition of PKC by sphingosine seem to mediate the sphingomyelin pathway to apoptosis. 4. A putative target protein appears to be p34cdc2 which is regulated by a network of kinases and phosphatases. The uncoupling of timing for p34cdc2 activation and the completion of DNA replication results in the so-called "mitotic catastrophe" that shares some features with apoptosis.

Animals

[Cytomegalovirus-induced colitis in HIV infection. Considerations on its diagnosis, treatment and complications].

The diagnosis of cytomegalovirus intestinal disease in patients with HIV (human immunodeficiency virus) infection frequently raises diagnostic problems in view of the absence of definite pathological, serological or virological markers of active CMV infection. We describe the case of a 47-year-old man with a CMV colitis which illustrates several diagnostic and therapeutic problems and that was complicated by an intestinal perforation. We emphasize that in HIV+ patients with chronic diarrhea, the presence of abdominal pain should suggest the possibility of a CMV colitis and that in such cases a colonoscopy with biopsies of the right colon should be performed, in view of the higher frequency of the typical histopathological changes at this level. On the other hand, this case presented a marked thickening of the colon wall, simulating pseudotumoral images on CAT scans, as recently described in literature. The therapeutic possibilities as well as the complications of CMV colitis are discussed in the context of the occurrence of an ileal perforation, which represents the first report of this complication in Portuguese literature and which had the particularity of having a long survival after surgery in comparison with the previous cases described in international literature.

AIDS-Related Opportunistic Infections

Receptor-independent mechanisms are involved in the priming of neutrophil's oxidase by vasoactive intestinal peptide.

Vasoactive intestinal peptide (VIP) primed the respiratory burst of human neutrophils induced by phorbol myristate acetate (PMA) and by the chemotactic peptide N-formyl-Met-Leu-Phe (fMLP). The sigmoidal-shaped curve of the priming effect of VIP differs for both agonist since the Hill coefficient was close to three in the case of neutrophil activation by fMLP whereas the corresponding value for PMA was close to one. The priming effect of VIP was enhanced when neutrophils were stimulated by FMLP in the presence of sphinganine, a protein kinase C inhibitor, at concentrations which almost abolished the response to PMA. VIP failed to increase resting cytosolic free calcium and to modify the transient increase in [Ca2+]i induced by fMLP. The described results point out that the mechanism of the priming of neutrophils by VIP is also independent of calcium and protein kinase C. The absence of VIP receptors in plasma membrane of neutrophils suggests that a receptor-independent mechanism modulates the agonist-triggered signaling pathway. The priming of neutrophils by VIP can not be considered as a pharmacological effect, as may be deduced from the required VIP concentration; it should be rather considered that the enhancement of the formation of reactive oxygen metabolites by VIP may be interesting in the understanding of the neuroimmune axis.

Calcimycin

Pancreastatin activates protein kinase C by stimulating the formation of 1,2-diacylglycerol in rat hepatocytes.

We describe here the stimulation by pancreastatin of 1,2-diacylglycerol production and protein kinase C activity in liver plasma membrane and isolated hepatocytes. The dose-dependency for the stimulation of both processes was similar to the recently described pattern of glucose output and cytosolic Ca2+ transients produced by pancreastatin. The time course of diacylglycerol production at 30 degrees C showed a rapid increase within 5 min, reaching a maximum at 10 min. Protein kinase C from hepatocytes was dependent on Ca2+ and phosphatidylserine. Neither the pancreastatin-stimulated diacylglycerol production nor the activation of protein kinase C was affected by pretreatment with pertussis toxin. However, the presence of GTP partially inhibited this pancreastatin stimulation of 1,2-diacylglycerol in a dose-dependent manner, although GTP alone stimulates diacylglycerol accumulation. This inhibitory effect of GTP on pancreastatin stimulation of diacylglycerol synthesis was completely abolished by the pretreatment with pertussis toxin. In conclusion, this study provides evidence that pancreastatin stimulates the formation of 1,2-diacylglycerol by a pertussis-toxin-independent mechanism, which may be responsible for the pancreastatin activation of protein kinase C.

Adenosine Diphosphate Ribose

Coexpression of alpha and gamma enolase genes in neurons of adult rat brain.

