Biomedical subjects

M A Alonso

Publications and source records attributed to M A Alonso.

At least 73 records · Page 4Linked to original sources

PubMed1992

The occurrence of Salmonella serotypes in marine recreational waters of Valencia, Spain.

Salmonellae serotypes were studied in order to know their prevalence in marine recreational areas of Valencia. Two hundred eight strains were isolated. The strains belonging to serogroups B, C, D, E and G. The serotyping yielded twenty one different serotypes. The most frequent salmonellae serotypes were S. anatum and S. bredeney. Our results were compared with those reported by other authors in Spain.

Animals↗

PubMed1991

Cell type determines the relative proportions of (-) and (+) strand RNA during poliovirus replication.

To examine the influence of the host cell type on poliovirus RNA synthesis we compared the levels of (-) and (+) strand poliovirus RNA during infection of epithelial (HeLa and HEp-2), leukocytic (U-937, HL-60 and K-562) and nerve (IMR-32) cells. The levels of (-) strand RNA were higher in IMR-32, U-937, K-562 or HL-60 cells than those in HeLa or HEp-2 cells. By contrast, (+) strand RNA content was greater in HeLa or HEp-2 cells. Although significant levels of (+) strand RNA were detected in U-937, K-562 and HL-60 cells, no viral protein synthesis was detected by polyacrylamide gel electrophoretic analysis of metabolically labelled proteins. The molar ratio of poliovirus (-) and (+) RNAs was 2-3 fold higher in IMR-32, U-937 and K-562 cells than in HeLa or HL-60 cells and 5-6 fold higher than in HEp-2 cells. Differentiation of HL-60 cells with a variety of inducers produced differential effects on poliovirus (-) and (+) RNA content and modified the molar ratio of (-)/(+) strand RNAs. These findings indicate that host cell components play a critical role in the regulation of the amount of poliovirus (-) and (+) strand RNAs synthesized during infection.

Cell Line↗

PubMed1991

Effects of poliovirus replication on undifferentiated and differentiated monocytic U937 cells: comparative studies with human macrophages.

Poliovirus infection of either undifferentiated or differentiated U937 cells produced a decrease in the percentage of cells positive for the surface expression of the CD4, CD11c, CD14, or 8E11 antigens. The number of 4F2 surface molecules per cell increased in infected normal U937 cells, but was unaffected in differentiated cells. The level of O2- production in infected differentiated U937 cells was approximately 50% of that found when not infected. Finally, poliovirus RNA levels and infectious particle production were similar in either cell type.

Antigens, CD↗

PubMed1990

[Ewing's sarcoma in childhood: the therapeutic results].

We evaluated 27 patients with localized Ewing's sarcoma diagnosed in our hospital. In 9 cases the lesion was centroaxial and in 18 it was located in the limbs. The treatment consisted of: 1) chemotherapy with 6 drugs (vincristine, cyclophosphamide, methotrexate, bleomycin, actinomycin D and adriamycin) during one year in all patients, and 2) radiotherapy in 24 patients with a dose of 30-40 Gy in the involved bone with a 10-15 Gy superimpression on the tumor area. In 7 patients nonradical surgery followed by radiotherapy was carried out. In 2 cases with rib localization and in 1 with tibial localization and large bone destruction radical surgery was undertaken. One patient had a local relapse (femur) and six had distant metastases. The remaining 20 patients (74%) are alive and without evidence of disease, with follow-up periods ranging from 17 to 80 months (median 38 months). The estimated actuarial free from disease survival was 64% after 58 months. Sequelae of therapy were evaluated in the 12 survivors with primary localization in the limbs. In 7 cases (58%) there were minimal sequelae, in two moderate and in three there was a severe limitation, but amputation was not required in any patient. The results of our study show that the local control of the disease can be achieved in most cases without radical surgery and with an acceptable limb function.

