Biomedical subjects

F Miralles

Publications and source records attributed to F Miralles.

At least 37 records · Page 2Linked to original sources

PubMed1998

UV irradiation induces the murine urokinase-type plasminogen activator gene via the c-Jun N-terminal kinase signaling pathway: requirement of an AP1 enhancer element.

UV irradiation leads to severe damage, such as cutaneous inflammation, immunosuppression, and cancer, but it also results in a gene induction protective response termed the UV response. The signal triggering the UV response was thought to originate from DNA damage; recent findings, however, have shown that it is initiated at or near the cell membrane and transmitted via cytoplasmic kinase cascades to induce gene transcription. Urokinase-type plasminogen activator (uPA) was the first protein shown to be UV inducible in xeroderma pigmentosum DNA repair-deficient human cells. However, the underlying molecular mechanisms responsible for the induction were not elucidated. We have found that the endogenous murine uPA gene product is transcriptionally upregulated by UV in NIH 3T3 fibroblast and F9 teratocarcinoma cells. This induction required an activator protein 1 (AP1) enhancer element located at -2.4 kb, since deletion of this site abrogated the induction. We analyzed the contribution of the three different types of UV-inducible mitogen-activated protein (MAP) kinases (ERK, JNK/SAPK, and p38) to the activation of the murine uPA promoter by UV. MEKK1, a specific JNK activator, induced transcription from the uPA promoter in the absence of UV treatment, whereas coexpression of catalytically inactive MEKK1(K432M) and of cytoplasmic JNK inhibitor JIP-1 inhibited UV-induced uPA transcriptional activity. In contrast, neither dominant negative MKK6 (or SB203580) nor PD98059, which specifically inhibit p38 and ERK MAP kinase pathways, respectively, could abrogate the UV-induced effect. Moreover, our results indicated that wild-type N-terminal c-Jun, but not mutated c-Jun (Ala-63/73), was able to mediate UV-induced uPA transcriptional activity. Taken together, we show for the first time that kinases of the JNK family can activate the uPA promoter. This activation links external UV stimulation and AP1-dependent uPA transcription, providing a transcription-coupled signal transduction pathway for the induction of the murine uPA gene by UV.

3T3 Cells↗

PubMed1998

Follistatin regulates the relative proportions of endocrine versus exocrine tissue during pancreatic development.

In this study, we have investigated the role of the embryonic mesenchyme in the development of the pancreas. We have compared the development in vitro of E12.5 rat pancreatic rudiments grown in the presence or absence of mesenchyme. When the E12.5 pancreatic epithelial rudiment is cultured in the presence of its surrounding mesenchyme, both morphogenesis and cytodifferentiation of the exocrine component of the pancreas are completely achieved, while only a few immature endocrine cells develop. The pancreatic rudiments grown in the absence of mesenchyme develop in a completely different way; the exocrine tissue develops poorly and fails to undergo acinar morphogenesis, while the endocrine tissue develops actively. Four times more insulin-positive cells develop after removal of the mesenchyme than in the cultures performed in the presence of mesenchyme. Moreover, the insulin-expressing cells developed in the mesenchyme-depleted rudiments appear mature since they do not coexpress glucagon, express the glucose transporter Glut-2 and express Rab3A, a molecule associated with the secretory granules. Moreover, these endocrine cells are able to associate and form true islets. Both the inductive effect of the mesenchyme on the proper development of the exocrine tissue and its repressive effect on the development of the endocrine cells are mediated by soluble factors. Follistatin, which is expressed by E12.5 pancreatic mesenchyme, can mimic both inductive and repressive effects of the mesenchyme. Follistatin could thus represent one of the mesenchymal factors required for the development of the exocrine tissue while exerting a repressive role on the differentiation of the endocrine cells.

Amylases↗

PubMed1998

Expression of nerve growth factor and its high-affinity receptor Trk-A in the rat pancreas during embryonic and fetal life.

