Contribution of glutathione to the control of cellular redox homeostasis under toxic metal and metalloid stress.

The accumulation of toxic metals and metalloids, such as cadmium (Cd), mercury (Hg), or arsenic (As), as a consequence of various anthropogenic activities, poses a serious threat to the environment and human health. The ability of plants to take up mineral nutrients from the soil can be exploited to develop phytoremediation technologies able to alleviate the negative impact of toxic elements in terrestrial ecosystems. However, we must select plant species or populations capable of tolerating exposure to hazardous elements. The tolerance of plant cells to toxic elements is highly dependent on glutathione (GSH) metabolism. GSH is a biothiol tripeptide that plays a fundamental dual role: first, as an antioxidant to mitigate the redox imbalance caused by toxic metal(loid) accumulation, and second as a precursor of phytochelatins (PCs), ligand peptides that limit the free ion cellular concentration of those pollutants. The sulphur assimilation pathway, synthesis of GSH, and production of PCs are tightly regulated in order to alleviate the phytotoxicity of different hazardous elements, which might induce specific stress signatures. This review provides an update on mechanisms of tolerance that depend on biothiols in plant cells exposed to toxic elements, with a particular emphasis on the Hg-triggered responses, and considering the contribution of hormones to their regulation.

Early transcriptional responses to mercury: a role for ethylene in mercury-induced stress.

Understanding the cellular mechanisms of plant tolerance to mercury (Hg) is important for developing phytoremediation strategies of Hg-contaminated soils. The early responses of alfalfa (Medicago sativa) seedlings to Hg were studied using transcriptomics analysis. A Medicago truncatula microarray was hybridized with high-quality root RNA from M. sativa treated with 3 µM Hg for 3, 6 and 24 h. The transcriptional pattern data were complementary to the measurements of root growth inhibition, lipid peroxidation, hydrogen peroxide (H2 O2 ) accumulation and NADPH-oxidase activity as stress indexes. Of 559 differentially expressed genes (DEGs), 91% were up-regulated. The majority of DEGs were shared between the 3 and 6 h (60%) time points, including the 'stress', 'secondary metabolism' and 'hormone metabolism' functional categories. Genes from ethylene metabolism and signalling were highly represented, suggesting that this phytohormone may be relevant for metal perception and homeostasis. Ethylene-insensitive alfalfa seedlings preincubated with the ethylene signalling inhibitor 1-methylcyclopronene and Arabidopsis thaliana ein2-5 mutants confirmed that ethylene participates in the early perception of Hg stress. It modulates root growth inhibition, NADPH-oxidase activity and Hg-induced apoplastic H2 O2 accumulation. Therefore, ethylene signalling attenuation could be useful in future phytotechnological applications to ameliorate stress symptoms in Hg-polluted plants.

Polyamines in human breast milk for preterm and term infants.

Maternal milk is the first source of exogenous polyamines for the newborn. Polyamines modulate gut maturation in neonates, but no studies are available on polyamine concentration in human milk of preterm babies, even though they could be important for their immature gut. The present study aimed to determine polyamine concentration in human breast milk of mothers with preterm or term infants during the first month of lactation. Human milk samples were obtained during the first month of lactation from twenty-seven mothers with preterm babies and twelve mothers with babies born at term. The polyamine concentration in human milk was quantified by HPLC. During the first month of lactation, the total polyamine concentration was significantly higher in preterm milk than in term milk samples (7590 (SD 4990) v. 4660 (SD 4830) nmol/l, respectively (P » 0·034)), as well as individual polyamine concentrations. Polyamine concentration in mature milk for preterm babies was significantly higher than that in mature milk for babies at term, and a similar trend was observed in colostrum and transition human milk. The spermidine/spermine ratio was higher in transition milk in preterm v. term samples, while in mature milk, the ratio was significantly lower in preterm than in term babies. In conclusion, the polyamine concentration was significantly higher in human milk for preterm than for term infants. This and the different spermidine/spermine ratios could influence the gut development of premature babies.

Bacillus H47 triggers Olea europaea metabolism activating DOXP and shikimate pathways simultaneously and modifying leaf extracts' antihypertensive activity.

