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Biomedical subjects

F Lozano

Publications and source records attributed to F Lozano.

At least 55 records · Page 3Linked to original sources
PubMed1998

CD43 interacts with moesin and ezrin and regulates its redistribution to the uropods of T lymphocytes at the cell-cell contacts.

Chemokines as well as the signaling through the adhesion molecules intercellular adhesion molecule (ICAM)-3 and CD43 are able to induce in T lymphocytes their switching from a spherical to a polarized motile morphology, with the formation of a uropod at the rear of the cell. We investigated here the role of CD43 in the regulation of T-cell polarity, CD43-cytoskeletal interactions, and lymphocyte aggregation. Pro-activatory anti-CD43 monoclonal antibody (MoAb) induced polarization of T lymphocytes with redistribution of CD43 to the uropod and the CCR2 chemokine receptor to the leading edge of the cell. Immunofluorescence analysis showed that all three ezrin-radixin-moesin (ERM) actin-binding proteins localized in the uropod of both human T lymphoblasts stimulated with anti-CD43 MoAb and tumor-infiltrating T lymphocytes. Radixin localized at the uropod neck, whereas ezrin and moesin colocalized with CD43 in the uropod. Biochemical analyses showed that ezrin and moesin coimmunoprecipitated with CD43 in T lymphoblasts. Furthermore, in these cells, the CD43-associated moesin increased after stimulation through CD43. The interaction of moesin and ezrin with CD43 was specifically mediated by the cytoplasmic domain of CD43, as shown by precipitation of both ERM proteins with a GST-fusion protein containing the CD43 cytoplasmic tail. Videomicroscopy analysis of homotypic cell aggregation induced through CD43 showed that cellular uropods mediate cell-cell contacts and lymphocyte recruitment. Immunofluorescence microscopy performed in parallel showed that uropods enriched in CD43 and moesin localized at the cell-cell contact areas of cell aggregates. The polarization and homotypic cell aggregation induced through CD43 was prevented by butanedione monoxime, indicating the involvement of myosin cytoskeleton in these phenomena. Altogether, these data indicate that CD43 plays an important regulatory role in remodeling T-cell morphology, likely through its interaction with actin-binding proteins ezrin and moesin. In addition, the redistribution of CD43 to the uropod region of migrating lymphocytes and during the formation of cell aggregates together with the enhancing effect of anti-CD43 antibodies on lymphocyte cell recruitment suggest that CD43 plays a key role in the regulation of cell-cell interactions during lymphocyte traffic.

Antigens, CD↗
PubMed1998

[Electromagnetic interference between automatic defibrillators and digital and analog cellular telephones].

BACKGROUND AND OBJECTIVES: Functional pacemaker interference by mobile telephones has been described with analogical systems and with possible greater influence, digital systems, including inhibition and inadequate pacing. The influence of both system has not been extensively studied in patients with implantable cardioverter defibrillators (ICD). PATIENTS AND METHODS: We studied the influence of mobile phones, both digital and analogic network, on the performance of several models of defibrillators, in a standardised test set up designed to provide high sensitivity. The purpose of our study was to establish whether there are any influences on ICD functions, both in in vivo and in in vitro models. Several mobile phones, with different transmission powers, were moved towards the defibrillator and the electrode, under continuous documentation of defibrillator sensing and interrogation afterwards. The experimental model was performed with the aid of an arrhythmia simulator (Intersim) and demo-defibrillators. The tests were repeated both in and out of a solution of saline water with an impedance within normal human limits. RESULTS: Partial loss of telemetry was found in 14 patients, 8 with analogical phones and 6 with digital phones. Fourteen patients showed alterations only on the surface electrocardiogram channel and five on the intracavitary channel. The same results were reproduced in the in vitro model. However, the in vitro test allowed us to simulate multiple ventricular arrhythmias, and demonstrate the normal sensing and functioning of the defibrillator during a "spontaneous" arrhythmia. After testing, we demonstrate that no real oversensing/undersensing was documented in any device. There was no evidence of ICD reprogramming or pacing inhibition. In particular, no inadequate therapies were delivered. CONCLUSIONS: a) in our series, we have not demonstrated clinically significant electromagnetic interferences with mobile phones of digital or analogical networks: b) the in vitro model allowed us to conclude that even if a spontaneous arrhythmia appears, the function of the defibrillator is not altered; c) the use of mobile phones seems to be safe for defibrillator patients, and d) however, some basic rules, such as to maintain the phone at least 15 cm away from the defibrillator, are advised.

