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M Iturralde

Publications and source records attributed to M Iturralde.

At least 19 recordsLinked to original sources
PubMed2005

Characterization of the lipolytic pathways that mediate free fatty acid release during Fas/CD95-induced apoptosis.

We have undertaken a study to characterize the lipolytic pathway responsible for the generation of free fatty acids (FFA) during Fas/CD95-induced apoptosis in Jurkat cells. It was initially shown that the cellular lipid fraction that suffered the major quantitative decrease during Fas-induced apoptosis was that of phosphatidylcholine (PC). In addition, the secretion of palmitic acid-derived FFA was largely prevented by D609, an inhibitor of PC-specific phospholipase C (PC-PLC) and also by the diacylglycerol lipase (DAGL) inhibitor RHC-80267, suggesting that the secretion of these FFA during Fas-induced apoptosis is mediated by the generation of DAG by a PC-PLC activity and, sequentially, by a 1-DAGL activity which generates the FFA from its sn-1 position. The endocannabinoid 2-arachidonoyl glycerol (2-AG) should be generated as a sub-product of this pathway, but it did not accumulate inside the cells nor was secreted into the supernatant. Interestingly, the complete inhibition of free AA secretion during Fas-induced apoptosis was only achieved by using the AA trifluoromethylketone, which not only inhibits all types of phospholipase-A(2) (PLA(2)) activities, but also the described lytic activities on 2-AG. Using a combination of RHC-80267 and the iPLA(2)-specific inhibitor bromoenol lactone, it was shown that the DAGL pathway also cooperates with iPLA(2) in the generation of free arachidonate.

Apoptosis↗
PubMed2005

The concentration of apolipoprotein A-I decreases during experimentally induced acute-phase processes in pigs.

In this work, apolipoprotein A-I (ApoA-I) was purified from pig sera. The responses of this protein after sterile inflammation and in animals infected with Actinobacillus pleuropneumoniae or Streptococcus suis were investigated. Decreases in the concentrations of ApoA-I, two to five times lower than the initial values, were observed at 2 to 4 days. It is concluded that ApoA-I is a negative acute-phase protein in pigs.

Actinobacillus Infections↗
PubMed2004

ITIH4 (inter-alpha-trypsin inhibitor heavy chain 4) is a new acute-phase protein isolated from cattle during experimental infection.

We have isolated from calf serum a protein with an apparent M(r) of 120,000. The protein was detected by using antibodies against major acute-phase protein in pigs with acute inflammation. The amino acid sequence of an internal fragment revealed that this protein is the bovine counterpart of ITIH4, the heavy chain 4 of the inter-alpha-trypsin inhibitor family. The response of this protein in the sera was determined for animals during experimental bacterial and viral infections. In the bacterial model, animals were inoculated with a mixture of Actinomyces pyogenes, Fusobacterium necrophorum, and Peptostreptococcus indolicus to induce an acute-phase reaction. All animals developed moderate to severe clinical mastitis and exhibited remarkable increases in ITIH4 concentration in serum (from 3 to 12 times the initial values, peaking at 48 to 72 h after infection) that correlated with the severity of the disease. Animals with experimental infections with bovine respiratory syncytial virus (BRSV) also showed increases in ITIH4 concentration (from two- to fivefold), which peaked at around 7 to 8 days after inoculation. Generally, no response was seen after a second infection of the same animals with the virus. Because of the significant induction of the protein in the animals in the mastitis and BRSV infection models, we can conclude that ITIH4 is a new positive acute-phase protein in cattle.

Amino Acid Sequence↗
PubMed2001

Differential secretion of Fas ligand- or APO2 ligand/TNF-related apoptosis-inducing ligand-carrying microvesicles during activation-induced death of human T cells.