Improvement of plant adaptation by beneficial bacteria (PGPB) may be achieved by triggering multiple pathways to overcome the environmental stress on plant's growth cycle, activating plant's metabolism. The present work reports the differential ability of three Bacillus strains to trigger olive tree metabolism, among which, only H47 was outstanding increasing iridoid and flavonol concentration. One-year old olive seedlings grown open air, under harsh conditions of water shortage in saline soils, were root-inoculated with three Bacillus PGPB strains throughout a 12-month period after which, photosynthesis was determined; photosynthetic pigments and bioactive secondary metabolites (iridoids and flavonols) were analyzed, and a study of gene expression of both pathways involved was undertaken to unravel molecular targets involved in the activation. All three strains increased plant fitness based on photosynthetic values, increasing energy dissipation capacity to lower oxidative stress; only H47 increased CO2 fixation and transpiration. Bacillus H47 was found to trigger synthases in the DOXP pathway (up to 5-fold in DOXP-synthase, 3.5-fold in Iridoid synthase, and 2-fold in secologanin synthase) associated to a concomitant increase in iridoids (up to 5-fold in oleuropein and 2-fold in its precursor secologanin). However, despite the 2-fold increases detected in the two predominant flavonols, gene expression was not enhanced, suggesting involvement of a pulse activation model proposed for innate immunity. Furthermore, the activity of leaf extracts to inhibit Angiotensin Converting Enzyme was evaluated, to explore further uses of plant debris with higher added value. Despite the increases in iridoids, leaf extracts from H47 did not increase ACE inhibition, and still, increased antihypertensive potential in oil obtained with this strain is to be explored, as leaves are the source for these metabolites which further translocate to fruits. In summary, Bacillus H47 is an effective strain to increase plant adaptation to dry and saline environments, activates photosynthesis and secondary metabolism in olive tree.

Effects of a high-fat-diet supplemented with probiotics and ω3-fatty acids on appetite regulatory neuropeptides and neurotransmitters in a pig model.

The pig is a valuable animal model to study obesity in humans due to the physiological similarity between humans and pigs in terms of digestive and associated metabolic processes. The dietary use of vegetal protein, probiotics and omega-3 fatty acids is recommended to control weight gain and to fight obesity-associated metabolic disorders. Likewise, there are recent reports on their beneficial effects on brain functions. The hypothalamus is the central part of the brain that regulates food intake by means of the production of food intake-regulatory hypothalamic neuropeptides, as neuropeptide Y (NPY), orexin A and pro-opiomelanocortin (POMC), and neurotransmitters, such as dopamine and serotonin. Other mesolimbic areas, such as the hippocampus, are also involved in the control of food intake. In this study, the effect of a high fat diet (HFD) alone or supplemented with these additives on brain neuropeptides and neurotransmitters was assessed in forty-three young pigs fed for 10 weeks with a control diet (T1), a high fat diet (HFD, T2), and HFD with vegetal protein supplemented with Bifidobacterium breve CECT8242 alone (T3) or in combination with omega-3 fatty acids (T4). A HFD provoked changes in regulatory neuropeptides and 3,4-dihydroxyphenylacetic acid (DOPAC) in the hypothalamus and alterations mostly in the dopaminergic system in the ventral hippocampus. Supplementation of the HFD with B. breve CECT8242, especially in combination with omega-3 fatty acids, was able to partially reverse the effects of HFD. Correlations between productive and neurochemical parameters supported these findings. These results confirm that pigs are an appropriate animal model alternative to rodents for the study of the effects of HFD on weight gain and obesity. Furthermore, they indicate the potential benefits of probiotics and omega-3 fatty acids on brain function.

[Home and Ambulatory Artificial Nutrition (NADYA) Group Report. Home parenteral nutrition in Spain, 2018]. / Nutrición parenteral domiciliaria en España 2018. Informe del Grupo de Nutrición Artificial Domiciliaria y Ambulatoria NADYA.

