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Biomedical subjects

G Suarez

Publications and source records attributed to G Suarez.

At least 19 recordsLinked to original sources
PubMed2004

Class II MHC-expressing myofibroblasts play a role in the immunopathogenesis associated with staphylococcal enterotoxins.

Food poisoning due to staphylococcal enterotoxins (SEs) affects hundreds of thousands of people each year. Little is known about how SEs initiate immune responses and cause pathogenesis. Here, we demonstrate that cultured human intestinal myofibroblasts (IMFs) bind SEs in an MHC class II-dependent fashion. IMFs respond to SE exposure with increased secretion of IL-6, IL-8, and TNF-alpha. A significant proliferative T cell response was observed when MHC class II-expressing IMFs were pulsed with SEA and cocultured with human CD4(+) T cells. In conclusion, our findings support the hypothesis that IMFs may play an important role in pathology associated with staphlococcocal enterotoxigenic disease.

Cell Line↗
PubMed2004

Intestinal myofibroblasts and immune tolerance.

Stromal cells, such as myofibroblasts and fibroblasts, represent a significant fraction of MHC class II-positive cells in the normal human colonic lamina propria, suggesting they may play an important role in CD4(+) T cell regulation in a tolerogenic environment. The aim of this study was to examine whether human colonic myofibroblasts (CMFs) phenotypically and functionally resemble conventional antigen-presenting cells (APCs). Our results support the hypothesis that intestinal myofibroblasts are a novel, nonprofessional APC phenotype important in modulating mucosal T cell responses. Given their strategic location, we propose that intestinal myofibroblasts play a critical role in mediating tolerance to luminal antigens.

Cells, Cultured↗
PubMed2001

Cellular distribution of bovine leukemia virus proteins gp51SU, Pr72(env), and Pr66(gag-pro) in persistently infected cells.

Monoclonal antibodies (mAbs) against bovine leukemia virus (BLV) mature proteins and precursors were used to map the localization of these proteins in persistently infected non-lymphocytic cell lines using immunofluorescence assay (IFA) and immuno-electron microscopy. IFA staining was observed in the basolateral surface of live FLK-BLV cells. When using a mAb against Pr66(gag-pro), mottled pinpoint fluorescence was seen in the cell surface of polarized cells, but no reaction was observed in cells undergoing mitosis. However, a mAb against Pr72(env) stained only mitotic cells and cellular fragments. Additionally, in these dividing cells, this envelope (Env) precursor polyprotein was not evenly distributed but concentrated predominantly in only one daughter cell. To the best of our knowledge, this observation has not been reported previously, either for BLV or for other retroviruses. The results of immunogold electron microscopy confirmed the specificity of the mAbs in the intracellular level. In infected cells, Pr72(env) and gp51SU were seen in proximity at the plasma membrane in incipient budding sites. Additionally, the mAb against Pr72(env) also reacted with Env precursor polyproteins in the mitochondria of BLV-bat(2) ultrathin sections. These mAbs may be used as a tool for mapping virus excretion sites in the cell surface of naturally or in vitro infected cells in the different stages of the cell cycle.

Animals↗
PubMed2001

Mineralocorticoid receptor-mediated signaling regulates the ion gated sodium channel in vascular endothelial cells and requires an intact cytoskeleton.

The PCR analysis followed by sequence alignment showed that both the mineralocorticoid receptor (MCR) and the epithelial sodium channel (ENaC) genes were expressed in the human vascular endothelial cell line (ECV). The growth and multiplication of the ECV in culture were influenced by both aldosterone and the MCR-specific antagonist ZK 91587. Following double labelled immunofluorescence recorded by confocal microscopy, both the MCR and the ENaC were found to colocalize with the tubulin filaments in ECV cells in situ; no association was observed with cellular actin. ZK 91587 not only eliminated the basal expression, but it also impaired the transactivation of the ENaC gene by aldosterone. The disruption of actin and tubulin by cytochalasin D and colchicine, respectively, resulted in the total elimination of ENaC induction by aldosterone. These studies suggest that (i) the transcriptional regulation of the ENaC gene by the MCR-mediated signalling is not restricted to epithelial cells and requires cytoskeleton integrity in ECV cells in situ, (ii) tubulin may form a new and novel mediator in cell regulation, and (iii) the vascular tone may actually be regulated via transactivation of the ion gated sodium channel in the endothelial cell of the blood vessels under direct, receptor-mediated action of aldosterone.

