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Obesity induced by a leptin deficiency causes extensive damage in brain due to a large increase in oxidative stress. Mitochondria have a central role in this neural damage. Leptin receptors have a wide expression in the brain and its absence is associated with reduced mitochondrial respiration by a decoupled electron transport chain and a significantly increased complex II activity which is major player in mitochondrial free radical generation. The consequences are an abrupt reduction in ATP production and a reduced activity of the tricarboxylic acid (TCA) cycle, evidence of dysfunction of processes related to energy production, the reduction of which contributes to multiple brain pathologies. Melatonin, a major protector of mitochondria against free radicals with a significant influence on glucose metabolism, has been shown to counteract these conditions. In the present study the main respirasome expression was recovered by melatonin, with a reduction in complex II activity and the complex II dependent free radical generation. Additionally, melatonin normalized the TCA cycle. Reduction in ATP synthesis was caused by UCP2 activation. The uninterrupted sensation of starvation due to leptin deficiency involves impairments in glucose metabolism, which was reversed by melatonin via negatively acting on hexokinase II, a key regulator of glycolysis and major contributor to the Warburg effect. Hexokinase II reduction was accompanied by a significant Bcl-2 reduction, inducing a delicate readjustment of pro and anti-apoptotic proteins in the mitochondria to preserve cell survival, which was associated with a marked reduction of the Bax activator, Puma, observed in obese animals treated by melatonin.

Annual Scientific Meeting of the European Society for Clinical Investigation “Precision medicine for healthy ageing” (52nd. 2018. Barcelona)

Objectives There are no tools or biomarkers for a quantitative analysis of sarcopenia. Study design Cross-sectional study of the diagnosis of sarcopenia in 200 independent adults aged 70 years or over. Main outcome measures Sarcopenia was defined as loss of muscle mass together with low strength and/or loss of physical performance. We considered different clinical parameters and assayed potential blood biomarkers (cell energetic metabolism, muscle performance, inflammation, infection and oxidative stress). Results The prevalence of sarcopenia was 35.3% in women and 13.1% in men, and it was significantly associated with advanced age, a low functional performance in the lower extremities, deficient weekly consumption of kilocalories, risk of malnutrition, and drug use for the digestive system. A close relationship was found between sarcopenia, pre-frailty and depressed mood. With these confounding variables, we observed that products of lipid peroxidation were closely associated with sarcopenia in independent older adults (frail participants and those with severe dependence had been excluded from the sample). The best multivariate model proposed was able to predict 67.6% of the variance in sarcopenia, with a power of discrimination of 93.5%. Additional analyses considering lipid levels, fat mass, dyslipidemia, use of lipid-lowering drugs and hypertension confirmed this close association between lipid peroxidation and sarcopenia. Conclusions Given the difficulty in the diagnosis of sarcopenia in clinical practice, we suggest the use of blood circulating products of lipid peroxidation as potential biomarkers for an early diagnosis of sarcopenia in independent older adults.

Sarcopenia is the gradual loss of skeletal muscle mass, strength and quality associated with aging. Changes in body composition, especially in skeletal muscle and fat mass are crucial steps in the development of chronic diseases. We studied the effect of overweight on skeletal muscle tissue in elderly people without reaching obesity to prevent this extreme situation. Overweight induces a progressive protein breakdown reflected as a progressive withdrawal of anabolism against the promoted catabolic state leading to muscle wasting. Protein turnover is regulated by a network of signaling pathways. Muscle damage derived from overweight displayed by oxidative and endoplasmic reticulum (ER) stress induces inflammation and insulin resistance and forces the muscle to increase requirements from autophagy mechanisms. Our findings showed that failure of autophagy in the elderly deprives it to deal with the cell damage caused by overweight. This insufficiently efficient autophagy leads to an accumulation of p62 and NBR1, which are robust markers of protein aggregations. This impaired autophagy affects myogenesis activity. Depletion of myogenic regulatory factors (MRFs) without links to variations in myostatin levels in overweight patients suggest a possible reduction of satellite cells in muscle tissue, which contributes to declined muscle quality. This discovery has important implications that improve the understanding of aged-related atrophy caused by overweight and demonstrates how impaired autophagy is one of the main responsible mechanisms that aggravate muscle wasting. Therefore, autophagy could be an interesting target for therapeutic interventions in humans against muscle impairment diseases.