INTRODUCTION: Aim: to communicate home parenteral nutrition (HPN) data obtained from the HPN registry of the NADYA-SENPE group (www.nadya-senpe.com) for the year 2018 Material and methods: descriptive analysis of the data collected from adult and pediatric patients with HPN in the NADYA-SENPE group registry from January 1st, 2018 to December 31st, 2018. Results: there were 278 patients from 45 Spanish hospitals (54.7% women), 23 children and 255 adults, which represent a prevalence rate of 5.95 patients/million inhabitants/year 2018. The most frequent diagnosis in adults was "palliative cancer" (22.0%), followed by "others". In children it was Hirschsprung's disease together with necrotizing enterocolitis, with four cases (17.4%). The first indication was short bowel syndrome in both children (60.9%) and adults (35.7%). The most frequently used type of catheter was tunneled in both children (81.0%) and adults (41.1%). Ending 75 episodes, the most frequent cause was death (52.0%) and change to oral feeding (33.3%). Conclusions: the number of centers and collaborating professionals in the registry of patients receiving HPN remains stable, as well as the main indications and reasons for termination of HPN.

INTRODUCCIÓN: Objetivo: comunicar los datos de nutrición parenteral domiciliaria (NPD) obtenidos del registro del grupo NADYA-SENPE (www.nadya-senpe.com) del año 2018. Material y métodos: análisis descriptivo de los datos recogidos de pacientes adultos y pediátricos con NPD en el registro NADYA-SENPE del 1 de enero al 31 de diciembre de 2018. Resultados: se registraron 278 pacientes (54,7% mujeres), 23 niños y 255 adultos, procedentes de 45 hospitales españoles, lo que representa una tasa de prevalencia de 5,95 pacientes/millón de habitantes/año 2018. El diagnóstico más frecuente en adultos fue "oncológico paliativo" (22,0%), seguido de "otros". En niños fue la enfermedad de Hirschsprung junto con la enterocolitis necrotizante, con cuatro casos (17,4%). El primer motivo de indicación fue síndrome de intestino corto tanto en niños (60,9%) como en adultos (35,7%). El tipo de catéter más utilizado fue el tunelizado tanto en niños (81,0%) como en adultos (41,1%). Finalizaron 75 episodios, la causa más frecuente fue el fallecimiento (52,0%) y el paso a vía oral (33,3%). Conclusiones: el número de centros y profesionales colaboradores en el registro de pacientes que reciben NPD se mantiene estable, así como las principales indicaciones y los motivos de finalización de la NPD.

[Home and Ambulatory Artificial Nutrition (NADYA) Report. Home Parenteral Nutrition in Spain, 2017]. / Nutrición parenteral domiciliaria en España 2017. Informe del Grupo de Nutrición Artificial Domiciliaria y Ambulatoria NADYA.

AIM: to communicate HPN data obtained from the HPN registry of the NADYA-SENPE group (www.nadya-senpe.com) for the year 2017. MATERIAL AND METHODS: descriptive analysis of the data collected from adult and pediatric patients with HPN in the NADYA-SENPE group registry from January 1st, 2017 to December 31st, 2017. RESULTS: there were 308 patients from 45 Spanish hospitals (54.5% women), 38 children and 270 adults, with 3,012 episodes, which represent a prevalence rate of 6.61 patients/million inhabitants/year 2017. The most frequent diagnosis in adults was "palliative cancer" (25.6%), followed by "others". In children, it was Hirschsprung's disease with six cases (15.8%). The first indication was short bowel syndrome in both children (55.3%) and adults (33.7%). The most frequently used type of catheter was tunneled in both children (73.4%) and adults (38.2%). Ending 81 episodes, the most frequent cause was death (62.9%) and transition to oral feeding (34.7%). CONCLUSIONS: the progressive increase of collaborating centers and professionals in the registry of patients receiving NPD is maintained. The main indications of HPN and the motive for ending have remained stable.

