Antonio Gonzalez

Institute of Molecular Pathology Biomarkers; University of Extremadura · Department of Physiology

One of the most important constituents of the cell membrane is arachidonic acid. Lipids forming part of the cellular membrane can be metabolized in a variety of cellular types of the body by a family of enzymes termed phospholipases: phospholipase A2, phospholipase C and phospholipase D. Phospholipase A2 is considered the most important enzyme type...

We have investigated the effects of melatonin on major pathways related with cellular proliferation and energetic metabolism in pancreatic stellate cells. In the presence of melatonin (1 mM, 100 µM, 10 µM, or 1 µM), decreases in the phosphorylation of c-Jun N-terminal kinase and of p44/42 and an increase in the phosphorylation of p38 were observed....

Pancreatic stellate cells (PSCs) are major players in the fibrosis that develops in pancreatic cancer. Due to the rapid growth of the tumoral tissue, a hypoxia condition is established, under which cells included in the mass can proliferate. Additionally, melatonin has gained attention as an agent with therapeutic potential against pancreatic cance...

Pancreatic stellate cells (PSCs) play a pivotal role in the fibrosis that develops in pancreatic diseases such as chronic pancreatitis or pancreatic cancer. In the tumour microenvironment, a hypoxia condition develops under which cancer cells are able to proliferate. The growth of fibrotic tissue contributes to hypoxia. In this study, the effect of...

Pancreatic stellate cells (PSCs), the main cell type responsible for the development of fibrosis in pancreatic cancer, proliferate actively under hypoxia. Melatonin has received attention as a potential antifibrotic agent due to its anti-proliferative actions on PSCs. In this work, we investigated the activation of the PI3K/Akt/mTOR pathway and the...

In solid tumors, the development of vasculature is, to some extent, slower than the proliferation of the different types of cells that form the tissue, both cancer and stroma cells. As a consequence, the oxygen availability is compromised and the tissue evolves toward a condition of hypoxia. The presence of hypoxia is variable depending on where th...

Introduction: In rapid growing tumoral tissue a hypoxia condition is established, under which cells included in the mass proliferate. Pancreatic stellate cells (PSCs) are major players in the fibrosis that develops in pancreatic cancer. Additionally, melatonin exerts potential therapeutic actions against pancreatic cancer. Aims:We investigated the...

In certain diseases of the pancreas, pancreatic stellate cells form an important part of fibrosis and are critical for the development of cancer cells. A hypoxic condition develops within the tumor, to which pancreatic stellate cells adapt and are able to proliferate. The consequence is the growth of the tumor. Melatonin, the product of the pineal...

Pancreatic stellate cells (PSC) play a major role in the formation of fibrotic tissue in pancreatic tumors. On its side, melatonin is a putative therapeutic agent for pancreatic cancer and inflammation. In this work, the actions of melatonin on PSC subjected to hypoxia were evaluated. Reactive oxygen species (ROS) generation reduced (GSH) and oxidi...

This manuscript is a review focused onto the role of astrocytes in the protection of neurons against oxidative stress and how melatonin can contribute to the maintenance of brain homeostasis. The first part of the review is dedicated to the dependence of neurons on astrocytes by terms of survival under oxidative stress conditions. Additionally, the...

The harmful effects of food-occurring oxidized amino acids, namely, aminoadipic acid (AAA), dityrosine (DTYR), L-kynurenine (KN), kynurenic acid (KA) and 3-nitrotyrosine (3NT), were studied on differentiated CACO-2 cells by flow cytometry and quantification of glutathione (GSH), and allysine. Cells were exposed to food-relevant doses (200 μM) of ea...

Background information Pancreatic stellate cells play a key role in the fibrosis that develops in diseases such as pancreatic cancer. In the growing tumor, a hypoxia condition develops under which cancer cells are able to proliferate. The growth of fibrotic tissue contributes to hypoxia. In this study the effect of hypoxia (1% O2) on pancreatic ste...