Correctly distinguishing preeclampsia (PE), gestational hypertension (GH), and intrauterine growth retardation (IUGR) is a challenge for clinicians due to existing similarities. In our previous study, we showed that serum strontium (Sr) levels were elevated in preeclamptic women compared to healthy and GH pregnant women at the end of pregnancy. The main aim of this study was to evaluate Sr and oxidative stress in PE at the time of symptoms onset and before and compare it with IUGR/GH.Samples collected at symptoms onset included 77 preeclamptic women and 72 women diagnosed with IUGR/GH divided into two groups according to the gestational extraction week (34 and ≥ 34). Fifteen patients were also serialized until delivery. Samples collected before symptoms onset included 140 women who developed early-onset PE (E-PE, n = 9), late-onset PE (L-PE, n = 13), IUGR (n = 9), GH (n = 32) and no pathologies (n = 77). Strontium, placental growth factor (PlGF), soluble fms-like tyrosine kinase 1 (sFlt-1), uric acid (UA), creatinine, lipid peroxidation, and total antioxidant activity (TAA) were measured.Mean Sr, sFlt-1/PIGF ratio, UA, and lipid peroxidation/TAA ratio levels were significantly higher (p = 0.002,0.0001,0.0001 and = 0.03, respectively) and estimated glomerular filtration rate (eGFR) and TAA significantly lower (p = 0.0008 and 0.0001, respectively) in E-PE vs other pathologies when gestational extraction week was34. There was a significant correlation between Sr and eGFR (r = 0.43, p = 0.02), sFlt-1/PIGF ratio (r = 0.56, p = 0.002), TAA and gestational week of sampling (r = -0.45, p = 0.02) and UA (r = -0.82, p 0.0001) in the E-PE serial samples. No differences were found in Sr levels before symptoms onset.Serum Sr concentration and oxidative status are increased in E-PE when compared to other pathologies at the time of symptoms onset. More studies are needed to elucidate the causes of Sr levels elevation and its role in the pathophysiology of PE.

Leptin, as a nutritional inhibitor by repressing food intake, is critical compromised in the major common forms of obesity. Skeletal muscle is the main effector tissue for energy expenditure modifications by the effect of endocrine axes, such as leptin signaling. Our study has been carried out using skeletal muscle from leptin-deficient animal model, in order to ascertain the importance of this hormone in eating disorders. Here we report that leptin-deficiency stimulates an uncontrolled oxidative phosphorylation metabolism, resulting in an excess of energy production that culminates in mitochondrial dysfunction. Thus, different nutrient sensing pathways are perturbed, loosing proteostasis and promoting lipid anabolism, that induces myofiber degeneration and drives oxidative type I fiber conversion. Melatonin treatment plays a significant role in regulating energy homeostasis and fuel utilization. This study reveals melatonin to be a decisive mitochondrial function-fate regulator, with implications for resembling physiological energy requirements and targeting glycolytic type II fibers recovery.

// Rute Loureiro 1 , Silvia Magalhaes-Novais 1, 2 , Katia A. Mesquita 1, 2 , Ines Baldeiras 1, 3 , Isabel S. Sousa 1, 2 , Ludgero C. Tavares 1, 2 , Ines A. Barbosa 1 , Paulo J. Oliveira 1 , Ignacio Vega-Naredo 1, 4 1 CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal 2 Department of Life Sciences, University of Coimbra, Coimbra, Portugal 3 School of Medicine, University of Coimbra, Coimbra, Portugal 4 Department of Morphology and Cell Biology, University of Oviedo, Oviedo, Spain Correspondence to: Ignacio Vega-Naredo, e-mail: naredo@gmail.com Keywords: melatonin, mitochondria, metabolism, dichloroacetate, cancer stem cells Received: January 19, 2015 Accepted: May 08, 2015 Published: May 20, 2015 ABSTRACT Although melatonin oncostatic and cytotoxic effects have been described in different types of cancer cells, the specific mechanisms leading to its antitumoral effects and their metabolic context specificity are still not completely understood. Here, we evaluated the effects of melatonin in P19 embryonal carcinoma stem cells (CSCs) and in their differentiated counterparts, cultured in either high glucose medium or in a galactose (glucose-free) medium which leads to glycolytic suppression and increased mitochondrial metabolism. We found that highly glycolytic P19 CSCs were less susceptible to melatonin antitumoral effects while cell populations relying on oxidative metabolism for ATP production were more affected. The observed antiproliferative action of melatonin was associated with an arrest at S-phase, decreased oxygen consumption, down-regulation of BCL-2 expression and an increase in oxidative stress culminating with caspase-3-independent cell death. Interestingly, the combined treatment of melatonin and dichloroacetate had a synergistic effect in cells grown in the galactose medium and resulted in an inhibitory effect in the highly resistant P19 CSCs. Melatonin appears to exert its antiproliferative activity in P19 carcinoma cells through a mitochondrially-mediated action which in turn allows the amplification of the effects of dichloroacetate, even in cells with a more glycolytic phenotype.