OBJETIVO: comunicar los datos de nutrición parenteral domiciliaria (NPD) obtenidos del registro del grupo NADYA-SENPE (www.nadya-senpe. com) del año 2017. MATERIAL Y MÉTODOS: análisis descriptivo de los datos recogidos de pacientes adultos y pediátricos con NPD en el registro NADYA-SENPE desde el 1 de enero al 31 de diciembre de 2017. RESULTADOS: se registraron 308 pacientes (54,5% mujeres), 38 niños y 270 adultos, procedentes de 45 hospitales españoles, en total 312 episodios, lo que representa una tasa de prevalencia de 6,61 pacientes/millón de habitantes/año 2017. El diagnóstico más frecuente en adultos fue "oncológico paliativo" (25,6%), seguido de "otros". En niños fue la enfermedad de Hirschsprung, con seis casos (15,8%). El primer motivo de indicación fue síndrome de intestino corto tanto en niños (55,3%) como en adultos (33,7%). El tipo de catéter más utilizado fue el tunelizado tanto en niños (74,3%) como en adultos (38,2%). Finalizaron 81 episodios; la causa más frecuente fue el fallecimiento (62,9%) y que pasaron a vía oral (34,7%). CONCLUSIONES: se mantiene el incremento progresivo de centros y profesionales colaboradores en el registro de pacientes que reciben NPD. Las principales indicaciones de NPD y de motivo de finalización se mantienen estables.

Fast atom transport and launching in a nonrigid trap.

We study the shuttling of an atom in a trap with controllable position and frequency. Using invariant-based inverse engineering, protocols in which the trap is simultaneously displaced and expanded are proposed to speed up transport between stationary trap locations as well as launching processes with narrow final-velocity distributions. Depending on the physical constraints imposed, either simultaneous or sequential approaches may be faster. We consider first a perfectly harmonic trap, and then extend the treatment to generic traps. Finally, we apply this general framework to a double-well potential to separate different motional states with different launching velocities.

[Home and Ambulatory Artificial Nutrition (NADYA) Group Report - Home parenteral nutrition in Spain, 2016]. / Nutrición parenteral domiciliaria en España 2016; informe del Grupo de Nutrición Artificial Domiciliaria y Ambulatoria NADYA.

OBJECTIVE: To communicate HPN data obtained from the HPN registry of the NADYA-SENPE group (www.nadya-senpe.com) for the year 2016. MATERIAL AND METHODS: Descriptive analysis of the data collected from adult and pediatric patients with HPN in the NADYA-SENPE group registry from January 1st, 2016 to December 31st, 2016. RESULTS: There were 286 patients from 42 Spanish hospitals (54.2% women), 34 children and 252 adults, with 294 episodes, which represent a prevalence rate of 6.16 patients / million inhabitants / year 2016. The most frequent diagnosis in adults was "palliative cancer" (25.8%), followed by "others". In children it was "motility alterations" with 6 cases (17.6%), Hirschsprung's disease and necrotising enterocolitis, both with 5 children (14.7%). The first indication was short bowel syndrome in both children (64.7%) and adults (37.3%), followed by intestinal obstruction in 28.6% adults and 14.7% in children. The most frequently used type of catheter was tunnelled in both children (70.6%) and adults (37.9%). The most frequent complication in adults was infection related to the catheter, which presented a rate of 0.48 infections / 1,000 days of NPD. During this period, 71 episodes ended in adults and the main cause was death (57.7%) followed by resuming the oral route (31%). CONCLUSIONS: There is a progressive increase of centers and professional collaborators in the registry who report patients receiving parenteral nutrition at home. The main indications of HPN and the motive for ending have remained stable.

Influence of dietary source of docosahexaenoic and arachidonic acids on their incorporation into membrane phospholipids of red blood cells in term infants.

Here we studied whether the chemical structure of dietary arachidonic and docosahexaenoic acids in full-term infant diets affects their incorporation into erythrocyte membrane phospholipids. From birth to 3 months, infants were fed breast milk (n = 9) or formula milk containing arachidonic acid and docosahexaenoic acid provided by egg phospholipids (n = 10) or by low-eicosapentaenoic acid fish oil and fungal triglycerides (n = 10). We compared the fatty acid composition of erythrocyte phosphatidylethanolamine, phosphatidylcholine and sphingomyelin before and after administration of the experimental diet. At 3 months, infants on formula milk showed lower concentrations of docosahexaenoic acid (in phosphatidylcholine and in phosphatidylethanolamine) and arachidonic acid (in phosphatidylcholine) than those receiving breast milk. We conclude the incorporation of the two fatty acids into erythrocyte phospholipids depends mainly on the lipid composition of the diet received rather than the chemical form in which they are delivered.

