Dietary α‐Linolenic Acid, Marine ω‐3 Fatty Acids, and Mortality in a Population With High Fish Consumption: Findings From the PREvención con DIeta MEDiterránea (PREDIMED) Study

Background: Epidemiological evidence suggests a cardioprotective role of α‐linolenic acid (ALA), a plant‐derived ω‐3 fatty acid. It is unclear whether ALA is beneficial in a background of high marine ω‐3 fatty acids (long‐chain n‐3 polyunsaturated fatty acids) intake. In persons at high cardiovascular risk from Spain, a country in which fish consumption is customarily high, we investigated whether meeting the International Society for the Study of Fatty Acids and Lipids recommendation for dietary ALA (0.7% of total energy) at baseline was related to all‐cause and cardiovascular disease mortality. We also examined the effect of meeting the society's recommendation for long‐chain n‐3 polyunsaturated fatty acids (≥500 mg/day). Methods and Results: We longitudinally evaluated 7202 participants in the PREvención con DIeta MEDiterránea (PREDIMED) trial. Multivariable‐adjusted Cox regression models were fitted to estimate hazard ratios. ALA intake correlated to walnut consumption (r=0.94). During a 5.9‐y follow‐up, 431 deaths occurred (104 cardiovascular disease, 55 coronary heart disease, 32 sudden cardiac death, 25 stroke). The hazard ratios for meeting ALA recommendation (n=1615, 22.4%) were 0.72 (95% CI 0.56–0.92) for all‐cause mortality and 0.95 (95% CI 0.58–1.57) for fatal cardiovascular disease. The hazard ratios for meeting the recommendation for long‐chain n‐3 polyunsaturated fatty acids (n=5452, 75.7%) were 0.84 (95% CI 0.67–1.05) for all‐cause mortality, 0.61 (95% CI 0.39–0.96) for fatal cardiovascular disease, 0.54 (95% CI 0.29–0.99) for fatal coronary heart disease, and 0.49 (95% CI 0.22–1.01) for sudden cardiac death. The highest reduction in all‐cause mortality occurred in participants meeting both recommendations (hazard ratio 0.63 [95% CI 0.45–0.87]). Conclusions: In participants without prior cardiovascular disease and high fish consumption, dietary ALA, supplied mainly by walnuts and olive oil, relates inversely to all‐cause mortality, whereas protection from cardiac mortality is limited to fish‐derived long‐chain n‐3 polyunsaturated fatty acids. Clinical Trial Registration URL: http://www.Controlled-trials.com/. Unique identifier: ISRCTN35739639

Utilización de pregabalina en neuralgia postherpética refractaria a terapia convencional

Following an episode of acute herpes zoster, pain lasting more than 3 months and extending beyond the skin lesions is classified as postherpetic neuralgia (PHN) (1). Several drugs have been used for the treatment of PHN, including nonopiate analgesics, nonsteroidal anti-inflammatory drugs (NSAIDs), tricyclic antidepressants, anticonvulsants, phenothiazines, anti-arrhythmics and opiate agents, with varying rates of success (2). Tricyclic antidepressants and gabapentin are currently considered the firstline treatment of PHN (2); however, the inclusión of pregabalin as firstline therapy approved for the treatment of neuropathic pain has opened up new perspectives (3). The purpose of this study is to demónstrate the efficacy and safety of pregabalin (LYRICA®) in the treatment of neuropathic pain in PHN refractory to standard therapy. We conducted a study of the efficacy and safety of pregabalin therapy in 10 patients with PHN lasting more than 3 months and poor response to standard medical therapy. The primary endpoint was the Visual Analogue Scale (VAS) at baseline and every month throughout the 4 months of data collection. Medications were uptitrated monthly if a reduction of more than 30% was not achieved with the previous dosage. After the assessment of the first month of treatment with pregabalin, a reduction of VAS pain by more than 30% from baseline was achieved in half of the patients, with similar results being obtained at the second and third months of treatment. Pregabalin proved to be a highly efficient drug for the reduction of pain in PHN refractory to standard therapy, with a low level of side effects.Tras un episodio de Herpes Zóster agudo, un dolor de duración superior a 3 meses cuya extensión supere las lesiones cutáneas, se clasifica como Neuralgia postherpética (NPH) (1). En su tratamiento se han utilizado diversos fármacos como analgésicos no opiáceos, antiinflamatorios no esteroideos (AINES), antidepresivos tricíclicos, anticonvulsivantes, fenotiacinas, antiarrítmicos y opiáceos con un éxito variable (2). Actualmente los antidepresivos tricíclicos y la gabapentina son considerados de primera línea en el tratamiento de la NPH (2); sin embargo, la inclusión de la pregabalina como tratamiento de primera línea autorizado en el tratamiento del dolor neuropático ha abierto nuevas perspectivas (3). El objetivo de este estudio es demostrar la eficacia y seguridad de la Pregabalina (LYRICA®) en el tratamiento del dolor neuropático de la NPH refractaria a la terapia convencional. Se realizó un estudio de eficacia y seguridad de la terapia con pregabalina en 10 pacientes con NPH de más de 3 meses de evolución con escasa respuesta a tratamiento médico convencional. La variable fundamental era la Escala Visual Analógica (EVA) al inicio y cada mes durante los 4 meses que duró la recogida de datos. Los ascensos de fármacos se realizaban mensualmente si no se obtenía una reducción superior al 30% con la dosis previa. Tras la evaluación del primer mes de tratamiento con pregabalina se obtuvo en la mitad de los pacientes una reducción del dolor superior al 30% del EVA inicial, obteniéndose resultados similares para el segundo y tercer mes de tratamiento. La Pregabalina resultó ser un fármaco de elevada eficacia en la reducción del dolor en la NPH refractaria a terapia convencional, con un bajo nivel de efectos secundarios