Actins↗
PubMed2001

Evaluation of virus excretion by cells persistently infected with the bovine leukaemia virus (BLV) using monoclonal antibodies.

BACKGROUND: bovine leukaemia virus (BLV) is the causative agent of enzootic bovine leukaemia. Studies in vitro usually require the use of infected cell lines, mostly to produce antigen. Two of the most widely used cell lines are FLK-BLV and BLV-bat2. OBJECTIVE: the dynamics of the excretion of BLV proteins and whole virus by the persistently BLV-infected cell lines mentioned above was studied using an indirect ELISA in combination with eight monoclonal antibodies (mAbs) and cow and rabbit serum. STUDY DESIGN: tissue culture flasks were seeded with different concentrations of cells (13000-67000 cells per cm2, corresponding to 1-5 million cells per 75 cm2 flask) and were studied for 20 days. Samples (1.5 ml) were removed every 24 h and the presence of BLV proteins was determined using an indirect ELISA assay in which the antigen reaction with the monoclonal antibodies was evidenced by peroxidase labeled anti-mouse immunoglobulins. RESULTS: cell line FLK-BLV produced a complete monolayer as early as 4 days after passage, 3 days earlier than BLV-bat2. Using mAbs, the amount of viral proteins in the supernatant showed a cyclic pattern, with two evident peaks at days ca. 8 and 16. These peaks occurred even in the absence of passage or medium change, which causes depletion of essential nutrients and acidity. In comparison to polyclonal serum, mAbs gave more clear and defined values and are useful for determining the dynamics of viral production. CONCLUSION: when aiming for high viral yield, BLV should be harvested between days 6 and 8 after passage, when viral shedding is at its maximum. These results are very useful for preparing antigen for monoclonal antibody production, or for techniques such as ELISA or Western blot.

Animals↗
PubMed2001

Molecular typing by pulsed-field gel electrophoresis of Spanish animal and human Listeria monocytogenes isolates.

A total of 153 strains of Listeria monocytogenes isolated from different sources (72 from sheep, 12 from cattle, 18 from feedstuffs, and 51 from humans) in Spain from 1989 to 2000 were characterized by pulsed-field gel electrophoresis. The strains of L. monocytogenes displayed 55 pulsotypes. The 84 animal, 51 human, and 18 feedstuff strains displayed 31, 29, and 7 different pulsotypes, respectively, indicating a great genetic diversity among the Spanish L. monocytogenes isolates studied. L. monocytogenes isolates from clinical samples and feedstuffs consumed by the diseased animals were analyzed in 21 flocks. In most cases, clinical strains from different animals of the same flock had identical pulsotypes, confirming the existence of a listeriosis outbreak. L. monocytogenes strains with pulsotypes identical to those of clinical strains were isolated from silage, potatoes, and maize stalks. This is the first study wherein potatoes and maize stalks are epidemiologically linked with clinical listeriosis.

Animal Feed↗
PubMed2000

Production and characterization of monoclonal antibodies against bovine leukaemia virus using various crude antigen preparations: a comparative study.

A total of 59 monoclonal antibodies (mAbs) specific against the bovine leukaemia virus (BLV) using different antigen preparations was produced. The five antigen preparations for immunizing BALB/c mice were: live cells (CEL), sonicated and ultracentrifuged cells (SOC), cell lysates (LYS), semi-purified BLV (PV), and formalin-treated cells (FOR) from two cell lines permanently infected with BLV (FLK-BLV and BLV-bat2). These viral component presentations were selected to obtain mAbs against specific BLV proteins: located on the cell surface (FOR and CEL), in free virus particles (PV) and intracellular viral proteins (SOC and LYS). Two antigen preparations (SOC and LYS) were lethal to the mice following the intravenous and intrasplenic routes. Six fusions were performed in this study that rendered specific antibodies against BLV. The highest number of hybridomas was produced with SOC; however, the majority of the hybridomas produced (> 90%) were against cellular proteins. Even though immunization with PV gave the lowest number of hybridomas, the majority of them were specific against BLV. Based on the reactivity of the mAbs in Western blot (WB), we classified the mAbs into five groups, namely anti-gp51SU (39 mAbs), anti-gp30TM (six mAbs), anti-Pr72env (nine mAbs), anti-Pr66gag-pro (one mAb) and anti-Prgag (four mAbs). A very high percentage of the mAbs produced (48 of 59) reacted with gp51SU, suggesting that this is the most immunogenic and accessible BLV protein presented in the different antigen preparations. The majority of our mAbs recognized more than one band in WB, suggesting that, aside from reacting with mature proteins, the mAbs also recognized viral precursors.

