Importancia del estrés oxidativo en la diferencia de longevidad entre machos y hembras

RESUMEN Uno de los mayores logros del siglo XX es el aumento de la esperanza de vida de la población humana. En Europa, ésta se ha duplicado entre 1900 y 1992. En todos los casos la esperanza de vida de las mujeres supera a la de los hombres. De hecho, en España en 1900, la esperanza de vida para los hombres era de 33.7 años y para las mujeres de 35.1, es decir, un 3.8% superior en mujeres frente a hombres. En 1992 la esperanza de vida de los hombres era de 73.7 años, mientras que la de las mujeres era 83.8, lo cual supone un 9.9% más en las mujeres que en los hombres. La base de la diferencia de esperanza de vida entre machos y hembras está aún por esclarecer. Este fenómeno no es atribuible a diferencias sociológicas, puesto que también se reproduce en otras especies. En nuestro laboratorio, las ratas Wistar hembras muestran un 16% más de vida media que las ratas Wistar machos. En la presente tesis demostramos que las diferencias de longevidad entre machos hembras tienen una base biológica: los estrógenos suponen una ventaja ante la supervivencia de las hembras, puesto que inducen la expresión de enzimas antioxidantes, y ello las protege frente al estrés oxidativo, y por tanto les confiere una mayor longevidad. Así pues, el estrés oxidativo encontrado en los machos es superior al de las hembras. Se determinó la producción de peróxido de hidrógeno en mitocondrias hepáticas y cerebrales (sinápticas y no sinápticas) aisladas de machos y hembras y observamos que las procedentes de los machos producen una cantidad significativamente superior de peróxido de hidrógeno. Asimismo, los niveles del antioxidante endógeno glutatión reducido son mayores en las mitocondrias procedentes de las hembras en comparación con los machos. A consecuencia del mayor estrés oxidativo en los machos, el DNA mitocondrial de los mismos está hasta cuatro veces más oxidado que el de las hembras. El aumento de este parámetro está asociado a una menor longevidad. Todo ello se puede explicar porque las hembras se comportan como dobles transgénicos, que sobreexpresan las enzimas antioxidantes glutatión peroxidasa y manganeso superóxido dismutasa. Además la expresión del marcador de envejecimiento 16S rRNA está disminuida en los machos en comparación con las hembras, lo cual supone que con una misma edad cronológica, las hembras poseen una edad biológica menos que los machos, es decir son más jóvenes que los machos. Otro parámetro relacionado con la longevidad que determinamos es la actividad telomerasa, la cual además posee un elemento de respuesta a estrógenos, y obtuvimos que las hembras poseen una actividad telomerasa superior en comparación con los machos. Por otro lado, los estudios in vitro realizados en la presente tesis, empleando un cultivo celular de células MCF7, que provienen de carcinoma mamario, indican que el mecanismo por el cual los estrógenos inducen la expresión de enzimas antioxidantes está mediado tanto por los receptores estrogénicos, como por la activación de la vía de señalización de las MAP Kinasas. Además, los efectos antioxidantes del estradiol son mimetizados por la genisteína, el fitoestrógeno más abundante presente en la soja. La importancia de este hecho recae en que en este caso, la genisteína no sólo no posee efectos cancerígenos, sino que previene el desarrollo de diversos cánceres como el mamario o el de próstata. __________________________________________________________________________________________________Females live longer than males in many species, including humans. This can be explained on the basis of the mitochondrial theory of aging. We have investigated the differential mitochondrial oxidative stress between males and females to understand the molecular mechanisms enabling females to live longer than males. Mitochondria are a major source of free radicals in cells. Those from female rats generate half the amount of peroxides than those of males. Mitochondria from females have higher levels of reduced glutathione than those from males. Oxidative damage to mitochondrial DNA in males is four fold higher than that of females. This is due to a higher expression and activities of Mn-superoxide dismutase and of glutathione peroxidase in females, which behave as double transgenics overexpressing superoxide dismutase, and glutathione peroxidase, conferring protection against free radical mediated damage in aging. Moreover, 16S rRNA expression, which decreases significantly with aging, is four times higher in mitochondria from females than in those from males of the same chronological age. Studies in vitro show that oestradiol is an antioxidant, but that estrogenic effects must be transcriptional, by inducing the antioxidant enzyme gene expression. The facts reported here provide molecular evidence to explain the different life span in males and females. The challenge for the future is to find molecules that have the beneficial effects of estradiol, but without its feminizing effects. Phytoestrogens or phytoestrogen-related molecules may be good candidates to meet this challenge

Medical physiology 3

Material docente fisiología médica 3 en inglé

Influence of different types of pulp treatment during isolation in the obtention of human dental pulp stem cells