We are members of the cROS group and the INPROTEOLYS and INEUROPA networks. This work was supported by FISS-13-RD12/0043/0030, FISS-13-RD12/0043/0017 and FISS-14-PI13/02741 (Instituto de Salud Carlos III, Spanish Ministry of Economy and Competitiveness), FC-15-GRUPIN14-071 (FICYT, PCTI, Principado de Asturias), and FEDER funds. A. Rubio-González and Z. Pérez-Martínez are pre-doctoral hired (FC-15-GRUPIN14-071) from FICYT, PCTI (Principado de Asturias). Y. Potes is a FISS pre-doctoral fellow (FI14/00405) from the Instituto de Salud Carlos III (Spanish Ministry of Economy and Competitiveness)., Rubio-González, A., Bermejo-Millo, J.C., de Luxán-Delgado, B., Potes, Y., Pérez-Martínez, Z., Boga, J.A., Vega-Naredo, I., Caballero, B., Solano, J.J., Coto-Montes, A.

Although numerous studies have demonstrated the harmful effect of excessive fructose consumption at the systemic level, there is little information on its effects in the central nervous system. The purpose of the present work was to study the cellular alterations related to oxidative stress and protein quality control systems induced by a high-fructose diet in the brain of Syrian hamsters and their possible attenuation by exogenous melatonin. High-fructose intake induced type II diabetes together with oxidative damage, led to alterations of the unfolded protein response by activating the eIF2α branch, and impaired the macroautophagic machinery in the brain, favoring the accumulation of aggregates labeled for selective degradation and neurodegeneration markers such as β-amyloid (1-42), tau-p-S199, and tau-p-S404. Melatonin attenuated the manifestation of type II diabetes and reduced oxidative stress, deactivated eIF2α, and decreased tau-p-S404 levels in the brain of animals fed a high-fructose diet.

The Syrian hamster Harderian gland exhibits sexually dimorphic porphyrin biosynthesis, wherein the female glands display an extraordinarily high concentration of porphyrins. Damage derived from this production of porphyrins, mediated by reactive oxygen species, causes the glands to develop autophagic processes, which culminate in detachment-derived cell death; these cells normally play a central role in the secretory activity of the gland. The main aim of this study was to analyze how a change in the redox state impacts autophagy. Female Syrian hamsters were treated daily with melatonin (25 μg, subcutaneously) at ZT 10 for 1-2 months (N-acetyl-5-methoxytryptamine), an endogenous antioxidant that ameliorates the deleterious effects of free radicals via a variety of mechanisms. The length of treatment affected the redox balance, the autophagy machinery, and the activation of p53 and NF-κB. One-month treatment displaces redox balance to the antioxidant side, promotes autophagy through a p53-mediated mechanism, and increases cell detachment. Meanwhile, 2-month treatment restores redox balance to the oxidant side, activates NF-κB reducing autophagy to basal levels, increases number of type II cells, and reduces number of detached cells. Our results conclude that the redox state can modulate autophagy through redox-sensitive transcriptions factors. Additionally, these findings support a hypothesis that ascribes differences in the autophagic-lysosomal pathway to epithelial cell types, thereby restricting detachment-induced autophagic cell death to epithelial cell type I.

This work was supported by the Instituto de Salud Carlos III (Spanish Ministry of Economy and Competitiveness) (PI13/02741 and PI17/02009); the PCTI (Principado de Asturias) (GRUPIN14-071); and FEDER funds. J.C. B-M. is a pre-doctoral fellow at the Instituto de Investigación Sanitaria del Principado de Asturias (ISPA). Y.P. is a FISS pre-doctoral fellow at the Instituto de Salud Carlos III (Spanish Ministry of Economy and Competitiveness) (FI14/00405)., Potes, Y., Pérez-Martinez, Z., Bermejo-Millo, J.C., Rubio-Gonzalez, A., Fernandez-Fernández, M., Bermudez, M., Arche, J.M., Solano, J.J., Boga, J.A., Oliván, M., Caballero, B., Vega-Naredo, I., Coto-Montes, A.

