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Biomedical subjects

M Rocha

Publications and source records attributed to M Rocha.

At least 55 records · Page 3Linked to original sources
PubMed1996

A role for sialoadhesin-positive tissue macrophages in host resistance to lymphoma metastasis in vivo.

Sialoadhesin (SER) is a newly described macrophage-restricted adhesion molecule with a sequence similarity to CD22 on B cells and to myelin-associated glycoprotein on Schwann cells. We describe here a functional role of SER+ spleen macrophages in antigen processing and presentation to T lymphocytes. In two syngeneic murine tumour systems (ESb-MP and lacZ transduced ESbL T-lymphoma cells), the activation state of SER+ macrophages (tested by activity of marker enzymes adenosine deaminase and 5'-nucleotidase) correlated with the arrest of lymphoma metastasis. Furthermore, this macrophage subpopulation became activated upon anti-tumour immunization as well as upon adoptive transfer of immune T lymphocytes into tumour-bearing hosts. We suggest that in situ-activated SER+ macrophages contribute to host resistance against metastasis.

Animals↗
PubMed1996

Death in a dish: controls of apoptosis within the developing retinal tissue.

Studies of programmed cell death in the developing retina in vitro are currently reviewed. The results of inhibiting protein synthesis in retinal explants indicate two mechanisms of apoptosis. One mechanism depends on the synthesis of positive modulators ('killer proteins'), while a distinct, latent mechanism appears to be continuously blocked by negative modulators. Extracellular modulators of apoptosis include the neurotrophic factors NT-4 and BDNF, while glutamate may have either a positive or a negative modulatory action on apoptosis. Several protein kinases selectively modulate apoptosis in distinct retinal layers. Calcium and nitric oxide were also shown to affect apoptosis in the developing retinal tissue. The protein c-Jun was found associated with apoptosis in various circumstances, while p53 seems to be selectively expressed in some instances of apoptosis. The results indicate that the sensitivity of each retinal cell to apoptosis is controlled by multiple, interactive, cell type- and context-specific mechanisms. Apoptosis in the retina depends on a critical interplay of extracellular signals delivered through neurotrophic factors, neurotransmitters and neuromodulators, several signal transduction pathways, and the expression of a variety of genes.

Animals↗
PubMed1996

Dynamic expression changes in vivo of adhesion and costimulatory molecules determine load and pattern of lymphoma liver metastasis.

Although intradermal primary tumor growth and spontaneous liver metastasis of ESbL-lacZ lymphoma in syngeneic DBA/2 mice are progressive and malignant, they are characterized by a transient plateau period with a constant tumor diameter and a low number of metastasized cells in the liver. This period, which was shown to be immune dependent, was followed by a second expansion phase characterized by a preferential localization of tumor cells in the periportal areas of liver lobules (mosaic phenotype). To elucidate possible mechanisms leading to the plateau period as well as for the mosaic-like metastasis pattern, we investigated, using flow cytometry analysis, alterations in costimulatory and adhesion molecule expression in liver sinusoidal cells as well as in tumor cells isolated directly ex vivo throughout the kinetics of metastasis. In tumor and sinusoidal cells, we found up-regulation in the expression of MHC class II and B7 molecules during the plateau period. These molecules, which facilitate cell-mediated immune responses, were again down-regulated during the final exponential tumor growth and metastasis. In the final expansion phase, in which the mosaic phenotype of liver metastasis is seen, we detected a significant increase of leukocyte function-associated antigen-1/intercellular adhesion molecule-1 expression in both tumor and sinusoidal cells, suggesting tumor cell-sinusoidal cell interactions. vascular cell adhesion molecule-1/very late activated antigen-4 did not show any modification during the whole metastatic process. In vivo application of monoclonal antibodies directed to leukocyte function-associated antigen-1 and intercellular adhesion molecule-1 appeared to block the spread of metastasis, while no effect was seen with monoclonal antibodies directed to vascular cell adhesion molecule-1 and very late activated antigen-4. This study reveals in situ expression changes of cell surface molecules in tumor and host cells during metastasis. The changes seen during the plateau phase and during the second expansion phase differ, suggesting associations with mechanisms of immune control and tumor immune evasion, respectively.

