Hydroxytyrosol as a Promising Ally in the Treatment of Fibromyalgia.

Fibromyalgia (FM) is a chronic and highly disabling syndrome, which is still underdiagnosed, with controversial treatment. Although its aetiology is unknown, a number of studies have pointed to the involvement of altered mitochondrial metabolism, increased oxidative stress and inflammation. The intake of extra virgin olive oil, and particularly of one of its phenolic compounds, hydroxytyrosol (HT), has proven to be protective in terms of redox homeostatic balance and the reduction of inflammation. In this context, using a proteomic approach with nanoscale liquid chromatography coupled to tandem mass spectrometry, the present study analysed: (i) Changes in the proteome of dermal fibroblasts from a patient with FM versus a healthy control, and (ii) the effect of the treatment with a nutritional relevant dose of HT. Our results unveiled that fibroblast from FM show a differential expression in proteins involved in the turnover of extracellular matrix and oxidative metabolism that could explain the inflammatory status of these patients. Moreover, a number of these proteins results normalized by the treatment with HT. In conclusion, our results support that an HT-enriched diet could be highly beneficial in the management of FM.

Hydroxytyrosol, the Major Phenolic Compound of Olive Oil, as an Acute Therapeutic Strategy after Ischemic Stroke.

Stroke is one of the leading causes of adult disability worldwide. After ischemic stroke, damaged tissue surrounding the irreversibly damaged core of the infarct, the penumbra, is still salvageable and is therefore a target for acute therapeutic strategies. The Mediterranean diet (MD) has been shown to lower stroke risk. MD is characterized by increased intake of extra-virgin olive oil, of which hydroxytyrosol (HT) is the foremost phenolic component. This study investigates the effect of an HT-enriched diet directly after stroke on regaining motor and cognitive functioning, MRI parameters, neuroinflammation, and neurogenesis. Stroke mice on an HT diet showed increased strength in the forepaws, as well as improved short-term recognition memory probably due to improvement in functional connectivity (FC). Moreover, mice on an HT diet showed increased cerebral blood flow (CBF) and also heightened expression of brain derived neurotrophic factor (Bdnf), indicating a novel neurogenic potential of HT. This result was additionally accompanied by an enhanced transcription of the postsynaptic marker postsynaptic density protein 95 (Psd-95) and by a decreased ionized calcium-binding adapter molecule 1 (IBA-1) level indicative of lower neuroinflammation. These results suggest that an HT-enriched diet could serve as a beneficial therapeutic approach to attenuate ischemic stroke-associated damage.

Angiogenesis is VEGF-independent in the aged striatum of male rats exposed to acute hypoxia.

Brain hypoxia is involved in many diseases. The activation of angiogenesis is one of the major adaptive mechanisms to counteract the adverse effects of hypoxia. In a previous work, we have shown that the adult rat striatum promotes angiogenesis in response to hypoxia via upregulation of the most important proangiogenic factor, the vascular endothelial growth factor (VEGF). However, the effects of hypoxia on angiogenesis in the aged striatum remain unknown and constitute our aim. Here we show the upregulation of hypoxia-inducible factor-1α in the striatum of aged (24-25 months old) Wistar rats exposed to acute hypoxia and analysed during a reoxygenation period ranging from 0 h to 5 days. While the mRNA expression of the proangiogenic factors VEGF, transforming growth factor-ß1 (TGF-ß1), and adrenomedullin dropped at 0 h post-hypoxia compared to normoxic control, no changes were detected at the protein level, showing an impaired response of these proangiogenic factors to hypoxia in the aged striatum. However, the striatal blood vessel network increased at 24 h of reoxygenation, suggesting that mechanisms independent from these proangiogenic factors may be involved in hypoxia-induced angiogenesis in the striatum of aged rats. A thorough understanding of the factors involved in the response to hypoxia is essential to guide the design of therapies for hypoxia-related diseases in the aged brain.

Nitric oxide averts hypoxia-induced damage during reoxygenation in rat heart.

