Isolation of quiescent and nonquiescent cells from yeast stationary-phase cultures

Quiescence is the most common and, arguably, most poorly understood cell cycle state. This is in part because pure populations of quiescent cells are typically difficult to isolate. We report the isolation and characterization of quiescent and nonquiescent cells from stationary-phase (SP) yeast cultures by density-gradient centrifugation. Quiescent cells are dense, unbudded daughter cells formed after glucose exhaustion. They synchronously reenter the mitotic cell cycle, suggesting that they are in a G0 state. Nonquiescent cells are less dense, heterogeneous, and composed of replicatively older, asynchronous cells that rapidly lose the ability to reproduce. Microscopic and flow cytometric analysis revealed that nonquiescent cells accumulate more reactive oxygen species than quiescent cells, and over 21 d, about half exhibit signs of apoptosis and necrosis. The ability to isolate both quiescent and nonquiescent yeast cells from SP cultures provides a novel, tractable experimental system for studies of quiescence, chronological and replicative aging, apoptosis, and the cell cycle

Identification of Circulating Proteins associated With General Cognitive Function among Middle-Aged and Older adults

Identifying circulating proteins associated with cognitive function may point to biomarkers and molecular process of cognitive impairment. Few studies have investigated the association between circulating proteins and cognitive function. We identify 246 protein measures quantified by the SomaScan assay as associated with cognitive function (p \u3c 4.9E-5, n up to 7289). Of these, 45 were replicated using SomaScan data, and three were replicated using Olink data at Bonferroni-corrected significance. Enrichment analysis linked the proteins associated with general cognitive function to cell signaling pathways and synapse architecture. Mendelian randomization analysis implicated higher levels of NECTIN2, a protein mediating viral entry into neuronal cells, with higher Alzheimer\u27s disease (AD) risk (p = 2.5E-26). Levels of 14 other protein measures were implicated as consequences of AD susceptibility (p \u3c 2.0E-4). Proteins implicated as causes or consequences of AD susceptibility may provide new insight into the potential relationship between immunity and AD susceptibility as well as potential therapeutic targets

52 Genetic Loci Influencing Myocardial Mass.

BACKGROUND: Myocardial mass is a key determinant of cardiac muscle function and hypertrophy. Myocardial depolarization leading to cardiac muscle contraction is reflected by the amplitude and duration of the QRS complex on the electrocardiogram (ECG). Abnormal QRS amplitude or duration reflect changes in myocardial mass and conduction, and are associated with increased risk of heart failure and death. OBJECTIVES: This meta-analysis sought to gain insights into the genetic determinants of myocardial mass. METHODS: We carried out a genome-wide association meta-analysis of 4 QRS traits in up to 73,518 individuals of European ancestry, followed by extensive biological and functional assessment. RESULTS: We identified 52 genomic loci, of which 32 are novel, that are reliably associated with 1 or more QRS phenotypes at p < 1 × 10(-8). These loci are enriched in regions of open chromatin, histone modifications, and transcription factor binding, suggesting that they represent regions of the genome that are actively transcribed in the human heart. Pathway analyses provided evidence that these loci play a role in cardiac hypertrophy. We further highlighted 67 candidate genes at the identified loci that are preferentially expressed in cardiac tissue and associated with cardiac abnormalities in Drosophila melanogaster and Mus musculus. We validated the regulatory function of a novel variant in the SCN5A/SCN10A locus in vitro and in vivo. CONCLUSIONS: Taken together, our findings provide new insights into genes and biological pathways controlling myocardial mass and may help identify novel therapeutic targets

Trends in the incidence of dementia: design and methods in the Alzheimer Cohorts Consortium.

