Activin A Plays a Critical Role in Proliferation and Differentiation of Human Adipose Progenitors

International audienceAbstractObjective: Growth of white adipose tissue takes place in normal development and in obesity. A pool of adipose progenitors is responsible for the formation of new adipocytes and for the potential of this tissue to expand in response to chronic energy overload. However, factors controlling self-renewal of human adipose progenitors are largely unknown. We investigated the expression profile and the role of activin A in this process. Research Design and Methods: Expression of INHBA/activin A has been investigated in three types of human adipose progenitors. We then analyzed at the molecular level the function of activin A during human adipogenesis. We finally investigated the status of activin A in adipose tissues of lean and obese subjects and analyzed macrophage-induced regulation of its expression. Results: INHBA/activin A is expressed by adipose progenitors from various fat depots and its expression dramatically decreases as progenitors differentiate into adipocytes. Activin A regulates the number of undifferentiated progenitors. Sustained activation or inhibition of the activin A pathway impairs or promotes respectively adipocyte differentiation via C/EBPbeta-LAP and Smad2 pathway in an autocrine/paracrine manner. Activin A is expressed at higher levels in adipose tissue of obese patients compared to lean subjects. Indeed, activin A levels in adipose progenitors are dramatically increased by factors secreted by macrophages derived from obese adipose tissue. Conclusions: Altogether, our data show that activin A plays a significant role in human adipogenesis. We propose a model in which macrophages which are located in adipose tissue regulate adipose progenitor self-renewal through activin A

Cellules souches induites à la pluripotence (modèle d'étude des étapes précoces du développement adipocytaire humain)

Si, aujourd hui, les étapes les plus tardives de l adipogénèse humaine ont été bien caractérisées, les étapes précoces qui correspondent à la génération des précurseurs adipocytaires blancs et bruns restent encore à définir. Les cellules souches humaines induites à la pluripotence (hiPS) constituent un bon modèle d étude de l adipogénèse précoce. Le but de ce travail était de générer des cellules hiPS à partir des cellules souches multipotentes du tissu adipeux (hMADS), de les caractériser et de les utiliser comme un outil d étude de l apparition des précurseurs adipeux blancs et bruns in vitro. Nous avons également utilisé des cellules hiPS générées à partir des cellules souches neurales humaines (hNSC) en utilisant un vecteur PiggyBac. Nous avons mis au point un protocole de différenciation des cellules iPS-hMADS et iPS-hNSC en adipocytes. C une façon intéressante, notre protocole de différenciation permet l apparition d adipocytes blancs et bruns. Nous avons montré que l activation de la voie de l acide rétinoïque (RA) au cours des étapes précoces de différenciation augmente fortement la génération d adipocytes blancs, et diminue celle des adipocytes bruns. En revanche, l utilisation du SB431542, un inhibiteur sélectif de la voie TGFb/Activine, induit une augmentation de la génération es adipocytes blancs et une inhibition de l apparition des adipocytes bruns. L ensemble de nos résultats soutient un modèle dans lequel les progéniteurs adipeux blancs et bruns apparaissent précocement au cours du développement embryonnaire humain. Dans ce processus, la voie de l acide rétinoïque et la voie du TGFb/Activine régulent la génération des Pas blancs et bruns.The terminal steps of adipocyte differentiation are well established ; however the earliest steps controlling brown and white adipogenic lineage specification remain unknown in humans. We have investigated the human induced Pluripotent Stem cells (hips) as a model to study the early steps of brown and white adipogenesis. We will present the generation of hips cells from human multipotent adipose derived stem cells (hMADS). We have also used iPS cells reprogrammed from human neural stem cells generated using the PiggyBac technology. We provide an efficient protocol to differentiate iPS-hMADS and iPS-hNSC cells into adipocytes. Interestingly, our data show that hips cells are able to differentiate both into white and brown adipocytes. We show that Retinoic Acid (RA) pathway activation at an early phase of hips development dramatically enhanced generation of white adipocytes and inhibited generation of Brown adipocytes. In contrast, the use of SB431542, a selective inhibitor of TGFb/Activin pathway, indicated that this pathway was required for the generation of brown adipocytes. Altogether, these data support a model in which brown and white adipocytes progenitors diverge early during human embryonic development. RA and TGFb/Activin pathways regulate the generation of white APs and brown APs respectively.NICE-BU Sciences (060882101) / SudocSudocFranceF

Characterization of brown adipose tissue in the human perirenal depot.

