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Biomedical subjects

L Carrasco

Publications and source records attributed to L Carrasco.

At least 91 records · Page 5Linked to original sources
PubMed1998

Myocarditis associated with Theileria spp. in calves.

Two 3-week-old Limousin x Retinta calves from extensive farms in the south of Spain died suddenly. Microscopic examination of the spleen and lymph nodes revealed an intense lymphoid proliferation; the germinal centres were prominent and were surrounded by areas of haemorrhage. Parasitic forms, similar to Koch's bodies, were observed in only a small number of lymphoid cells and macrophages. In the myocardium a severe myocarditis was found. Inflammatory cells were in close contact with damaged myocytes, sometimes surrounding fragments of cells. In some areas the vessels had intraluminal deposits of fibrin. The endothelial damage and thrombosis may have contributed to the damage observed in the myocytes, together with the cytotoxic mechanism associated with the presence of an intense inflammatory infiltrate. The active multiplication of parasites may be due to the age of the animals and their breed, since they are not an autochthonous breed and present low resistance to infection.

Animals↗
PubMed1998

Effect of nitric oxide on poliovirus infection of two human cell lines.

The role of nitric oxide after poliovirus infection of the human HeLa (carcinoma) and U937 (promonocytic) cell lines has been analyzed. Both types of cells produced detectable levels of nitric oxide after poliovirus infection. However, this production was not sufficient to limit viral productivity. On the other hand, pretreatment with the nitric oxide donor glycerine trinitrate lengthened the course of poliovirus infection.

HeLa Cells↗
PubMed1998

Identification of regions of poliovirus 2BC protein that are involved in cytotoxicity.

The expression of poliovirus 2BC protein in yeast and mammalian cells leads to a number of metabolic and morphological alterations, such as growth inhibition, intracellular membrane proliferation, blockade of the exocytic pathway, and enhanced membrane permeability. Yeast cells that express poliovirus 2BC in an inducible manner were used to identify the regions of 2BC implicated in the modifications of these cellular functions. Several 2BC deletion mutants were generated to define the minimal portion of 2BC required to alter these activities. Additional deletion mutants that were obtained by random mutagenesis followed by selection in yeast cells provided new insights into the structure and mechanism of action of 2BC. The activity responsible for membrane proliferation is located in 2C, while the activities responsible for membrane permeabilization and inhibition of the exocytic pathway are located in 2B. Several regions of 2B and 2C required for the different functions of 2BC were identified. Thus, the integrity of the N termini of both 2B and 2C is necessary for 2BC-induced cytotoxicity. It is also possible to separate the different cellular alterations provoked by 2BC by the use of several 2BC variants. Deletion of amino acids 52 to 65 in 2B generates a 2BC deletion variant, 2bC deltaAvrII, that still blocks yeast growth but is unable to enhance membrane permeability or to inhibit the exocytic pathway. On the other hand, 2Bcl28*.32b and 2Bcl28*.3c, which contain only 73 and 77 amino acids of 2B, interfere with yeast division and enhance membrane permeability but affect the exocytic pathway only weakly and do not induce membrane proliferation. Our findings indicate that Saccharomyces cerevisiae represents a useful model system to analyze the functions of poliovirus 2BC and show the feasibility of separating the activities assigned to this protein.

Amino Acid Sequence↗
PubMed1998

Ultrastructural pathology of the bone marrow in pigs inoculated with a moderately virulent strain (DR'78) of African swine fever virus.

Interpretation of changes in bone marrow during infectious processes is quite complex. This paper reports bone marrow lesions observed in pigs inoculated with a moderately virulent ASF virus strain and studies their relationship to the pathogenesis of the disease. In this work, we have carried out the structural and ultrastructural study of the bone marrow of 14 Large White x Landrace pigs that were inoculated by the intramuscular route with 10(5) 50% hemodsorbing doses (HAD50) of the Dominican Republic'78 ASF virus strain. The inoculated pigs were killed at 3, 5, 7, 9, 11, 13, 15 and 17 days postinjection. Analysis of cells and structures belonging to the two main bone-marrow compartments, the hematopoietic cells and the hematopoietic micro-environment, showed that after inoculation with a moderately virulent strain, the most significant changes occurred in macrophages and megakaryocytes, consisting in virus replication in these cell populations and apoptosis of megakaryocytes, related with the sudden and transitory thrombocytopenia detected in the subacute ASF.