Preformed Fas ligand (FasL) and APO2 ligand (APO2L)/TNF-related apoptosis-inducing ligand (TRAIL) are stored in the cytoplasm of the human Jurkat T cell line and of normal human T cell blasts. The rapid release of these molecules in their bioactive form is involved in activation-induced cell death. In this study, we show by confocal microscopy that FasL and APO2L/TRAIL are mainly localized in lysosomal-like compartments in these cells. We show also by immunoelectron microscopy that FasL and APO2L/TRAIL are stored inside cytoplasmic compartments approximately 500 nm in diameter, with characteristics of multivesicular bodies. Most of these compartments share FasL and APO2L/TRAIL, although exclusive APO2L/TRAIL labeling can be also observed in separate compartments. Upon PHA activation, the mobilization of these compartments toward the plasma membrane is evident, resulting in the secretion of the internal microvesicles loaded with FasL and APO2L/TRAIL. In the case of activation with anti-CD59 mAb, the secretion of microvesicles labeled preferentially with APO2L/TRAIL predominates. These data provide the basis of a new and efficient mechanism for the rapid induction of autocrine or paracrine cell death during immune regulation and could modify the interpretation of the role of FasL and APO2L/TRAIL as effector mechanisms in physiological and pathological situations.

Antibodies, Monoclonal↗
PubMed2001

Binding of a surface protein of Staphylococcus aureus to cultured ovine mammary gland epithelial cells.

Staphylococcus aureus is the most persistent pathogen causing ovine mastitis. This study investigated S. aureus binding to cultured epithelial cells obtained from the mammary gland. A staphylococcal 145kDa cell wall adhesin, originally isolated from a bovine mastitis strain, was detected in lysostaphin-solubilized ovine mastitis strains and in the encapsulated strain A. This adhesin was able to bind to cultured ovine mammary gland epithelial cells (MGEC) and to a rat intestinal epithelial cell line (RIE-1), exhibiting different electrophoretic mobilities that could be attributable to protein polymorphism. Inhibition assays using antibodies against 145kDa adhesin and against whole bacteria showed the specificity of the binding to cells. The role of this protein in adherence was assessed by adherence inhibition tests carried out in vitro with radiolabeled bacteria and cultured epithelial cells. Preincubation of bacteria with antibodies against adhesin 145kDa or against strain c195 resulted in a statistically significant decrease of adherence. These experiments suggest that adherence of S. aureus to MGEC may be critical for colonization.

Adhesins, Bacterial↗
PubMed2001

Effect of slime on adherence of Staphylococcus aureus isolated from bovine and ovine mastitis.

The interactions between slime, Staphylococcus aureus and ovine mammary gland epithelial cells (MGEC) were studied in vitro. Suspensions of radiolabelled bacteria incubated with slime significantly increased the ability of S. aureus strains to adhere to a filter. When suspensions of radiolabelled bacteria were incubated with MGEC treated with trypsin, the ability of slime to improve S. aureus adherence was also shown, indicating that it was not dependent on cell membrane proteins. The interaction of radiolabelled bacteria with slime prior to the adherence test with MGEC demonstrated that the adherence process requires the interaction between slime and bacteria. This interaction is inhibited by anti-slime antibodies. This study provides evidence that a specific interaction between bacteria coated with slime and MGEC could be a critical part of mammary gland infection.

Animals↗
PubMed2000

Pig MAP/ITIH4 and haptoglobin are interleukin-6-dependent acute-phase plasma proteins in porcine primary cultured hepatocytes.

The acute-phase expression of pig MAP (major acute-phase protein)/ITIH4 (inter-alpha-trypsin inhibitor heavy chain 4) and haptoglobin were analysed in primary cultures of isolated pig hepatocytes in response to recombinant human (rh) cytokines: tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6), as well as to bacterial lipopolysaccharide (LPS). Analysis of pig MAP/ITIH4 and haptoglobin mRNAs was carried out by RT-PCR amplification. Secreted proteins from the cytokine-treated hepatocytes were quantified by immunochemical techniques. Time-course and dose-response experiments show that pig MAP/ITIH4 and haptoglobin belong to the type II acute-phase proteins, as they are specifically induced by rhIL-6 and not by rhTNF-alpha or rhIL-1. Stimulation of cultured pig hepatocytes with rhIL-6 for 48 h at doses of 1000 U.mL-1 showed a fourfold to fivefold increase in pig MAP/ITIH4 concentration in the medium, while the concentration of haptoglobin only increased twofold. A similar increase in the concentration of pig MAP/ITIH4 was also observed in media of LPS-treated hepatocytes with the simultaneous generation of IL-6 by the Kupffer cells present in the cultures. Albumin secretion decreased after stimulation with doses of 100 or 1000 U.mL-1 rhTNF-alpha, rhIL-1 or rhIL-6. Therefore, it can be concluded that pig MAP/ITIH4 behaves as a major acute-phase protein produced by porcine hepatocytes under the effect of inflammatory cytokines.

