Senescence detection using reflected light.

Senescence is an important cellular program occurring in development, tissue repair, cancer, and aging. Increased senescence is also associated with disease states, including obesity and Type 2 diabetes (T2D). Characterizing and quantifying senescent cells at a single cell level has been challenging and particularly difficult in large primary cells, such as human adipocytes.

White adipocyte dysfunction and obesity-associated pathologies in humans.

The prevalence of obesity and associated chronic diseases continues to increase worldwide, negatively impacting on societies and economies. Whereas the association between excess body weight and increased risk for developing a multitude of diseases is well established, the initiating mechanisms by which weight gain impairs our metabolic health remain surprisingly contested. In order to better address the myriad of disease states associated with obesity, it is essential to understand adipose tissue dysfunction and develop strategies for reinforcing adipocyte health.

A DNA methylation atlas of normal human cell types.

DNA methylation is a fundamental epigenetic mark that governs gene expression and chromatin organization, thus providing a window into cellular identity and developmental processes. Current datasets typically include only a fraction of methylation sites and are often based either on cell lines that underwent massive changes in culture or on tissues containing unspecified mixtures of cells. Here we describe a human methylome atlas, based on deep whole-genome bisulfite sequencing, allowing fragment-level analysis across thousands of unique markers for 39 cell types sorted from 205 healthy tissue samples.

The regulation of adipocyte growth in white adipose tissue.

Adipocytes can increase in volume up to a thousand-fold, storing excess calories as triacylglycerol in large lipid droplets. The dramatic morphological changes required of adipocytes demands extensive cytoskeletal remodeling, including lipid droplet and plasma membrane expansion. Cell growth-related signalling pathways are activated, stimulating the production of sufficient amino acids, functional lipids and nucleotides to meet the increasing cellular needs of lipid storage, metabolic activity and adipokine secretion.

Elevated brain-derived cell-free DNA among patients with first psychotic episode - a proof-of-concept study.

Schizophrenia is a common, severe, and debilitating psychiatric disorder. Despite extensive research there is as yet no biological marker that can aid in its diagnosis and course prediction. This precludes early detection and intervention.

Liquid biopsy reveals collateral tissue damage in cancer.

Cancer inflicts damage to surrounding normal tissues, which can culminate in fatal organ failure. Here, we demonstrate that cell death in organs affected by cancer can be detected by tissue-specific methylation patterns of circulating cell-free DNA (cfDNA). We detected elevated levels of hepatocyte-derived cfDNA in the plasma of patients with liver metastases originating from different primary tumors, compared with cancer patients without liver metastases.

Obesity and hyperinsulinemia drive adipocytes to activate a cell cycle program and senesce.

Obesity is considered an important factor for many chronic diseases, including diabetes, cardiovascular disease and cancer. The expansion of adipose tissue in obesity is due to an increase in both adipocyte progenitor differentiation and mature adipocyte cell size. Adipocytes, however, are thought to be unable to divide or enter the cell cycle.

Cellular senescence and its role in white adipose tissue.

Cell senescence is defined as a state of irreversible cell cycle arrest combined with DNA damage and the induction of a senescence-associated secretory phenotype (SASP). This includes increased secretion of many inflammatory agents, proteases, miRNA's, and others. Cell senescence has been widely studied in oncogenesis and has generally been considered to be protective, due to cell cycle arrest and the inhibition of proliferation.

Practical Guidelines for Optimization and Characterization of the Beckman Coulter CytoFLEX™ Platform.

Cytometer characterization is critical to define operational bounds within which the data generated are reliable and reproducible. Existing instrument optimization and characterization protocols were developed for cytometers relying on photomultiplier tubes (PMTs) for photon detection. Recently, instrument manufacturers have begun incorporating avalanche photodiodes (APDs) in place of PMTs.

Mature Human White Adipocytes Cultured under Membranes Maintain Identity, Function, and Can Transdifferentiate into Brown-like Adipocytes.

White adipose tissue (WAT) is a central factor in the development of type 2 diabetes, but there is a paucity of translational models to study mature adipocytes. We describe a method for the culture of mature white adipocytes under a permeable membrane. Compared to existing culture methods, MAAC (membrane mature adipocyte aggregate cultures) better maintain adipogenic gene expression, do not dedifferentiate, display reduced hypoxia, and remain functional after long-term culture.

Comprehensive human cell-type methylation atlas reveals origins of circulating cell-free DNA in health and disease.

Methylation patterns of circulating cell-free DNA (cfDNA) contain rich information about recent cell death events in the body. Here, we present an approach for unbiased determination of the tissue origins of cfDNA, using a reference methylation atlas of 25 human tissues and cell types. The method is validated using in silico simulations as well as in vitro mixes of DNA from different tissue sources at known proportions.

Transforming Growth Factor-β3 Regulates Adipocyte Number in Subcutaneous White Adipose Tissue.

White adipose tissue (WAT) mass is determined by adipocyte size and number. While adipocytes are continuously turned over, the mechanisms controlling fat cell number in WAT upon weight changes are unclear. Herein, prospective studies of human subcutaneous WAT demonstrate that weight gain increases both adipocyte size and number, but the latter remains unaltered after weight loss.

