Tala Mohsen-Kanson

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

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.

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

OBJECTIVE 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 was 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 the C/EBPβ-LAP and Smad2 pathway in an autocrine/paracrine manner. Activin A is expressed at higher levels in adipose tissue of obese patients compared with the expression levels in 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 that are located in adipose tissue regulate adipose progenitor self-renewal through activin A.

Differentiation of Human Induced Pluripotent Stem Cells into Brown and White Adipocytes: Role of Pax3

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 (BAP) and white adipocyte progenitors (WAPs) in humans, highlighting the need for an in vitro model of human adipocyte development. Here, we report a procedure to selectively derive BAP and WAPs 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 WAPs, whereas expression of PRDM16, Dio2, and Pax3 marked BAPs. 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 WAPs into brown‐like APs. Together, these data support a model in which Pax3 is a new marker of human BAPs and a molecular mediator of their fate. The findings of this 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 2014;32:1459–1467

Characterization of brown adipose tissue in the human perirenal depot.

To characterize brown adipose tissue (BAT) in the human perirenal adipose tissue depot.

Tumor microenvironment modifications induced by soluble VEGF receptor expression in a rat liver metastasis model

Vascular endothelial growth factor is a potent pro-angiogenic growth factor which is also known to alter tumor microenvironment by inhibiting dendritic cell differentiation and promoting accumulation of myeloid-derived suppressor cells. In the present study, we analyzed the modifications induced by intratumoral expression of sFLT-1, a soluble VEGF receptor, in a rat metastatic colon carcinoma model. We generated colon cancer cell lines stably expressing sFLT-1 or a mock construct. Human umbilical vein endothelial cells cultured with conditioned medium from sFLT-1-expressing tumor cells exhibit a significantly decreased survival, demonstrating the functionality of the secreted sFLT-1. Invivo, sFLT-1 expression induced a 30% decrease in microvessel density in 15-day old experimental liver metastasis from colon carcinoma. Tumor growth was inhibited by 63% and 52% in left and right liver lobes respectively within 25days. In these tumors, sFLT-1 expression was associated with a decreased myeloid cell infiltration and a modification in the expression of several cytokines/chemokines. Altogether, these results suggest that VEGF trapping by sFLT-1 intratumoral expression results in reduced vascularization, tumor growth inhibition and modification of immune tumor microenvironment.

Expression of cell surface markers during self-renewal and differentiation of human adipose-derived stem cells

Human adipose-derived stem cell populations express cell surface markers such as CD105, CD73, CD146 and CD140a/PDFGRα. However, it was unclear whether these markers could discriminate subpopulations of undifferentiated cells and whether the expression of these markers is modulated during differentiation. To address this issue, we analysed the immunophenotype of cultured human multipotent adipose derived stem (hMADS) cell populations at different adipocyte differentiation steps. We found that 100% of undifferentiated cells expressed CD73 and CD105. In contrast, CD146 and CD140a/PDFGRα marked two different subpopulations of cells. CD140a/PDGFRα subpopulation was regulated by FGF2, a critical factor of human adipose-derived stem cell self-renewal. During differentiation, CD73 was maintained and marked lipid-laden cells, whereas CD105 expression was inhibited in fully differentiated cells. The percentage of CD146 and CD140a/PDFGRα-positive cells declined as soon as cells had undergone differentiation. Altogether, these data support the notion that expanded adipose-derived stem cells are heterogeneous mixtures of cells and cell surface markers studied can discriminate subpopulations.

Development of Adipose Cells

The current epidemic of obesity and overweight has caused a surge of interest in the study of adipose tissue formation. Much progress has been made in defining the transcriptional networks controlling the terminal differentiation of preadipocytes into mature adipocytes. However, the earliest steps that direct mesenchymal stem cells located in adipose tissue (adipose-derived stem cells (ASCs)) down the adipocyte lineage remain largely unknown. In this chapter, we first focus on the identification of intrinsic and extrinsic factors regulating thebalance between human ASC proliferation and differentiation. We point out the role of factors secreted by undifferentiated ASCs, such a FGF2, activin A, Hedgehog molecules, or secreted by adipose tissue macrophages. We also outline the role of miRNAs in these processes. In a second part, the developmental origin of adipocytes during embryogenesis is discussed. We summarize the surprising findings that have recently emerged from in vivo lineage tracing studies, unravelling unsuspected developmental origins for white adipocytes. We examine the possibility that the differential developmental origin of adipocytes could also reflect functional site-specific regulations of adipose tissue. Finally, we present human-induced pluripotent stem cells as a novel model for investigating the earliest steps of human adipocyte development.

The generation and the manipulation of human multipotent adipose-derived stem cells.

In this chapter, we describe a method to isolate and to expand multipotent adipose-derived stem (hMADS) cells from human adipose tissue. We also describe culture conditions to differentiate them into adipocytes at a high rate. This culture system provides a powerful means for studying the first steps of human adipose cell development and a route for investigating effects of drugs on the biology of adipocytes. Finally, we provide a protocol to investigate gene function during proliferation and differentiation of hMADS cells by means of siRNA-mediated gene silencing approaches or forced expression by transducing hMADS cells permissive to infection with murine retrovirus vectors.

Differentiation of Brown Adipocyte Progenitors Derived from Human Induced Pluripotent Stem Cells

In this chapter, we describe a protocol to induce at a high rate the differentiation of brown/brown like adipocyte progenitors (BAPs) derived from human induced pluripotent stem cells (hiPSCs). We also describe culture conditions to maintain hiPSCs and to derive hiPSC-BAPs.This novel culture system provides an unlimited source of human brown adipocytes and a unique means for studying events regulating the generation and recruitment of human BAPs.