Acute-Phase Proteins↗
PubMed2000

Metabolism of n -9, n -6 and n -3 fatty acids in hepatoma Morris 7777 cells. Preferential accumulation of linoleic acid in cardiolipin.

The objective of this study was to investigate, using a pulse-chase technique, the different incorporation of (1-(14)C) n -9, n -6 and n 3 fatty acids into hepatoma lipids and their secretion to the culture medium. Docosahexaenoic acid (DHA) accumulated preferentially into the triacylglycerol while arachidonic acid (AA) did into the phospholipid fraction. DHA was poorly secreted to the culture medium whereas AA was secreted to a large extent. The fatty acids were initially esterified mainly into phosphatidylcholine and phosphatidylethanolamine. During the 24 h chase, a general shift from phosphatidylcholine to phosphatidylethanolamine was observed. Linoleic acid was esterified in cardiolipin to a much greater extent than any other fatty acid and it was not converted to more polyunsaturated fatty acids. The supplementation of the culture medium with polyunsaturated fatty acids had no inhibitory effect on the growth of the hepatoma cells, in marked contrast to observations made in other tumoral cells. The reasons for the resistance of the hepatoma cells to polyunsaturated fatty acid toxicity, including the possible antioxidant effect of linoleic acid accumulation in cardiolipin, are also discussed.

Albumins↗
PubMed1999

Fatty acid desaturation: effect of alphafetoprotein on alpha-linolenic acid conversion by fetal rat hepatocytes.

Freshly isolated fetal hepatocytes transformed 4.3, 8.5 and 19.2 pmol/min/10(6) cells of stearic, linoleic and alpha-linolenic acids, respectively, complexed to albumin or alpha-fetoprotein (AFP), to more unsaturated derivatives. Thus, fetal hepatocytes displayed high elongase and delta9, delta6, delta5-desaturase activities, as well as an ability to synthesize hexaene derivatives. Desaturase activities decreased when the time of culture of fetal hepatocytes (previous to incubation with the substrate) was prolonged, being practically undetectable after 24 h of culture. However, the rate of fatty acid uptake remained nearly constant. When AFP was used as the carrier the amount of hexaene fatty acid derivatives of alpha-linolenic acid recovered in cells was reduced up to 50% by albumin. This effect was associated with an increase of radioactivity found in the culture medium of hepatocytes incubated with AFP compared to albumin. Both observations taken together could be explained by an efflux of hexaene derivatives from cells caused by AFP.

Animals↗
PubMed1993

Adherence of ruminant mastitis Staphylococcus aureus strains to epithelial cells from ovine mammary gland primary cultures and from a rat intestinal cell line.

Staphylococcus aureus isolated from mastitis (14 bovine and 11 ovine strains) exhibited an ability to adhere to epithelial primary cultures from ovine mammary gland and to a rat epithelial cell line, RIE-1. Strain differences in the degree of adherence were observed in both cases. These differences were maintained when comparing different epithelial sources (rat vs. ovine). RIE-1 cells can thus be used as a model for studying staphylococcal adherence to epithelial cells. Changes in bacterial adherence were observed according to the bacterial growth phase. The magnitude of these changes differed among strains. Bacterial cell surface hydrophobicity was not related to the degree of adherence to mammalian epithelial cells.

Animals↗
PubMed1992

Effect of iron and retinoic acid on the control of transferrin receptor and ferritin in the human promonocytic cell line U937.

