It has long been presumed impossible to measure telomeres in vertebrate DNA by PCR amplification with oligonucleotide primers designed to hybridize to the TTAGGG and CCCTAA repeats, because only ...primer dimer-derived products are expected. Here we present a primer pair that eliminates this problem, allowing simple and rapid measurement of telomeres in a closed tube, fluorescence-based assay. This assay will facilitate investigations of the biology of telomeres and the roles they play in the molecular pathophysiology of diseases and aging.
Humans choose which foods they will eat from multiple options. The use of cafeteria-style diets with rodent models has increased our understanding of how a multichoice food environment affects eating ...and health. However, the wide variances in energy density, texture, and the content of micronutrients, fiber, and protein can be interpretatively problematic when human foodstuffs are used to create rodent cafeteria diets. We minimized these differences with a custom rodent cafeteria diet (ROD) that varied similarly to a previously used human-foods cafeteria diet (HUM) in fat and sugar content. Here, we used our custom Five-Item Food Choice Monitor to compare the intake and meal patterns of rats offered ROD and HUM in a crossover design. Compared with chow, rats consumed more calories, sugar, and fat and less protein and carbohydrate while on either of the choice diets (
< 0.05). While energy intake was similar between HUM and ROD, there were differences in the responses. Rats consumed more of the low-fat, low-sugar choice on the ROD compared with the nutritionally similar choice on the HUM leading to differences in fat and carbohydrate intake between the diets (
< 0.05). The stability of macronutrient intake while on either choice diet suggests macronutrient intake is determined by the available foods and is strongly regulated. Therefore, interpretative consideration must be given to the nature of food choices in the context of available options when interpreting cafeteria-diet intake.
Obesity and shorter telomeres are commonly associated with elevated risk for age-related diseases and mortality. Whether telomere length (TL) may be associated with obesity or variations in adiposity ...is not well established. Therefore, we set out to test the hypothesis that TL may be a risk factor for increased adiposity using data from a large population-based cohort study.
Levels of adiposity were assessed in six ways (obesity status, body mass index (BMI), the percentage of body fat or % body fat, leptin, visceral and subcutaneous fat mass) in 2721 elderly subjects (42% black and 58% white). Associations between TL measured in leukocytes at baseline and adiposity traits measured at baseline, and three of these traits after 7 years of follow-up were tested using regression models adjusting for important covariates. Additionally, we look at weight changes and relative changes in BMI and % body fat between baseline and follow-up.
At baseline, TL was negatively associated with % body fat (ß=-0.35±0.09, P=0.001) and subcutaneous fat (ß=-2.66±1.07, P=0.01), and positively associated with leptin after adjusting for % body fat (ß=0.32±0.14, P=0.001), but not with obesity, BMI or visceral fat. Prospective analyses showed that longer TL was associated with positive percent change between baseline and 7-year follow-up for both BMI (ß=0.48±0.20, P=0.01) and % body fat (ß=0.42±0.23, P=0.05).
Our study suggests that shorter TL may be a risk factor for increased adiposity. Coupling with previous reports on their reversed roles, the relationship between adiposity and TL may be complicated and may warrant more prospective studies.
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DOBA, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Objectives
Human age‐dependent telomere attrition and telomere shortening are associated with several age‐associated diseases and poorer overall survival. The aim of this study was to determine ...longitudinal leucocyte telomere length dynamics and identify factors associated with temporal changes in telomere length.
Design and Methods
Leucocyte telomere length was measured by quantitative polymerase chain reaction in 8074 participants from the Prevention of Renal and Vascular End‐stage Disease (PREVEND) study, an ongoing community‐based prospective cohort study initiated in 1997. Follow‐up data were available at two time‐points up to 2007. Leucocyte telomere length was measured, on between one and three separate occasions, in a total of 16 783 DNA samples. Multilevel growth models were created to identify the factors that influence leucocyte telomere dynamics.
Results
We observed an average attrition rate of 0.47 ± 0.16 relative telomere length units (RTLUs) per year in the study population aged 48 (range 39–60) years at baseline. Annual telomere attrition rate increased with age (P < 0.001) and was faster on average in men than in women (P for interaction 0.043). The major independent factors determining telomere attrition rate were active smoking (approximately tripled the loss of RTLU per year, P < 0.0001) and multiple traits of the metabolic syndrome (waist–hip ratio, P = 0.007; blood glucose level, P = 0.045, and HDL cholesterol level, P < 0.001).
