Bcl3 Couples Cancer Stem Cell Enrichment With Pancreatic Cancer Molecular Subtypes

[Background & Aims]: The existence of different subtypes of pancreatic ductal adenocarcinoma (PDAC) and their correlation with patient outcome have shifted the emphasis on patient classification for better decision-making algorithms and personalized therapy. The contribution of mechanisms regulating the cancer stem cell (CSC) population in different subtypes remains unknown. [Methods]: Using RNA-seq, we identified B-cell CLL/lymphoma 3 (BCL3), an atypical nf-κb signaling member, as differing in pancreatic CSCs. To determine the biological consequences of BCL3 silencing in vivo and in vitro, we generated bcl3-deficient preclinical mouse models as well as murine cell lines and correlated our findings with human cell lines, PDX models, and 2 independent patient cohorts. We assessed the correlation of bcl3 expression pattern with clinical parameters and subtypes. [Results]: Bcl3 was significantly down-regulated in human CSCs. Recapitulating this phenotype in preclinical mouse models of PDAC via BCL3 genetic knockout enhanced tumor burden, metastasis, epithelial to mesenchymal transition, and reduced overall survival. Fluorescence-activated cell sorting analyses, together with oxygen consumption, sphere formation, and tumorigenicity assays, all indicated that BCL3 loss resulted in CSC compartment expansion promoting cellular dedifferentiation. Overexpression of BCL3 in human PDXs diminished tumor growth by significantly reducing the CSC population and promoting differentiation. Human PDACs with low BCL3 expression correlated with increased metastasis, and BCL3-negative tumors correlated with lower survival and nonclassical subtypes. [Conclusions]: We demonstrate that bcl3 impacts pancreatic carcinogenesis by restraining CSC expansion and by curtailing an aggressive and metastatic tumor burden in PDAC across species. Levels of BCL3 expression are a useful stratification marker for predicting subtype characterization in PDAC, thereby allowing for personalized therapeutic approaches.This work was supported by the Deutsche Forschungsgemeinschaft (grants AL 1174/4-1, AL1174/4-2, and Collaborative Research Center 1321 “Modeling and Targeting Pancreatic Cancer” to Hana Algül; SFB824 Z2 to Katja Steiger), the Deutsche Krebshilfe (grant 111646 to Hana Algül), a Ramon y Cajal Merit Award from the Ministerio de Economía y Competitividad, Spain (to Bruno Sainz Jr), a Coordinated Grant from Fundación Asociación Española Contra el Cáncer (GC16173694BARB to Bruno Sainz Jr), funding from The Fero Foundation (to Bruno Sainz Jr), and a Proyecto de Investigacion de Salud, ISCIII, Spain (no. PI18/00757 to Bruno Sainz Jr). Jiaoyu Ai is supported by the “China Scholarship Council” grant program

A Roadmap for HEP Software and Computing R&D for the 2020s

Particle physics has an ambitious and broad experimental programme for the coming decades. This programme requires large investments in detector hardware, either to build new facilities and experiments, or to upgrade existing ones. Similarly, it requires commensurate investment in the R&D of software to acquire, manage, process, and analyse the shear amounts of data to be recorded. In planning for the HL-LHC in particular, it is critical that all of the collaborating stakeholders agree on the software goals and priorities, and that the efforts complement each other. In this spirit, this white paper describes the R&D activities required to prepare for this software upgrade.Peer reviewe

Hyperphagia of female UCP1-deficient mice blunts anti-obesity effects of FGF21

Abstract Increasing energy expenditure through uncoupling protein 1 (UCP1) activity in thermogenic adipose tissue is widely investigated to correct diet-induced obesity (DIO). Paradoxically, UCP1-deficient male mice are resistant to DIO at room temperature. Recently, we uncovered a key role for fibroblast growth factor 21 (FGF21), a promising drug target for treatment of metabolic disease, in this phenomenon. As the metabolic action of FGF21 is so far understudied in females, we aim to investigate potential sexual dimorphisms. Here, we confirm that male UCP1 KO mice display resistance to DIO in mild cold, without significant changes in metabolic parameters. Surprisingly, females gained the same amount of body fat as WT controls. Molecular regulation was similar between UCP1 KO males and females, with an upregulation of serum FGF21, coinciding with beiging of inguinal white adipose tissue and induced lipid metabolism. While energy expenditure did not display significant differences, UCP1 KO females significantly increased their food intake. Altogether, our results indicate that hyperphagia is likely counteracting the beneficial effects of FGF21 in female mice. This underlines the importance of sex-specific studies in (pre)clinical research for personalized drug development

Genetic disruption of uncoupling protein 1 in mice renders brown adipose tissue a significant source of FGF21 secretion