Flow Cytometry of Mouse and Human Adipocytes for the Analysis of Browning and Cellular Heterogeneity.

Adipocytes, once considered simple lipid-storing cells, are rapidly emerging as complex cells with many biologically diverse functions. A powerful high-throughput method for analyzing single cells is flow cytometry. Several groups have attempted to analyze and sort freshly isolated adipocytes; however, using an adipocyte-specific reporter mouse, we demonstrate that these studies fail to detect the majority of white adipocytes.

Impact of fat mass and distribution on lipid turnover in human adipose tissue.

Differences in white adipose tissue (WAT) lipid turnover between the visceral (vWAT) and subcutaneous (sWAT) depots may cause metabolic complications in obesity. Here we compare triglyceride age and, thereby, triglyceride turnover in vWAT and sWAT biopsies from 346 individuals and find that subcutaneous triglyceride age and storage capacity are increased in overweight or obese individuals. Visceral triglyceride age is only increased in excessively obese individuals and associated with a lower lipid removal capacity.

Transplanted Bone Marrow-Derived Cells Contribute to Human Adipogenesis.

Because human white adipocytes display a high turnover throughout adulthood, a continuous supply of precursor cells is required to maintain adipogenesis. Bone marrow (BM)-derived progenitor cells may contribute to mammalian adipogenesis; however, results in animal models are conflicting. Here we demonstrate in 65 subjects who underwent allogeneic BM or peripheral blood stem cell (PBSC) transplantation that, over the entire lifespan, BM/PBSC-derived progenitor cells contribute ∼10% to the subcutaneous adipocyte population.

Adult Neurogenesis in Humans.

Adult neurogenesis appears very well conserved among mammals. It was, however, not until recently that quantitative data on the extent of this process became available in humans, largely because of methodological challenges to study this process in man. There is substantial hippocampal neurogenesis in adult humans, but humans appear unique among mammals in that there is no detectable olfactory bulb neurogenesis but continuous addition of new neurons in the striatum.

Adrenergically stimulated blood flow in brown adipose tissue is not dependent on thermogenesis.

Brown adipose tissue (BAT) thermogenesis relies on blood flow to be supplied with nutrients and oxygen and for the distribution of the generated heat to the rest of the body. Therefore, it is fundamental to understand the mechanisms by which blood flow is regulated and its relation to thermogenesis. Here, we present high-resolution laser-Doppler imaging (HR-LDR) as a novel method for noninvasive in vivo measurement of BAT blood flow in mice.

Maintenance of white adipose tissue in man.

Obesity is increasing in an epidemic manner in most countries and constitutes a public health problem by enhancing the risk for diseases such as diabetes, fatty liver disease and atherosclerosis. Together these diseases form a cluster referred to as the metabolic syndrome. Despite the negative health consequences associated with excess adipose tissue, very little is known about the origin and maintenance of white adipose tissue in man.

Analysis of radiocarbon, stable isotopes and DNA in teeth to facilitate identification of unknown decedents.

The characterization of unidentified bodies or suspected human remains is a frequent and important task for forensic investigators. However, any identification method requires clues to the person's identity to allow for comparisons with missing persons. If such clues are lacking, information about the year of birth, sex and geographic origin of the victim, is particularly helpful to aid in the identification casework and limit the search for possible matches.

Dynamics of hippocampal neurogenesis in adult humans.

Adult-born hippocampal neurons are important for cognitive plasticity in rodents. There is evidence for hippocampal neurogenesis in adult humans, although whether its extent is sufficient to have functional significance has been questioned. We have assessed the generation of hippocampal cells in humans by measuring the concentration of nuclear-bomb-test-derived ¹⁴C in genomic DNA, and we present an integrated model of the cell turnover dynamics.

Personal identification of cold case remains through combined contribution from anthropological, mtDNA, and bomb-pulse dating analyses.

In 1968, a child's cranium was recovered from the banks of a northern Canadian river and held in a trust until the "cold case" was reopened in 2005. The cranium underwent reanalysis at the Centre for Forensic Research, Simon Fraser University, using recently developed anthropological analysis, "bomb-pulse" radiocarbon analysis, and forensic DNA techniques. Craniometrics, skeletal ossification, and dental formation indicated an age-at-death of 4.

The age of olfactory bulb neurons in humans.

Continuous turnover of neurons in the olfactory bulb is implicated in several key aspects of olfaction. There is a dramatic decline postnatally in the number of migratory neuroblasts en route to the olfactory bulb in humans, and it has been unclear to what extent the small number of neuroblasts at later stages contributes new neurons to the olfactory bulb. We have assessed the age of olfactory bulb neurons in humans by measuring the levels of nuclear bomb test-derived (14)C in genomic DNA.

Dynamics of human adipose lipid turnover in health and metabolic disease.

Adipose tissue mass is determined by the storage and removal of triglycerides in adipocytes. Little is known, however, about adipose lipid turnover in humans in health and pathology. To study this in vivo, here we determined lipid age by measuring (14)C derived from above ground nuclear bomb tests in adipocyte lipids.