Defibrillators, Implantable↗
PubMed1998

Isolation of two CD50 (ICAM-3)-negative Jurkat T-cell clones and their application for analysis of CD50 function.

The leukocyte differentiation antigen CD50 (intercellular adhesion molecule-3, ICAM-3), mediates cell-cell adhesion through its ligand LFA-1 and is a transducting receptor molecule during T-cell activation. Since CD50 homologues in other species have not yet been identified, the role of this molecule can only be analyzed in human cell models. Thus, to better study CD50 function in T cells, we have obtained two CD50-negative T-cell clones, named CAMY.1 and CAMY.2. These clones were derived from the Jurkat T-cell variant PPL.1. Data from analysis of protein expression, specific mRNA content and calcium mobilization assays have confirmed the absence of functional CD50 molecules on these two clones. Thus, CAMY.1 and CAMY.2 show no CD50 expression by phenotypical and immunoprecipitation analysis. CD50-specific mRNA content is undetectable by Northern blot analysis in these clones and, only, when RT-PCR was performed could specific mRNA be detected. Additionally, CD50 cross-linking on theses clones shows no increase in intracellular calcium. Transfection of CD50 cDNA on CAMY cells restores not only CD50 surface expression, but its functional ability to induce calcium mobilization, CD69 upregulation and cell morphological changes. The CAMY.1 and CAMY.2 clones provide useful model systems to analyze CD50 function in T cells.

Animals↗
PubMed1998

Multicriteria decision analysis for determining drug therapy for intermittent claudication.

Drugs of various classes are prescribed for intermittent claudication. However, there is some discrepancy between medical practice and the scientific basis for drug selection. We have developed a quantitative criteria-based decision analysis to evaluate all implications of drug treatment choices for intermittent claudication. Pentoxifylline, buflomedil, naftidrofuryl and ticlopidine were the drugs selected for analysis. The evaluation criteria were 1) therapeutic efficacy, 2) safety, 3) patient acceptance and 4) cost. A review panel of experts determined the relative importance of each criterion by assigning points (or utility values) to each one. The points were 48, 20, 14 and 18, respectively, for criteria 1, 2, 3 and 4. A probability value, or numerical estimate of how well a drug meets a criterion, was assigned to each drug for each of the 4 criteria. The probability value was multiplied by the utility value to determine the score for each drug and criterion. The criteria points for each drug were added for a total score for the drug. The drug with the highest overall score was pentoxifylline, with 69 points out of an ideal score of 100. The rank order for the other drugs was buflomedil, ticlopidine and naftidrofuryl. A sensitive analysis showed that the relative ranking of the drugs remained unchanged over a series of data modifications.

Decision Support Techniques↗
PubMed1998

[Infections by Candida and cryptococci].