An in vitro model of ischemic-like stress in retinal pigmented epithelium cells: protective effects of antioxidants.

We have developed a model of in vitro cell oxidative stress in bovine retinal pigment epithelium cells exposed to a ischemia-like condition obtained by interference with glucose utilization through both oxidative phosphorylation and glycolysis. This resulted in a statistically significant decrease of the intracellular ATP levels, which reflects a bioenergetic decline similar to that associated with mitochondrial damage or loss in normal post-mitotic cells aging in vivo. This new model of cellular oxygen stress seems adequate for investigation of the protective action of antioxidants, in agreement with our finding of a statistically significant increase in the ATP levels over the values of the non-treated samples in retinal pigment epithelium cells exposed to the above oxygen stress in medium supplemented with 300 microM vitamin C or 10 mM N-acetylcysteine.

Bilateral deep brain stimulation of the subthalamic nucleus in Parkinson's disease.

Deep brain stimulation (DBS) of the subthalamic nucleus (STN) is rapidly becoming the preferred surgical choice for the treatment of advanced Parkinson's disease (PD). We report initial results in 15 patients after 12 months and in nine patients evaluated between 30 and 36 months postoperatively. Our experience confirms the robust antiparkinsonian effect of DBS of the STN in advanced PD. The severity of "off" episodes, as assessed by the Unified Parkinson Disease Rating Scale (UPDRS), was drastically reduced by 74% at 12 months, and dyskinesia scores (Dyskinesia Rating Scale) decreased. The levodopa daily dose was reduced by 55% at 12 months. A double-blind assessment to determine the effect of stimulation performed in nine patients at 3 months in the "off" medication condition was very significant (p<0.05). Nine patients have been followed for 3 years with maintained efficacy in the UPDRS "off" score and the dyskinesia score. The experience of other groups using a similar technique is reviewed. The overall assessment indicates a high antiparkinsonian effect of DBS of the STN even in advanced patients. The existence of a learning curve for this procedure should be taken into account when initial results are evaluated.

Comparison of four lymphocyte isolation methods applied to rodent T cell subpopulations and B cells.

The aim of this study was to establish the validity of four lymphocyte isolation methods. The effects of three different erythrocyte lysing methods commonly used in the analysis of human cells, namely, lysis by ammonium chloride (AC), Becton Dickinson lysis (BDL) and the Coulter Q-Prep (CQP) preparation system were established by flow cytometry on rat lymphocyte subsets. The results were compared with those obtained with a Ficoll-Isopaque (FI) density gradient procedure adapted for use with rat cells. Lymphocyte isolation by AC or FI gradient was performed before labelling the lymphocyte subpopulations, whereas the BDL and CQP methods were performed after staining the cells in whole blood. The FI gradient yielded the lowest CD5+, CD4+ and CD25+ cell percentages. On the other hand AC lysis produced higher percentages of T cells and lower percentages of B cells than the other methods studied. The percentages obtained after BDL or CQP methods for T lymphocyte subsets and B cells were found to be reproducible. The commercial methods (BDL and CQP) are faster but rather expensive, whereas AC lysis and FI gradient separations are cheap and particularly useful when there is a requirement to culture the cells.

Separate [3H]-nitrendipine binding sites in mitochondria and plasma membranes of bovine adrenal medulla.

