Papers by ROSALBA FONTERIZ
European Journal of Anaesthesiology, 2010
The aims were to analyze the temporal evolution of neutrophil apoptosis, to determine the differe... more The aims were to analyze the temporal evolution of neutrophil apoptosis, to determine the differences in neutrophil apoptosis among 28-day survivors and nonsurvivors, and to evaluate the use of neutrophil apoptosis as a predictor of mortality in patients with septic shock. Materials and Methods: Prospective multicenter observational study carried out between July 2006 and June 2009. The staining solution study included 80 patients with septic shock and 25 healthy volunteers. Neutrophil apoptosis was assessed by fluorescein isothiocyanate (FITC)-conjugated annexin V and aminoactinomycin D staining. Results: The percentage of neutrophil apoptosis was significantly decreased at 24 hours, 5 days, and 12 days after the diagnosis of septic shock (14.8% ± 13.4%, 13.4% ± 8.4%, and 15.4% ± 12.8%, respectively; P b .0001) compared with the control group (37.6% ± 12.8%). The difference in apoptosis between 28-day surviving and nonsurviving patients was nonsignificant (P N .05). The mortality rate at 28 days was 53.7%.
Biomedicines, 2022
Alzheimer’s disease (AD) is the most frequent cause of dementia. After decades of research, we kn... more Alzheimer’s disease (AD) is the most frequent cause of dementia. After decades of research, we know the importance of the accumulation of protein aggregates such as β-amyloid peptide and phosphorylated tau. We also know that mutations in certain proteins generate early-onset Alzheimer’s disease (EOAD), and many other genes modulate the disease in its sporadic form. However, the precise molecular mechanisms underlying AD pathology are still unclear. Because of ethical limitations, we need to use animal models to investigate these processes. The nematode Caenorhabditis elegans has received considerable attention in the last 25 years, since the first AD models overexpressing Aβ peptide were described. We review here the main results obtained using this model to study AD. We include works studying the basic molecular mechanisms of the disease, as well as those searching for new therapeutic targets. Although this model also has important limitations, the ability of this nematode to gener...
Resumen del trabajo presentado al VI Spanish Worm Meeting, celebrado en Valencia del 9 al 10 de m... more Resumen del trabajo presentado al VI Spanish Worm Meeting, celebrado en Valencia del 9 al 10 de marzo de 2017.
Frontiers in Pharmacology, 2021
We have reported recently that the mitochondrial Na+/Ca2+ exchanger inhibitor CGP37157 extends li... more We have reported recently that the mitochondrial Na+/Ca2+ exchanger inhibitor CGP37157 extends lifespan in Caenorhabditis elegans by a mechanism involving mitochondria, the TOR pathway and the insulin/IGF1 pathway. Here we show that CGP37157 significantly improved the evolution with age of the sarcomeric regular structure, delaying development of sarcopenia in C. elegans body wall muscle and increasing the average and maximum speed of the worms. Similarly, CGP37157 favored the maintenance of a regular mitochondrial structure during aging. We have also investigated further the mechanism of the effect of CGP37157 by studying its effect in mutants of aak-1;aak-2/AMP-activated kinase, sir-2.1/sirtuin, rsks-1/S6 kinase and daf-16/FOXO. We found that this compound was still effective increasing lifespan in all these mutants, indicating that these pathways are not involved in the effect. We have then monitored pharynx cytosolic and mitochondrial Ca2+ signalling and our results suggest that...
International Journal of Molecular Sciences, 2020
Mitochondrial [Ca2+] plays an important role in the regulation of mitochondrial function, control... more Mitochondrial [Ca2+] plays an important role in the regulation of mitochondrial function, controlling ATP production and apoptosis triggered by mitochondrial Ca2+ overload. This regulation depends on Ca2+ entry into the mitochondria during cell activation processes, which is thought to occur through the mitochondrial Ca2+ uniporter (MCU). Here, we have studied the mitochondrial Ca2+ dynamics in control and MCU-defective C. elegans worms in vivo, by using worms expressing mitochondrially-targeted YC3.60 yellow cameleon in pharynx muscle. Our data show that the small mitochondrial Ca2+ oscillations that occur during normal physiological activity of the pharynx were very similar in both control and MCU-defective worms, except for some kinetic differences that could mostly be explained by changes in neuronal stimulation of the pharynx. However, direct pharynx muscle stimulation with carbachol triggered a large and prolonged increase in mitochondrial [Ca2+] that was much larger in contro...