The introduction of highly active antiretroviral therapy (HAART) has reduced dramatically the incidence of mucosal candidiasis and cryptococcosis in AIDS patients. Fluconazole is the drug of choice for candidiasis. The duration of antifungal treatment is based on response, but typically 7 to 14 days are required for oropharyngeal forms and up to 21 days for esophageal disease (200 the first day and 100 mg thereafter). Resistant candidiasis tends to occur in persons with advanced HIV disease and previous fluconazole therapy who have been noted to result in clinical improvement with HAART. HAART must be considered the therapy of choice for refractory candidiasis. The preferred treatment for cryptococcal meningitis includes two weeks induction treatment with amphotericin B (0.7 mg/kg/d IV) with or without flucytosine (25 mg/kg qid) followed by 8 weeks of fluconazole (200-400 mg PO qd). Long-term maintenance therapy with fluconazole (200-400 mg PO qd) is required to prevent relapses. In patients with elevated intracranial pressure who had focal neurologic deficits or mental status changes, serial lumbar punctures should be performed. In refractory cases the immediate placement of CSF drains must be considered.

AIDS-Related Opportunistic Infections↗
PubMed1997

The cytoplasmic domain of CD5 mediates both TCR/CD3-dependent and -independent diacylglycerol production.

CD5 is a 67-kDa surface glycoprotein found in association with the Ag receptor complex on both T and B lymphocytes. CD5 modulates Ag receptor-mediated immune responses, but the molecular mechanisms of its action remain unclear. In this respect, the assessment of the relative and unique contribution of CD5 in cell signaling events is a crucial point. We have used Jurkat variants and anti-CD5 mAbs to show that the CD5 signaling pathway is distinct from that used by the TCR/CD3 complex. We hereby identify two independent mechanisms of CD5-mediated diacylglycerol release by virtue of their different kinetics: 1) an early and transient diacylglycerol increase that results from the activation of a phosphatidylcholine-specific phospholipase C, and 2) a late and sustained increase that requires de novo phospholipid synthesis. Studies performed on a TCR/CD3-deficient Jurkat cell variant indicate that only the CD5-mediated phosphatidylcholine-specific phospholipase C activation is dependent on TCR/CD3 expression. Mutational analyses of CD5 demonstrate that both mechanisms are dependent on the integrity of the CD5 distal cytoplasmic region. Our results show that CD5 is a signaling molecule per se that uses mechanisms resembling those used by some cytokine receptors (such as IL-1 or TNF receptors) to modulate lymphocyte activation.

Animals↗
PubMed1997

Moesin interacts with the cytoplasmic region of intercellular adhesion molecule-3 and is redistributed to the uropod of T lymphocytes during cell polarization.

During activation, T lymphocytes become motile cells, switching from a spherical to a polarized shape. Chemokines and other chemotactic cytokines induce lymphocyte polarization with the formation of a uropod in the rear pole, where the adhesion receptors intercellular adhesion molecule-1 (ICAM-1), ICAM-3, and CD44 redistribute. We have investigated membrane-cytoskeleton interactions that play a key role in the redistribution of adhesion receptors to the uropod. Immunofluorescence analysis showed that the ERM proteins radixin and moesin localized to the uropod of human T lymphoblasts treated with the chemokine RANTES (regulated on activation, normal T cell expressed, and secreted), a polarization-inducing agent; radixin colocalized with arrays of myosin II at the neck of the uropods, whereas moesin decorated the most distal part of the uropod and colocalized with ICAM-1, ICAM-3, and CD44 molecules. Two other cytoskeletal proteins, beta-actin and alpha-tubulin, clustered at the cell leading edge and uropod, respectively, of polarized lymphocytes. Biochemical analysis showed that moesin coimmunoprecipitates with ICAM-3 in T lymphoblasts stimulated with either RANTES or the polarization- inducing anti-ICAM-3 HP2/19 mAb, as well as in the constitutively polarized T cell line HSB-2. In addition, moesin is associated with CD44, but not with ICAM-1, in polarized T lymphocytes. A correlation between the degree of moesin-ICAM-3 interaction and cell polarization was found as determined by immunofluorescence and immunoprecipitation analysis done in parallel. The moesin-ICAM-3 interaction was specifically mediated by the cytoplasmic domain of ICAM-3 as revealed by precipitation of moesin with a GST fusion protein containing the ICAM-3 cytoplasmic tail from metabolically labeled Jurkat T cell lysates. The interaction of moesin with ICAM-3 was greatly diminished when RANTES-stimulated T lymphoblasts were pretreated with the myosin-disrupting drug butanedione monoxime, which prevents lymphocyte polarization. Altogether, these data indicate that moesin interacts with ICAM-3 and CD44 adhesion molecules in uropods of polarized T cells; these data also suggest that these interactions participate in the formation of links between membrane receptors and the cytoskeleton, thereby regulating morphological changes during cell locomotion.