African Swine Fever↗
PubMed1998

Pancreaticoduodenectomy due to cancer in the elderly.

The elderly have traditionally been excluded from pancreaticoduodenectomy due to the high morbimortality of this procedure. Six cases of pancreaticoduodenectomy) 5 cephalic and 1 total) for periampullary tumors in patients over 70 are reported. There was no mortality. We conclude that, in selected cases, pancreaticoduodenectomy can be performed safely in the elderly.

Age Factors↗
PubMed1997

Membrane permeability changes induced in Escherichia coli by the SH protein of human respiratory syncytial virus.

The small hydrophobic (SH) protein of human respiratory syncytial virus (HRSV) has been efficiently expressed in Escherichia coli. In analogy to small hydrophobic proteins encoded by other RNA viruses, membrane permeability changes to low-molecular-weight compounds were detected in bacteria expressing HRSV SH protein. These changes implied, at least, the entry of both the protein synthesis inhibitor hygromycin B and the beta-galactoside substrate o-nitrophenyl-beta-d-galactopyranoside and the exit of preloaded [3H]uridine from bacterial cells. Site-directed mutagenesis indicated that the C-terminal end of SH is needed for induction of membrane permeability changes. In addition, amino acid substitution at residue 32 (Ile to Lys) abolished that activity. This was correlated with a drastic increase in SH electrophoretic mobility and a decrease of the predicted values of alpha-helix for all residues of the SH transmembrane domain. Other sequence changes have either partial effect or no effect on the membrane permeability changes induced by the SH protein. However, none of the mutations abrogated the association of SH protein with bacterial membranes, indicating that incorporation of SH protein to membranes is not sufficient to induce the observed changes. Membrane permeability changes then might provide a useful test for the identification of key amino acid residues in this unique HRSV gene product.

Amino Acid Sequence↗
PubMed1997

Cleavage of p220 by purified poliovirus 2A(pro) in cell-free systems: effects on translation of capped and uncapped mRNAs.

Poliovirus protease 2A(pro) has been obtained in soluble form as a fusion protein with maltose binding protein (MBP). Addition of MBP-2A(pro) to rabbit reticulocyte cell-free systems gives rise to efficient cleavage of the initiation factor of translation p220 (eIF-4G). Translation of capped mRNA encoding the influenza virus NP protein is severely impaired in lysates in which p220 has been proteolytically cleaved. This inhibition is dependent on the concentration of mRNA added to the lysate. Thus, increasing the concentrations of mRNA substantially overcomes the blockade of NP synthesis after p220 cleavage. Notably, translation of uncapped NP mRNA is also compromised in p220-deficient rabbit reticulocyte lysates, suggesting that p220 participates in the translation of both capped and uncapped NP mRNAs. The effects of p220 proteolysis by poliovirus 2A(pro) have also been assayed on luciferase mRNA translation. Three types of mRNAs encoding for luciferase have been examined: capped, uncapped, and mRNA bearing the poliovirus 5' leader region (leader luc mRNA). Synthesis of luciferase directed by any of these mRNAs was inhibited after cleavage of p220 in rabbit reticulocyte lysates. Interestingly, supplementation of the lysate with HeLa cell extracts stimulates leader luc mRNA translation by poliovirus 2A(pro). These results indicate that activation of translation of mRNAs bearing the poliovirus leader region promoted by this poliovirus protease requires a factor present in HeLa cell extracts. These findings agree well with recent experiments implicating p220 not only in protein synthesis directed by capped mRNAs but also in the translation of naturally uncapped mRNAs.

Carrier Proteins↗
PubMed1997

The N-terminal Arg-rich region of human immunodeficiency virus types 1 and 2 and simian immunodeficiency virus Nef is involved in RNA binding.