Conclusions
Smoking and variables linked to the metabolic syndrome are modifiable lifestyle factors that accelerate telomere attrition in humans. The higher rate of cellular ageing may mediate the link between smoking and the metabolic syndrome to an increased risk of several age‐associated diseases.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
•Both depletion models (antibiotic depletion and germ free) led to receiver rats displaying microbiota profile characteristic of their respective donors.•Colonization with a dysbiotic microbiota led ...to an increase in body weight gain and food intake.•Microbiota composition modulates Iba1+ immune cell presence along the gut-brain axis•Colonization with a dysbiotic microbiota led to a reduction on vagal innervation to the NTS associated with a loss of CCK-induced satiety.
Vagal afferent neurons (VAN), located in the nodose ganglion (NG) innervate the gut and terminate in the nucleus of solitary tract (NTS) in the brainstem. They are the primary sensory neurons integrating gut-derived signals to regulate meal size. Chronic high-fat diet (HFD) consumption impairs vagally mediated satiety, resulting in overfeeding. There is evidence that HFD consumption leads to alterations in both vagal nerve function and structural integrity. HFD also leads to marked gut microbiota dysbiosis; in rodent models, dysbiosis is sufficient to induce weight gain. In this study, we investigated the effect of microbiota dysbiosis on gut-brain vagal innervation independently of diet. To do so, we recolonized microbiota-depleted rats with gastrointestinal (GI) contents isolated from donor animals fed either a HFD (45 or 60% fat) or a low fat diet (LFD, 13% fat). We used two different depletion models while maintaining the animals on LFD: 1) conventionally raised Fischer and Wistar rats that underwent a depletion paradigm using an antibiotic cocktail and 2) germ free (GF) raised Fischer rats. Following recolonization, receiver animals were designated as ConvLF and ConvHF. Fecal samples were collected throughout these studies and analyzed via 16S Illumina sequencing. In both models, bacteria that were identified as characteristic of HFD were successfully transferred to recipient animals. Three weeks post-colonization, ConvHF rats showed significant increases in ionized calcium-binding adapter molecule-1 (Iba1) positive immune cells in the NG compared to ConvLF animals. Additionally, using isolectin B4 (IB4) staining to identify c-fibers, we found that, compared to ConvLF animals, ConvHF rats displayed decreased innervation at the level of the medial NTS; c-fibers at this level are believed to be primarily of vagal origin. This alteration in vagal structure was associated with a loss in satiety induced by the gut peptide cholecystokinin (CCK). Increased presence of immunocompetent Iba1+ cells along the gut-brain axis and alterations in NTS innervation were still evident in ConvHF rats compared to ConvLF animals 12 weeks post-colonization and were associated with increases in food intake and body weight (BW). We conclude from these data that microbiota dysbiosis can alter gut-brain vagal innervation, potentially via recruitment and/or activation of immune cells.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The current quantitative polymerase chain reaction (QPCR) assay of telomere length measures telomere (T) signals in experimental DNA samples in one set of reaction wells, and single copy gene (S) ...signals in separate wells, in comparison to a reference DNA, to yield relative T/S ratios that are proportional to average telomere length. Multiplexing this assay is desirable, because variation in the amount of DNA pipetted would no longer contribute to variation in T/S, since T and S would be collected within each reaction, from the same input DNA. Multiplexing also increases throughput and lowers costs, since half as many reactions are needed. Here, we present the first multiplexed QPCR method for telomere length measurement. Remarkably, a single fluorescent DNA-intercalating dye is sufficient in this system, because T signals can be collected in early cycles, before S signals rise above baseline, and S signals can be collected at a temperature that fully melts the telomere product, sending its signal to baseline. The correlation of T/S ratios with Terminal Restriction Fragment (TRF) lengths measured by Southern blot was stronger with this monochrome multiplex QPCR method (R² = 0.844) than with our original singleplex method (R² = 0.677). Multiplex T/S results from independent runs on different days were highly reproducible (R² = 0.91).