1. Two binding sites for the 1,4-dihydropyridine (DHP) derivative [3H]-nitrendipine have been found in the bovine adrenal medulla. The high-affinity site (Kd = 0.48 nM and Bmax = 128 fmol mg-1 protein) was specifically located in purified plasma membranes. The low-affinity site (Kd = 252 nM and Bmax = 169 pmol mg-1 protein) was located only in mitochondria. Chromaffin granule membranes lacked specific binding sites for [3H]-nitrendipine. 2. Kinetic analysis of the rates of association and dissociation of [3H]-nitrendipine, saturation isotherms and displacement experiments with unlabelled nitrendipine and PN200-110 revealed single, homogeneous populations of high- and low-affinity sites in plasma and mitochondrial membranes, respectively. 3. The high affinity site was sensitive to Ca2+ deprivation and heating; it was practically unaffected by changes in ionic strength of the medium and its optimal pH was slightly alkaline. This site exhibited a strong DHP stereoselectivity; diltiazem increased and verapamil decreased the affinity of [3H]-nitrendipine. 4. In contrast, binding of [3H]-nitrendipine to the low affinity site was more heat resistant and less affected by Ca2+ removal. Its optimal pH was slightly acid and the increase in ionic strength enhanced the number of available sites. The site had no DHP stereoselectivity. Verapamil decreased the dissociation constant of [3H]-nitrendipine acting in a non-competitive manner; diltiazem did not affect equilibrium binding parameters of [3H]-nitrendipine. 5. These results suggest that both biding sites reflect different receptor entities. The high-affinity binding site corresponds to the dihydropyridine receptor associated with the L-type calcium channel. The function of the mitochondrial, low-affinity binding site is, at present, unknown.

The low-affinity dihydropyridine receptor and Na+/Ca2+ exchanger are associated in adrenal medullary mitochondria.

The effect of Ca2+ channel-acting drugs on bovine adrenal mitochondria Ca2+ movements was investigated. Mitochondrial Ca2+ uptake is performed by an energy-driven Ca2+ uniporter with a Km of 20.9 +/- 3.2 microM and Vmax of 148.1 +/- 7.2 nmol 45Ca2+ min-1 mg-1. Ca2+ release is performed through an Na+/Ca2+ antiporter with a Km for Na+ of 4.2 +/- 0.5 mM, a Vmax of 7.5 +/- 0.4 nmol 45Ca2+ min-1 mg-1, and a Hill coefficient of 1.4 +/- 0.2 Ca2+ efflux through the mitochondrial Na+/Ca2+ exchanger was inhibited by several dihydropyridines (nitrendipine, felodipine, nimodipine, (+)isradipine) and by the benzothiazepine diltiazem with similar potencies. In contrast, neither CGP 28392, Bay-K-8644, amlodipine, nor verapamil had any effect on Ca2+ efflux. Nitrendipine at 20 microM modified neither the Km nor the Hill coefficient for Na+, whereas the Vmax was reduced to 2.9 nmol 45Ca2+ min-1 mg-1, thus demonstrating noncompetitive modulation of the Na+/Ca2+ exchanger. None of the Ca2+ channel-acting drugs assayed at 100 microM affected Ca2+ influx through the uniporter. Ca2+ channel blockers inhibited the Na+/Ca2+ antiporter and displaced the specific binding of [3H]nitrendipine to intact mitochondria with Ki values similar to the IC50s obtained for the inhibition of the Ca2+ efflux. Ca2+ channel-acting drugs that did not inhibit the Na+/Ca2+ exchanger (amlodipine, CGP 28392, Bay-K-9644, and verapamil, at concentrations of 100 microM or higher) had no effect on [3H]nitrendipine binding. These results suggest that the adrenomedullary mitochondrial dihydropyridine receptor is associated with the Na+/Ca2+ exchanger.

Determination of sterol content in different food samples by capillary gas chromatography.

An accurate method for the determination of sterols by capillary gas chromatography was developed and applied to the analysis of food. The procedure includes the following steps: dichloromethane-methanol (2:1, v/v) lipid extraction, saponification at 80 degrees C and separation of the unsaponifiable matter with cyclohexane, derivatization to form trimethylsilyl ethers and gas chromatography using 5 alpha-cholestane as the internal standard. The method shows good accuracy, precision and sensitivity and is suitable for the determination of sterols in food.

Comparison of two methods for the determination of fatty acid profiles in plasma and erythrocytes.