Adolescent↗

PubMed1990

BiP expression is not increased by the accumulation of PiZ alpha 1-antitrypsin in the endoplasmic reticulum.

PiZ, a mutant human alpha 1-antitrypsin, is associated with liver and pulmonary disease and is characterized by defective secretion and accumulation of the protein in the endoplasmic reticulum. We tested the hypothesis that BiP (a protein that binds newly synthesized protein in the endoplasmic reticulum, prevents secretion of incorrectly folded protein, and solubilizes protein aggregates), could play a part in the retention of PiZ alpha 1-antitrypsin in the endoplasmic reticulum. Subcellular fractions from PiM (normal) and PiZ livers were prepared and analyzed by immunoblotting. No increase of BiP was detected in the PiZ liver. In addition, when total RNA from the same livers were analyzed by slot and Northern blot hybridization, no difference was found in the level of BiP mRNA between PiM and PiZ livers. Similar results were found in clones of CHO and MDCK cells transfected with PiM of PiZ alpha 1-antitrypsin cDNAs. These results indicate that BiP does not play a part in the retention of PiZ alpha 1-antitrypsin and suggest that PiZ protein is not misfolded.

Animals↗

PubMed1990

Synergy of tumor necrosis factor with protein kinase C activators on T cell activation.

The ability of tumor necrosis factor (TNF)-alpha to activate T lymphocytes in combination with other stimuli has been studied. TNF was strongly co-mitogenic with low doses of anti-CD3 antibodies or phorbol esters (those which are strong activators of protein kinase C, PKC) but poorly with phytohemagglutinin or concanavalin A. No synergism was seen with the calcium ionophore A23187. TNF was co-mitogenic with several phorbol esters known to activate PKC but was uneffective with inactive phorbol esters such as methyl-phorbol 12-myristate 13-acetate. Furthermore, H-7 a known inhibitor of PKC, inhibited the proliferative response of T cells induced by esters plus TNF. This effect took place at low doses of TNF and was also observed with purified T lymphocytes indicating that the effect of TNF was not dependent on accessory cells. This proliferative effect of TNF was inhibited by an anti-interleukin 2 receptor (IL2R) antibody, MAR 108, which blocks IL2 binding to its receptor. Although PKC activation induced CD25 (IL2R) expression but very little IL2 synthesis, TNF did not synergize by augmenting the synthesis of this lymphokine in peripheral blood lymphocytes stimulated with phorbol esters. By contrast, TNF strongly increased the membrane level of CD25 and to a lesser extent that of the activation antigen, 4F2, over the levels already induced by phorbol esters on T cells. More interestingly, TNF significantly increased the number of high-affinity IL2R on purified T cells in the presence of phorbol 12,13-dibutyrate. Our results indicate that TNF is co-mitogenic with those stimuli which strongly activate PKC and suggest that TNF may play a role on T cell activation increasing the number of effective IL2/IL2R interactions when these are limiting.

Antigens, CD↗

PubMed1990

Role of soluble cytokines on the restricted replication of poliovirus in the monocytic U937 cell line.

Poliovirus infection of monocytic U937 cells is characterized by an incomplete shut-off of host protein synthesis occurring at late times postinfection and by a slow accumulation of viral mRNA which is not efficiently translated. By using exogenous cytokines, neutralizing antibodies to different cytokines and specific cDNA probes, we have studied the involvement of soluble factors on the restricted replication of poliovirus in U937 cells. Our results show that both IFN-gamma and TNF-alpha mRNA levels increased in U937 cells upon poliovirus infection. Moreover, treatment of infected U937 cells with antibodies to IFN-gamma induced a remarkably stronger shut-off of host protein synthesis than that found in control cultures. A role of IFN-gamma in protecting the infected cells from an extensive blockade of host mRNA translation is proposed.

Biological Factors↗

PubMed1989

Restriction of poliovirus RNA translation in a human monocytic cell line.