Polμ deficiency increases resistance to oxidative damage and delays liver aging

Polμ is an error-prone PolX polymerase that contributes to classical NHEJ DNA repair. Mice lacking Polμ (Polμ−/−) show altered hematopoiesis homeostasis and DSB repair and a more pronounced nucleolytic resection of some V(D)J junctions. We previously showed that Polμ−/− mice have increased learning capacity at old ages, suggesting delayed brain aging. Here we investigated the effect of Polμ−/− deficiency on liver aging. We found that old Polμ−/− mice (>20 month) have greater liver regenerative capacity compared with wt animals. Old Polμ−/− liver showed reduced genomic instability and increased apoptosis resistance. However, Polμ−/− mice did not show an extended life span and other organs (e.g., heart) aged normally. Our results suggest that Polμ deficiency activates transcriptional networks that reduce constitutive apoptosis, leading to enhanced liver repair at old age

Polμ deficiency increases resistance to oxidative damage and delays liver aging

Polμ is an error-prone PolX polymerase that contributes to classical NHEJ DNA repair. Mice lacking Polμ (Polμ−/−) show altered hematopoiesis homeostasis and DSB repair and a more pronounced nucleolytic resection of some V(D)J junctions. We previously showed that Polμ−/− mice have increased learning capacity at old ages, suggesting delayed brain aging. Here we investigated the effect of Polμ−/− deficiency on liver aging. We found that old Polμ−/− mice (>20 month) have greater liver regenerative capacity compared with wt animals. Old Polμ−/− liver showed reduced genomic instability and increased apoptosis resistance. However, Polμ−/− mice did not show an extended life span and other organs (e.g., heart) aged normally. Our results suggest that Polμ deficiency activates transcriptional networks that reduce constitutive apoptosis, leading to enhanced liver repair at old age

Evaluation of SCE in different cellular models of Polμ deficiency.

<p>(<b>A</b>) Illustrative example of sister chromatid exchange (SCE) between green chromatids (labeled by BrdU incorporation) and blue chromatids (DAPI stained). The enlarged image to the left shows a chromosome with 3 crossovers (indicated by arrows) in which green and blue are combined in the same chromatid. (<b>B–D</b>) Sister chromatid exchange (SCE) in three models of Polμ deficiency: (<b>B</b>) MEFs, (<b>C</b>) B lymphocytes and (<b>D</b>) CHO-KS4 cells expressing DN-Polμ (ΔN); EV = empty vector.</p

Improved liver function in old Polμ^−/− mice.

<p>(<b>A</b>) Analysis of 53P1-positive cells in liver of old (>20 months) wt (n = 6; black bars) and Polμ^−/− mice (n = 6; gray bars). (<b>B</b>) Cell cycle analysis of freshly prepared liver suspensions from wt (black bars) and Polμ^−/− mice (gray bars), after staining with PI. (<b>C</b>) TUNEL assay in cryopreserved liver tissue from adult (14 m) mice. Percentages of positive (apoptotic) cells per field are shown. Three livers per genotype were analyzed (4771 wt cells and 8289 Polμ^−/− cells) and individual values (relative to wt) and means ± SEM are shown. (<b>D</b>) Percentage of proliferating (PH3-positive) cells in livers of old (18–23 m) wt (black) and Polμ^−/− mice (gray) after partial hepatectomy (PH). Percentages of PH3-positive cells are presented as the mean ± SEM of two-three animals per condition; n = 11. Polμ^−/− animals survived surgery better than wt animals (82%; 9/11 vs 64%; 7/11). (<b>E, F</b>) Representative images of immunostaining for PH3 (E) and CK19 (F) in sections of old (18–23 m) Polμ^−/− and wt liver at the indicated times after PH. Control indicates negative controls without the primary antibody.</p

Liver from old Polμ^−/− mice shows fewer aging-related features than liver from age-matched wt controls.