Animals↗
PubMed1999

Rapid detection of specific polyclonal and monoclonal antibodies against bovine leukemia virus.

ELISA and Western blot have been used for detecting specific antibodies or antigens for routine diagnostic laboratory tests and experimental protocols, as well as for screening hybridomas secreting antibodies. Although these techniques are sensitive, some slow growing hybridomas are identified as positive only when they are grown slowly long time. We standardized the dot-ELISA, a more sensitive technique, for the detection of antibodies against BLV. The main advantages of the dot-ELISA described in this study are (a) its sensitivity, detecting hybridomas which would otherwise be considered negative and discarded from the results of indirect ELISA and/or Western blot; and (b) the possibility of economizing reagents using as little as 1 microl of the antigen and 0.5 microl of antibody and conjugate. Different BLV-antigen preparations were bound to nitrocellulose membranes (NC), including cells lysed chemically (LYS) or by sonication (SOC), semi-purified virus (PV), and supernatant from infected cultures, either without treatment (SUP) or sonicated (SOS). The antigen preparations most adequate for detecting monoclonal antibodies against BLV and polyclonal antibodies in cattle sera were undiluted cell lysates (LYS) and semi-purified BLV (PV). When testing bovine sera, the supernatant (SUP) and sonicated supernatant (SOS) antigens gave a high background due to the presence of FCS which reacted with the anti-bovine labeled antibodies. In this study, 59 BLV specific antibody secreting hybridomas were identified using the dot-ELISA, compared to only 20 detected using iELISA, and doubtful reactions due to nonspecific binding to fetal calf serum (FCS) and cellular components were measured. The results of the improved dot-ELISA described may be stored at room temperature for future reference. Results were consistently reproducible in coated nitrocellulose membranes kept at different storage temperatures (-20 degrees C, 4 degrees C, and 25-30 degrees C) 48 h, 1 week and 5 months.

Animals↗
PubMed1996

Microbiological diagnoses of chronic otitis externa in the dog.

The microbiological characteristics of otic exudates from 26 dogs with chronic otitis externa was studied with special reference to the implication of yeasts in the aetiology of the disease. A high frequency of yeasts and Staphylococcus aureus were isolated, alone or in association. In reference to the yeasts, there was a clear predominance of the genus Candida (48% of the total yeasts). Malassezia (Pytirosporum) represented only 3% of the isolates. It can be concluded that yeasts have an important role in the pathogenicity of this disease. For the microbiological diagnosis of otitis externa, we recommend the simultaneous use of Columbia/5% Sheep Blood Agar and Sabouraud-Dextrose without antibiotic addition, the use of 37 degrees C as the incubation temperature and direct microscopic observation of the sample before culture.

Animals↗
PubMed1995

ABO-haemolytic disease of the newborn (ABO-HDN): factors influencing its severity and incidence in Venezuela.

The objective of this study was to determine the incidence of the Haemolytic Disease of the Newborn due to ABO blood group incompatibility (ABO-HDN) in the population of Caracas attending the Maternity Hospital 'Concepción Palacios'. The relationship between A and B antigens density of cord blood erythrocytes and the cytotoxic activity of antibodies in mothers' sera with the severity of the haemolytic disease, was also studied. From a sample of 245 blood group 'O' mothers, 68 gave birth to full term 'A' or 'B' blood group infants. The evolution of serum bilirubin and the routine haematological values, were followed in all the babies during 72 h after birth, allowing the diagnosis of ABO-HDN in 21 infants. Taking into account that in Venezuela the frequency of blood group 'O' is 59 per cent, it was concluded that in the general population of newborns, 16 per cent present foeto-maternal ABO incompatibility, and the incidence of ABO-HDN was near to 5 per cent. The density of the 'A' and 'B' antigens in cord red cells was studied using an immunoenzymatic assay. No statistically significant association between antigen maturity and severity of the ABO-HDN could be shown. A positive association was found between cytotoxic capacity of mothers' sera and development of ABO-HDN (P < 0.05). Twenty ABO incompatibles children presented moderate late anaemia at 3 weeks of age.

ABO Blood-Group System↗
PubMed1995

Simple and rapid detection of Mycobacterium tuberculosis complex organisms in bovine tissue samples by PCR.