Enolase (EC 4.2.1.11) is a glycolytic enzyme active as a dimer. In adult brain extracts, three forms, alpha alpha, alpha gamma and gamma gamma, have been described, with the alpha gamma hybrid accounting for 30% of total enolase activity (Fletcher et al., Dev Biol 65:462-475, 1978; Lucas et al., Dev Neurosci 10:91-98, 1988). Previous biochemical studies strongly suggest that this hybrid is not generated artefactually during the extraction procedures (Keller et al., J Neurochem 36:1389-1397, 1981; Shimizu et al., BBA 748:278-284, 1983). Immunocytological observations have demonstrated the cell specific localization of the alpha subunit in astrocytes and of the gamma subunit in neurons at the adult stage, but failed to identify a cell type containing both the alpha and gamma subunits necessary for the formation of the alpha gamma hybrid isoform (Ghandour et al., Exp Brain Res 41:271-279, 1981; Vinores et al., J Histochem Cytochem 32:1295-1302, 1984; Iwanaga et al., Arch Histol Cytol [Suppl] 52:13-24, 1989). We sought to approach this question by performing in situ hybridization studies in order to visualize the alpha and gamma mRNAs. In agreement with the immunocytological reports, we observe a specific accumulation of the gamma enolase transcripts in neurons and a high accumulation of alpha enolase transcripts in some glial cells such as the ependymocytes lining the ventricles. Our observations, following hybridization with 35S labeled oligonucleotide specific probes on adjacent thin sections, demonstrate for the first time that transcription of both alpha and gamma enolase genes occurs in many neurons of different brain regions. These results render highly probable the formation of the alpha gamma hybrid in mature neurons. Furthermore, we observe a differential expression of the genes encoding the alpha and gamma enolase subunits in various neuronal populations of the brain. The implications of these observations are discussed.

Animals

[Effectiveness of the treatment of spontaneous pneumothorax using small caliber pleural catheter].

BACKGROUND: The aim of this study was to evaluate the efficacy of the treatment of spontaneous pneumothorax (PT) by small caliber pleural catheter (SCPC) carried out by physicians with no surgical experience in the emergency department. METHODS: Forty-eight patients with spontaneous PT (23 total, 4 of them tensional) were included in the study. An 8 French caliber polyethylene SCPC was inserted in all the patients requiring pleural drainage. RESULTS: Thirty-two patients (67%) required pleural drainage. The catheter was carried for a mean of 4 days (1-7) resolving PT in 27 (84%) with no resolution being achieved in 5 (15%) and posterior surgical treatment being required. No major complications were observed during the evolution. CONCLUSIONS: The use of small caliber pleural catheters in the treatment of spontaneous pneumothorax (both total and to pressure) performed by physicians with no previous surgical knowledge is safe and effective.

Adolescent

Protein kinase C activation promotes cell survival in mature lymphocytes prone to apoptosis.

The putative protein kinase C (PKC) inhibitors polymyxin B and staurosporine were used to test the influence of PKC activity on the viability of lymphocytes. The cytotoxic effect of polymyxin B was characterized and it was found to be both time and dose dependent, with an LD50 in micromolar range, and counteracted by phorbol myristate acetate (PMA). To explore further the possible mechanism of action involved in polymyxin B-induced cell death, PKC activity and intracellular calcium were measured in polymyxin B-challenged lymphocytes. Polymyxin B inhibited PKC activity in both resting (25% inhibition) and PMA-stimulated (50% inhibition) cells, and increased intracellular calcium without disruption of the plasma membrane, a signal which is known to trigger apopotosis. Additionally, a number of experiments were conducted to assess the effect of staurosporine on PKC activity, cell growth, cell death and survival of mature lymphocytes. Staurosporine inhibited PKC activity in a dose-dependent manner (Ki close to 1 microM) and this effect correlated to some extent with the inhibition of [3H]thymidine incorporation and the breakdown of DNA into oligonucleosome-sized fragments. These results support the hypothesis that PKC is involved in the survival of mature lymphocytes undergoing apoptosis.

Alkaloids

LIFE: Learning Informally From Elders.

The Better Elder Services Today (BEST) Project helps frail elders maintain healthy, independent living through partnership with their formal and informal caregivers. This article reports on achieving one particular project goal: the development of community forums in which elders engaged in problem-solving with health care professionals and policy makers. The methodology involved community assessment and qualitative analysis of themes in 102 elders' responses to the question, "What has it been like for you trying to remain as independent as possible in the community?" Five predominant themes emerged: safety; problems communicating with health care providers; dissatisfaction with hospital services; complexity in access to community services; and fears about losses.

Aged

Serum PAPP-A measurements in first-trimester screening for Down syndrome.