Animals↗
PubMed1995

Liver endothelial cells participate in T-cell-dependent host resistance to lymphoma metastasis by production of nitric oxide in vivo.

Tumor growth and metastasis of lacZ-transduced murine lymphoma ESbL cells inoculated into syngeneic DBA/2 mice are characterized by a transient plateau phase with a constant tumor diameter and low metastatic load, indicating a host response against the tumor. Here we show that endothelial cells participate in a T-cell-dependent, anti-metastatic response by producing NO in situ. Liver endothelial cells were isolated and examined directly ex vivo without further manipulation. NO production in liver endothelial cells reached the highest level during the plateau phase but declined toward the end of it, followed by an overall breakdown of host response, leading to progressive tumor growth and high load of liver metastasis. Mice subjected to anti-tumor immunization and subsequent challenge with a tumorigenic dose of ESbL-lacZ cells showed, in comparison to non-immunized challenged controls, reduced liver metastasis and increased endothelial NO production. Adoptive transfer of anti-tumor immune spleen cells from semi-allogeneic B10.D2 mice into tumor-bearing animals during the plateau phase caused a regression of primary tumor and metastases, together with a preservation of the high level of NO synthesis in endothelial cells. In immuno-incompetent (SCID) mice, tumor growth and metastasis were progressive and there was no endothelial NO response. Pre-immunization of immuno-competent mice with both live and irradiated tumor cells at different sites of the body led to an induction of NO production by liver endothelial cells. These results reveal a novel role of endothelial cells in the suppression of lymphoma metastasis in the liver. The inducible endothelial cell NO response is apparently dependent and induced by mature T lymphocytes.

Animals↗
PubMed1995

Rapid acquisition of dendritic spines by visual thalamic neurons after blockade of N-methyl-D-aspartate receptors.

N-Methyl-D-aspartate (NMDA) receptors play an important role in the development of retinal axon arbors in the mammalian lateral geniculate nucleus (LGN). We investigated whether blockade of NMDA receptors in vivo or in vitro affects the dendritic development of LGN neurons during the period that retinogeniculate axons segregate into on-center and off-center sublaminae. Osmotic minipumps containing either the NMDA receptor antagonist D-2-amino-5-phosphonovaleric acid (D-APV) or saline were implanted in ferret kits at postnatal day 14. After 1 week, LGN neurons were intracellularly injected with Lucifer yellow. Infusion of D-APV in vivo led to an increase in the number of branch points and in the density of dendritic spines compared with age-matched normal or saline-treated animals. To examine the time course of spine formation, crystals of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate were placed in the LGN in brain slices from 14- to 18-day-old ferrets. Labeled LGN cell dendrites were imaged on-line in living slices by confocal microscopy, with slices maintained either in normal perfusion medium or with the addition of D-APV or NMDA to the medium. Addition of D-APV in vitro at doses specific for blocking NMDA receptors led to a > 6-fold net increase in spine density compared with control or NMDA-treated slices. Spines appeared within a few hours of NMDA receptor blockade, indicating a rapid local response by LGN cells in the absence of NMDA receptor activation. Thus, activity-dependent structural changes in postsynaptic cells act together with changes in presynaptic arbors to shape projection patterns and specific retinogeniculate connections.

2-Amino-5-phosphonovalerate↗
PubMed1995

Estimation of body fat and lean tissue distribution by dual energy X-ray absorptiometry and abdominal body fat evaluation by computed tomography in Cushing's disease.