Background: Different methods have been used in order to isolate dental pulp stem cells. The aim of this study was to study the effect of different types of pulp treatment during isolation, under 3% O 2 conditions, in the time needed and the efficacy for obtaining dental pulp stem cells. Material and Methods: One hundred and twenty dental pulps were used to isolate dental pulp stem cells treating the pulp tissue during isolation using 9 different methods, using digestive, disgregation, or mechanical agents, or combining them. The cells were positive for CD133, Oct4, Nestin, Stro-1, CD34 markers, and negative for the hematopoietic cell marker CD-45, thus confirming the presence of mesenchymal stem cells. The efficacy of dental pulp stem cells obtention and the minimum time needed to obtain such cells comparing the 9 different methods was analyzed. Results: Dental pulp stem cells were obtained from 97 of the 120 pulps used in the study, i.e. 80.8% of the cases. They were obtained with all the methods used except with mechanical fragmentation of the pulp, where no enzymatic digestion was performed. The minimum time needed to isolate dental pulp stem cells was 8 hours, digesting with 2mg/ml EDTA for 10 minutes, 4mg/ml of type I collagenase, 4mg/ml of type II dispase for 40 minutes, 13ng/ ml of thermolysine for 40 minutes and sonicating the culture for one minute. Conclusions: Dental pulp stem cells were obtained in 97 cases from a series of 120 pulps. The time for obtaining dental pulp stem cells was reduced maximally, without compromising the obtention of the cells, by combining digestive, disgregation, and mechanical agent

Application of mesenchymal stem cells in bone regenerative procedures in oral implantology. A literature review

Objective: The aim of this work was to review de literature about the role of mesenchymal stem cells in bone regenerative procedures in oral implantology, specifically, in the time require to promote bone regeneration. Study Desing: A bibliographic search was carried out in PUBMED with a combination of different key words. Animal and human studies that assessed histomorphometrically the influence of mesenchymal stem cells on bone regeneration procedures in oral implantology surgeries were examined. Reults: - Alveolar regeneration: Different controlled histomorphometric animal studies showed that bone regeneration is faster using stem cells seeded in scaffolds than using scaffolds or platelet rich plasma alone. Human studies revealed that stem cells increase bone regeneration. - Maxillary sinus lift: Controlled studies in animals and in humans showed higher bone regeneration applying stem cells compared with controls. - Periimplantary bone regeneration and alveolar distraction: Studies in animals showed higher regeneration when stem cells are used. In humans, no evidence of applying mesenchymal stem cells in these regeneration procedures was found. Conclusion: Stem cells may promote bone regeneration and be useful in bone regenerative procedures in oral implantology, but no firm conclusions can be drawn from the rather limited clinical studies so far performed

PTEN Mediates the Antioxidant Effect of Resveratrol at Nutritionally Relevant Concentrations

Introduction. Antioxidant properties of resveratrol have been intensively studied for the last years, both in vivo and in vitro. Its bioavailability after an oral dose is very low and therefore it is very important to make sure that plasma concentrations of free resveratrol are sufficient enough to be active as antioxidant. Aims. In the present study, using nutritionally relevant concentrations of resveratrol, we aim to confirm its antioxidant capacity on reducing peroxide levels and look for the molecular pathway involved in this antioxidant effect. Methods. We used mammary gland tumor cells (MCF-7), which were pretreated with different concentrations of resveratrol for 48 h, and/or a PTEN inhibitor (bpV: bipy). Hydrogen peroxide levels were determined by fluorimetry, PTEN levels and Akt phosphorylation by Western Blotting, and mRNA expression of antioxidant genes by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Results. Resveratrol treatment for 48 h lowered peroxide levels in MCF-7, even at low nutritional concentrations (1 nM). This effect was mediated by the activation of PTEN/Akt pathway, which resulted in an upregulation of catalase and MnSOD mRNA levels. Conclusion. Resveratrol acts as an antioxidant at nutritionally relevant concentrations by inducing the expression of antioxidant enzymes, through a mechanism involving PTEN/Akt signaling pathway

Small extracellular vesicles from young adipose-derived stem cells prevent frailty, improve health span, and decrease epigenetic age in old mice.

Aging is associated with an increased risk of frailty, disability, and mortality. Strategies to delay the degenerative changes associated with aging and frailty are particularly interesting. We treated old animals with small extracellular vesicles (sEVs) derived from adipose mesenchymal stem cells (ADSCs) of young animals, and we found an improvement in several parameters usually altered with aging, such as motor coordination, grip strength, fatigue resistance, fur regeneration, and renal function, as well as an important decrease in frailty. ADSC-sEVs induced proregenerative effects and a decrease in oxidative stress, inflammation, and senescence markers in muscle and kidney. Moreover, predicted epigenetic age was lower in tissues of old mice treated with ADSC-sEVs and their metabolome changed to a youth-like pattern. Last, we gained some insight into the microRNAs contained in sEVs that might be responsible for the observed effects. We propose that young sEV treatment can promote healthy aging