The expression of functional receptors for nerve growth factor in insulin-producing cell lines grown in vitro has recently been demonstrated. The possible importance of signals transduced via these receptors in the control of islet maturation has been proposed based on data obtained using an in vitro culture system. To further support this hypothesis, we have studied the expression of Trk-A, the high-affinity receptor for NGF, in vivo during the embryonic and fetal development of the rat pancreas. We have also examined the expression of NGF during the same period. Immunohistological analysis shows that at embryonic day 11 (E11), Trk-A is expressed by the epithelial cells of the presumptive pancreas. The few pancreatic endocrine cells present at that stage express Trk-A. At E12 and E16, Trk-A expression was detected in the developing ductal network. The endocrine cells located in the ducts express Trk-A while those that have migrated into the surrounding mesenchyme now stain negative for Trk-A. By E20, Trk-A expression by ductal cells has considerably decreased and can be detected only in small ducts closely associated with islet-like structures. These islet-like structures stain negative for Trk-A. After birth, insulin-positive cells arranged into islets re-express Trk-A. During the same period, NGF mRNA is found to be expressed in the developing pancreas. The expression of Trk-A and its ligand NGF in the pancreas during embryonic and fetal life suggests that NGF and its receptor could play an important role in the development of the pancreas.

Animals↗

PubMed1998

Fibroblast growth factor 2 promotes pancreatic epithelial cell proliferation via functional fibroblast growth factor receptors during embryonic life.

Several investigators have postulated that soluble growth factors are involved in the early development of the pancreas. In many tissues in which soluble factors are implicated in development, these factors act on their target cells through tyrosine kinase receptors. Because we had some preliminary evidence that fibroblast growth factor receptors (FGFRs) were expressed in the early pancreas, we investigated the effect of fibroblast growth factors (FGFs) during embryonic pancreatic development. For that purpose, we first studied the distribution and the functionality of FGFRs during pancreatic organogenesis. FGFR1 and FGFR4 were shown to be expressed at a high level during early pancreatic development before embryonic day 16, their levels of expression decreasing thereafter. The functionality of FGFR was studied next. It was demonstrated in vitro that both FGF1 and FGF2 induce the expression of NGFI-A mRNA, a useful indicator of functional growth factor-signaling pathways. Finally, the effect of FGF2 on embryonic pancreatic epithelial cell proliferation was studied. It was shown that FGF2 induces the proliferation of pancreatic epithelial cells during embryonic life. Taken together, these data strongly suggest that FGFs are implicated in pancreatic development during embryonic life.

Animals↗

PubMed1997

Purification and characterization of ferritin from alfalfa seeds.

Ferritin from alfalfa (Medicago sativa) seeds was isolated, purified, and characterized. The apparent molecular mass of the native protein was found to be 560 kDa. Electrophoresis in denaturing gradient polyacrylamide-SDS gels revealed subunits of 28-26.5 kDa. The average iron cores were 4 nm in diameter and contained about 1400 iron atoms, with an iron-to-phosphorus ratio of 4:1. N-terminal amino acid sequencing of the 28 kDa subunit revealed close homology with other plant proteins. Immunochemical analysis using polyclonal antibodies raised against pea-seed ferritin has confirmed, in agreement with previous reports, that plant proteins share common epitopes.

Amino Acid Sequence↗

PubMed1997

A simple method for the eradication of Legionella pneumophila from potable water systems.

In this paper we describe a simple method, noncorrosive to pipes, for the eradication of Legionella pneumophila from potable water systems. This method is based on the systematic purging of the pipe networks with cold water containing 1-1.5 mg residual chlorine/L. In the hot water system, a new pipe bypassing the water heater was installed, whereas in the air conditioning system, the circuit is purged with water from the tap water system. The feasibility of this method was studied in two hotels in which the presence of Legionella was detected despite treatment of the water by the hyperchlorination method. The evolution of the presence of Legionella was studied by culture and polymerase chain reaction. Eighty samples from hotel A and sixty-seven samples from hotel B were analyzed during the time that the eradication method was applied. Our results showed that this method permitted the effective elimination of L. pneumophila after 5 months in hotel A and 7 months in hotel B.

Chlorine↗

PubMed1997

Inhibition of urokinase-type plasminogen activator (uPA) abrogates myogenesis in vitro.