Aging is commonly defined as a physiological phenomenon associated with morphological and functional deleterious changes in which oxidative stress has a fundamental impact; therefore, readjusting the oxidative balance should have beneficial effects. In our study, we tested the antioxidant melatonin in old mouse brains and showed positive effects at the cellular and mitochondrial levels. Melatonin attenuated β-amyloid protein expression and α-synuclein deposits in the brain compared to aged group. Furthermore, oxidative stress was increased by aging and induced the nuclear translocation of nuclear factor-kappa B (NF-κB), which was suppressed by melatonin treatment. The antioxidant mitochondrial expression, superoxide dismutase 2 (SOD2), was increased in both control and melatonin-treated old mice, despite the different activation states of the NF-κB pathway. The NF-κB pathway was activated in the old mice, which may be explained by this group’s response to the increased oxidative insult; this insult was inhibited in melatonin-treated animals, showing this group an increase in active mitochondria population that was not observed in old group. We also report that melatonin is capable of restoring the mitochondrial potential of age-damaged neurons. In conclusion, melatonin’s beneficial effects on brain aging are linked to the increase in mitochondrial membrane potential and SOD2 expression, which probably reduces the mitochondrial contribution to the oxidative stress imbalance.

Oxidative stress has been reported to increase during aging and conditions of hypoxia. Although low oxygen saturation has a key role in the development of several age-related diseases, the underlying mechanisms are still unknown. We analyzed the relationship between aging and hypoxia by examining oxidative stress and inflammation-related cytokines. We collected blood samples from three volunteer experimental groups, consisting of one group of normoxic middle-aged people and two groups of individuals older than 75 years, which comprised a subgroup of normoxic subjects and another with oxyhemoglobin saturation lower than 95% (hypoxic). Our results showed a fall in antioxidant defenses in older people with hypoxia. TNF-alpha, the first element in the cytokine cascade, was significantly increased in the aged population, implying that aging is accompanied by a gradual increase in this inflammatory biomarker. IL-6 was not associated with aging, but it was highly elevated under hypoxia conditions in elderly subjects. Thus, these parameters could be used as biological markers of different inflammatory processes triggered by oxidative stress induced by a decrease in antioxidant defenses in the elderly population, with TNF-alpha as an indicator of chronic processes, such as aging, and IL-6 as a marker for acute responses, such as hypoxia.

Aged spleens from senescence-accelerated prone mice 8 (SAMP8) and senescence-accelerated resistant mice 1 (SAMR1) were examined to determine whether sex or melatonin had an effect on oxidative stress-related immune impairments. We observed that the immunosenescence of SAMP8 mice was associated with a redox imbalance, leading to an age-related increase in oxidative damage, resulting from a decrease in antioxidant defense and protease activity. Moreover, increased apoptotic cell death, a decrease in proliferative activity and the loss of NF-kappaB activation were also related to the immunodeficiency seen in SAMP8 compared to SAMR1 mice. Females demonstrated higher oxidative stress-related alterations in the immune response, and subsequent, melatonin treatment provided the best protective effects. Pathways involved in autophagy were upregulated in SAMP8 as an adaptive response to oxidative stress, in an attempt to rescue the cell from increased apoptosis and age-related immunodeficiency. However, the NF-kappaB signaling and autophagic processes were unaffected by treatment with melatonin. Therefore, we propose a key role for NF-kappaB signaling and autophagy in the oxidative stress-related immunosenescent spleens of SAMP8 mice.

The sedentary lifestyle of modern society along with the high intake of energetic food has made obesity a current worldwide health problem. Despite great efforts to study the obesity and its related diseases, the mechanisms underlying the development of these diseases are not well understood. Therefore, identifying novel strategies to slow the progression of these diseases is urgently needed. Experimental observations indicate that melatonin has an important role in energy metabolism and cell signalling; thus, the use of this molecule may counteract the pathologies of obesity. In this study, wild-type and obese (ob/ob) mice received daily intraperitoneal injections of melatonin at a dose of 500 μg/kg body weight for 4 weeks, and the livers of these mice were used to evaluate the oxidative stress status, proteolytic (autophagy and proteasome) activity, unfolded protein response, inflammation and insulin signalling. Our results show, for the first time, that melatonin could significantly reduce endoplasmic reticulum stress in leptin-deficient obese animals and ameliorate several symptoms that characterize this disease. Our study supports the potential of melatonin as a therapeutic treatment for the most common type of obesity and its liver-associated disorders.