Nitric oxide (NO), synthesized by the hemoproteins NO synthases (NOS), is known to play important roles in physiological and pathological conditions in the heart, including hypoxia/reoxygenation (H/R). This work investigates the role that endogenous NO plays in the cardiac H/R-induced injury. A follow-up study was conducted in Wistar rats subjected to 30 min of hypoxia, with or without prior treatment using the nonselective NOS inhibitor L-NAME (1.5 mM). The rats were studied at 0 h, 12 h, and 5 days of reoxygenation, analysing parameters of cell, and tissue damage (lipid peroxidation, apoptosis, and protein nitration), as well as in situ NOS activity and NO production (NOx). The results showed that after L-NAME administration, in situ NOS activity was almost completely eliminated in all the experimental groups, and consequently, NOx levels fell. Contrarily, the lipid peroxidation level and the percentage of apoptotic cells rose throughout the reoxygenation period. These results reveal that NOS inhibition exacerbates the peroxidative and apoptotic damage observed before the treatment with L-NAME in the hypoxic heart, pointing to a cardioprotective role of NOS-derived NO against H/R-induced injury. These findings could open the possibility of future studies to design new therapies for H/R-dysfunctions based on NO-pharmacology.

Endogenous nitric oxide can act as beneficial or deleterious in the hypoxic lung depending on the reoxygenation time.

Nitric oxide (NO) has been implicated in many pathophysiological situations in the lung, including hypoxia/reoxygenation. This work seeks to clarify the current controversy concerning the double protective/toxic role of endogenous NO under hypoxia/reoxygenation situations in the lung by using a nitric oxide synthase (NOS) inhibitor, in a novel approach to address the problems raised from assaults under such circumstances. A follow-up study was conducted in Wistar rats submitted to hypoxia/reoxygenation (hypoxia for 30 min; reoxygenation of 0 h, 48 h, and 5 days), with or without prior treatment using the nonselective NOS inhibitor L-NAME (1.5 mM, in drinking water). Lipid peroxidation, apoptosis level, protein nitration, in situ NOS activity and NO production (NOx) were analyzed. This is the first work to focus on the time-course effects of L-NAME in the adult rat lung submitted to hypoxia/reoxygenation. The results showed that after L-NAME administration, in situ NOS activity was almost completely eliminated and consequently, NOx levels fell. Lipid peroxidation and the percentage of apoptotic cells rose at the earliest reoxygenation time (0 h), but decreased in the later period (48 h and 5 days). Also nitrated protein expression decreased at 48 h and 5 days posthypoxia. These results suggest that NOS-derived NO exerts two different effects on lung hypoxia/reoxygenation injury depending on the reoxygenation time: NO has a beneficial role just after the hypoxic stimulus and a deleterious effect in the later reoxygenation times. Moreover, we propose that this dual role of NO depends directly on the producer NOS isoform.

Polyphenol oxidase and its relationship with oleuropein concentration in fruits and leaves of olive (Olea europaea) cv. 'Picual' trees during fruit ripening.

Oleuropein, the main phenolic compound of olive fruit, has important antioxidant properties that are responsible for some of the nutritional properties of fruits and the defence mechanism of leaves. Polyphenol oxidase (PPO) activity changes during fruit ripening in many plants. We studied the kinetics and molecular properties of PPO in fruits and leaves of olive (Olea europaea L.) cv. 'Picual' trees and the relationship between PPO and oleuropein concentration during fruit ripening. Polyphenol oxidase showed hyperbolic kinetics in fruits and leaves. Significant increases in PPO specific activity, V(max), K(m )and catalytic efficiency occurred during fruit ripening. Based on SDS-PAGE under partially denaturing conditions and in-gel staining with DL-3,4-dihydroxyphenylalanine, PPO activity was found in one major protein of 55 and 50 kDA in fruits and leaves, respectively. During the last stages of fruit maturation, a second 36 kDa protein was observed in fruits but not in leaves, indicating that this protein could serve as a marker of the final phase of fruit maturation. Under fully denaturing conditions, only one 27.7 kDa immunoreactive band was detected in fruits. Both the amount of PPO activity and the amount of PPO protein increased significantly during fruit maturation. Immunohistochemical studies indicated that PPO is located in the epidermis, parenchyma and companion vascular cells of leaves as well as in the epidermis of fruit. During fruit maturation, oleuropein concentration measured by HPLC significantly decreased in fruits and increased in leaves.