Mycobacterium bovis is a slowly growing microorganism, and confirmation of the diagnosis by conventional culture is a lengthy process. A simple, rapid method for the extraction of DNA from bovine tissue samples was developed and used in a PCR designed for the diagnosis of tuberculosis. Tissues from 81 cattle from tuberculosis-infected herds (group 1) and 19 cattle from tuberculosis-free herds (group 2) were tested in this PCR, and the results were compared with those of conventional culture. The PCR assay detected 71.4% of the culture-positive animals from group 1. Tissue from all animals in group 2 were negative in the PCR assay and by culture. The described method could be used as a rapid screening technique which would be complementary to culture of tissue specimens for the routine diagnosis of bovine tuberculosis. The PCR technique is much faster than culture and reduces the time for diagnosis from several months to 2 days. It also provides for the detection of M. bovis when rapidly growing Mycobacterium spp. are present in the sample and may be able to detect the presence of M. bovis in samples even when organisms have become nonviable.

Animals↗
PubMed1994

Treatment of ABO hemolytic disease with synthetic blood group trisaccharides.

Thirteen infants, 10 with A-O and 3 with B-O hemolytic disease of the newborn (ABO-HDN), were treated with synthetic A or B blood group trisaccharides (ATS, BTS) which cause dissociation of maternal antibody bound to infant red cells. The clinical outcome was compared with that of a control group of 21 infants treated with phototherapy during the preceding year. Exchange transfusion was required in 2 out of 13 infants in the experimental group and in 7 in the control group. A randomized prospective controlled study is necessary to confirm these results.

ABO Blood-Group System↗
PubMed1993

Solubility and stability of sterigmatocystin in different organic solvents.

In the present work we studied the solubility of sterigmatocystin (ST) in different organic solvents and the stability of these solutions during cold and frozen storage. ST was more soluble in chloroform and in pyridine. In cold storage ST was more stable in chloroform after both 7 and 30 days. In pyridine, ST was stable on day 7 but not on day 30. In frozen storage, 90% recovery was achieved only in chloroform. We conclude that chloroform is the most suitable organic solvent of those studied to solubilize and store standard solutions of ST.

Drug Stability↗
PubMed1993

Clinical experience with Mentor Alpha I inflatable penile prosthesis. Report on 333 cases.

Penile prostheses are a well-established form of therapy for erectile dysfunction. We report our experience with 333 cases using the Mentor Alpha I inflatable penile prosthesis. With a follow-up period of 15.4 months, there were 25 (7.5%) complications with 15 (4.5%) major complications that required reoperation. We also review the advantages and the procedures for implanting the penile prosthesis on an outpatient basis or a one-day hospital stay.

Adult↗
PubMed1993

Application of a diphasic dialysis technique to the extraction of aflatoxins in dairy products.

A new method is described for the extraction of aflatoxins from milk and milk products based on a diphasic dialysis technique, followed by detection and quantification of aflatoxins by TLC. Recovery was 65 to 99% for aflatoxins B1, G1, and M1, and the detection limit in milk and yogurt was .01 to .02 ppb for all aflatoxins studied. In cheese, the detection limit was .03 to .04 ppb for aflatoxins B1 and G1 and .1 ppb for aflatoxin M1. The main advantages of this technique are its simplicity; sensitivity; very economical use of reagents, thereby minimizing environmental pollution; efficiency, allowing the quick processing of a high number of samples; use by laboratories with little equipment; and a sensitivity comparable with that of much more sophisticated techniques.

Aflatoxin M1↗
PubMed1992

Effect of six organic acids on staphylococcal growth and enterotoxin production.

Four Staphylococcus aureus strains were incubated at 37 degrees C for 24 h in broth progressively acidified with lactic, citric, ascorbic, acetic, pyruvic and propionic acids, and their survival rate and enterotoxin producing ability was studied. Acids were chosen based on their frequent use by the food industry. Periodically, samples were withdrawn to determine counts, pH and the presence of enterotoxins A, B, C, and D. For a given acid, the effect on growth and enterotoxin synthesis was different. The most inhibitory acid for the growth of strains FRI-100 and FRI-472 was pyruvic acid, for strain FRI-137 was lactic acid, all six acids were equally effective on strain S6. Lactic acid was very inhibitory to enterotoxin synthesis, but the effect on this parameter of acetic and citric acids was almost nil. Enterotoxins were seen to be inactivated at acid pH values; enterotoxin B was the most resistant to inactivation.

Acetates↗