To assess whether oxidative damage in some tissues was related to their melatonin concentration, endogenous melatonin levels and the age-linked protein and lipid damage in spleen, thymus and liver in 5-month-old SAM P8 mice were examined. The results show that high levels of melatonin in spleen and thymus correlate with lower protein and lipid damage. The liver, which had much lower melatonin concentrations than the other two tissues, had much higher levels of oxidatively damaged protein, as measured by carbonyl values. These results add new evidence concerning the protective role of endogenous melatonin as an antioxidant agent, and suggest that a treatment with this molecule might help to reduce age-associated functional deficits in many organs, including those of the immune system.

The blind subterranean mole rat Spalax ehrenbergi superspecies has evolved adaptive strategies to cope with underground stress. Hypoxia is known to stimulate reactive oxygen species generation; however, mechanisms by which Spalax counteracts oxidative damage have not been investigated before. We studied in Spalax the oxidative status of the Harderian gland (HG), an organ which is particularly vulnerable to oxidative stress in many rodents. With regard to the sexual dimorphism found in this gland, differences between males and females were determined and compared to the surface-dwelling Syrian hamster. Our results show, for the first time, that Spalax exhibits remarkably low biomolecular damage, which implies the existence of physiological strategies to avoid oxidative damage under fluctuating O(2) and CO(2) levels existing in the mole rat's subterranean niche. Correspondingly, main antioxidant enzymes, such as superoxide dismutase (SOD), catalase, and glutathione reductase (GR), exhibited high activities in both genders; in particular, remarkably high levels were measured in SOD. SOD and GR activities showed statistically significant differences between sexes. Melatonin, an important circadian agent is also a very important antioxidant molecule and is synthesized in the Harderian glands (HGs) of Spalax. Therefore, the possible interaction between antioxidant enzymes and melatonin is suggested.

The senescence-accelerated mouse (SAM) is a useful animal model to study aging or age-associated disorder. In the present study, we have used a multidisciplinary approach to the characterization of changes that occur in aging and in the modelling of brain aging. The SAMP8 mouse at 5 months of age exhibited an increase in gliosis and molecular oxidative damage. Likewise, we found that superoxide dismutase activity decreased compared with age-matched SAMR1 while there were no differences in activity of catalase and glutathione reductase. These results indicate that the decrease of superoxide dismutase may be involved in the increase of oxidative stress in brain of SAMP8 at younger stages. This suggestion is supported by an increase in the expression of alpha-synuclein together with phosphorylated tau protein, which is concurrent with the decline of that antioxidant enzyme. Alpha-synuclein aggregates are invariably associated with tau pathologies and our results demonstrate that alpha-synuclein accumulation is a potent inducer of tau pathologies not only in neurodegenerative diseases but also in normal aging. These results also imply that SAMP8 are exposed to elevated levels of oxidative stress from an early age, and that could be a very important cause of the senescence-related impairments and degeneration in the brain seen in this strain.

Double-muscled syndrome in cattle improves meat tenderness. However, the nature of the proteolytic processes associated with this phenomenon remains unknown. The aim of this study was to monitor changes in the activity of cathepsins (B, B + L, D and H) during meat aging and their gradual release from lysosomes to the cytosol in the longissimus muscle of yearling bulls of two breeds from northern Spain (Asturiana de los Valles and Asturiana de la Montana) showing three genotypes for muscular hypertrophy (mh/mh, mh/+ and + / +). The data showed that the pattern of cathepsin activity during meat aging paralleled variations in tenderness in the different genotypes studied. Maximal cathepsin D activity and minimal cathepsin H activity were recorded during meat aging times ranging from 3 to 21 days. The activities of cathepsins B and B + L were lower than that of cathepsin D at the established time points (3, 7, 14 and 21 days post-slaughter). The role of these enzymes in the activation of cathepsin D is discussed. All cathepsins showed similar action patterns, with high levels early on in the aging process and lower levels at later times. This pattern depended on the genotype and was significantly faster (P≤0.05) in meat from mh/mh animals, intermediate in meat from mh/+ animals and slower in meat from normal (+/+) animals of both breeds. Copyright © 2006 Society of Chemical Industry