The effect of changes in iron availability and induction of differentiation on transferrin receptor expression and ferritin levels has been examined in the promonocytic cell line U937. Addition of iron (as 200 micrograms/ml saturated transferrin) or retinoic acid (1 microM) both caused approx. 70% reduction in the average number of surface transferrin receptors, while the iron chelator desferrioxamine caused an 84% increase. Comparable changes also occurred in the levels of transferrin receptor mRNA. Neither iron nor retinoic acid significantly altered the half-life of transferrin receptor mRNA in the presence of actinomycin D (approx. 75 min) but a 10-fold increase in stability occurred in the presence of desferrioxamine. Iron and retinoic acid both caused an increase in intracellular ferritin levels (approx. 4-and 3-fold, respectively), while desferrioxamine reduced ferritin levels by approx. two-thirds. The effect of iron and retinoic acid added together did not differ greatly from that of each agent alone. None of the treatments greatly affected levels of L-ferritin mRNA. Virtually no H-ferritin mRNA was detected in U937 cells. These results show that changes in ferritin and transferrin receptor caused by treatment with retinoic acid are similar to those induced by excess iron, and suggest that changes in these proteins during cell differentiation are due to redistribution of intracellular iron into the regulatory pool(s), rather than to iron-independent mechanisms.

Cell Differentiation↗
PubMed1992

An efficient microtest to study adherence of bacteria to mammalian cells.

A simple, fast and highly reproducible microtest was developed for in vitro adherence studies. A rat epithelial cell line was investigated for the adherence of clinical and subclinical ovine and bovine Staphylococcus aureus strains isolated from mastitis. Staphylococcus aureus strains differed in their ability to adhere to epithelial cells, the degree of adherence being dependent on the concentration of bacteria used in the test.

Animals↗
PubMed1991

Effect of alpha-fetoprotein and albumin on the uptake of polyunsaturated fatty acids by rat hepatoma cells and fetal rat hepatocytes.

The uptake of polyunsaturated fatty acids by rat hepatoma cells and fetal hepatocytes has been studied using albumin and alpha-fetoprotein (AFP) as carrier proteins. Both types of cells took up linoleic (18:2(n-6)) and linolenic (18:3(n-3)) fatty acids at the same rate when they were added complexed either to albumin or AFP at a 1:1 molar ratio. At 37 degrees C a greater incorporation of arachidonic acid (20:4(n-6)) and mainly docosahexaenoic acid (22:6(n-3)) was observed when these acids were bound to albumin (6.5 nmol 20:4(n-6); 6.4 nmol 22:6(n-3) /million cells) as compared to AFP (5.5 nmol 20:4(n-6); 4.3 nmol 22:6(n-3)/million cells). The 20:4(n-6) and 22:6(n-3) uptake seems to be inversely related to the apparent association constants (k'a) between these fatty acids and both proteins (1.3 and 1.5 x 10(-7) M-1 20:4(n-6) and 22:6(n-3) for albumin; 6.4 and 54.0 x 10(-7) M-1 20:4(n-6) and 22:6(n-3) for AFP). Experiments carried out at 4 degrees C indicated that binding of 20:4(n-6) (0.83 nmol/million cells in presence of albumin; 2.16 nmol/million cells in presence of AFP) and 22:6(n-3) (0.83 nmol/million cells in presence of albumin; 1.32 nmol/million cells in presence of AFP) by cell membranes was also inversely related to the k'a of these proteins. At 4 degrees C, the k'a of AFP and albumin for 20:4(n-6) and 22:6(n-3) changed considerably (12.7 and 9.6 x 10(-7) M-1 20:4(n-6) and 22:6(n-3) for albumin; 3.9 and 14.6 x 10(-7) M-1 20:4(n-6) and 22:6(n-3) for AFP) with respect to the k'a calculated at 37 degrees C. Hence, k'a values were higher for albumin and lower for AFP than the corresponding values at 37 degrees C. It was concluded that uptake by cells and interaction of fatty acids with cell membranes depend mainly on the k'a of fatty acids and carrier proteins at equilibrium.

Animals↗
PubMed1990

Linoleate and linolenate desaturation by rat hepatoma cells.

Morris 7777 rat hepatoma cells in culture possess high delta 6 and delta 5 desaturase activities over linolenic acid added to the medium as albumin or alpha-fetoprotein complexes. After 2 hours incubation with [1-14C] linolenic acid (7 microM), around 40% of the radioactivity was recovered in other polyene fatty acids, mainly pentaenes. After 24 hours incubation with this substrate the polyene derivatives raised to more than 60%. However, [1-14C] linoleic acid was poorly converted to other polyene fatty acids. Linoleic acid up to 58 microM concentration in the medium do not inhibited linolenic acid desaturation. Long-term supplementation with 50 microM linoleic or linolenic acid, which modified the fatty acid profile of hepatoma lipids, enhanced the desaturase activities against linoleic acid. Desaturase activities were not affected by the fatty acid protein carrier, alpha-fetoprotein or albumin.