Urokinase-type plasminogen activator (uPA) is one of the components of blood's fibrinolytic cascade. uPA acts as a broad spectrum proteolytic enzyme involved in different physio-pathological processes including cellular fibrinolysis, adhesion, migration, invasion and remodeling. Here, we present evidence that uPA participates in myogenesis, a process which requires drastic cell membrane reorganization, leading to the plurinucleated myotube from the progenitor myoblast. We have dissected the expression of uPA throughout the different myogenic compartments and found an increase in uPA enzymatic activity associated with myotube formation in C2C12 myoblast cells, with uPA mRNA increasing prior the onset of fusion and differentiation. When both fusion and differentiation were blocked by specific inhibitors (DMSO, cytochalasin B) the levels of uPA were strongly downregulated. This process was reversible and specific: the removal of the inhibitors immediately restored the levels of uPA mRNA while the specific inhibition of uPA enzymatic activity by an anti-uPA antibody resulted in a 50% reduction of the extent of fusion and in the abrogation of muscle-specific gene products, such as alpha-actin and MyoD. Moreover, the conversion of fibroblasts to muscle-like cells upon acquisition of MyoD resulted in a dramatic increase of uPA mRNA, which was partially due to transcriptional activation of the uPA gene. These results indicate that the increase in uPA expression prior to fusion and differentiation occurs via a MyoD-mediated mechanism whereas the normal MyoD expression requires the plasminogen activation-dependent activity of this protease. Therefore, these studies extend the sphere of influence of myogenic factors to fibrinolysis, an intrinsic component of the hematological system. Taken together, one mechanism used by the myoblast cell to become a differentiated myotube, involving the inductive extracellular proteolysis of urokinase, is proposed.

Actins↗

PubMed1997

[Muscarinic receptors on lymphocytes].

INTRODUCTION: The presence of muscarinic receptors on lymphocytes and possible modifications in their number and affinity may be related to cholinergic changes found in muscarinic receptors at a central level. OBJECTIVE: The objective of this paper is to bring the technique for the determination of lymphocyte muscarinic receptors by means of their union with radioligands for application in the study of disorders involving the cholinergic system, such as Alzheimer's disease, up to date. MATERIAL AND METHODS: We therefore made a density gradient isolation of lymphocytes from healthy subjects and made a trial of union or binding with N-methyl-escopolamine marked with tritium (3H-NMS) on intact lymphocytes. RESULTS: We were unable to specifically label the muscarinic receptors. However, we have shown the usefulness of the technique, using rat anterior cerebral homogenate. CONCLUSIONS: We consider whether the absence of specific union with the lymphocyte muscarinic receptor is due to technical problems as is often described in the literature, or possibly due to the absence of these receptors on the lymphocyte as suggested by other authors and proved using molecular biology techniques to be the case for the M2 receptor subtype.

Animals↗

PubMed1996

Activity-dependent modulation of the presynaptic potassium current in the frog neuromuscular junction.

1. Changes in the electrical properties of frog motor nerve endings caused by the invasion of an action potential were studied by the perineural recording technique. Two equal supramaximal stimuli separated by a variable time interval were applied to the nerve trunk. The latency and amplitude of the deflections associated with the nodal Na+ current and presynaptic K+ current elicited by the second pulse were compared with control currents. 2. The deflection associated with the presynaptic K+ current elicited in response to the second stimulus was absent at the shortest interstimulus interval and showed a progressive increase in its amplitude as the interstimulus interval was lengthened, reaching values greater than control in most terminals. During the same period the nodal Na+ current did not change. 3. The experimental results were compared with a computer model of the distal axonal segment and its terminal. Response of the model to twin-pulse stimulation was in marked disagreement with the experimental results unless an inactivating K+ channel, with properties derived ad hoc, was incorporated into the simulation. 4. These results suggest that in the first 6-7 ms after a nerve impulse has invaded a frog motor nerve ending, maximal K+ conductance remains below the value at rest due to the fast inactivation of a K+ conductance. Following this, there is a period in which K+ conductance is greater than control values although the basis for this is unknown.

4-Aminopyridine↗

PubMed1996

Niflumic acid-induced increase in potassium currents in frog motor nerve terminals: effects on transmitter release.