Antigens, CD↗
PubMed1997

Desialylation of T lymphocytes overcomes the monocyte dependency of pokeweed mitogen-induced T-cell activation.

Activation of T lymphocytes by pokeweed mitogen (PWM) is strictly monocyte (Mo)-dependent and results in T-cell mitogenesis and interleukin-2 (IL-2) secretion, coupled with an inability to utilize IL-2 due to an impaired expression of functional IL-2 receptor (IL-2R). Such IL-2R impairment could arise in PWM-activated T cells themselves or, alternatively, be the result of Mo-derived influences, as it is known that PWM binds Mo strongly and does not or poorly binds lymphocytes, and Mo becomes rapidly destroyed in PWM-stimulated cultures of blood mononuclear cells or T cells plus Mo. The present study investigated these possibilities. The results show for the first time that desialylation of T lymphocytes strongly increases their PWM-binding capacity and, in addition, overcomes the Mo requirement for PWM to induce T-cell mitogenesis and IL-2 secretion. Such secreted IL-2 levels were even higher that those found in cultures of Mo-dependent PWM-activated T lymphocytes but similarly to the latter, PWM-activated desialylated purified T lymphocytes exhibited negligible high-affinity IL-2 binding capacity and an inability to utilize the IL-2 they produced. These effects were not due to desialylation itself, as indicated by data obtained with peanut agglutinin, a lectin that becomes strongly reactive with desialylated T lymphocytes. The data clearly indicate the existence of PWM-related events capable of impairing the expression of functional IL-2R without affecting IL-2 secretion, and indicate that such events are due to mechanisms arising at the level of PWM-activated T cells themselves.

Adult↗
PubMed1996

Conformation of the hypervariable region L3 without the key proline residue.

The refined structure of the Fab fragment of the monoclonal antibody CRIS-I (IgG2a kappa) against the leukocyte differentiation antigen CD5, determined at 1.9 A resolution with an agreement R-factor of 18.3%, reveals a variant of the canonical conformations proposed for the light chain complementarity determining region L3 (CDR-L3). This is the first Fab structure available with a kappa light chain in which the CDR-L3 lacks the key proline residue in either position 94 or 95. The conformation found could be significant for about 10% of the murine IgG molecules with kappa light chains without proline in their CDR-L3 sequences.

Animals↗
PubMed1996

Prospective evaluation of fever of unknown origin in patients infected with the human immunodeficiency virus. Grupo Andaluz para el Estudio de las Enfermedades Infecciosas.

The aim of this study was to determine the frequency and aetiology of fever of unknown origin (FUO) in patients infected with the human immunodeficiency virus (HIV), to assess the value of the tests used in its diagnosis, and to evaluate possible models of diagnosis for the causes found most frequently. One hundred twenty-eight (3.5%) of 3603 hospitalised HIV-positive patients evaluated from October 1992 to December 1993 had FUO, defined by established criteria. Eighty-six percent of patients with FUO had previously progressed to AIDS. The median CD4+ cell count was 46/mm3. A definite diagnosis was made in 96 (75%) of the 128 patients and a possible diagnosis in 24 (18.7%). whilst no diagnosis was made in eight cases (6.2%). Tuberculosis (48.3%), visceral leishmaniasis (16%), and infection by Mycobacterium avium complex (6.9%) were the diseases found most frequently. The most useful diagnostic tests were liver biopsy (68.9%) and bone marrow aspirate/biopsy (39.7%). It is not possible to predict clinically the cases of FUO due to tuberculosis, whilst thrombocytopaenia < 100,000 cells/mm3 alone is useful for differentiating the cases of visceral leishmaniasis, with a negative predictive value of 95.2%.