Several studies have reported a decline in incidence of dementia which may have large implications for the projected burden of disease, and provide important guidance to preventive efforts. However, reports are conflicting or inconclusive with regard to the impact of gender and education with underlying causes of a presumed declining trend remaining largely unidentified. The Alzheimer Cohorts Consortium aggregates data from nine international population-based cohorts to determine changes in the incidence of dementia since 1990. We will employ Poisson regression models to calculate incidence rates in each cohort and Cox proportional hazard regression to compare 5-year cumulative hazards across study-specific epochs. Finally, we will meta-analyse changes per decade across cohorts, and repeat all analysis stratified by sex, education and APOE genotype. In all cohorts combined, there are data on almost 69,000 people at risk of dementia with the range of follow-up years between 2 and 27. The average age at baseline is similar across cohorts ranging between 72 and 77. Uniting a wide range of disease-specific and methodological expertise in research teams, the first analyses within the Alzheimer Cohorts Consortium are underway to tackle outstanding challenges in the assessment of time-trends in dementia occurrence

The genetics of blood pressure regulation and its target organs from association studies in 342,415 individuals

To dissect the genetic architecture of blood pressure and assess effects on target-organ damage, we analyzed 128,272 SNPs from targeted and genome-wide arrays in 201,529 individuals of European ancestry and genotypes from an additional 140,886 individuals were used for validation. We identified 66 blood pressure loci, of which 17 were novel and 15 harbored multiple distinct association signals. The 66 index SNPs were enriched for cis-regulatory elements, particularly in vascular endothelial cells, consistent with a primary role in blood pressure control through modulation of vascular tone across multiple tissues. The 66 index SNPs combined in a risk score showed comparable effects in 64,421 individuals of non-European descent. The 66-SNP blood pressure risk score was significantly associated with target-organ damage in multiple tissues, with minor effects in the kidney. Our findings expand current knowledge of blood pressure pathways and highlight tissues beyond the classic renal system in blood pressure regulation

Statistical methods in microarrays and high-throughput flow cytometry

Abstract-I Background: Heterogeneous cell populations have previously been described as noisy. However, recent studies have demonstrated that heterogeneity can be biologically significant. We present here an approach for rapid and complete identification of heterogeneous cell populations from high-throughput flow cytometry data. We have developed a novel measure Slope Differentiation Identification (SDI) using flow cytometry-based protein expression, quantifying the rate of change in protein expression between two conditions (exponential and stationary phase) of yeast cells, as a function of cell size or cell granularity. Results: SDI had superior Gene Ontology enrichment when compared with other approaches such as k-means clustering and an approach based on the bi-modality of the fluorescence intensity distribution. Cell populations were also validated using gradient-separation followed by microscopy, where proteins with high SDI measure showed significant levels of differentiation between high and low density cells. Conclusion: Overall, our approach has identified novel protein expression patterns that differentiate quiescent and non-quiescent cell populations. Abstract-II Background: With the advent of genomics, there has been a rapid increase in the use of two and onecolor microarrays, used to measure mRNA abundance for the entire genome. Variability in microarray analysis undermines its utility in identifying the entire subset of differentially expressed mRNAs. Recent microarray studies have shown that, although it is assumed that variances are constant for every hybridized spot within a microarray, variances may differ for each biological sample analyzed (Ritchie, Diyagama et al. 2006). Another common assumption is that log-intensity values for any given gene have a Normal distribution. For many datasets, both assumptions have been shown to be incorrect, resulting in distortions in the significance when testing for differential expression of each gene (Bar-Even, Paulsson et al. 2006; Wentzell, Karakach et al. 2006). Approach: To overcome the limitations of existing approaches in identifying significant, differentially expressed genes, we have developed a novel unsupervised statistical approach called Calibration Regression Analysis of Microarrays (CRAM) that uses a combination of empirical Bayes and regression calibration. The main novelty of our approach is the modeling of gene expression variances as a function of the log-intensity within each sample. Another version was later developed CRAM-GS in which the association between genes is captured using an adjusted gene correlation measure. Results: CRAM was compared to four existing approaches for identifying differentially expressed genes. Performance was based on the ability to identify co-regulated genes in the same Gene Ontology process. CRAM exhibited a marginal improvement in GO process enrichment compared with the other approaches. To the original datasets, three more were included in which the later version CRAM-GS, showed a significant improvement compared to CRAM, suggesting a major additional benefit of incorporating gene correlations into the model. All versions of CRAM were two orders of magnitude faster than the existing approaches. Overall, CRAM provides an adaptive, computationally efficient approach for accurate identification of differentially expressed genes.National Science Foundation (NSF) grant MCB-0645854 and National Institutes of Health (NIH) grant GM-67593 (to M.W.W) and NIH grant 1U54MH084690-01 (to L.S). P.H.T was supported by NIH/IMSD grant GM-060201. R.J was supported by NIH grant GM-075149.BiologyDoctoralUniversity of New Mexico. Biology Dept.Werner-Washburne, MaggieToolson, EricNatvig, DonaldWearing, Hele