Body composition determined by dual energy x-ray absorptiometry and the abdominal visceral fat component determined by computed tomographic scanning were examined in women with Cushing's disease and compared with those in obese women with the same anthropometric parameters and those in nonobese women. Patients with Cushing's had no increase in total body fat or the trunk region (android) component, but had a higher intraabdominal fat area compared to the obese subjects. The total lean tissue mass was slightly reduced in Cushing's compared to that in the obese subjects due to a significant decrease in the muscle of the legs and arms; the reduced amounts of fat and lean tissue masses in the arms were the most significant findings in hypercortisolism. The body mineral and bone calcium contents were slightly reduced in Cushing's compared to those in the obese controls. Thus, although obese subjects had more fat and lean tissue and mineral masses than their normal weight counterparts, the Cushing's patients, with the same total fat mass and its components (except in the arms) as obese individuals, present total lean tissue and fractions, including body mineral and bone calcium contents, similar to those in nonobese subjects due to the depletion of the protein depots, as seen in hypercortisolism.

Absorptiometry, Photon↗
PubMed1995

Phagocytosis of apoptotic bodies by liver endothelial cells.

Using electron microscopy and cytofluorimetry we studied the role of carbohydrate-specific recognition systems in the interaction of apoptotic bodies with normal and interleukin 1-activated sinusoidal endothelial cells. Microfluorimetric observation of liver tissue sections revealed octadecylrhodamine B-labelled apoptotic body binding to the sinusoidal wall of mouse liver, when they were injected intraportally. Plate-scanning cytofluorimetry demonstrated that about 20-25% of Acridine Orange-labelled apoptotic bodies could adhere specifically to cultured endothelial cells after 15 minutes of incubation. Adhesion increased to 30% when the cells were incubated for 60 minutes. Using a mixture of galactose/N-acetylglucosamine/mannose as competition solution apoptotic body adhesion was significantly reduced especially after longer times of incubation, when the percentage of inhibition reached 50%. Following 4 hours exposure of liver endothelial cells to 1 ng/ml human recombinant interleukin-1 beta adhesion markedly increased after 60 minutes of incubation, whereas the co-incubation of interleukin-1 beta with the inhibitors brings down the adhesion to basal values obtained in controls. Electron microscopic observation of the adhesion process showed that the number of endothelial cells binding apoptotic bodies gradually increased from low to high values with time. After 60 minutes of incubation, the majority of apoptotic bodies were seen inside phagosomes and only a few remained at the cell surface. Liver endothelial cells bound and endocytosed apoptotic bodies through carbohydrate-specific receptors. Moreover, this scavenger action was interleukin-1 enhanced, thus suggesting its possible activation during inflammatory and immune processes.

Animals↗
PubMed1995

[Randomized comparative study of induced labor with oxytocin and misoprostol in prolonged pregnancies].

UNLABELLED: The efficacy of oxytocin for induction of labor at 41 to 42 weeks of gestational age, was compared with misoprostol (a PGE1 analog) in 100 pregnant women. Half a tablet containing 100 mcg of misoprostol was placed intravaginally in 50 women and the remaining 50 received iv oxytocin (2-32 mU/min). The obstetric features were similar in both groups. Delivery within 24 hours was achieved in 88% of the patients induced with misoprostol and in 72% with oxytocin (p < 0.05). The mean time from induction to delivery was similar, 10.3-10.9 hrs (p NS) and no differences in obstetric and perinatal outcomes were found. Polisystoly with misoprostol, was the only side effect observed in the study (11.4%), and was without neonatal consequences. CONCLUSION: intravaginal misoprostol (100 mcg), is more effective, cheaper and less stressful than IVI oxytocin in post term inductions. The safety profiles of both were similar.

Abortifacient Agents, Nonsteroidal↗
PubMed1994

Pattern and load of spontaneous liver metastasis dependent on host immune status studied with a lacZ transduced lymphoma.