We have examined the effects of α-aminoadipic acid, an oxidized derivative from the amino acid lysine, on the physiology of mouse pancreatic acinar cells. Changes in intracellular free-Ca²⁺ concentration, the generation of reactive oxygen species, the levels of carbonyls and thiobarbituric-reactive substances, cellular metabolic activity and trypsi...

In this study, we have examined the effects of luzindole, a melatonin receptor‐antagonist, on cultured pancreatic stellate cells. Intracellular free‐Ca²⁺ concentration, production of reactive oxygen species (ROS), activation of mitogen‐activated protein kinases (MAPK), endoplasmic reticulum stress and cell viability were analyzed. Stimulation of ce...

In this study, the effects of melatonin (1 μM–1 mM) on pancreatic stellate cells (PSC) have been examined. Cell viability and proliferation, caspase-3 activation, and the expression of cyclin A and cyclin D were analyzed. Our results show that melatonin decreased PSC viability in a time- and concentration-dependent manner. This effect was not inhib...

In this work we have studied the effects of pharmacological concentrations of melatonin (1 µM–1 mM) on pancreatic stellate cells (PSC). Cell viability was analyzed by AlamarBlue test. Production of reactive oxygen species (ROS) was monitored following CM-H2DCFDA and MitoSOX Red-derived fluorescence. Total protein carbonyls and lipid peroxidation we...

Tartrazine (E102) is a synthetic food coloring, which belongs to the class of mono azo dyes and is known to cause numerous health problems. The current research aimed to evaluate the effect of this food dye on the enzymatic activity of amylase, lipase and proteases after a subchronic ingestion in Swiss mice. Additionally, an in vitro digestion mode...

Background In this work we studied the effects of the melatonin receptor-antagonist luzindole (1 μM–50 μM) on isolated mouse pancreatic acinar cells. Methods Changes in intracellular free-Ca²⁺ concentration, reactive oxygen species production and trypsin secretion were analyzed. Results Luzindole induced increases in [Ca²⁺]i that diminished CCK-8...

In this study, the effects of pharmacological concentrations of melatonin (1 μM–1 mM) on human pancreatic stellate cells (HPSCs) have been examined. Cell type–specific markers and expression of melatonin receptors were analyzed by western blot analysis. Changes in intracellular free Ca²⁺ concentration were followed by fluorimetric analysis of fura-...

It was already described that genetic inhibition of NUPR1 induces tumor growth arrest. In this paper we studied the metabolism changes after NUPR1 downregulation in pancreatic cancer cells, which results in a significant decrease of OXPHOS activity with a concomitant lower ATP production which precedes the necrotic cell death. We demonstrated that...

Desde los inicios de la investigación científica, los animales han estado íntimamente relacionados con el desarrollo de los experimentos y la consecución de resultados que, a posteriori, han permitido llegar a conclusiones extrapolables a la realidad humana. Este tipo de uso de los animales ha sido, no obstante, objeto de controversia y polémicas d...

In the present study we employed cultured pancreatic stellate cells to study the effect of the organophosphorus insecticide dimethoate on pancreatic cell physiology. Esterase activity, cell viability, reactive oxygen species generation and Ca2+ mobilization were examined. Our results show that dimethoate (0.1, 1 and 10 μM) induced a concentration-d...

We studied the effects of the tartrazine-metabolite sulfanilic acid on the physiology of pancreatic AR42J cells. Sulfanilic acid (1 μM-1 mM) induced a slow and progressive increase in intracellular free-calcium concentration that reached a plateau. The effect of sulfanilic acid was not concentration-dependent. Stimulation of cells with thapsigargin...

Genetic inhibition of NUPR1 induces tumor growth arrest. Inactivation of NUPR1 expression in pancreatic cancer cells results in lower ATP production, higher consumption of glucose with a significant switch from OXPHOS to glycolysis followed by necrotic cell death. Importantly, induction of necrosis is independent of the caspase activity. We demonst...

Resumen: Desde los inicios de la investigación científica, los animales han estado íntimamente relacionados con el desarrollo de los experimentos y la consecución de resultados que, a posteriori, han permitido llegar a conclusiones extrapolables a la realidad humana. Este tipo de uso de los animales ha sido, no obstante, objeto de controversia y po...

Ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) is an organoselenium radical scavenger compound, which has strong antioxidant and anti-inflammatory effects. However, evidence suggests that this compound could exert deleterious actions on cell physiology. In this study we have analyzed the effect of ebselen on rat pancreatic AR42J cells. Cytosolic...

1. Introduction - Is a common practice amongst professors to put an exam, grade it and forget the assessment until the next call. Nevertheless, the analysis of students’ answers is essential to improve the quality of the examination procedures. Being aware of the importance that assessment has, universities are introducing exams quality evaluation...

Ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) is an organoselenium radical scavenger compound, which has strong antioxidant and anti-inflammatory effects. Because of its properties, it may be protective against injury to the nervous tissue. However, evidence suggests that its glutathione peroxidase activity could underlie certain deleterious ac...

Copper is essential for several important cellular processes, but an excess of copper can also lead to oxidative damage. In brain, astrocytes are considered to play a pivotal role in the copper homeostasis and antioxidative defence. To investigate whether antioxidants and copper chelators can modulate the uptake and the toxicity of copper ions in b...

The goal of this study was to evaluate the potential activation of the nuclear factor erythroid 2-related factor and the antioxidant-responsive element (Nrf2-ARE) signaling pathway in response to melatonin in isolated mouse pancreatic acinar cells. Changes in intracellular free-Ca(2+) concentration were followed by fluorimetric analysis of fura-2-l...

Resveratrol (3,4',5-trihydroxy-trans-stilbene) is an antioxidant widely employed in cell physiology studies. It has been reported that it interferes with fura-2-derived fluorescence, making the employment of this dye nonviable. In this work, the interference of resveratrol with fura-2 determinations of intracellular free-Ca(2+) concentration ([Ca(2...

Neurotransmission unavoidably increases mitochondrial reactive oxygen species. However, the intrinsic antioxidant defense of neurons is weak and hence the mechanism whereby these cells are physiologically protected against oxidative damage is unknown. Here we found that the antioxidant defense of neurons is repressed owing to the continuous protein...

Melatonin, the product of the pineal gland, possesses antioxidant, anti-inflammatory, and antitumor properties in different tissues, in addition to its role as regulator of biological rhythms. In this study, the effects of pharmacological concentrations of melatonin (1 μM-1 mM) on pancreatic stellate cells (PSCs) have been examined. Cell viability...

Melatonin, a derivative of the amino acid tryptophan, was initially thought to be mainly produced and secreted by the pineal gland; with time, it was also found in other tissues and organs, and even in plants. Since its discovery, the study of the role of the indole in cellular homeostasis has generated impressive data, which have led researchers t...

Intracellular Ca(2+) overload has been considered a common pathological precursor of pancreatic injury. In this study, the effects of melatonin on Ca(2+) mobilization induced by cholecystokinin octapeptide (CCK-8) in freshly isolated mouse pancreatic acinar cells have been examined. Changes in intracellular free Ca(2+) concentration were followed b...

Ebselen is a seleno-organic compound that causes cell death in several cancer cell types. The mechanisms underlying its deleterious effects have not been fully elucidated. In this study, the effects of ebselen (1μM-40μM) on AR42J tumor cells have been examined. Cell viability was studied using AlamarBlue(®) test. Cell cycle phase determination was...

In this study we studied Ca2+ responses to different stimuli in pancreatic acinar cells subjected to culture conditions. Cells were isolated from adult mice pancreas and were subjected to culture conditions along one week. Changes in intracellular free Ca2+ concentration were monitored by single cell fluorescence analysis of fura-2-loaded cells. Mi...

In the present work, we have evaluated the effect of an acute addition of melatonin on cholecystokinin octapeptide (CCK-8)-evoked Ca(2+) signals and amylase secretion in mouse pancreatic acinar cells. For this purpose, freshly isolated mouse pancreatic acinar cells were loaded with fura-2 to study intracellular free Ca(2+) concentration ([Ca(2+)]c)...