Association of Serum Klotho with Loss of Bone Mineral Density and Fracture Risk in Older Adults

ObjectivesKlotho deficiency has been previously linked to aging-like phenotypes such as osteoporosis, cognitive impairment, and sarcopenia. Low serum klotho was shown to be related to grip strength and disability. Nonetheless, no previous study has explored the association between serum klotho and fractures. The purpose of this report is to examine the relationship of serum klotho with bone mineral density (BMD) loss and fractures in older adults.DesignThe Health, Aging, and Body Composition (Health ABC) Study is a longitudinal cohort study of 3,075 community-dwelling older adults.SettingUS clinical centers.ParticipantsTwo thousand seven hundred and seventy six well-functioning black and white adults aged 70 to 79 years with serum klotho measurements were followed up for a median of 5 years.MeasurementsPercent annualized BMD change and fracture risk were compared across klotho quartiles. A Poisson distribution was used to calculate age-adjusted fracture incidence rates, and Cox proportional hazards models for multivariable-adjusted hazard ratios.ResultsThe annualized percent changes in hip, femoral neck, and vertebral BMD were similar across klotho quartiles. Participants experienced 507 nonspine fractures, 203 hip fractures, and 135 vertebral fractures. The Incidence rate (IR) of nonspine fractures was 17 per 1,000 person-years. The most frequent site was hip (IR = 6 per 1,000 person-years) and the IR of vertebral fractures was 3 per 1,000 person-years. There was no association between the lowest quartile of plasma klotho and nonspine (hazard ratio (HR) = 1.19, 95% confidence interval (CI) = 0.86-1.65), hip (HR = 1.34, 95% CI = 0.79-2.27), or vertebral fractures (HR = 1.17, 95% CI = 0.65-2.11).ConclusionAlthough klotho gene is a susceptible gene for reduced BMD, klotho blood concentration does not appear to be a predictor of bone loss or fracture risk in well-functioning older adults

N-Terminal pro-Brain Natriuretic Peptide and Associations With Brain Magnetic Resonance Imaging (MRI) Features in Middle Age: The CARDIA Brain MRI Study

ObjectiveAs part of research on the heart–brain axis, we investigated the association of N-terminal pro-brain natriuretic peptide (NT-proBNP) with brain structure and function in a community-based cohort of middle-aged adults from the Brain Magnetic Resonance Imaging sub-study of the Coronary Artery Risk Development in Young Adults (CARDIA) Study.Approach and resultsIn a cohort of 634 community-dwelling adults with a mean (range) age of 50.4 (46–52) years, we examined the cross-sectional association of NT-proBNP to total, gray (GM) and white matter (WM) volumes, abnormal WM load and WM integrity, and to cognitive function tests [the Digit Symbol Substitution Test (DSST), the Stroop test, and the Rey Auditory–Verbal Learning Test]. These associations were examined using linear regression models adjusted for demographic and cardiovascular risk factors and cardiac output. Higher NT-proBNP concentration was significantly associated with smaller GM volume (β = −3.44; 95% CI = −5.32, −0.53; p = 0.003), even after additionally adjusting for cardiac output (β = −2.93; 95% CI = −5.32, −0.53; p = 0.017). Higher NT-proBNP levels were also associated with lower DSST scores. NT-proBNP was not related to WM volume, WM integrity, or abnormal WM load.ConclusionIn this middle-aged cohort, subclinical levels of NT-proBNP were related to brain function and specifically to GM and not WM measures, extending similar findings in older cohorts. Further research is warranted into biomarkers of cardiac dysfunction as a target for early markers of a brain at risk