The infection of the human monocytic cell line U-937 by poliovirus was characterized by a low level of virus production and a slow progression of the cytopathic effect. Infection took place in greater than 99% of the cells as revealed by a limiting dilution assay. No viral protein synthesis was evident in the infected U-937 cells when analyzed by polyacrylamide gel electrophoresis. However, a low level of poliovirus RNA translation was detected by immunofluorescence analysis using a mixture of polyclonal antisera against non-structural proteins. Although there was only a low level of viral protein synthesis, a gradual accumulation of viral mRNA took place in U-937 cells as revealed by RNA blot analysis. Similar results were obtained when the erythroleukemic cell line K-562 was used as a host cell for poliovirus. RNA extracted from infected U-937 cells was efficiently translated in rabbit reticulocyte extracts giving rise to a pattern of viral polypeptides similar to that detected when virion-purified RNA was the template used for the in vitro translation assay, suggesting that the poliovirus RNA present in infected U-937 cells is functional. The existence in U-937 cells of a discriminatory mechanism which differentially interferes with poliovirus RNA translation is discussed.

Cell Line↗

PubMed1989

Poliovirus infection interferes with the phorbol ester-induced differentiation of the monocytic U937 cell line.

Monocytic U937 cells can differentiate in vitro into macrophage-like cells by treatment with phorbol esters such as phorbol 12-myristate 13-acetate (PMA). We have analyzed the effect of poliovirus infection in this pathway of differentiation. Poliovirus RNA replication took place in both untreated and PMA-treated U937 cells infected before or after PMA addition, although a slight reduction in poliovirus RNA levels was observed in PMA-treated cells at late times postinfection. Total protein synthesis remained unchanged during the first 5 h of infection both in normal and PMA-treated cells. However, an inhibition on total RNA synthesis was observed early in infection. PMA-induced c-myc mRNA expression was abolished when infection took place 1 h before PMA addition but was just partially inhibited when poliovirus was added 1 h after PMA stimulation. Fluorescence flow cytometry analysis revealed that poliovirus infection induced an increase in the number of 4F2 molecules per cell in normal U937 cells and a slight decrease in the number of positive cells for the antigens CD14, CD4 and CD11c in both untreated or PMA-treated U937 cells. These findings suggest that poliovirus infection of U937 cells interferes at various levels with monocyte maturation yielding cells which are unable to undergo the complete pathway of differentiation to macrophages.

Antigens, Surface↗

PubMed1989

Human gamma interferon and tumor necrosis factor exert a synergistic blockade on the replication of herpes simplex virus.

The replication of herpes simplex virus type 1 (HSV-1) is not inhibited in either HeLa or HEp-2 cells treated with human alpha interferon (HuIFN-alpha), particularly when high multiplicities of infection are used. However, HuIFN-gamma partially inhibits HSV-1 translation in HEp-2 cells infected at low multiplicities. Under these conditions, the transcription of genes alpha 22, TK, and gamma 0 is greatly diminished. The combined addition of human tumor necrosis factor (TNF) and HuIFN-gamma to HEp-2 cells exerts a synergistic inhibition of HSV-1 translation. Cells treated with both cytokines continue synthesizing cellular proteins, even 20 h after HSV-1 infection. As little as 10 U of IFN-gamma per ml blocked HSV-1 DNA replication, provided that TNF was also present in the medium. Analyses of HSV-1 gene transcription suggest that the action of both TNF and IFN-gamma blocked a step that comes at or prior to early HSV-1 gene expression. This early step in HSV-1 replication inhibited by TNF and IFN-gamma occurs after virus attachment and entry into cells, since the internalization of radioactive HSV-1 virion particles was not blocked by the presence of the two cytokines. Therefore, we conclude that the synergistic action of TNF plus IFN-gamma affects a step in HSV-1 replication that comes after virus entry but before or at the transcription of immediate-early genes.

DNA, Viral↗

PubMed1989

Local control and survival of Ewing's sarcoma in children with radiotherapy and chemotherapy without radical surgery.