The seleno-organic compound and radical scavenger ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) have been extensively employed as an anti-inflammatory and neuroprotective compound. However, its glutathione peroxidase activity at the expense of cellular thiols groups could underlie certain deleterious actions of the compound on cell physiology....

Resveratrol (3,4',5-trihydroxy-trans-stilbene), a phytoalexin naturally found in grapes and red wine, is a redox-active compound endowed with significant positive activities. In this study, the effects of resveratrol on intracellular free Ca(2+) concentration ([Ca(2+)](c)) and on cell viability in tumoral AR42J pancreatic cells are examined. The re...

This work was designed in order to gain an insight on the mechanisms by which antioxidants prevent pancreatic disorders. We have examined the properties of cinnamtannin B-1, which belongs to the class of polyphenols, against the effect of hydrogen peroxide (H(2)O(2)) in mouse pancreatic acinar cells. We have studied Ca(2+) mobilization, oxidative s...

In this study we have used rat hippocampal astrocytes in culture to investigate the effect of ethanol on kainate-induced glutamate secretion. Our results show that kainate (10 μM to 500 μM) stimulated glutamate release from astrocytes. Preincubation of astrocytes in the presence of ethanol induced a concentration-dependent (1mM-50mM) inhibition of...

Calcium flux through L-type voltage-activated calcium (Cav1) channels is crucial for regulating brain functions including memory formation and behavior. Alterations in Ca²+ homeostasis have been linked to many cognitive disorders, and understanding the regulation of this process is crucial for their remedy. Therefore, here, we have evaluated the ef...

Melatonin reduces proliferation in many different cancer cell lines. Thus, melatonin is considered a promising antitumor agent, promoting apoptosis in tumor cells while preserving viability of normal cells. Herein, we examined the effects of melatonin on the pancreatic AR42J tumor cell line. We have analyzed cytosolic-free Ca(2+) concentration ([Ca...

1. Cinnamtannin B-1 is a naturally occurring A-type proanthocyanidin that belongs to a class of polyphenols widely distributed throughout the plant kingdom and exhibiting anti-oxidant properties. 2. In the present study, we examined the effects of cinnamtannin B-1 on cholecystokinin octapeptide (CCK-8)-evoked Ca2+ mobilization, reactive oxygen spec...

Melatonin is produced following circadian rhythm with high levels being released at night and has been implicated in the regulation of physiological processes in major tissues, including the pancreas. The aim of our study was to examine the effects of melatonin on intracellular free Ca(2+) concentration ([Ca(2+) ](c)) in AR42J pancreatic cells. Our...

Alcohol abuse is a major cause of pancreatitis, a condition that can manifest as both acute necroinflammation and chronic damage (acinar atrophy and fibrosis). Pancreatic acinar cells can metabolize ethanol via the oxidative pathway, which generates acetaldehyde and involves the enzymes alcohol dehydrogenase and possibly cytochrome P4502E1. Additio...

In the central nervous system (CNS), both neurones and astrocytes play crucial roles. On a cellular level, brain activity involves continuous interactions within complex cellular circuits established between neural cells and glia. Although it was initially considered that neurones were the major cell type in cerebral function, nowadays astrocytes a...

The effects of alcohol on Ca(2+) signalling remains poorly understood. Here we have investigated the effects of acute ethanol exposure on Ca(2+) influx in mouse pancreatic acinar cells. Cells were loaded with fura-2 and the changes in fluorescence were monitored by spectrofluorimetry and imaging analysis. Stimulation of cells with 20 pM cholecystok...

A significant percentage of patients with pancreatitis often presents a history of excessive alcohol consumption. Nevertheless, the patho-physiological effect of ethanol on pancreatitis remains poorly understood. In the present study, we have investigated the early effects of acute ethanol exposure on CCK-8-evoked Ca2+ signals in mouse pancreatic a...

In the central nervous system both neurons and astrocytes play crucial roles. On a cellular level, brain activity involves continuous interactions within complex cellular circuits established between neural cells and glia. Despite it was initially considered that neurons were the major cell type in cerebral function, nowadays astrocytes are conside...