Serum PAPP-A measurements taken from 254 women in the first trimester are reported. Eleven chromosomal abnormalities were detected. The mean serum PAPP-A levels in cases of Down syndrome were 0.44 MOM at 9 weeks gestation, 0.15 MOM at 10 weeks, and 0.29 MOM at 11 weeks. The PAPP-A level at 10 weeks was below those of pregnancies which aborted spontaneously. At 11 weeks, the pregnancies with Down syndrome recorded the lowest PAPP-A levels at that gestation. On this small sample, offering chorionic villus sampling to women with singleton pregnancies and a PAPP-A level below 0.3 MOM (approximately 6.5 per cent of this at-risk group) would have detected all the Down syndrome fetuses at 10 weeks and 50 per cent at 11 weeks without selecting those cases destined to abort. This suggests that serum PAPP-A should continue to be investigated as a potential first-trimester screening test for Down syndrome.

Adult

Sensitivity of insulin-secreting RIN m5F cells to undergoing apoptosis by the protein kinase C inhibitor staurosporine.

This study shows the sensitivity of insulin-secreting RIN m5F cells to undergoing apoptosis without modifying intracellular free calcium concentrations. The culture of cells in the absence of serum during 6 h failed to produce spontaneous DNA fragmentation. However, when cultures were carried out in the presence of the putative protein kinase C inhibitor staurosporine (1 microM), cells underwent apoptosis. Tumor-promoting phorbol ester failed to inhibit this effect. The presence of the chemotherapeutic drug bleomycin (0.6 mg/ml) in the culture medium reproduced the same pattern of cleaved DNA in nucleosomal pieces. Lower doses of staurosporine (0.1-1 nM) inhibited DNA synthesis but were unable to trigger apoptosis in 6 h culture. Higher doses of staurosporine (0.1-1 microM), which abolished DNA synthesis almost completely, were needed to trigger apoptosis. Short incubation of RIN m5F cells in the presence of staurosporine did not produce any change in the concentration of intracellular calcium or in the integrity of the plasma membrane. These results suggest the involvement of protein kinase C in RIN m5F cell survival.

Alkaloids

Takayasu's disease presenting as a nephrotic syndrome due to amyloidosis.

We report an 18 year old black woman who presented with nephrotic syndrome in whom the investigations led to the diagnosis of diffuse Takayasu's disease, renal amyloidosis of AA type and interstitial lung disease. Proteinuria in Takayasu's disease is usually ascribed to hypertension or more rarely to glomerulonephritis. This case suggests that amyloidosis should be considered also in the investigation of proteinuria in these patients in view of the serious prognostic implications. This case represents further evidence that Takayasu's disease can be the cause of systemic reactive amyloidosis which may also be the presenting feature.

Adolescent

Coelenterazine is a superoxide anion-sensitive chemiluminescent probe: its usefulness in the assay of respiratory burst in neutrophils.

The oxidation of free coelenterazine by superoxide anion was analyzed and compared to the oxidation by the semisynthetic photoprotein obelin, prepared by incorporation of synthetic coelenterazine into apoobelin. The oxidation of bound coelenterazine was triggered upon binding of calcium to the reconstituted photoprotein. The oxidation of free synthetic coelenterazine, in the absence of the apoprotein, was triggered by superoxide anion. The production of reactive oxygen metabolites by fMet-Leu-Phe- and 4b-phorbol 12b-myristate 13a-acetate-stimulated neutrophils was studied by means of the luminescence of synthetic coelenterazine. The features of this chemiluminescent probe were compared with those of luminol and are summarized as follows: (a) coelenterazine-dependent chemiluminescence was inhibited by superoxide dismutase; (b) coelenterazine was as sensitive as luminol in detecting the oxidative burst of neutrophils; (c) azide failed to inhibit coelenterazine chemiluminescence; (d) in contrast with luminol, which requires the catalytic removal of hydrogen peroxide, coelenterazine chemiluminescence did not depend on the activity of cell-derived myeloperoxidase. These results indicate the usefulness of coelenterazine as a very sensitive and specific chemiluminescence probe of superoxide anion.

Aequorin

Glycogenolytic effect of pancreastatin in isolated rat hepatocytes is mediated by a cyclic-AMP-independent Ca(2+)-dependent mechanism.