Animals↗
PubMed1989

Interaction of rat alpha-fetoprotein and albumin with polyunsaturated and other fatty acids: determination of apparent association constants.

The interaction of fatty acids with rat alpha-fetoprotein and albumin was measured using a partition equilibrium method. alpha-Fetoprotein (AFP) displays one high-affinity binding site for fatty acids and albumin near two binding sites. The AFP association constants for most fatty acids were similar to those of albumin (in the 10(7) M-1 range) whereas for docosahexaenoic acid it was 9.7 x 10(8) M-1, about 50-fold higher than that corresponding to albumin. This difference justifies docosahexaenoic acid in fetal or neonatal serum being mainly bound to AFP and can indicate a highly specific role of AFP in the transport of this fatty acid.

Animals↗
PubMed1988

The role of nuclear cardiology procedures in the evaluation of cardiac function following heart transplantation.

Heart transplantation is, today, an accepted and recommended modality in the management of selected patients suffering from terminal heart disease. However, acute rejection and infection remain the major complications of this operation. Serial endomyocardial biopsy (EB), considered as the standard for diagnosis of cardiac rejection, is an invasive and delicate operation, not free of complications, even when done by skilled personnel in specialized centers. The object of this study was to compare and correlate between radionuclide ventriculography (RNV) and the histologic findings of EB. Furthermore, to validate the use of nuclear cardiology techniques that allow noninvasive, reliable, and rapid quantitation of ventricular function and myocardial perfusion for the diagnosis and management of rejection in patients with heart transplants. Radionuclide studies of left ventricular function were performed in 3 heterotopic heart transplant patients (HHT) with long term survival and early after the operation in 5 patients with HHT, 12 orthotopic heart transplants (OHT) and in 2 heart and lung transplants (HLT). Simultaneous EBs were performed in the early posttransplant patients and a histologic score for acute rejection was obtained. First pass (FP) and multigated equilibrium blood pool ventriculography, using the in vivo 99mTc-labelling of RBCs was used to measure left ventricular volumes (LVV) such as stroke volume (SV), end-diastolic volume (EDV), end-systolic volume (ESV), and both global and regional ejection fraction (EF, REF). The histological grading of acute rejection was classified into four groups: (1) no rejection, (2) mild rejection, (3) moderate rejection, and (4) severe rejection. The median of each LVV parameter was calculated and correlated with the EB using a nonparametric one way analysis of variance. A percentage change of LVVs was used rather than the difference of the calculated LVVs. During moderate acute rejection, SV had the highest correlation in P less than 0.004, followed by the EDV (P less than 0.05), and finally ESV (P less than 0.02). During severe acute rejection the correlation was SV (P less than 0.0008), EDV (P less than 0.001), and ESV (P less than 0.006). Myocardial perfusion scintigraphy using 201T1 was performed in the HHT patients, although, at this stage we have not attempted a correlation with the histologic findings. In one patient with long term survival OHT, increased 131I-metaiodobenzylguanidine (MIBG) myocardial uptake was evident during a rejection episode.

Adolescent↗
PubMed1985

Radionuclide studies in upper gastro-intestinal ulceration--are they reliable? A pilot study of 6 patients.

Endoscopy is at present the diagnostic technique of choice in the evaluation and detection of upper gastro-intestinal tract ulceration. Because of the physical discomfort suffered by patients during endoscopic examination, the search for better and less invasive methods of examination (especially in the unco-operative and seriously ill patient) continues. According to reports from the Orient, sucralfate (Ulsanic; Continental Ethicals) has prominent ulceravid properties. These properties are being used in conjunction with a tagging agent, in this case technetium-99m, as a diagnostic method for the detection and localization of upper gastro-intestinal ulceration. In this pilot study on 6 patients the positive findings of others regarding the specificity and promise of this method could not be confirmed.

Aged↗