Comparison of the amino acid sequences of human immunodeficiency virus (HIV) Nef protein and several RNA-binding proteins shows similarities in some regions of these proteins. Thus, poliovirus protein 2C, an RNA-binding protein, shares with Nef the sequence YXQQ...MDD...DXXD. In addition, both proteins contain an Arg-rich motif that, in the case of poliovirus 2C, is involved in RNA-binding activity. Moreover, the RNA-binding, anti-terminator N proteins of lambda, phi21 and P22 phages show sequence similarities with HIV Nef at the Arg-rich motif. To assess the significance of this motif, native and deletion variants of Nef protein were assayed for RNA-binding activity. The N-terminal 35 amino acids of HIV-1 Nef that comprise the Arg-rich motif are sufficient for RNA binding. Point mutations engineered at the Arg-rich motif of HIV-1 Nef revealed that basic amino acid residues are essential for RNA-binding activity. The Nef proteins from HIV-2 and SIV can also interact with RNA, while the same proteins with the N-terminal Arg-rich domain truncated fail to interact with RNA. These findings indicate that all three Nef proteins from HIV-1, HIV-2 and simian immunodeficiency virus belong to the RNA-binding family of proteins. The three proteins contain an Arg-rich region at the N-terminus which is necessary to interact with RNA.

Amino Acid Sequence↗
PubMed1997

The yeast Saccharomyces cerevisiae as a genetic system for obtaining variants of poliovirus protease 2A.

The inducible expression of poliovirus protease 2A (2Apro) blocks the growth of Saccharomyces cerevisiae. A number of yeast colonies that grow after 2Apro induction have been isolated. The majority of these clones express 2Apro to control levels, suggesting that their ability to divide is not due to the loss of 2Apro gene inducibility. The sequences of the 2Apro genes isolated from 22 clones were determined. Most of the 2Apro sequences from these colonies contain point mutations in the poliovirus protease. The different variant protease sequences were transferred to an infectious poliovirus cDNA clone. Translation of genomic RNA obtained from these poliovirus mutants in cell-free systems revealed that some of them had defects in their ability to cleave P1-2A in cis. In addition, several of these variants cleaved the translation initiation factor eIF-4G inefficiently. Transfection of the RNA generated from the full-length poliovirus genomes mutated in 2Apro yielded five viable polioviruses with a small plaque phenotype. These five polioviruses efficiently cleaved p220 but showed defects in viral protein synthesis, transactivation of a leader-luciferase mRNA, and 3CD cleavage to 3C' and 3D'. All 2Apro mutant sequences, including those that did not yield viable viruses, were cloned in pTM1 vector under a T7 promoter. Only the 2Apro variants that have activity to cleave 3CD produced viable poliovirus. Our findings indicate that S. cerevisiae represents a useful system for obtaining poliovirus 2Apro variants that may provide further insight into the role of this protease during the poliovirus replication cycle.

Amino Acid Sequence↗
PubMed1997

Permeabilization of mammalian cells to proteins: poliovirus 2A(pro) as a probe to analyze entry of proteins into cells.

Two hybrid protein molecules containing the poliovirus protease 2A (MBP-2A(pro)) (maltose-binding protein-2A(pro) and MBP-Pseudomonas exotoxin A-2A(pro)) have been constructed and purified. Both hybrid proteins efficiently cleave the translation initiation factor eIF-4G when they are co-internalized into cells with adenovirus particles. Almost no intact eIF-4G can be detected in cells incubated with these proteins following this method. Reovirus infectious subviral particles also promote the delivery of MBP-2A(pro) into cells, although less efficiently than adenovirus particles. None of the other methods employed to permeabilize cells to MBP-2A(pro) achieves the degree of eIF-4G cleavage observed with adenovirus particles. By comparison about 30% of cells electroporated with MBP-2A(pro) still contain intact eIF-4G. More drastic electroporation conditions lead to a significant decrease of cell survival. Osmotic lysis of pinocytic vesicles resulted in 30% of the eIF-4G being cleaved in cells treated in suspension. Delivery of MBP-2A(pro) by pH-sensitive liposomes leads to poor hydrolysis of eIF-4G. Taken together our results indicate that permeabilization of cells with adenovirus particles is the most efficient method for introducing MBP-2A(pro) into cells.

ADP Ribose Transferases↗
PubMed1997

Entry of Semliki forest virus into cells: effects of concanamycin A and nigericin on viral membrane fusion and infection.