Detection of disseminated leukemia within organ is often very difficult and might lead to underestimation of the metastatic load. Therefore, we transduced the mouse ESb T lymphoma with the bacterial lacZ gene, which allowed us to follow metastasis at the single cell level. Intradermal primary tumor growth of lacZ transduced ESbL cells (L-CI.5s) comprised three phases: an initial expansion phase (day 0 to 9, increase from 0 to 8 mm, tumor diameter), a plateau phase (day 9 to 20, constant diameter of 8 mm and necrosis), and a second expansion phase (day 20 to 30, increase from 8 to 15 mm). Liver metastasis could already be detected at day 3 and maintained at that level until day 23, where exponential expansion started. A distinct mosaic-like metastasis pattern developed, with preferential localization of tumor cells to the periportal areas of the liver in immunocompetent animals. In contrast, in immunocompromised mice, primary tumor growth and metastasis were progressive and metastasis appeared as diffuse or focal/clustered. Healthy animals surviving a tumor cell inoculum of a variant cell ESbL-CI.5) with a reduced metastatic potential carried low levels of possibly dormant tumor cells in the bone marrow. Thus, this study showed that host immunocompetence determines to a large extent kinetics and load of spontaneous liver metastases and even influences the pattern and localization of disseminated lymphoma cells.

Animals↗
PubMed1993

Isolation and enrichment of two sublobular compartment-specific endothelial cell subpopulations from liver sinusoids.

Similar to the well-recognized phenotypical heterogeneity of hepatocytes, in situ sublobular variations have recently been detected in the cell structure, fenestration patterns, filtrating efficiency, surface glycosylation, scavenger function and pathological responses of the sinusoidal lining endothelium. However, unlike other liver cell populations, until now no endothelial cell subpopulations had been isolated or defined with clarity, much less with sublobular/acinar zone-related differential properties. On the basis of our previous studies showing that periportal segments of mouse liver sinusoids express a significantly higher number of wheat germ agglutinin-binding sites than do perivenous ones, we used this differential feature for in vitro labeling of the specific sublobular derivation of isolated sinusoidal lining endothelial cells to correlate their original lobular position with other features determined on flow cytometry, centrifugal elutriation, discontinuous arabinogalactan density gradients and electron microscopy. Our results revealed additional heterogeneous properties whose association with high or low wheat germ agglutinin-binding capacity made it possible to define in vitro two dominant endothelial cell subpopulations that appear similar to the differential features in the periportal and perivenous sinusoidal segments. Type 1 endothelial cells had low forward angle light scatter and high integrated side scatter, low cytoplasmic porosity index (12% +/- 5%) and high wheat germ agglutinin-binding efficiency (160 +/- 35 fluorescence intensity units/cell size); these findings are similar to what was observed in situ in the periportal sinusoidal endothelium. On the other hand, type 2 endothelial cells, with high forward angle light scatter and low integrated side scatter, had a high cytoplasmic porosity index (25% +/- 8%) and low wheat germ agglutinin-binding efficiency (60 +/- 15 fluorescence intensity units/cell size), findings similar to in situ observations of the perivenous sinusoidal lining endothelium. Moreover, these physical and morphological differences entail different cell sedimentation behaviors: type 1 endothelial cell sedimented at high centrifugal elutriation counterflow rates (23 to 37 ml/min) and high arabinogalactan density gradient levels (10% to 15%), whereas type 2 endothelial cell sedimented at low counterflow rates (18 to 23 ml/min) and low density levels (6% to 10%). The combination of these separation procedures made it possible to isolate a 90%-enriched type 1 endothelial cell population in the 12% to 15% interphase of the 23 and 37 ml/min elutriation flow rates and a 75%-enriched type 2 endothelial cell population in the 6% to 10% interphase of the 18 and 23 ml/min flow rates.

Animals↗
PubMed1991

Differences in the lectin-binding patterns of the periportal and perivenous endothelial domains in the liver sinusoids.