The present study was undertaken in 27 children with localized Ewing's sarcoma treated at our hospital. In 9 cases the lesion was central and in 18 peripheral. Treatment consisted of: 1) Chemotherapy with 6 drugs (vincristine, cyclophosphamide, methotrexate, bleomycin, dactinomycin and doxorubicin) for one year in all patients; 2) Radiotherapy in 24 patients at a dose of 30-40 Gy in the entire bone with a booster dose of 10-15 Gy in the lesion. In 7 patients debulking surgery was performed followed by irradiation. Two cases with tumor in expendable bones and one with extensive destructive lesion in the tibia underwent radical surgery. One patient suffered local recurrence (femur) and 6 developed distant metastases. The remaining 20 (74%) patients were alive and tumor-free after follow-up periods of 17-80 months. Among the 12 patients with extremity lesions who were alive, 7 (58%) had minimal or mild treatment sequelae, 2 moderate, and 3 severe sequelae but no amputation has been required. Our study shows that local control can be achieved in almost all patients without radical surgery and with acceptable extremity function in the majority.

Adolescent↗

PubMed1988

Assignment of the T-cell differentiation gene MAL to human chromosome 2, region cen----q13.

A cDNA clone has recently been isolated that encodes a protein expressed only in the intermediate and late stages of T-cell differentiation, termed MAL. The polypeptide deduced from the MAL cDNA sequence contains four potential transmembrane domains. We have used a panel of 28 human X rodent hybrid cell lines to assign the MAL gene to the proximal long arm of human chromosome 2, region cen----q13. The significance of the MAL map position and its relationship with other genes on chromosome 2 are discussed.

Antigens, Differentiation, T-Lymphocyte↗

PubMed1988

Selective expression of an erythroid-specific isoform of protein 4.1.

We have conducted comparative analysis of nucleotide sequences encoding erythroid and lymphoid protein 4.1 isoforms. The lymphoid protein 4.1 isoforms exhibit several nucleotide sequence motifs that appear to be either inserted into or deleted from the mRNA sequence by alternative splicing of a common mRNA precursor. One of these motifs, located within the spectrin-actin binding domain, is found only in erythroid cells and is specifically produced during erythroid cell maturation. The selective expression of the alternatively spliced mRNA during erythroid maturation implies the existence of a lineage-specific splicing mechanism whose activity is triggered by terminal maturation.

Amino Acid Sequence↗

PubMed1987

cDNA cloning and sequence of MAL, a hydrophobic protein associated with human T-cell differentiation.

We have isolated a human cDNA that is expressed in the intermediate and late stages of T-cell differentiation. The cDNA encodes a highly hydrophobic protein, termed MAL, that lacks a hydrophobic leader peptide sequence and contains four potential transmembrane domains separated by short hydrophilic segments. The predicted configuration of the MAL protein resembles the structure of integral proteins that form pores or channels in the plasma membrane and that are believed to act as transporters of water-soluble molecules and ions across the lipid bilayer. The presence of MAL mRNA in a panel of T-cell lines that express both the T-cell receptor and the T11 antigen suggests that MAL may be involved in membrane signaling in T cells activated via either T11 or T-cell receptor pathways.

Amino Acid Sequence↗

PubMed1985

The regulation of translation in reovirus-infected cells.

The regulation of translation in reovirus-infected cells has been investigated by double-infection experiments. Different cell lines were able to translate uncapped encephalomyocarditis (EMC) virus mRNA and capped vesicular stomatitis virus (VSV) mRNAs both early and late during reovirus infection. These results are not fully in agreement with a previously suggested model in which, in the early phase of reovirus infection, only capped mRNAs are translated, whereas in the late phase the cells are modified to translate exclusively uncapped mRNAs. The observations that EMC virus and VSV shut down host protein synthesis more efficiently than reovirus translation in the late phase in double-infections indicate that competition for a message-discriminatory factor may not be involved in the shut-off of host protein synthesis in these animal virus-infected cells.