We have studied the effect of pig pancreastatin on glucose and lactate production in freshly isolated rat hepatocytes. Pancreastatin stimulated the rate of glucose output, whereas, in contrast with glucagon, it failed to modify the rate of lactate production. The effective concentration of pancreastatin was in the range 0.1-100 nM, with half-maximal rate close to 1 nM. The ability of pancreastatin to increase glucose output was abolished by chelation of the calcium in the medium. By itself, pancreastatin did not increase cyclic AMP (cAMP) levels and had no influence on cAMP levels in glucagon-stimulated hepatocytes. Our results point out a possible role of pancreastatin in glycogenolysis. This appears to be mediated by a cAMP-independent Ca(2+)-dependent mechanism.

1-Methyl-3-isobutylxanthine

Preparation of a protein-free total brain white matter lipid fraction: characterization of liposomes.

A method of preparing a total lipid extract (TLE), free of protein, by extracting brain white matter with tetrahydrofuran is presented. The optimal conditions of extraction were found to be 50 ml of THF per gram of lyophilized tissue, though fresh tissue can also be used if larger volumes of solvent are employed. The method allowed, in a short time and in a single step, a yield of TLE of 50% on a dry weight basis. Its analytical characterization revealed a qualitative and quantitative composition very similar to the lipid composition of CNS myelin, including all the phospholipid and galactolipid species, cholesterol and gangliosides, but it contained only traces (0.1%) of protein. TLE has been used to prepare liposomes, either multilamellar (MLVs) or unilamellar (LUVs, SUVs), characterized by freeze-fracture electron microscopy. A multilayered, heterogeneous population of liposomes is observed in the MLVs preparation. When these samples were submitted to a freezing and thawing procedure the resulting liposomes were single-walled, and their intravesicular volume was increased. They were quite impermeable to the monovalent cation 86Rb+ and, by contrast, rather permeable to 45Ca+ +. Their complex lipid composition, together with their permeability properties and their response to ionophores, make them very useful to study protein-lipid interactions occurring within the myelin membrane as well as the functional properties of myelin proteins in reconstitution experiments.

Animals

Decreased protein kinase C activity is associated with programmed cell death (apoptosis) in freshly isolated rat hepatocytes.

Apoptosis of freshly isolated rat hepatocytes was induced by either the omission of fetal bovine serum in the culture medium or addition of the protein kinase C inhibitors polymyxin B or staurosporine. The time-course of DNA breakdown into oligonucleosome-sized fragments and the activity of protein kinase C was determined. Hepatocytes were found to be sensitive to bleomycin which induced a high degree of DNA breakdown even within 30 min incubation. Both staurosporine and polymyxin B induced DNA degradation in hepatocytes after three hours incubation, an effect that was partially prevented by phorbol myristate acetate (PMA). After eight hours incubation, PMA failed to counteract this action and itself produced the apoptosis of rat hepatocytes. The results suggest the involvement of protein kinase C in hepatocyte survival.

Alkaloids

European brown hare syndrome in the U.K.; a calicivirus related to but distinct from that of viral haemorrhagic disease in rabbits.

The virus recovered from cases of European brown hare syndrome in the U.K. contains one major capsid protein of approximately 60 k molecular weight and morphologically resembles known caliciviruses. It has been compared with a European isolate of rabbit haemorrhagic disease calicivirus and, although it shows some antigenic similarity, it is not identical. In transmission and protection studies the virus from U.K. hares failed to produce disease in rabbits and did not effectively protect against subsequent challenge with the rabbit calicivirus.

Animals

Vasoactive intestinal peptide enhances phorbol myristate acetate-induced chemiluminescence in human lymphocytes.

Phorbol-myristate-acetate (PMA) induced in lymphocytes the production or reactive oxygen intermediates in a process which was stimulated by the presence of vasoactive intestinal peptide (VIP) in a dose-dependent response at VIP concentrations in the range 10(-11)-10(-8) M. The dissociation constant for the high-affinity receptors of VIP agreed with the ID50 of the activation of adenylate cyclase, and the ID50 for the stimulation by VIP of PMA-induced chemiluminescence, which were close to 0.2 nM VIP. Forskolin produced in lymphocytes an effect quite similar to VIP. A comparison of the response to VIP and forskolin of lymphocytes and monocytes showed that, in contrast to forskolin, VIP failed to induce the above described effect in monocytes. A possible mechanism involving protein kinase C, which is activated by PMA, and an intracellular signal linked to VIP receptors is pointed out. This study further supports a role for VIP as a mediator in the neuroimmune system.

Adenylyl Cyclases