Alcohol consumption has long been associated with cell damage, and it is thought that it is involved in approximately 40% of cases of acute pancreatitis. In the present study, we have investigated the early effects of acute ethanol exposure on cholecystokinin octapeptide (CCK-8)-evoked calcium (Ca2+) signals in mouse pancreatic acinar cells. Cells...

In this study we have investigated the effect of ethanol on [Ca2+]c by microfluorimetry and glutamate secretion using an enzyme-linked system, in rat hippocampal astrocytes in culture. Our results show that ethanol (1-200 mM) evoked a dose-dependent increase in glutamate secretion. 50 mM ethanol, a concentration within the range of blood alcohol le...

We have investigated the effect of the seleno-organic compound and radical scavenger ebselen on rat hippocampal astrocytes in culture. Throughout our study we carried out determinations of [Ca2+](c) in fura-2-loaded cells by single cell imaging, glutamate secretion employing an enzymatic-based assay and GFAP expression, which was monitorized by imm...

We have employed rat hippocampal astrocytes in culture to investigate the effect of ethanol on reactive oxygen species (ROS) production as well as its effect on [Ca2+]c and GFAP expression. Cells were loaded with the fluorescent probes fura-2 and H2DCFDA for the determination of changes in [Ca2+]c and ROS production respectively, employing spectrof...

In the present study we have employed single cell imaging analysis to monitor the propagation of cholecystokinin-evoked Ca(2+) waves in mouse pancreatic acinar cells. Stimulation of cells with 1 nM CCK-8 led to an initial Ca(2+) release at the luminal cell pole and subsequent spreading of the Ca(2+) signal towards the basolateral membrane in the fo...

The effect of hydrogen peroxide (H2O2) on cytosolic free calcium concentration ([Ca2+]c) as well as its effect on glutamate secretion in rat hippocampal astrocytes have been the aim of the present research. Our results show that 100 microM H2O2 induces an increase in [Ca2+]c, that remains at an elevated level while the oxidant is present in the per...

Reactive oxygen species and related oxidative damage have been implicated in the initiation of acute pancreatitis, a disease characterised in its earliest stages by disruption of intracellular Ca2+ homeostasis. The present study was carried out in order to establish the effect of the organic pro-oxidant, tert-butylhydroperoxide (tBHP), on the mobil...

We have employed confocal laser scanning microscopy to investigate how intracellular free calcium concentration ([Ca2+]i) is influenced by hydrogen peroxide (H2O2) in collagenase-dispersed mouse pancreatic acinar cells. In the absence of extracellular calcium, treatment of cells with increasing concentrations of H2O2 resulted in an increase in [Ca2...

In the present study, we have investigated the effect of ethanol on amylase release in response to cholecystokinin octapeptide (CCK-8). We have also studied the effect of ethanol on cytosolic free Ca(2+) concentration ([Ca(2+)](c)) and reactive oxygen species (ROS) production by loading of cells with fura-2 and 5-(and-6)-chloromethyl-2',7'-dichloro...

This paper studies the effect of H(2)O(2) on mitochondrial responses evoked by CCK-8 (cholecystokinin 8) in mouse pancreatic acinar cells. Cytosolic ([Ca(2+)](c)) and mitochondrial ([Ca(2+)](m)) free-calcium concentrations, mitochondrial inner membrane potential (psi(m)) and FAD autofluorescence were monitored using confocal laser scanning microsco...

This study employed confocal laser scanning microscopy to monitor the effect of H2O2 on cytosolic as well as mitochondrial calcium (Ca2+) concentrations, mitochondrial inner membrane potential (ψm ) and flavine adenine dinucleotide (FAD) oxidation state in isolated mouse pancreatic acinar cells. The results show that incubation of pancreatic acinar...

We evaluated the long-term effects of a fat-enriched diet (virgin olive oil) on calcium mobilization and amylase secretion induced by cholecystokinin-octapeptide (CCK-8) in rat pancreatic acinar cells. Olive oil is a major component of the Mediterranean diet, and its role in human health is actively being debated. Weaning male Wistar rats (21 d old...