Cells, 2020
Ca2+ is a ubiquitous second messenger that plays an essential role in physiological processes suc... more Ca2+ is a ubiquitous second messenger that plays an essential role in physiological processes such as muscle contraction, neuronal secretion, and cell proliferation or differentiation. There is ample evidence that the dysregulation of Ca2+ signaling is one of the key events in the development of neurodegenerative processes, an idea called the “calcium hypothesis” of neurodegeneration. Caenorhabditis elegans (C. elegans) is a very good model for the study of aging and neurodegeneration. In fact, many of the signaling pathways involved in longevity were first discovered in this nematode, and many models of neurodegenerative diseases have also been developed therein, either through mutations in the worm genome or by expressing human proteins involved in neurodegeneration (β-amyloid, α-synuclein, polyglutamine, or others) in defined worm tissues. The worm is completely transparent throughout its whole life, which makes it possible to carry out Ca2+ dynamics studies in vivo at any time, ...
American Journal of Physiology-Cell Physiology, 1992
Stimulation of fura-2-loaded bovine chromaffin cells with the nicotinic agonist 1,1-dimethyl-4-ph... more Stimulation of fura-2-loaded bovine chromaffin cells with the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP; 10 microM) or depolarization with high [K+] (50 mM) accelerated the entry of both Ca2+ and Mn2+, used here as a Ca2+ surrogate for Ca2+ channels. Removal of extracellular Na+ prevented the effects of DMPP but did not modify the effects of K+, indicating that Na+ is necessary for coupling of Ca2+ entry to the nicotinic receptor activation and that the ionophore associated with it is functionally impermeable to divalent cations. DMPP- as well as K(+)-evoked Ca2+ and Mn2+ influx were blocked completely by Ni2+ but only partially by dihydropyridines, suggesting that, in addition to L-type Ca2+ channels, other Ca2+ entry pathways may be present. Inactivation of Ca2+ channels, followed by comparing the rates of Mn2+ uptake at different time periods after the addition of DMPP or high K+, did not happen in the absence of extracellular Ca2+. When 1 mM Ca2+ was prese...
The FASEB Journal, 1992
We have studied the effects of cytochrome P450 inhibitors on the entry of Ca2+ and Mn2+, used her... more We have studied the effects of cytochrome P450 inhibitors on the entry of Ca2+ and Mn2+, used here as a Ca2+ surrogate for Ca2+ channels, in fura-2-loaded GH3 pituitary cells and bovine chromaffin cells depolarized with high-K+ solutions. Imidazole antimycotics were potent inhibitors (econazole greater than miconazole greater than clotrimazole greater than ketoconazole). alpha-Naphtoflavone and isosafrole, but not metyrapone, also inhibited the entry of Ca2+ and Mn2+ induced by depolarization. This inhibitory profile most resembles that reported for IA-type cytochrome P450. However, carbon monoxide (CO), a well-known cytochrome P450 antagonist, had no effect on Ca2+ (Mn2+) entry. Given the high selectivity of the imidazole antimycotics for the heme moiety, our results suggest that a hemoprotein closely related to cytochrome P450 (but insensitive to CO) might be involved in the regulation of voltage-gated Ca2+ channels. The inhibitory pattern was also similar to that previously reported for agonist-induced Ca2+ (Mn2+) influx in neutrophils and platelets, although CO was an efficient inhibitor in this case. These results pose the question of whether similarities in the sensitivity to cytochrome P450 inhibitors exhibited by receptor-operated and voltage-gated channels reflect unknown similarities either in structural features or regulation mechanisms.
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Oncotarget, Jan 22, 2017
Progressive decline in mitochondrial function is generally considered one of the hallmarks of agi... more Progressive decline in mitochondrial function is generally considered one of the hallmarks of aging. We have expressed a Ca(2+) sensor in the mitochondrial matrix of C. elegans pharynx cells and we have measured for the first time mitochondrial [Ca(2+)] ([Ca(2+)]M) dynamics in the pharynx of live C. elegans worms during aging. Our results show that worms stimulated with serotonin display a pharynx [Ca(2+)]M oscillatory kinetics that includes both high frequency oscillations (up to about 1Hz) and very prolonged "square-wave" [Ca(2+)]M increases, indicative of energy depletion of the pharynx cells. Mitochondrial [Ca(2+)] is therefore able to follow "beat-to-beat" the fast oscillations of cytosolic [Ca(2+)]. The fast [Ca(2+)]M oscillations kept steady frequency values during the whole worm life, from 2 to 12 days old, but the height and width of the peaks was progressively reduced. [Ca(2+)]M oscillations were also present with similar kinetics in respiratory chain c...