Semliki forest virus (SFV) was biosynthetically labeled with pyrene phospholipids and used to investigate two alternative routes of entry of SFV into BHK-21 cells: (1) receptor-mediated endocytosis followed by fusion of the viral envelope with the endosomal membrane and (2) direct fusion of SFV with the plasma membrane induced by low pH treatment. The selective inhibitor of the vacuolar proton-ATPase, concanamycin A, abolished fusion and subsequent infection only when the virus utilized the endocytic route to enter cells. The inhibitory effect of this macrolide antibiotic was bypassed by low pH treatment of cells. However, the ionophore nigericin was inhibitory irrespective of the route used by the virus to infect cells, suggesting the necessity of a transmembrane pH gradient for the entry process. According to our results, concanamycin A emerges as a suitable tool for selectively investigating the involvement of endosomal function in animal virus entry.

Animals↗
PubMed1997

Equine pulmonary mycosis due to Aspergillus niger and Rhizopus stolonifer.

Invasive pulmonary mycosis caused by Aspergillus niger and Rhizopus stolonifer is reported in a 2-year-old horse, one of three that died after being housed in a disused, uncleaned stable. Lesions were characterized by thrombosis of the blood vessels with haemorrhage and tissue necrosis. Fungal hyphae were observed both in thrombosed vessels and in adjacent necrotic tissue. In culture media inoculated with lung samples and samples from the bedding hay, two types of colony were recorded and identified as A. niger and R. stolonifer. This study is the first description of equine pulmonary mucormycosis and of A. niger as an aetiological agent in the horse.

Animals↗
PubMed1997

Focal mesangial-sclerosing glomerulonephritis and acute-spontaneous infectious canine hepatitis: structural, immunohistochemical and subcellular studies.

The glomerular alterations observed in a dog with acute spontaneous infectious canine hepatitis (ICH) are described. Histologic changes of the glomeruli were enlargement of the mesangium with presence of intranuclear inclusion bodies and without proliferation of mesangial cells. Electron microscopy revealed adenovirus replication sites in glomerular mesangial cells and in endothelial cells of glomerular capillaries, as well as a focal mesangial-sclerosing glomerulonephritis associated with electron dense deposits which were closely related with extracellular ICH viral particles and immunohistochemically reactive for immunoglobulin (Ig) G, IgA, IgM and C3c complement components.

Acute Disease↗
PubMed1997

High-resolution ultrasound associated with aspiration biopsy in the follow-up of patients with differentiated thyroid cancer.

OBJECTIVE: To assess the value of ultrasound in the follow-up of patients undergoing surgery for differentiated thyroid carcinoma. SUBJECTS: The study included 89 patients (70 women and 19 men) with differentiated thyroid carcinoma (76 papillary and 13 follicular cancer). METHODS: High-frequency ultrasound (US) was used in the evaluation of 89 subjects who underwent surgery for thyroid carcinoma. Fine-needle aspiration was performed in cases with positive US. In addition, determinations of thyroglobulin (Tgb) in serum, scintigraphy with (131)I, and cervical palpation were evaluated. We determined sensitivity, specificity, and overall accuracy for each of these diagnostic methods. RESULTS: Ultrasonography was positive in 22 subjects, 16 in the nodal area and 6 in the thyroid bed. Twenty-two subjects received fine-needle aspiration with US control; 13 (59%) of 22 were positive for cancer. The results of the US for detecting neoplastic disease showed a sensitivity of 65%, specificity of 86%, and overall accuracy of 82%. The overall accuracy for scintigraphy was 88% and for Tgb, 91%. CONCLUSION: We concluded that US can be included in the follow-up protocol for patients undergoing surgery for differentiated cancer of thyroid, as a valuable tool to localize the recurrence. This technique is particularly useful in the evaluation of patients who are found to have elevated Tgb levels.

Adenocarcinoma, Follicular↗
PubMed1997

Ultrastructural findings in natural canine hepatozoonosis.

The ultrastructure of several stages of Hepatozoon canis found in dogs with clinically and histologically diagnosed infections was determined using transmission electron microscopy. Merozoites, macro- and microschizonts and gamonts were found in spleen, liver, kidneys and lungs. Macro- and micromeronts were characterized by their size and by the presence of intracytoplasmic amylopectin granules. Gamonts, which provide the basis for clinical diagnosis of the disease, were observed within mononuclear cells (monocytes/macrophages); they were butterfly-shaped and exhibited varying electron densities. On the basis of the microscopic observations reported here, it is clear that the life cycle of H. canis is a complex one.

Animals↗