The actions of the nonsteroidal antiinflammatory drug niflumic acid were studied on frog neuromuscular preparations by conventional electrophysiological techniques. Niflumic acid reduced the amplitude and increased the latency of endplate potentials in a concentration-dependent manner. Neuromuscular junctions pretreated with niflumic acid (0.05-0.5 mM) showed much less depression than control when they were stimulated with trains of impulses. Inhibition of acetylcholine release was reverted by raising the extracellular Ca(2+) concentration but not by simply washing out the preparations with niflumic acid-free solutions. Pretreatment with indomethacin (0.1 mM), another nonsteroidal antiinflammatory drug, did not affect the niflumic acid-induced inhibition of evoked responses. Niflumic acid (0.1 mM) did not change the amplitude of miniature endplate potentials and had a dual action on the frequency of miniatures: it decreased their frequency at 0.1 mM whereas it produced an enormous increase in the rate of spontaneous discharge at 0.5 mM. Niflumic acid (0.1 - 1 mM) reversibly increased the amplitude and affected the kinetics of presynaptic voltage-activated K+ current and Ca(2+)-activated K(+) current in a concentration-dependent manner. Niflumic acid (0.1 - 1 mM) irreversibly decreased the amplitude and reversibly affected the kinetics of the nodal Na(+) current. Indomethacin (0.1 mM) had no effect on presynaptic currents. In conclusion, niflumic acid reduces acetylcholine release by increasing presynaptic K+ currents. This may shorten the depolarizing phase of the presynaptic action potential and may reduce the entry of Ca(2+) with each impulse.

Animals↗

PubMed1996

Action of suramin upon ecto-apyrase activity and synaptic depression of Torpedo electric organ.

1. The role of ATP, which is co-released with acetylcholine in synaptic contacts of Torpedo electric organ, was investigated by use of suramin. Suramin [8-(3-benzamido-4-methylbenzamido)naphthalene-1,3,5-trisulphoni c acid], a P2 purinoceptor antagonist, potently inhibited in a non-competitive manner the ecto-apyrase activity associated with plasma membrane isolated from cholinergic nerve terminals of Torpedo electric organ. The Ki was 30 microM and 43 microM for Ca(2+)-ADPase and Ca(2+)-ATPase respectively. 2. In Torpedo electric organ, repetitive stimulation decreased the evoked synaptic current by 51%. However, when fragments of electric organ were incubated with suramin the evoked synaptic current declined by only 14%. Fragments incubated with the selective A1 purinoceptor antagonist, DPCPX, showed 5% synaptic depression. 3. The effects of suramin and DPCPX on synaptic depression were not addictive. Synaptic depression may thus be linked to endogenous adenosine formed by dephosphorylation of released ATP by an ecto-apyrase. The final effector in synaptic depression, adenosine, acts via the A1 purinoceptor. 4. ATP hydrolysis is prevented in the presence of suramin. It slightly increased (20%) the mean amplitude of spontaneous miniature endplate currents. The frequency distribution of the amplitude of spontaneous events was shifted to the right, indicating that ATP, when not degraded, may modulate the activation of nicotinic acetylcholine receptors activated by the quantal secretion of acetycholine.

Acetylcholine↗

PubMed1995

Bile salt-dependent lipase transcripts in human fetal tissues.

In human fetal pancreas, we identified two cDNA transcripts of the bile salt-dependent lipase (BSDL) using reverse transcription followed by polymerase chain reaction (RT-PCR). The sequence of four overlapping segments obtained by RT-PCR matched the sequence of the 2.2 kb cDNA cloned from human adult pancreas (Reue et al. (1991) J. Lipid Res. 32, 267-276). A second RT-PCR product of approx. 1.1 kb was evidenced, the sequence of which corresponds to that of the BSDL-pseudogene transcript (Nilsson et al. (1993) Genomics, 17, 416-422). The short transcript is present in all tissues examined whereas the former one (2.2 kb) is either poorly (in liver and kidney) or not at all expressed in adult tissues, excepted in the pancreas. On the other hand, the 2.2 kb transcript specific of the BSDL gene was detected in all fetal tissues examined as early as the 6th week of gestation. Results also suggested that the fetal pancreas contains more 2.2 kb transcript than its adult counterpart. Therewith, BSDL was immuno-precipitated from fetal liver. The role of BSDL-gene expression during the fetal life is discussed.