<p>(<b>A</b>) Involvement of autophagy activity in the Polμ^−/− lifespan extension phenotype. Heart samples from old (18–25 m; o) and young (8–12 w; y) Polμ^−/− (gray bars) and wt mice (black bars) were analyzed by qRT-PCR for the expression of a panel of genes essential for autophagy (Atg2l, Atg5l, Atg7l and Map11c3b). Results, expressed as 2∧–(DCT), show mean values ± SD (n = 5). (<b>B</b>) Protein carbonyls (Carb) and peroxided lipids (PeroxL; thiobarbituric acid reactive substances, TBARS), as macromolecular oxidative damage, were determined (nmol/mg prot), as described in the supporting information, in perfused liver of old (>20 months) Polμ^−/− (black bars) and wt gray bars) mice Results are mean values ± SD (n = 3). (<b>C</b>) Heat map of microarray expression data for livers from adult (14 m) wt and Polμ^−/− mice. The relative gene expression scale is shown, with normalized scores ranging from 0 to 20. Gene cluster and case cluster dendrograms are plotted to the top and left. (<b>D</b>) Microarray data validation. mRNA levels were determined by TaqMan RT-PCR in livers from adult wt (black) and Polμ^−/− mice (gray). Each determination was performed in triplicate and normalized to β-actin expression. Data are the means ± SEM of 4 samples per genotype, represented relative to expression in age-matched wt liver. (<b>E</b>) Scheme of the interaction network among apoptosis-related or p53-inducible genes downregulated in old Polμ^−/− liver.</p

Genetic stability in Polμ^−/− mice.

<p>(<b>A</b>) Polyploidy (determined by PI staining) in hepatocytes from old (14 m) wt (black) and Polμ^−/− mice (gray). Data are presented as the relative amounts (arbitrary units) of 8n and ≥8n cells (means ± SEM, 4 samples per genotype. (<b>B</b>) Survival (min) of adult (8–12 w) mice after i.v. injection with paraquat (70 mg/kg). Individual data and means are shown for wt (black) and Polμ^−/− (gray) mice (n = 13). Data are means ± SEM. (<b>C, D</b>) Comparative mutation frequency (MF) in a panel of selected microsatellites (MS) in Polμ^−/− mice vs wt animals. The table (<b>D</b>) compares individual absolute MF values in the different MS sequences analyzed in Polμ^−/− mice relative to wt animals (100%); (*) indicates animals treated with paraquat.</p

Summary of different physiological parameters analyzed in old Polμ^−/− mice.

<p>Numbers indicates the ratio KO/WT for each parameter. Footnotes.</p>(a)<p>Corresponding specifically to G2/M cells;</p>(b)<p>Evaluation of senescence <i>ex vivo</i>, at atmospheric concentration of O₂;</p>(c)<p>Ploidy (≥4n) evaluation after PI staining;</p>(d)<p>Aneuploidy and translocation frequency evaluation;</p>(e)<p>Evaluated on CFU-GM progenitors;</p>(f)<p>Evaluated in cell lines;</p>(g)<p>ALT/GPT; AST/GOT; Billirubin and Albumin;</p>(h)<p>P-Hepatect. Recovery: partial hepatectomy recovery;</p>(i)<p>Protein carbonyls;</p>(j)<p>peroxided lipids;</p>(k)<p>Values obtained at atmospheric oxygen;</p>(l)<p>Paraquat Resistance, (70 mg/kg; ip);</p>(m)<p>Analysis of CHO-DN cell line, at atmospheric oxygen;</p>(n)<p>HR express. qRT-PCR analysis of selected functions involved in HR;</p>(o)<p>Tumor susceptibility. Evaluation of spontaneous tumor incidence and susceptibility to thymic lymphomas by low dose of radiation;</p>(p)<p>Evaluations in 129/BALBc and B6 backgrounds, respectively; Abbreviations: Thy, thymus; Sp, spleen; Br, brain, Sk, skin; Ov, ovary; Panc, Pancreas; PB, peripheral blood. Symbols; (<b>++</b>), (<b>+</b>), (<b>+/−</b>) and (−) indicates that values for KO animals were clearly higher, superior, similar and reduced, respectivelly, in comparison with control (wt) animals.</p

Heart molecular signature and cardiovascular phenotype of Polμ^−/− mice.

<p>(<b>A–C</b>) Cardiovascular parameters in old Polμ^−/− (gray) and wt (black) mice (16–18 m). (<b>A</b>) Electrocardiographic parameters; QRS complex and PR segment intervals are shown in miliseconds (msec). (<b>B</b>) Heart rate (beats per minute; bpm). Data are means ±SD of weekly readings collected over a 4-week period. (<b>C</b>) Systolic and diastolic blood pressure (mm Hg) in wt (black) and Polμ^−/− mice (white) over a 4-week period. Results are means ± SD (n = 3 mice). (<b>D</b>) After DNA array expression analysis of hearts from old (20–24 m) and young (3–4 m) wt mice, six genes showing an aging-related profile were selected and confirmed by qRT-PCR. Data from old mice (black) are presented relative the expression level of each gene in young mice (gray). (<b>E</b>) Comparative gene expression analysis in young (y; 8–12 weeks) and old (o; 20–24 m) heart tissue from wt (black) and Polμ^−/− mice (gray). Data were normalized to the expression level of each gene in young wt mice. Results, expressed as 2∧–(DCT), are mean values ± SD (n = 3).</p