Objective: Circulating fibroblast growth factor 21 (FGF21) is an important auto- and endocrine player with beneficial metabolic effects on obesity and diabetes. In humans, thermogenic brown adipose tissue (BAT) was recently suggested as a source of FGF21 secretion during cold exposure. Here, we aim to clarify the role of UCP1 and ambient temperature in the regulation of FGF21 in mice. Methods: Wildtype (WT) and UCP1-knockout (UCP1 KO) mice, the latter being devoid of BAT-derived non-shivering thermogenesis, were exposed to different housing temperatures. Plasma metabolites and FGF21 levels were determined, gene expression was analyzed by qPCR, and tissue histology was performed with adipose tissue. Results: At thermoneutrality, FGF21 gene expression and serum levels were not different between WT and UCP1 KO mice. Cold exposure led to highly increased FGF21 serum levels in UCP1 KO mice, which were reflected in increased FGF21 gene expression in adipose tissues but not in liver and skeletal muscle. Ex vivo secretion assays revealed FGF21 release only from BAT, progressively increasing with decreasing ambient temperatures. In association with increased FGF21 serum levels in the UCP1 KO mouse, typical FGF21-related serum metabolites and inguinal white adipose tissue morphology and thermogenic gene expression were altered. Conclusions: Here we show that the genetic ablation of UCP1 increases FGF21 gene expression in adipose tissue. The removal of adaptive nonshivering thermogenesis renders BAT a significant source of endogenous FGF21 under thermal stress. Thus, the thermogenic competence of BAT is not a requirement for FGF21 secretion. Notably, high endogenous FGF21 levels in UCP1-deficient models and subjects may confound pharmacological FGF21 treatments

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

Summary: 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. We define critical settings required for adipocyte flow cytometry and provide a rigid strategy for analyzing and sorting white and brown adipocyte populations. The applicability of our protocol is shown by sorting mouse adipocytes based on size or UCP1 expression and demonstrating that a subset of human adipocytes lacks the β2-adrenergic receptor, particularly in the insulin-resistant state. In conclusion, the present study confers key technological insights for analyzing and sorting mature adipocytes, opening up numerous downstream research applications. : Freshly isolated adipocytes are a notoriously difficult cell type to study. Hagberg et al. provide a detailed flow cytometry protocol for the analysis and sorting of mouse and human adipocytes by defining the critical factors and conditions required for studying this specialized cell type and pinpointing common pitfalls. Keywords: adipocyte, adipose tissue, flow cytometry, FACS, mouse, human, uncoupled protein 1, beta 2 adrenergic recepto

Long-Term Cold Adaptation Does Not Require FGF21 or UCP1

Brown adipose tissue (BAT)-dependent thermogenesis and its suggested augmenting hormone, FGF21, are potential therapeutic targets in current obesity and diabetes research. Here, we studied the role of UCP1 and FGF21 for metabolic homeostasis in the cold and dissected underlying molecular mechanisms using UCP1-FGF21 double-knockout mice. We report that neither UCP1 nor FGF21, nor even compensatory increases of FGF21 serum levels in UCP1 knockout mice, are required for defense of body temperature or for maintenance of energy metabolism and body weight. Remarkably, cold-induced browning of inguinal white adipose tissue (iWAT) is FGF21 independent. Global RNA sequencing reveals major changes in response to UCP1- but not FGF21-ablation in BAT, iWAT, and muscle. Markers of mitochondrial failure and inflammation are observed in BAT, but in particular the enhanced metabolic reprogramming in iWAT supports the thermogenic role of UCP1 and excludes an important thermogenic role of endogenous FGF21 in normal cold acclimation

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

International audienceObesity 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. We demonstrate that mature human adipocytes unexpectedly display a gene and protein signature indicative of an active cell cycle program. Adipocyte cell cycle progression associates with obesity and hyperinsulinemia, with a concomitant increase in cell size, nuclear size and nuclear DNA content. Chronic hyperinsulinemia in vitro or in humans, however, is associated with subsequent cell cycle exit, leading to a premature senescent transcriptomic and secretory profile in adipocytes. Premature senescence is rapidly becoming recognized as an important mediator of stress-induced tissue dysfunction. By demonstrating that adipocytes can activate a cell cycle program, we define a mechanism whereby mature human adipocytes senesce. We further show that by targeting the adipocyte cell cycle program using metformin, it is possible to influence adipocyte senescence and obesity-associated adipose tissue inflammation

Two-stage evolution of mammalian adipose tissue thermogenesis

Brown adipose tissue (BAT) is a heater organ that expresses thermogenic uncoupling protein 1 (UCP1) to maintain high body temperatures during cold stress. BAT thermogenesis is considered an overarching mammalian trait, but its evolutionary origin is unknown. We show that adipose tissue of marsupials, which diverged from eutherian mammals ~150 million years ago, expresses a nonthermogenic UCP1 variant governed by a partial transcriptomic BAT signature similar to that found in eutherian beige adipose tissue. We found that the reconstructed UCP1 sequence of the common eutherian ancestor displayed typical thermogenic activity, whereas therian ancestor UCP1 is nonthermogenic. Thus, mammalian adipose tissue thermogenesis may have evolved in two distinct stages, with a prethermogenic stage in the common therian ancestor linking UCP1 expression to adipose tissue and thermal stress. We propose that in a second stage, UCP1 acquired its thermogenic function specifically in eutherians, such that the onset of mammalian BAT thermogenesis occurred only after the divergence from marsupials