Quinolinic acid is a well-known excitotoxin that induces oxidative stress and damage. In the present study, oxidative damage to biomolecules was followed by measuring lipid peroxidation and protein carbonyl formation in rat brain tissue culture over a period of 24 hr of exposure to this prooxidant agent at a concentration of 0.5 mm. Quinolinic acid enhanced lipid peroxidation in an early stage of tissue culture, and protein carbonyl at a later stage. These data confirm and extend previous studies demonstrating that quinolinic acid can induce significant oxidative damage. Melatonin, an antioxidant and neuroprotective agent with multiple actions as a radical scavenger and signaling molecule, completely prevented these prooxidant actions of quinolinic acid at a concentration of 1 mm. Morphological lesions and neurotoxicity induced by quinolinic acid were evaluated by light microscopy. Quinolinic acid produced extensive apoptosis/necrosis which was significantly attenuated by melatonin. Cotreatment with melatonin exerted a profound protective effect antagonizing the neurotoxicity induced by quinolinic acid. Glutathione reductase and catalase activities were increased by quinolinic acid and these effects were antagonized by melatonin. Furthermore, melatonin induced superoxide dismutase activity. Quinolinic acid and melatonin acted independently and by different mechanisms in modulating antioxidant enzyme activities. Our findings using quinolinic acid and melatonin clearly demonstrate that such changes should always be seen in the context of oxidative neurotoxicity and antioxidant neuroprotection.

Melatonin acts through several specific receptors, including membrane receptors (MT(1) and MT(2)) and members of the RZR/ROR nuclear receptors family, which have been identified in a large variety of mammalian and nonmammalian cells types. Both membrane and nuclear melatonin receptors have been partially characterized in Harderian gland of the Syrian hamster. Nevertheless, the identities of these receptors were unknown until this study, where the coexistence of MT(1) and RORalpha(1) in this gland was determined by nested RT-PCR followed by amplicon sequencing and Western-blot. Furthermore, the cellular localization of both receptors was determined by immunohistochemistry. Thus, MT(1) receptor was localized exclusively at the basal side of the cell acini, supporting the hypothesis that this receptor is activated by the pineal-synthesized melatonin. On the contrary, although a RORalpha(1)-immunoreactivity was observed in nuclei of epithelial cells of both sexes, an extranuclear specific staining, which was more frequently among those cells of males, was also seen. The implication of this possible nuclear exclusion of RORalpha(1) on the role of this indoleamine against oxidative stress is discussed.

The Syrian hamster Harderian gland has as the remarkable feature of an extraordinary rate of porphyrin production, even higher than the liver. The low activity of the last enzyme of the route gives rise to the accumulation of the uncomplex porphyrins in the female glands. Moreover, due to the localization of the Harderian gland, porphyrins exposed to light produce reactive oxygen species and, thus, the gland presents a physiological oxidative stress, with a great number of sings of degeneration, but without compromising the gland integrity. The appearance of abnormal features in this gland was largely described in the past, but the significance is interpreted for the first time in this study. We have found that autophagic processes are the first result of an elevated porphyrin metabolism, as it is observed in both sexes. This mechanism is considered, in this case, as a constant renovation system that allows the normal gland activity to be sustained. Furthermore, there is a second procedure, invasive processes toward connective tissue, which even occasionally reach blood vessels with intravasation of damaged gland components into the bloodstream. This effect is a consequence of a strong oxidative stress environment that is mainly observed in the female gland, resembling to tumoral progression. Both mechanisms, autophagy and invasive processes, have to be implied in the maintenance of the gland integrity.