In the present study we have studied the changes in the intracellular reduction-oxidation state in mouse pancreatic acinar cells following stimulation with cholecystokinin octapeptide (CCK-8) and its dependence on Ca2+ mobilization. In our investigations cytosolic Ca2+ concentration and reactive oxygen species (ROS) production were determined by lo...

In the present study, we have employed confocal laser scanning microscopy to investigate the effect that stimulation of mouse pancreatic acinar cells with the secretagogue cholecystokinin (CCK) has on mitochondrial activity. We have monitored changes in cytosolic as well as mitochondrial Ca2+ concentrations, mitochondrial membrane potential and FAD...

Store-mediated Ca2+ entry (SMCE) is one of the main pathways for Ca2+ influx in non-excitable cells. Recent studies favour a secretion-like coupling mechanism to explain SMCE, where Ca2+ entry is mediated by an interaction of the endoplasmic reticulum (ER) with the plasma membrane (PM) and is modulated by the actin cytoskeleton. To explore this pos...

The present study investigates the effect of reactive oxygen species (ROS) on actin filament reorganisation and its relevance to exocytosis in pancreatic acinar cells. Treatment of pancreatic acini with cholecystokinin (CCK-8) induced spatial and temporal changes in actin filament reorganisation with an initial depolymerisation of the apical actin...

In fura-2 loaded isolated mouse pancreatic acinar cells, xanthine oxidase (XOD)-catalyzed reactive oxygen species (ROS) generation caused an increase in the cytosolic Ca(2+) concentration ([Ca(2+)](i)) by release of Ca(2+) from intracellular stores. The ROS-induced Ca(2+) signals showed large variability in shape and time-course and resembled in pa...

This minireview is an attempt to put together some of the recent advances regarding the implications of mitochondria in Ca2+ homeostasis. Although the main role of this cytoplasmic organelle is ATP supply to the cell, during the past years strong evidence has been accumulated supporting an active role of these organelles in Ca2+ handling by the cel...

In the present study we have employed fura-2 loaded isolated mouse pancreatic acinar cells to monitor the effect that xanthine oxidase (XOD)-catalyzed reactive oxygen species generation presents on Ca(2+) mobilization by the secretagogue cholecystokinin octapeptide (CCK-8). Our results show that perfusion of pancreatic acinar cells with CCK-8 at a...

In pancreatic acinar cells analysis of the propagation speed of secretagogue-evoked Ca2+waves can be used to examine coupling of hormone receptors to intracellular signal cascades that cause activation of protein kinase C or production of arachidonic acid (AA). In the present study we have investigated the role of cytosolic phospholipase A2(cPLA2)...

In the present study we have investigated cytosolic and mitochondrial Ca(2+) signals in isolated mouse pancreatic acinar cells double-loaded with the fluorescent probes fluo-3 and rhod-2. Stimulation of pancreatic acinar cells with 500 nm acetylcholine caused release of Ca(2+) from intracellular stores and produced cytosolic Ca(2+) signals in form...

Release of Ca(2+) from inositol (1,4,5)-trisphosphate-sensitive Ca(2+) stores causes "capacitative calcium entry," which is mediated by the so-called "Ca(2+) release-activated Ca(2+) current" (I(CRAC)) in RBL-1 cells. Refilling of the Ca(2+) stores or high cytoplasmic [Ca(2+)] ([Ca(2+)](cyt)) inactivate I(CRAC). Here we address the question if also...

We employed confocal laser-scanning microscopy to monitor cholecystokinin (CCK)-evoked Ca(2+) signals in fluo-3-loaded mouse pancreatic acinar cells. CCK-8-induced Ca(2+) signals start at the luminal cell pole and subsequently spread toward the basolateral membrane. Ca(2+) waves elicited by stimulation of high-affinity CCK receptors (h.a.CCK-R) wit...

In pancreatic acinar cells stimulation of different intracellular pathways leads to different patterns of Ca2+ signaling. Bombesin induces activation of both phosphatidylinositol 4,5-bisphosphate (PIP2)-specific phospholipase C (PLC) and phospholipase D (PLD). The latter leads to generation of diacylglycerol (DAG) in addition to that produced by ac...