Amino Acid Sequence↗

PubMed1995

Prevalence of thyroid autoantibodies is not increased in blood donors with hepatitis C virus infection.

Serum autoimmune reactions are found in many patients with hepatitis C. A high prevalence of thyroid dysfunction and antithyroid antibodies in patients with chronic hepatitis C was recently reported. We have compared the prevalence of thyroid dysfunction and antithyroid peroxidase antibodies in blood donors with hepatitis C virus (HCV) infection (study group) and in seronegative anti-HCV donors (control group). One hundred and ninety-two blood donors were studied: 96 were anti-HCV positive by ELISA 2 (48 males and 48 females; age 48 +/- 12.9 years, mean +/- SD), and 96 were HCV seronegative (55 males and 41 females; age 37 +/- 14.8 years). In all patients, serum TSH (0.25-4.2 mU/l) and fT4 (9-23 pmol/l) were measured by immunochemiluminiscent assays and antithyroid peroxidase antibodies (normal < 100 U/ml) by RIA. In all anti-HCV positive donors, hepatitis C viremia was tested using the nested polymerase chain reaction. Thyroid dysfunction was found in three females (3.1%) in the anti-HCV positive group (three cases of hypothyroidism), and in four (4.1%) anti-HCV negative blood donors (three cases of hypothyroidism, two females and one male; one case of hyperthyroidism, a female), (p = NS). Antithyroid peroxidase antibody titers were above normal values in 5 (5.2%) anti-HCV positive individuals and in eight (8.3%) anti-HCV negative blood donors (p = NS). These results do not show an increase prevalence of thyroid dysfunction and antithyroid peroxidase antibodies in blood donors with HCV infection when compared with a control group.

Adult↗

PubMed1995

Comparative study of two high doses of lymphoblastoid interferon in the treatment of chronic hepatitis C: influence on the levels of ALT, viraemia and histologic activity.

Ninety consecutive patients with chronic hepatitis C were included in a randomized, uncontrolled trial to compare the efficacy of two treatment regimens, 10 MU (group A) vs 5 MU (group B), of lymphoblastoid interferon, in a step-down schedule for 24 weeks. All of the patients had antibodies against the hepatitis C virus, and all but one were HCV RNA positive in serum. The origin of the infection was attributed to blood transfusion in 30 patients and classified as sporadic in 60 patients. During treatment reduction in the ALT levels as well as the elimination of viraemia was observed in both treated groups, although these changes did not correlate significantly with the interferon dose. Nine months after the end of therapy, a sustained response was achieved in 13.6% (12/88) of the patients. Relapse in group B (87.5%) was significantly higher than in group A (59.1%). The percentage of cases which remained with undetectable HCV RNA was significantly higher for the sustained responders (66.7%) than for the non-responders (11.8%) and relapser patients (2.4%). Repeated liver biopsies showed an overall significant reduction of all the subindices of histological activity from patients with sustained response, except for fibrosis. In short: the 10 MU dosing regimen of lymphoblastoid interferon was as efficient as the 5 MU dose as it brought about a similar improvement in ALT levels, histological activity and elimination of viraemia, albeit 10 MU proved significantly more effective in the prevention of a relapse among the responders after 24 weeks therapy.

Adult↗

PubMed1994

Insensitivity of cathepsin D gene to estradiol in endometrial cells is determined by the sequence of its estrogen responsive element.

In MCF7 cells, transcription of the lysosomal protease cathepsin D is stimulated by estrogens via a non-consensus estrogen responsive element (ERE). By contrast, in estrogen responsive Ishikawa endometrial cancer cells, the cathepsin D gene is unresponsive to estrogens. We now show that the transfected cathepsin D promoter, which can be induced by estrogens in several cell types, is insensitive in Ishikawa cells. The block is not due to a mutation in the cathepsin D promoter or estrogen receptor, but involves the cathepsin D ERE, and implies a C at position 3 of the ERE sequence. Our results suggest that in Ishikawa cells, cathepsin D insensitivity to estrogen most likely occurs through a specific interaction with the ER, or with an endometrial factor which may compete with the ER for binding to the cathepsin D ERE.