International audienceTo characterize brown adipose tissue (BAT) in the human perirenal adipose tissue depot. Perirenal adipose tissue biopsies were obtained from 55 healthy kidney donors. Expression analysis was performed using microarray, real-time PCR, immunoblotting and immunohistochemistry. Additional studies using human stem cells were performed. UCP1 gene expression analysis revealed a large intra-individual variation in the perirenal adipose tissue biopsies. Both multi- and unilocular UCP1-positive adipocytes were detected in several of the adipose tissue samples analyzed by immunohistochemical staining. Microarray analysis identified 54 genes that were overexpressed in UCP1-positive perirenal adipose tissue. Real-time PCR analysis of BAT candidate genes revealed a set of genes that were highly correlated to UCP1 and a set of three transcription factor genes (PRDM16, PGC1α, and RXRγ) that were highly correlated to each other. RXRγ displayed nuclear immunoreactivity in brown adipocytes and an increased gene expression during brown adipogenesis in human stem cells. Our data provides the first molecular characterization of BAT in the perirenal adipose tissue depot. Furthermore, it highlights the transcription factor RXRγ as a new player in BAT development

Differentiation of human induced pluripotent stem cells into brown and white adipocytes: Role of Pax3.

International audience: Identification of molecular mechanisms involved in generation of different types of adipocytes is progressing substantially in mice. However, much less is known regarding characterization of brown and white adipocyte progenitors (APs) in humans, highlighting the need for an in vitro model of human adipocyte development. Here we report a procedure to selectively derive brown and white APs from human induced pluripotent stem cells. Molecular characterization of APs of both phenotypes revealed that BMP4, Hox8, Hoxc9 and HoxA5 genes were specifically expressed in white APs, whereas expression of PRDM16, Dio2 and Pax3 marked brown APs. We focused on Pax3 and we showed that expression of this transcription factor was enriched in human perirenal white adipose tissue samples expressing UCP1 and in human classical brown fat. Finally, functional experiments indicated that Pax3 was a critical player of human AP fate as its ectopic expression led to convert white APs into brown-like APs. Together, these data support a model in which Pax3 is a new marker of human brown APs and a molecular mediator of their fate. The findings of the present study could lead to new anti-obesity therapies based on the recruitment of APs and constitute a platform for investigating in vitro the developmental origins of human white and brown adipocytes. Stem Cells 2013

A Western-like fat diet is sufficient to induce a gradual enhancement in fat mass over generations[S]

The prevalence of obesity has steadily increased over the last few decades. During this time, populations of industrialized countries have been exposed to diets rich in fat with a high content of linoleic acid and a low content of α-linolenic acid compared with recommended intake. To assess the contribution of dietary fatty acids, male and female mice fed a high-fat diet (35% energy as fat, linoleic acid:α-linolenic acid ratio of 28) were mated randomly and maintained after breeding on the same diet for successive generations. Offspring showed, over four generations, a gradual enhancement in fat mass due to combined hyperplasia and hypertrophy with no change in food intake. Transgenerational alterations in adipokine levels were accompanied by hyperinsulinemia. Gene expression analyses of the stromal vascular fraction of adipose tissue, over generations, revealed discrete and steady changes in certain important players, such as CSF3 and Nocturnin. Thus, under conditions of genome stability and with no change in the regimen over four generations, we show that a Western-like fat diet induces a gradual fat mass enhancement, in accordance with the increasing prevalence of obesity observed in humans