We have validated and compared two methods for the determination of fatty acid profiles in biological samples by capillary gas chromatography. Method I consisted of a previous lipid extraction and esterification of fatty acids using boron trifluoride-methanol. Method II was a direct method that combined extraction and esterification of freeze-dried samples in a single step, using acetyl chloride as the reagent. The two methods were applied to the analysis of plasma and erythrocyte samples. Both methods gave similar results in plasma, whereas in erythrocytes, the direct method gave significantly higher contents of total fatty acids. Precision and recovery rates were determined and the results were satisfactory. Detection and quantification limits showed that both methods had excellent sensitivity. It was concluded that the direct method has substantial advantages over the conventional method, such as higher values in erythrocytes, rapidity and less possibility of contamination or fatty acid losses. Therefore, it is preferable for the analysis of biological samples such as plasma and erythrocytes.

Comparison of two methods for the determination of fatty acid profiles in plasma and erythrocytes.

We have validated and compared two methods for the determination of fatty acid profiles in biological samples by capillary gas chromatography. Method I consisted of a previous lipid extraction and esterification of fatty acids using boron trifluoride-methanol. Method II was a direct method that combined extraction and esterification of freeze-dried samples in a single step, using acetyl chloride as the reagent. The two methods were applied to the analysis of plasma and erythrocyte samples. Both methods gave similar results in plasma, whereas in erythrocytes, the direct method gave significantly higher contents of total fatty acids. Precision and recovery rates were determined and the results were satisfactory. Detection and quantification limits showed that both methods had excellent sensitivity. It was concluded that the direct method has substantial advantages over the conventional method, such as higher values in erythrocytes, rapidity and less possibility of contamination or fatty acid losses. Therefore, it is preferable for the analysis of biological samples such as plasma and erythrocytes.

Administration of low doses of fish oil derived N-3 fatty acids to elderly subjects.

OBJECTIVES: To assess the incorporation of n-3 polyunsaturated fatty acids (PUFA) in plasma and erythrocyte lipids of elderly subjects after ingestion of very low doses of fish oil. The effects on alpha-tocopherol and retinol concentrations were also studied. SETTING: Municipal nursing home in Barcelona, Spain. SUBJECTS: Forty-five elderly subjects aged 60-92 y. DESIGN AND INTERVENTION: Subjects received a non-commercialized milk formula containing 1% fish oil for 15 months, which provided 0.40 g/d of n-3 PUFA. Fatty acid profiles and antioxidant concentrations were measured before and after the intervention period. RESULTS: Fish oil ingestion was associated with significant increases in total n-3 PUFA in plasma and erythrocytes by 32% and 18%, respectively. Eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid concentrations were higher after the ingestion period both in plasma and erythrocytes (P < 0.05), whereas linoleic and arachidonic acids remained unchanged. The n-6/n-3 ratio decreased by 21% in plasma and by 16% in erythrocytes (P < 0.05). Moreover, younger subjects showed a greater incorporation of EPA and DHA than older subjects. Plasma alpha-tocopherol and retinol concentrations did not vary significantly, whereas erythrocyte alpha-tocopherol was significantly higher after the intervention period. CONCLUSION: This study shows that low doses of n-3 PUFA supplemented with adequate amounts of alpha-tocopherol can be incorporated into blood lipids in elderly subjects without lowering their antioxidant concentrations.

Alterations of lymphocyte populations in lymph nodes but not in spleen during the latency period of adjuvant arthritis.

The aim of this study was to analyze potential imbalances in lymphocyte populations from regional lymph nodes (LN) and spleen occurring before the development of the outer inflammation of adjuvant arthritis (AA). Percentages and absolute numbers of CD5+, CD4+, CD8+, Ig+, I-A+, NKR-P1+ and TCRgammadelta+ cells were determined. No differences in percentages of gammadelta T or NK cells were found either in LN or spleen, thus ruling out an important role of these minor subpopulations in these early stages of AA. While no significant lymphocyte imbalances were observed in spleen, an increase in the percentage of B lymphocytes was found in regional LN. Moreover, a high proliferation of CD8+ cells was observed when measuring absolute numbers of LN lymphocytes, thus producing an imbalance in the CD4/CD8 ratio at very early stages of the inflammatory process. These findings suggest a role for CD8+ and B lymphocytes in the latency period of AA at the LN level. Our results indicate a primary role for lymph nodes in initiating the inflammation of AA, whereas cells from the spleen probably play a secondary role.