We have studied the distribution patterns of carbohydrate terminals on the endothelial surface of the mouse liver microvasculature. For this purpose, a wide battery of FITC lectins specific to glucose, mannose, galactose, fucose, N-acetyl-neuraminic acid, N-acetyl-galactosamine and N-acetyl-glucosamine residues were incubated on liver cryostat sections or intraportally perfused under physiological conditions. All the resulting hepatic sections were examined under fluorescent microscopy and confocal laser scanning microscopy. With the exception of N-acetyl-galactosamine- and fucose-binding lectins, all the perfused lectins specifically bound to the microvascular wall as confirmed by blocking methods using their corresponding sugars. A wide range of binding was, however, observed among the lectins, and the latter were classified into four groups according to their affinities for the different segments of the hepatic microvasculature: (a) equal affinity for all segments (concanavalin A); (b) different affinities depending on acinar zone (wheat germ agglutinin, Ricinus communis toxin, phytohemagglutinin E, Erythrina cristagalli agglutinin and Pisum sativum agglutinin); (c) preferential binding to the sinusoidal network (Lathyrus odoratus, phytohemagglutinin); and (d) lectins that fail to bind to the hepatic microvasculature (N-acetyl-galactosamine- and fucose-binding lectins). Sinusoidal segment walls in acinar zone 1 expressed a higher concentration of certain lectin-binding carbohydrate residues (N-acetyl-neuraminic acid, N-acetyl-galactosamine, galactose, mannose and glucose) than in acinar zone 3. The labeling patterns obtained through the incubation of liver sections or through in vivo perfusion with the different lectins did not always coincide. Only concanavalin A, wheat germ agglutinin and phytohemagglutinin E lectins proved to be concordant (i.e., they produced identical labeling patterns in both procedures).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Nucleotide sequence of the glnA-glnL intercistronic region of Escherichia coli.

The nucleotide (nt) sequence of a 682-bp fragment containing the 3' end of the glnA gene, the region between the glnA and glnL genes, and the 5' end of the glnL gene from Escherichia coli was determined. This segment contains the region coding for the last 107 amino acids (aa) of glutamine synthetase, including the adenylylation site of this enzyme. The analysis of this sequence revealed two REP sequences, a Rho-independent terminator, the putative glnL promoter and the possible binding site for the glnG product, NRI.

Amino Acid Sequence↗
PubMed1984

cis-Dominant, glutamine synthetase constitutive mutations of Escherichia coli independent of activation by the glnG and glnF products.

Mutants resistant to 80 microM L-methionine-DL-sulfoximine (MS) were isolated on glucose-minimal 15 mM NH4+ medium plates from Escherichia coli cells which were hypersensitive to this concentration of the analogue by virtue of their harboring glnG mutations. MS-resistant mutants derived from strain MX902 carried, in addition to its glnG74 ::Tn5 allele, mutations tightly linked to glnA, as shown by P1-mediated transduction experiments. One particular allele, gln-76, which suppressed the MS-sensitivity conferred by glnG74 ::Tn5 but not its Ntr- phenotype (inability to transport and utilize compounds such as arginine or proline as the only nitrogen sources), was shown to allow constitutive expression of glutamine synthetase in the absence not only of a functional glnG product but also of a functional glnF product. This behavior was found to be cis-dominant in complementation experiments with F'14 merogenotes . In an otherwise wild-type genetic background as in MX929 (gln-76 glnA+ glnL+ glnG+ glnF +), however, normal activation, mediated by the glnG and glnF products was preferred over that mediated by gln-76.

Bacterial Proteins↗
PubMed1980

Cloning and physical mapping of the glnA gene of Escherichia coli K-12.

We have isolated part of the glnA region of Escherichia coli K-12 as a 6.4 Md DNA fragment on the ColE1 hybrid plasmid pACR1. DNA fragments from pACR1 obtained by cleavage with certain restriction endonucleases were subcloned into the pBR322 cloning vehicle. Recognition sites for the endonucleases BamHI, SmaI, BglII, and EcoRI were localized inside the glnA gene sequence. De novo synthesized polypeptides, employing minicells that carried some of these plasmids, allowed us to determine the direction of transcription of the glnA gene relative to an adjoining gene that codes for a 65 000 dalton protein.