Oncotarget, 2016
Ca 2+ is a key signal transducer for muscle contraction. Continuous in vivo monitoring of intrace... more Ca 2+ is a key signal transducer for muscle contraction. Continuous in vivo monitoring of intracellular Ca 2+-dynamics in C. elegans pharynx muscle revealed surprisingly complex Ca 2+ patterns. Despite the age-dependent decline of pharynx pumping, we observed unaltered fast Ca 2+ oscillations both in young and old worms. In addition, sporadic prolonged Ca 2+ increases lasting many seconds or minutes were often observed in between periods of fast Ca 2+ oscillations. We attribute them to the inhibition of ATP-dependent Ca 2+-pumps upon energy depletion. Accordingly, food deprivation largely augmented the frequency of prolonged [Ca 2+ ] increases. However, paradoxically, prolonged [Ca 2+ ] increases were more frequently observed in young worms than in older ones, and less frequently observed in energy-deficient mitochondrial respiratory chain nuo-6 mutants than in wild-type controls. We hypothesize that young animals are more susceptible to energy depletion due to their faster energy consumption rate, while nuo-6 mutants may keep better the energy balance by slowing energy consumption. Our data therefore suggest that the metabolic state of the pharynx during feeding stimulation depends mainly on the delicate balance between the instant rates of energy production and consumption. Thus, in vivo monitoring of muscle Ca 2+ dynamics can be used as a novel tool to study cellular energy availability.
Advances in experimental medicine and biology, 2016
The role of mitochondria in intracellular Ca(2+) signaling relies mainly in its capacity to take ... more The role of mitochondria in intracellular Ca(2+) signaling relies mainly in its capacity to take up Ca(2+) from the cytosol and thus modulate the cytosolic [Ca(2+)]. Because of the low Ca(2+)-affinity of the mitochondrial Ca(2+)-uptake system, this organelle appears specially adapted to take up Ca(2+) from local high-Ca(2+) microdomains and not from the bulk cytosol. Mitochondria would then act as local Ca(2+) buffers in cellular regions where high-Ca(2+) microdomains form, that is, mainly close to the cytosolic mouth of Ca(2+) channels, both in the plasma membrane and in the endoplasmic reticulum (ER). One of the first targets proposed already in the 1990s to be regulated in this way by mitochondria were the store-operated Ca(2+) channels (SOCE). Mitochondria, by taking up Ca(2+) from the region around the cytosolic mouth of the SOCE channels, would prevent its slow Ca(2+)-dependent inactivation, thus keeping them active for longer. Since then, evidence for this mechanism has accum...
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2016
MICU1 and MICU2 are the main regulators of the mitochondrial Ca 2+ -uniporter (MCU), but their pr... more MICU1 and MICU2 are the main regulators of the mitochondrial Ca 2+ -uniporter (MCU), but their precise functional role is still under debate. We show here that MICU2 behaves as a pure inhibitor of MCU at low cytosolic [Ca 2+ ] ([Ca 2+ ] c ), though its effects decrease as [Ca 2+ ] c is increased an disappear above 7µM. Regarding MICU1, studying its effects is more difficult because knockdown of MICU1 leads also to loss of MICU2. However, while knockdown of MICU2 induces only a persistent increase in mitochondrial Ca 2+ uptake, knockdown of MICU1 induces also a peculiar use-dependent transient activation of MCU that cannot be attributed to the parallel loss of MICU2. Therefore, MICU1 is endowed with a specific inhibitory effect on MCU at low [Ca 2+ ] c , separate and kinetically different from that of MICU2. On the other hand, we and others have shown previously that MICU1 activates MCU at [Ca 2+ ] c above 2.5µM. Thus, MICU1 has a double role in MCU regulation, inhibitory at low [Ca 2+ ] c and activatory at high [Ca 2+ ] c . Summary MICU2 inhibits MCU at low [Ca 2+ ] c (<7µM) and produces no effect above that. MICU1 activates MCU at high [Ca 2+ ] c (>2.5µM) but also inhibits MCU at low [Ca 2+ ] c , and this inhibitory effect is genuine and kinetically different.