Downregulation in the expression of the serine dehydratase in the rat liver during chronic metabolic acidosis.

Blood pH controls the activity of important regulatory enzymes in the metabolism. Serine dehydratase (SerDH) transforms l-serine into pyruvate and ammonium and is involved in the regulation of gluconeogenesis from serine in the rat liver. In this work, we investigate the effect of chronic metabolic acidosis on the kinetics, specific protein level, tissue location, and mRNA levels of rat liver SerDH. Experimental acidosis was induced in rats by ingestion of 0.28 M ammonium chloride solution for 10 days. Acidosis significantly (P<0.05) decreased SerDH activity at all substrate concentrations assayed. Moreover, the Vmax value was 38.50+/-3.51 mU/mg (n=7) of mitochondrial protein in the acidotic rats and 92.49+/-6.79 mU/mg (n=7) in the control rats. Western blot analysis revealed a significant reduction (14%) in the level of SerDH protein content in the rat liver during acidosis. Immunohistochemical analysis showed that SerDH location did not change in response to chronic metabolic acidosis and confirmed previous results on SerDH protein levels. Moreover, the SerDH mRNA level, estimated by RT-PCR, was also significantly 33.8% lower than in control. These results suggest that during experimental acidosis a specific repression of rat-liver SerDH gene transcription could result, lowering the amount and activity of this enzyme. The changes found in SerDH expression are part of an overall metabolic response of liver to maintain acid-base homeostasis during acidosis.

Serine dehydratase expression decreases in rat livers injured by chronic thioacetamide ingestion.

Serine dehydratase (SerDH) is a gluconeogenic enzyme involved in the catabolism of serine, which is regulated by the composition of their diet and their hormonal status in rats. This study examines how chronic injury caused to the liver of rats by the ingestion of thioacetamide (TAA) affects SerDH protein, mRNA levels, enzyme kinetics and its tissue location. After 97 days' oral intake of TAA, the activity of SerDH at all substrate concentrations assayed was about 60% lower than in controls. No significant differences in Km values were found between the treated group and controls. Immunoblotting and immunohistochemistry revealed a significant reduction in the level of SerDH protein in the livers of the treated rats. SerDH was detected specifically in the periportal zone of the hepatic acinus and this location did not change in response to TAA treatment. The level of SerDH mRNA, quantified by reverse transcription and polymerase chain reaction, was significantly lower in treated rats than in the controls. The present findings suggest that the SerDH expression is rendered to be down regulatory during chronic liver injury induced by TAA. These results enhance our understanding about the biochemical mechanisms implied in the control and integration of serine catabolism during liver injury in rat.

Glutathione S-transferase isoenzymatic response to aging in rat cerebral cortex and cerebellum.

Aging is associated with increased oxidant generation. One mechanism involved in the defense of oxidative products is the family of glutathione transferases (GST). We have analyzed the activity, distribution and expression of GSTP1 and GSTA4 isoenzymes in the cerebral cortex and cerebellum of young, adult and aged rats. The total GST activity, measured with the universal substrate 1-chloro-2,4-dinitrobenzene (CDNB), increased only with the maturation process; however GSTA4 activity, using the specific substrate 4-hydroxynonenal (HNE), did show an age-dependent increase in both brain regions. Cellular location of GSTA4 in astrocytes was not changed except for young cerebral cortex and adult/aged cerebellum that also showed immunoreactivity in layer III pyramidal neurons and Bergman radial glia, respectively. Distribution of GSTP1 was similar among groups and only an increased number of positive oligodendrocytes was found in the Purkinje and granular layer of adult/aged cerebellum. The GSTA4 and GSTP1 expression increased from young to adult/aged brain and GSTA4 even augmented in the aged cerebral cortex. These results suggest a GST isoenzymatic response with aging, but above all with the maturation process.