Bacteriocin Plasmids↗
PubMed1978

Comparative study on the activity, pH optimum and thermal stability of some glycosidases and acid phosphatase from pig and mule leukocytes.

1. alpha-D-mannosidase, beta-D-galactosidase, alpha-L-fucosidase, beta-N-acetylgalactosaminidase, alpha-D-glucosidase and acid phosphatase activities were studied in circulating blood leukocytes from Sus scropha var. domestica L. (pig) and Equus asinus x caballus (mule) by spectrophotometric procedures using p-nitrophenyl derivatives as substrates and three different buffer solutions. 2. The highest specific activity corresponds to acid phosphatase. The specific activities of the glycosidases, all relatively close together in all cases, were low in comparison with that of phosphatase. 3. Generally, each of the above-mentioned enzymes shows one common peak for the pH optimum between 3.5 and 6.0, except alpha-D-glucosidase, which shows two peaks. 4. The pH optima values are generally similar in three buffer solutions employed. 5. Specific activities of the studied enzymes show a parallelism in leukocytes from both pig and mule. Also, this parallelism is observed in their pH optima values. 6. Thermal stability of alpha-D-mannosidase is high whereas that of acid phosphatase is low, in both materials. For other enzymes, differences in the thermal stability was observed according to their source.

Acid Phosphatase↗
PubMed1978

Isolation and chemical composition of the leucocytes from donkey, horse, mule and pig.

1. Leucocytes from the blood of adult and young donkeys (Equus asinus L.), adult horses (Equus caballus L.), adult mules (Equus asinus x Equus caballus) and adult pigs (Sus scrofa L.) were obtained in a high degree of purity (99.9%) using Na2-EDTA-dextrans mixtures. 2. Sialic acids were released, purified, identified and determined from both non-delipided and delipided leucocytes. 3. N-glycolylneuraminic was the predominant sialic acid. N-acetylneuraminic acid and N,O-diacetyl-neuraminic acid were also found in all materials. Except in pig, other unidentified sialic acid(s) were also detected. 4. The concentration of total sialic acids (microgram/mg protein) is different according to the species, and in donkey species according to the age. 5. Galactose, glucose, mannose, fucose and (in a less amount) ribose were determined. Their total content is about 2-3-fold that of hexosamines. 6. There is a higher cholesterol content in adult donkey leucocytes than in those of young ones. 7. Total lipids, cholesterol or phospholipid contents are similar among the leucocytes of the above-mentioned species. 8. The similarities are marked in the electrophoresis patterns of proteins and glycoproteins for the donkey, mule and horse samples. 9. The molecular weights for leucocytes proteins were estimated as ranging between 230,000 and 20,000; and for the main protein bands, between 120,000 and 22,000.

Animals↗
PubMed1978

[Comparative study of six methods for lymphocyte isolation from several mammalian sources and determination of their carbohydrate composition (author's transl)].

The present paper deals with a comparative study on six methods for isolation of peripheral blood lymphocytes from various mammalian sources: Bos taurus L. (adult cow), Equus caballus L. (adult horse), Equus asinus L. (adult and young donkeys) and Sus scropha L. (adult pig). The following systems were used: a) Filtration through sand columns (a modification of Blaszczyszyn's method); b) Sodium metrizoate and "Ficoll 400" c) "Lymphoprep"; d) "Urovison" and dextran T150 (a modification of GILI et al.'s method); e) "Urografin" and dextran T150; f) "Ficoll-Paque". The final preparation of lymphocytes obtained by "Urovison" and dextran T150 (d = 1.081) procedure was free from platelets and erythrocytes; lymphocytes degree of purity was found to be 98%. The sialic acids, hexoses and hexosamines contents were determined.

Animals↗