The Syrian hamster Harderian gland (HG) has a marked sexual dimorphism and exhibits an extraordinary rate of porphyrinogenesis. The physiological oxidative stress, derived from constant porphyrin production, is so high that the HG needs additional survival autophagic mechanisms to fight against this chronic exposure, provoking the triggering of a holocrine secretion in female glands that forms two types of secretory masses: intra-tubular-syncytial and inter-tubular-syncytial masses. The aim of this work was to study the development of this inter-tubular holocrine secretion. To approach this task, we have considered that the steps developed during the formation of the so-called invasive masses consist of the growth of epithelial cells, cell detachment from the basal lamina and invasion of surrounding tissues. The presence of these masses, particularly in the female HG, are closely linked to sexual dimorphism in redox balance and to alterations in the expression of certain factors such as cytokeratins, P-cadherin, matrix metalloproteinases, cathepsin H, proliferating cell nuclear antigen, p53, CD-31 and vascular endothelial growth factor, which seem to be involved in tissue remodeling. The results document unusual mechanisms of secretion in Syrian hamster HG: an extraordinary system of massive secretion through the conjunctive tissue, disrupting the branched structure of the gland.

The present study investigated the changes in several erythrocyte oxidative stress biomarkers in hypoxic elderly individuals to analyze the deleterious effects of low oxyhemoglobin saturation in an elderly population. We collected blood samples from one normoxic middle-aged group and two groups composed of individuals older than 75 years of age: one normoxic group and one hypoxic group. Aging appeared to provoke a defective erythrocyte antioxidant defense associated with increased oxidative damage in the elderly population. Acute hypoxia activated an insufficient antioxidant defense response as suggested by the oxidative damage observed. The oxidative imbalance presented in older participants and increased in hypoxia participants had a direct effect on glyceraldehyde-3-phosphate dehydrogenase cell distribution. Oxidative stress levels altered Band 3 protein and mediated caspase-3 activation in erythrocyte from the aged group although it was not extended to hypoxic individuals. Therefore, aged participants appeared to activate an insufficient antioxidant response against hypoxia-related oxidative stress. Language: en

The Syrian hamster Harderian gland (HG) has a large porphyrin metabolism with a sexual dimorphism, showing male HGs much lower porphyrin concentrations than female glands. Damage derived from this production of porphyrins, displayed by reactive oxygen species, forces the gland to develop morphological changes that must have a physiological significance. Thus, oxidative stress is present in two states: mild oxidative stress in male HGs and extreme oxidative stress in female HGs. Cathepsins data gave indirect indications about the presence of programmed cell death affecting the lysosomal pathway, especially in female HGs, which showed an accumulation of autophagic bodies. Our results showed different degrees of autophagy in Syrian hamster HGs depending on sex and probably controlled by the redox-sensitive transcription factors: NFkappaB and p53. The discovery of these sexual dimorphisms in redox signaling and in autophagy corroborates previous findings and underlines the key role of reactive oxygen species in the regulation of autophagy. In addition, in this paper we propose a physiological significance for these phenomena: male HGs develop a survival autophagy, while in female HGs, autophagy culminates in a detachment-derived cell death that plays a central role in its secretory activity, leading to a massive glandular secretion.

The Syrian hamster Harderian gland (HG) displays a huge porphyrins metabolism with sexual dimorphism. Even in male Syrian hamsters with much lower porphyrins concentration than female HG, this activity is higher than in the liver. The damage derived from constant porphyrin production, displayed by reactive oxygen species, forces the gland to develop mechanisms that allow it to continue with its normal physiology. The survival strategy of the Harderian gland is mainly based on autophagic processes that are considered as a constant renovation system. Our results show different autophagy mechanisms in Syrian hamster HG, macroautophagy and other lysosomal‐like processes such as chaperone‐mediated autophagy, depending on sex and probably related to oxidative stress status. This chapter describes the methods used by us to characterize the autophagic processes that are being physiologically developed by this organ under normal conditions.

We studied the effect of age and melatonin on cell death processes in brain aging. Senescence-accelerated prone mice 8 (SAMP8) and senescence-accelerated resistant mice (SAMR1) at 5 and 10 months of age were used as models of the study. Melatonin (10 mg/kg) or its vehicle (ethanol at 0.066%) was administered in the drinking water from 1 to 9 months of age. Neurodegeneration, previously shown in the aged brain of SAMP8 and SAMR1 at 10 months of age, may be due to a drop in age-related proteolytic activities (cathepsin D, calpains, and caspase-3). Likewise, lack of apoptotic and macroautophagic processes were found, without apparent modification by melatonin. However, the caspase-independent cell death, owing to high p53 and apoptosis-inducing factor (AIF) levels, might be an alternative pathway of cell death in the aged brain. The main effects of melatonin treatment were observed in the aged SAMR1 mice; in this strain we observed a marked increase in antioxidant activity (catalase and superoxide dismutase). Likewise, a key antioxidant role of apoptosis-related proteins, Bcl-2 and AIF, was suggested in the aged brain of SAM mice, which was clearly influenced by melatonin. Moreover, the age-related increase of lysosomal activity of cathepsin B and a lysosomal membrane-associated protein 2 supports the possibility of the maintenance of lysosomal viability in addition to age-related impairments of the proteolytic or macroautophagic activities. The effectiveness of melatonin against the oxidative stress-related impairments and apoptosis during the aging process is, once more, corroborated in this article.