Animals↗

PubMed1984

Synthesis of heat-shock proteins in HeLa cells: inhibition by virus infection.

Incubation of HeLa cells at supraoptimal temperatures induces the synthesis of a class of proteins known as heat-shock or stress-response proteins. After restoration of cells to physiological temperatures heat-shock protein synthesis continues for several hours. Normal cellular translation eventually recovers even if cells are treated with actinomycin D, indicating that neither normal cellular mRNAs nor components of the translation machinery are irreversibly modified by heat treatment. The synthesis of heat-shock proteins after poliovirus infection is more resistant to inhibition than normal host proteins. Nevertheless, their synthesis is also inhibited in infected cells, even in cells treated with human interferon under which conditions viral RNA replication and viral translation are blocked. Translation of heat-shock proteins is also resistant to hypertonic shock indicating that their mRNAs bind ribosomes with high affinity.

Electrophoresis, Polyacrylamide Gel↗

PubMed1982

Translation of capped viral mRNAs in poliovirus-infected HeLa cells.

HeLa cells doubly infected with Semliki Forest virus (SFV) and poliovirus synthesize either more poliovirus proteins or more SFV late proteins depending on the time of super-infection with poliovirus. Under some conditions, the infected cells translate uncapped poliovirus mRNA and capped 26S mRNA from SFV simultaneously, even though host protein synthesis has been shut down. Vesicular stomatitis virus (VSV) protein synthesis is depressed drastically when VSV-infected cells are super-infected with poliovirus. In cells doubly infected with VSV and encephalomyocarditis (EMC) virus or with VSV and SFV, dominance of one of the viruses depends on the time of addition of the challenge virus. The influence of external conditions on the relative translation of capped or uncapped viral mRNA in doubly infected cells has also been analysed.

Encephalomyocarditis virus↗

PubMed1982

Translation of capped virus mRNA in encephalomyocarditis virus-infected cells.

The pattern of protein synthesis in HeLa cells simultaneously infected with encephalomyocarditis virus (EMCV) and Semliki Forest virus (SFV) has been analysed throughout the time course of the infection. The ratio of the picornavirus protein gamma versus the togavirus late protein C increased when the m.o.i. of EMCV was raised, and the ratio of C/gamma increased with higher multiplicities of SFV. Under some conditions, the co-infected cells simultaneously synthesized the picornavirus and togavirus proteins, and the cells exclusively translated the capped 26S mRNA from SFV at the end of the co-infection experiment. The influence of the time of addition of the second virus on the relative translation of the capped and uncapped mRNAs was also studied. When HeLa cells were co-infected with 5 p.f.u./cell of EMCV and 200 p.f.u./cell of SFV, only the synthesis of SFV proteins was apparent, whereas if SFV was added 1 to 3 h later during the course of EMCV infection the cells synthesized picornavirus and togavirus proteins. If the cells were superinfected with SFV 1 h after EMCV addition, host protein synthesis was drastically inhibited after 3 h of EMCV infection. By 5 h post-infection both kinds of virus proteins were synthesized and at 7 h post-infection the cells preferentially translated the capped 26S mRNA from SFV. If cells were first infected with SFV (10 p.f.u./cell) and co-infected or superinfected at 1 h with EMCV (50 p.f.u./cell), shut-off of host protein synthesis occurred 3 h after infection and the cells synthesized both kinds of virus proteins. However, 9 h after infection the cells synthesized SFV proteins only. When double-infected HeLa cells were placed in a hypotonic medium, they mainly synthesized the togavirus late proteins, whereas under hypertonic conditions, they translated the picornavirus RNA exclusively. These results suggest that the two kinds of mRNAs (SFV 26S mRNA and EMCV 35S mRNA) are present in the infected cells and that the relative translation of each of them depends on the external ionic conditions.

Encephalomyocarditis virus↗