PLoS ONE, 2014
The effect of the intake of antioxidant polyphenols such as resveratrol and others on survival an... more The effect of the intake of antioxidant polyphenols such as resveratrol and others on survival and different parameters of life quality has been a matter of debate in the last years. We have studied here the effects of the polyphenols resveratrol and kaempferol added to the diet in a murine model undergoing long-term hypercaloric diet. Using 50 mice for each condition, we have monitored weight, survival, biochemical parameters such as blood glucose, insulin, cholesterol, triglycerides and aspartate aminotransferase, neuromuscular coordination measured with the rotarod test and morphological aspect of stained sections of liver and heart histological samples. Our data show that mice fed since they are 3-months-old with hypercaloric diet supplemented with any of these polyphenols reduced their weight by about 5-7% with respect to the controls fed only with hypercaloric diet. We also observed that mice fed with any of the polyphenols had reduced levels of glucose, insulin and cholesterol, and better marks in the rotarod test, but only after 1 year of treatment, that is, during senescence. No effect was observed in the rest of the parameters studied. Furthermore, although treatment with hypercaloric diets induced large changes in the pattern of gene expression in liver, we found no significant changes in gene expression induced by the presence of any of the polyphenols. Thus, our data indicate that addition of resveratrol or kaempferol to mice food produces an initial decrease in weight in mice subjected to hypercaloric diet, but beneficial effects in other parameters such as blood glucose, insulin and cholesterol, and neuromuscular coordination, only appear after prolonged treatments.
Oncogene, 2011
Pancreatic cancer remains as one of the most deadly cancers, and responds poorly to current thera... more Pancreatic cancer remains as one of the most deadly cancers, and responds poorly to current therapies. The prognosis is extremely poor, with a 5-year survival of less than 5%. Therefore, search for new effective therapeutic drugs is of pivotal need and urgency to improve treatment of this incurable malignancy. Synthetic alkyllysophospholipid analogues (ALPs) constitute a heterogeneous group of unnatural lipids that promote apoptosis in a wide variety of tumor cells. In this study, we found that the anticancer drug edelfosine was the most potent ALP in killing human pancreatic cancer cells, targeting endoplasmic reticulum (ER). Edelfosine was taken up in significant amounts by pancreatic cancer cells, and induced caspase-and mitochondrial-mediated apoptosis. Pancreatic cancer cells show a prominent ER, and edelfosine accumulated in this subcellular structure, inducing a potent ER stress response, with caspase-4, BAP31 and c-Jun NH 2-terminal kinase (JNK) activation, CHOP/GADD153 upregulation, and phosphorylation of eukaryotic translation initiation factor 2 α-subunit (elF2α), that eventually led to cell death. Oral administration of edelfosine in xenograft mouse models of pancreatic cancer induced a significant regression in tumor growth, and an increase in apoptotic index, as assessed by TUNEL assay and caspase-3 activation in the tumor sections. The ER stress-associated marker CHOP/GADD 153 was visualized in the pancreatic tumor isolated from edelfosine-treated mice, indicating a strong in vivo ER stress response. These results suggest that edelfosine exerts its proapoptotic action in pancreatic cancer cells, both in vitro and in vivo, through its accumulation in the ER, which leads to ER stress and apoptosis. Thus, we propose that endoplasmic reticulum could be a key target in pancreatic cancer, and edelfosine may constitute a prototype for the development of a new class of antitumor drugs targeting endoplasmic reticulum.
Neuroscience Letters, 1987
Calcium antagonist; Calcium channel; Chromaffin cell; Catecholamine release; Dihydropyridine rece... more Calcium antagonist; Calcium channel; Chromaffin cell; Catecholamine release; Dihydropyridine receptor; Adrenal medulla K-cvoked 4sCa uptake into, and catecholamine release from cat adrenomedullary tissues were potently inhibited by the dihydropyridine (DHP) Ca antagonist (+)-PN 200-110 (IC5~=0.8 nM). Verapamil and diltiazem were 2000-fold less potent and flunarizine behaved as the least potent blocker (ICs0 = 2980 riM): other DHP had a full range of potencies between (+)-PN 200-110 and verapamil. Thc ordcr of potencies for inorganic antagonists was Zn > Cd > La > Ni > Co > Mn > Mg. Since a great controversy exists on the sensitivity of Ca channels to various antagonists, this comprehensive study will facilitate the selection of appropiatc Ca antagonists to answer fundamental questions concerning chromaffin and neural ('a channels.