3T3 Cells↗

PubMed1994

Zinc ions block rectifier potassium channels and calcium activated potassium channels at the frog motor nerve endings.

The effect of Zn2+ on presynaptic currents was investigated on frog cutaneous pectoris nerve-muscle preparations. Nerve terminal spikes were recorded with extracellular electrodes placed in the perineurial sheaths of motor nerves. Zinc ions reversibly suppressed the component of the waveform associated with K+ currents--unmasking an inward current at the terminal--and induced repetitive firing when were applied to preparations perfused with calcium containing solutions. In experiments in which delayed rectifier channels were blocked by 3,4-diaminopyridine, Zn2+ caused a prolonged and reversible inward current associated with a slight decrease in the peak calcium current generated by 3,4-diaminopyridine. Zinc ions abolished the plateau calcium current produced by the simultaneous action of 3,4-diaminopyridine and tetraethylammonium. Finally, in all the preparations in which the exposure to Zn2+ was prolonged it was observed a dramatic and irreversible reduction of the presynaptic currents. These results suggest that Zn2+ has, at least, four different effects on presynaptic currents: (1) blockade of delayed rectifier potassium currents, (2) blockade of calcium-activated potassium currents, (3) blockade of calcium currents and (4) a delayed and irreversible disruption of all ionic conductances of the terminal.

4-Aminopyridine↗

PubMed1994

Analysis of urea distribution volume in hemodialysis.

According to the urea kinetic model it is considered that the urea distribution volume (V) is that of body water, and that it is distributed in only one compartment. Since the V value is different to measure, it is normal to use 58% of body weight, in spite of the fact that it may range from 35 to 75%. In this study, we have calculated the value of V by using an accurate method based on the total elimination of urea from the dialysate. We have studied the V, and also whether the different dialysis characteristics modify it. Thirty-five patients were included in this study, 19 men and 16 women, under a chronic hemodialysis programme. The dialysate was collected in a graduated tank, and the concentration of urea in plasma and in dialysate were determined every hour. Every patient received six dialysis sessions, changing the blood flow (250 or 350 ml/min), the ultrafiltration (0.5 or 1.5 l/h), membrane (cuprophane or polyacrylonitrile) and/or buffer (bicarbonate or acetate). At the end of the hemodialysis session, the V value ranged from 43 to 72% of body weight; nevertheless, this value was practically constant in every patient. The V value gradually increased throughout the dialysis session, 42.1 +/- 6.9% of body weight in the first hour, 50.7 +/- 7.5% in the second hour and 55.7 +/- 7.9% at the end of the dialysis session. The change of blood flow, ultrafiltration, membrane or buffer did not alter the results. The V value was significantly higher in men in comparison with women, 60.0 +/- 6.6% vs. 50.5 +/- 5.9% of body weight (p < 0.001).

Aged↗

PubMed1994

Is it easier to dialyze women than men?

Since the urea kinetic model is used as an objective method for monitoring dialysis, it has been possible to shorten reasonably its duration. From the usual practice, we have observed that it is easier to reduce the dialysis time in women as compared to men. The purpose of this study is to find out the reason for such observation and to corroborate it. Fourty-two patients, 25 males and 17 females on 3-weekly dialysis (3 h/session) were studied. All patients were dialyzed under the same dialysis characteristics: 3-hour sessions, blood flow 350 ml/min, ultrafiltration 1.5 liters/h, 1.5-m2 cuprophane membrane and bicarbonate buffer. The dialysate was collected in a graduated tank. Urea concentration in plasma and in the dialysate was measured. Then, the urea distribution volume, dialyzer clearance, the KT/V index and protein catabolic rate were calculated. The KT/V value was higher in women with respect to men, 1,017 +/- 0.10 versus 0.82 +/- 0.14 (p < 0.001). The urea distribution volume value was significantly higher in men as compared to women, 60.04 +/- 6.6 versus 51.48 +/- 5.88% (p < 0.001). There were no significant differences in dialyzer clearance, protein catabolic rate or body weight. In conclusion, under identical dialysis conditions, it is easier to dialyze women than men, because women's urea distribution volume is lower than men's.

Adult↗