We previously reported that psychological stress is linked to and possibly accelerates cellular aging, as reflected by lower PBMC telomerase and shortened telomeres. Psychological stress is a major ...risk factor for cardiovascular disease (CVD), with multiple behavioral and physiological mediators. Telomere shortness has been associated with CVD, but the relationship between low telomerase activity, a potential precursor to telomere shortening, and CVD risk factors has not been examined in humans. Here we examine whether telomere length and telomerase in leukocytes are associated with physiological signs of stress arousal and CVD risk factors in 62 healthy women. Low telomerase activity in leukocytes was associated with exaggerated autonomic reactivity to acute mental stress and elevated nocturnal epinephrine. Further, low telomerase activity was associated with the major risk factors for CVD —smoking, poor lipid profile, high systolic blood pressure, high fasting glucose, greater abdominal adiposity—as well as to a composite Metabolic Syndrome variable. Telomere length was related only to elevated stress hormones (catecholamines and cortisol). Thus, we propose that low leukocyte telomerase constitutes an early marker of CVD risk, possibly preceding shortened telomeres, that results in part from chronic stress arousal. Possible cellular mechanisms by which low telomerase may link stress and traditional risk factors to CVD are discussed. These findings may implicate telomerase as a novel and important mediator of the effects of psychological stress on physical health and disease.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
ObjectivesTo examine the association of employment and work schedule with shorter DNA telomeres, a marker of cellular ageing and disease risk factor, and consider whether differences were related to ...health, behaviours and sociodemographic factors, or varied by stress levels or menopausal status.MethodsThis cross-sectional analysis of 608 women aged 35–74 in the Sister Study examined determinants of relative telomere length (rTL) measured by quantitative PCR in leucocyte DNA. Age-adjusted regression models estimated base pair (bp) rTL differences for current and lifetime schedule characteristics (ie, part-time, full-time or overtime hours; multiple jobs; irregular hours; shiftwork; work at night). Covariates included race, smoking, perceived stress, sleep, physical activity, health and menopausal status, education, marital status, live births, children under 18, measured body mass index and urinary stress hormones.ResultsCompared with non-employed women with moderate or substantial past work histories (n=190), those currently working full-time (n=247; median 40 h/week) had a shorter rTL, an age-adjusted difference of −329 bp (95% CI −110 to −548). Longer-duration full-time work was also associated with shorter rTL (age-adjusted difference of −472 bp, 95% CI −786 to −158 for 20+ vs 1–5 years). Findings were not explained by health and demographic covariates. However, rTL differences for working at least full-time were greater in women with higher stress and epinephrine levels.ConclusionsCurrent and long-term full-time work were associated with shorter rTL, with differences of similar magnitude to smoking and history of heart disease or diabetes. Longitudinal data with specific stress measures are needed to further evaluate the impact of work schedule on rTL.
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BFBNIB, CMK, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Our goal is to investigate if microbiota composition modulates reward signaling and assess the role of the vagus in mediating microbiota to brain communication.
Male germ-free Fisher rats were ...colonized with gastrointestinal contents from chow (low fat (LF) ConvLF) or HF (ConvHF) fed rats.
Following colonization, ConvHF rats consumed significantly more food than ConvLF animals. ConvHF rats displayed lower feeding-induced extracellular DOPAC levels (a metabolite of dopamine) in the Nucleus Accumbens (NAc) as well as reduced motivation for HF foods compared to ConvLF rats. Dopamine receptor 2 (DDR2) expression levels in the NAc were also significantly lower in ConvHF animals. Similar deficits were observed in conventionally raised HF fed rats, showing that diet-driven alteration in reward can be initiated via microbiota. Selective gut to brain deafferentation restored DOPAC levels, DRD2 expression, and motivational drive in ConvHF rats.
We concluded from these data that a HF-type microbiota is sufficient to alter appetitive feeding behavior and that bacteria to reward communication is mediated by the vagus nerve.
•A HF-type microbiota is sufficient to alter dopamine signaling and appetitive feeding behavior, independently of diet.•The vagal afferent pathway mediates the effects of microbiota on the mesolimbic dopamine system.•Both mesolimbic and nigrostriatal dopamine pathways are influenced by gastrointestinal bacterial make up.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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