Adolescent↗
PubMed1995

Targeting of non-Ig sequences in place of the V segment by somatic hypermutation.

Affinity maturation of antibodies is characterized by localized hypermutation of the DNA around the V segment. Here we show, using mice containing single or multiple transgene constructs, that an immunoglobulin V kappa segment can be replaced by human beta-globin or prokaryotic neo or gpt genes without affecting the rate of hypermutation; the V gene itself is not necessary for recruiting hypermutation. The ability to target hypermutation to heterologous genes in vivo could find more general applications in biology.

Animals↗
PubMed1995

A reporter gene to analyse the hypermutation of immunoglobulin genes.

The affinity maturation of antibodies is driven by somatic hypermutation which is localized to specific segments of the coding genes. The information available on this process derives from studies in vivo. With the intention of developing new approaches, we have constructed a fusion gene between a kappa chain and a selectable neomycin resistance gene, neor. The neor gene, which includes the SV40 small t intron and polyadenylation site, but not the upstream elements nor its first 12 amino acids, is an in-frame substitution of the FR2-CDR3 fragment of a rearranged V kappa OX1-J kappa 5 gene. Expression of neor activity is therefore dependent on the upstream immunoglobulin sequence. A stop codon was placed in the CDR1 region so that only mutants survive treatment with geneticin sulphate (G418). The effectiveness of the system was tested by transfecting the NS0 myeloma cell line and isolating spontaneous mutants. Neomycin-resistant clones arose at an estimated rate of 1 x 10(-8)/cell division, and over 90% were authentic structural mutants. Unlike the somatic hypermutations, the majority arose by in-frame deletions including the stop codon, although up to 30% involved a point mutation. The reporter gene was then modified by substituting all the sequences downstream of the J kappa 5 with others known to be required for full hypermutation in vivo. Different cell lines were transfected and G418-resistant clones analyzed. No significant increase in the rate of reversion or in the generation of point mutations versus deletions was detected, even using conditioned culture medium. In the presence of azacytidine however, a mutant involving multiple events (single base addition and deletion plus two point mutations) was detected. The reporter gene system therefore seems suitable to test culture conditions and modifications of the host cells aimed at the derivation of an in vitro assay of somatic hypermutation.

Amino Acid Sequence↗
PubMed1995

Existence of a soluble form of CD50 (intercellular adhesion molecule-3) produced upon human lymphocyte activation. Present in normal human serum and levels are increased in the serum of systemic lupus erythematosus patients.

CD50 (ICAM-3) is a leukocyte differentiation Ag expressed almost exclusively on hemopoietic cells, with a key role in the first steps of immune response. To develop a specific sandwich ELISA to detect a soluble CD50 form (sCD50), two different mAbs (140-11 and 101-1D2) recognizing non-overlapping epitopes were used. sCD50 was detected in the supernatant of stimulated PBMCs, with the highest levels after CD3 triggering. Simultaneously, the CD50 surface expression diminished during the first 24 h. sCD50 isolated from culture supernatant and analyzed by immunoblotting showed an apparent m.w. of 95 kDa, slightly smaller than the membrane form. These data, together with Northern blot kinetics analysis, suggest that sCD50 is cleaved from cell membrane. Furthermore, we detect sCD50 in normal human sera and higher levels in sera of systemic lupus erythematosus (SLE) patients, especially in those in active phase. The sCD50 levels showed a positive correlation with sCD27 levels (r = 0.4213; p = 0.0026). Detection of sCD50, both after in vitro CD3 triggering of PBMCs and increased in SLE sera, suggests that sCD50 could be used as a marker of lymphocyte stimulation.

Adolescent↗