Journal of Structural Biology, 2010
A confocal study on the visualization of chromaffin cell secretory vesicles with fluorescent targ... more A confocal study on the visualization of chromaffin cell secretory vesicles with fluorescent targeted probes and acidic dyes.
Journal of Neurochemistry, 1989
Bovine adrenal medulla plasma membranes were purified by a differential centrifugation procedure ... more Bovine adrenal medulla plasma membranes were purified by a differential centrifugation procedure using sucrose and Urografin discontinuous density gradients; the membranes were enriched 10-12-fold in acetylcholinesterase activity and [3H]ouabain binding sites. Specific (+)- [3H]PN200-1 10 binding to these membranes amounted to 90% of total binding and was saturabie and of high affinity (KD = 41 pM; B,,, = 119 fmol/mg of protein) with a Hill coefficient close to I, a result suggesting the presence of a single, homogeneous population of dihydropyridine receptors. The association and dissociation rate constants were, respectively, 7.5 X 10' M -' rnin-l and 0.023 min-I. Unlabeled (+)-PN200-110 displaced (+)-[3H]PN200-1 10 binding with a potency 100-fold higher than (-)-PN200-1 10 (ICsO, 0.5 and 45 nM, respectively). Although the two enantiomers of BAY K 8644 completely displaced (+)-[3H]PN200-1 10 binding, they exhibited no stereoselectivity (ICsO, 69 and 83 nM, respectively). Whereas (i-)-nitrendipine very potently displaced (+)-[3H]PN200-l 10 binding (ICsO = 1.3 nM), verapamil and cinnarizine displaced the binding by only 30 and 40% at 1 F M , and diltiazem increased it by 20% at 10 pM. [3H]Ouabain bound to plasma membranes with a KD of 34 nM and a B,,, of 9.75 pmol/mg of protein, a figure 80-fold higher than the B,,, for (+)-PN200-1 10. t3H]Ouabain also bound to intact chromaffin cells with a B,,, of 244 fmol/ lo6 cells. This figure, together with the plasma membrane ouabain/(+)-PNZOO-I I0 ratio of 80, allows the calculation of 1,884 (+)-PN200-110 binding sites/cell. Assuming a 1: 1 stoichiometry, this means that intact chromaffin cells contain -1.5 L-type, dihydropyridine-associated, voltage-dependent Ca2+ channels/pm2 of surface area. Key Words: Chromaffin cell-Dihydropyridine receptor-Digitalis receptor-calcium channels. Castillo C. J. F. et al. (+)-PN200-110 and ouabain binding sites in purified bovine adrenomedullary plasma membranes and chromaffin cells.
International Journal of Cancer, 2000
Antitumor ether lipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH(3); edelfo... more Antitumor ether lipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH(3); edelfosine) induces apoptosis in cancer cells, sparing normal cells. We have found that the apoptotic action of ET-18-OCH(3) required drug uptake and Fas in the target cell. Failure to accomplish one of these requirements prevents cell killing by the ether lipid. In human lymphoid leukemic cells, ET-18-OCH(3) does not promote Fas or FasL expression and ET-18-OCH(3)-induced apoptosis is not inhibited by pre-incubation with an anti-Fas blocking antibody that abrogates cell killing mediated by Fas/FasL interactions. ET-18-OCH(3)-resistant normal human Fas-positive fibroblasts do not incorporate ET-18-OCH(3), but undergo apoptosis upon ET-18-OCH(3) microinjection. Murine fibroblasts L929 and L929-Fas, stably transfected with human Fas cDNA, do not incorporate ET-18-OCH(3) and are resistant to its action when added exogenously. Microinjection of ET-18-OCH(3) induces apoptosis in L929-Fas cells, but not in wild-type L929 cells. Confocal laser scanning microscopy shows that ET-18-OCH(3) induces Fas clustering and capping during triggering of ET-18-OCH(3)-induced apoptosis. Microinjection-induced apoptosis and Fas clustering are specific for the molecular structure of ET-18-OCH(3). Our data indicate that ET-18-OCH(3) induces apoptosis via Fas after the ether lipid is inside the cell, and this Fas activation is independent of the interaction of Fas with its natural ligand FasL. This explains the selective action of ET-18-OCH(3) on tumors since only cancer cells incorporate sufficient amounts of the drug.
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Papers by ROSALBA FONTERIZ