The flank organ of the Syrian hamster shows a biodynamic response to androgenic stimulation and is, therefore, a suitable model for the study of androgenic effects on hair and sebaceous glands. This organ is susceptible to programmed cell death (PCD), a prominent feature associated with sexual organ adjustment. In the present report, the type of PCD (apoptosis or autophagy) exhibited by this organ was evaluated. Caspase-3 activity, indicative of apoptosis, was not detectable in flank organ homogenates. Furthermore, cytokeratins, which are normally degraded during apoptosis, remained intact. On the other hand, Western blotting of Beclin 1 and light chain 3-II, both important autophagy markers, revealed autophagic processes in the flank organ in both sexes, especially in females. Cathepsin D activity, higher in males than in females, and procathepsin D expression were also consistent with autophagy and not apoptosis. Taken together, these data indicate that macroautophagy, and not apoptosis, is the main mechanism by which the flank organ responds to androgen. This is the first direct evidence establishing the relationship between autophagy and morphological changes in androgen-dependent organs.

Senescence-accelerated mice (SAMP8) and senescence-accelerated resistant mice (SAMR1) were studied at 5 and 10 months of age, respectively. In the animals, neurodegenerative processes and how they were influenced by melatonin were examined. Melatonin (10 mg/kg) or vehicle (ethanol at 0.066%) treatments were administrated from the age of 1 to 9 months in the drinking water. Differences in the neurodegenerative markers examined were found between the two strains with a more damaged protein, phosphorylated Tau at Ser392, increased neurofibrillary tangles (NT) and higher alpha-synuclein expression in SAMP8 versus SAMR1 mice overall, when the mice were 10 months of age. Changes in density of receptors and oxidative stress-related signaling with age were found in the brains of SAM strains at 10 months as shown by a marked decrease in the level of MT-1 melatonin receptor and retinoic acid receptor-related orphan receptor (ROR)-alpha1. This diminution was earlier and more pronounced in SAMP8 mice. Likewise, the levels of nuclear factor-kappa B (NF-kB) transcriptional factor were higher in SAMP8 mice compared with SAMR1 mice regardless of age confirming the direct role of oxidative stress in the aging process. Treatment with melatonin in SAMP8 and SAMR1 mice reduced the neurodegenerative changes with an increase of ROR-alpha1 levels without an apparent influence in the levels of MT-1 receptor. However, different melatonin effects on NF-kB signaling were observed suggesting that NF-kB could trigger inflammatory processes in a different way, being SAM strain-dependent and associated with age-related oxidative stress levels. The effectiveness of melatonin in improving age-related neural impairments is corroborated.

The prevalence of liver diseases emphasizes the need of animal models to research on the mechanism of disease pathogenesis. Furthermore, most of the liver pathologies have the oxidative stress as an important component. The senescence-accelerated mouse strain SAMP8 was proposed as a valuable animal model for the study of liver diseases. To gain a better understanding of the mechanisms underlying degenerative processes in SAMP8 mice livers, we studied the oxidative-induced damage in 5-month-old SAMP8 mice and SAMR1, senescence-accelerated-resistant mice. We found profound differences in the antioxidant response to aging between sexes, with males displaying lowest levels of main antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione reductase (GR) in SAMP8; whereas females had no difference in their activities, except for GR, when compared with their SAMR1 controls. The results obtained show the binomial SOD/CAT as an important factor for counteracting reactive oxygen species-dependent damage. There were not pathological differences at the morphological level between both strains, although the decay in protection against free radicals had an immediate response by increasing lipid and protein oxidative damage in SAMP8 mice liver. At 5 months, both male and female SAMP8 mice confront the oxidative stress challenge to different extents. Indeed, proteins seem to be the most vulnerable biomolecule in SAMP8 male mice.