Genistein effect on cognition in prodromal Alzheimer's disease patients : the GENIAL clinical trial

Delaying the transition from minimal cognitive impairment to Alzheimer's dementia is a major concern in Alzheimer's disease (AD) therapeutics. Pathological signs of AD occur years before the onset of clinical dementia. Thus, long-term therapeutic approaches, with safe, minimally invasive, and yet effective substances are recommended. There is a need to develop new drugs to delay Alzheimer's dementia. We have taken a nutritional supplement approach with genistein, a chemically defined polyphenol that acts by multimodal specific mechanisms. Our group previously showed that genistein supplementation is effective to treat the double transgenic (APP/PS1) AD animal model

Functional transcriptomic analysis of centenarians' offspring reveals a specific genetic footprint that may explain that they are less frail than age-matched non-centenarians' offspring

Centenarians exhibit extreme longevity and compression of morbidity and display a unique genetic signature. Centenarians' offspring seem to inherit centenarians' compression of morbidity, as measured by lower rates of age-related pathologies. We aimed to ascertain whether centenarians' offspring are less frail and whether they are endowed with a "centenarian genetic footprint" in a case-control study, matched 1:1 for gender, age ±5 years, and place of birth and residence. Cases must have a living parent aged 97 years or older, aged 65-80 years, community dwelling, not suffering from a terminal illness, or less than 6 months of life expectancy. Controls had to meet the same criteria as cases except for the age of death of their parents (not older than 89 years). Centenarians were individuals 97 years or older. Frailty phenotype was determined by Fried's criteria. We collected plasma and peripheral blood mononuclear cells from 63 centenarians, 88 centenarians' offspring, and 88 noncentenarians' offspring. miRNA expression and mRNA profiles were performed by the GeneChip miRNA 4.0 Array and GeneChip Clariom S Human Array, respectively. We found a lower incidence of frailty among centenarians' offspring when compared with their contemporaries' noncentenarians' offspring (p < .01). Both miRNA and mRNA expression patterns in centenarians' offspring were more like those of centenarians than those of noncentenarians' offspring (p < .01). In conclusion, centenarians' offspring are less frail than age-matched noncentenarians' offspring, and this may be explained by their unique genetic endowment

Centenarians overexpress pluripotency-related genes

Human mesenchymal cells can become pluripotent by the addition of Yamanaka factors OCT3/4, SOX2, c-MYC, KLF4. We have recently reported that centenarians overexpress BCL-xL, which has been shown to improve pluripotency; thus, we aimed to determine the expression of pluripotency-related genes in centenarians. We recruited 22 young, 32 octogenarian, and 47 centenarian individuals and determined the mRNA expression of Yamanaka factors and other stemness-related cell surface marker genes (VIM, BMP4, NCAM, BMPR2) in peripheral blood mononuclear cells by reverse transcription polymerase chain reaction. We found that centenarians overexpress OCT3/4, SOX2, c-MYC, VIM, BMP4, NCAM, and BMPR2, when compared with octogenarians (p < .05). We further tested the functional role of BCL-xL in centenarians' ability to express pluripotency-related genes: lymphocytes from octogenarians transduced with BCL-xL overexpressed SOX2, c-MYC, and KLF4. We conclude that centenarians overexpress Yamanaka Factors and other stemness-related cell surface marker genes, which may contribute to their successful aging

Relation between genetic factors and frailty in older adults

Objectives Frailty is a geriatric syndrome that identifies individuals at higher risk of disability, institutionalization, and death. We previously reported that frailty is related to oxidative stress and cognitive impairment-related biomarkers. The aim of this study was to determine whether frailty is associated with genetic variants. Design Longitudinal population-based cohort of 2488 community-dwelling people from Toledo, Spain, aged 65 years or older. Setting and participants We obtained blood samples from 78 individuals with frailty and 74 nonfrail individuals who were nonfrail (according to Fried criteria) from the Toledo Study of Healthy Ageing and extracted DNA using the Chemagic DNA blood kit. Measures Sample genotyping was carried out by means of Axiom Exome 319 Genotyping Array (Thermo Fisher Scientific), which contains 295,988 markers [single-nucleotide polymorphisms (SNPs) and rare variants], and transferred to the GeneTitan Instrument (Affymetrix). Results We found 15 SNPs (P < .001), 18 genes (P < .005), and 4 pathways (P < .05) related to cytokine-cytokine receptor interaction, natural killer cell-mediated cytotoxicity, regulation of autophagy, and renin-angiotensin system as the most strongly associated with frailty. Conclusions/Implications The specific genetic features related to energy metabolism, biological processes regulation, cognition, and inflammation highlighted by this preliminary analysis offer useful insights for finding biologically meaningful biomarkers of frailty that allow early diagnosis and treatment. Further research is needed to confirm our novel findings in a larger population. Indeed, the EU-funded FRAILOMICS research effort will address this question