Mammalian adipogenesis regulator (Areg) cells use retinoic acid signalling to be non- and anti-adipogenic in age-dependent manner

Adipose stem and precursor cells (ASPCs) give rise to adipocytes and determine the composition and plasticity of adipose tissue. Recently, several studies have demonstrated that ASPCs partition into at least three distinct cell subpopulations, including the enigmatic CD142(+) cells. An outstanding challenge is to functionally characterise this population, as discrepant properties, from adipogenic to non- and anti-adipogenic, have been reported for these cells. To resolve these phenotypic ambiguities, we characterised mammalian subcutaneous CD142(+) ASPCs across various experimental conditions, demonstrating that CD142(+) ASPCs exhibit high molecular and phenotypic robustness. Specifically, we find these cells to be firmly non- and anti-adipogenic both in vitro and in vivo, with their inhibitory signals also impacting adipogenic human cells. However, these CD142(+) ASPC-specific properties exhibit surprising temporal phenotypic alterations, and emerge only in an age-dependent manner. Finally, using multi-omic and functional assays, we show that the inhibitory nature of these adipogenesis-regulatory CD142(+) ASPCs (Aregs) is driven by specifically expressed secretory factors that cooperate with the retinoic acid signalling pathway to transform the adipogenic state of CD142(-) ASPCs into a non-adipogenic, Areg-like state.

The role and transcriptional regulation of FAM46A in adipocyte differentiation

Ann-Kristin Hov

Obesity is currently a leading global health concern. Treatments include undergoing a weight-loss regimen. However, successful weight-loss is variable, where the heritability of obesity is considered part of the cause. This patient variability to weight-loss was addressed by Carayol et al. (2017), where protein plasma level changes was associated to genetic variability and changes in BMI under a low-calorie diet intervention. A highly significant association region was found to regulate downstream gene FAM46A; coding for a nucleotidyl-transferase protein. Loss of function mutations in FAM46A have previously been discovered in multiple human diseases, such as osteogenesis imperfecta. However, there is a lack of understanding of FAM46Aâs function, especially in the context of adipose tissue and metabolism. This thesis aimed to follow up on the Carayol et al. study by investigating the transcriptional regulation of FAM46A and how the associated region is influencing this regulation, as well as how FAM46A is involved in adipose tissue function. When mapping the physical DNA interaction network around the associated region using chromosome conformation capture (3C) in human subcutaneous adipocytes (SGBS cells), no interactions between the FAM46A promoter and selected loci of the associated region were found. Interestingly, significant interaction peaks downstream of the promoter overlapped with regions of open chromatin and were enriched for transcription factor (TF) motifs that are involved in stem cell differentiation and TGFÎ² signalling. SMAD4, part of the TGFÎ² signalling pathway, binds to this region, indicating a potential regulator of FAM46A. To address FAM46Aâs potential role in adipocyte function, FAM46A expression was disrupted in vitro in differentiating adipocytes. Reduction of FAM46A expression in SGBS cells resulted in decreased expression of PPARG and CEBPA, the master regulators of adipogenesis, complemented by a reduction in lipid accumulation. Follow up studies in murine mesenchymal stem cells where Fam46a was over-expressed were controversial as this led to a decrease in adipogenic markers (Adipoq and PPARðŸ). Overall, disruption of FAM46A expression led to opposing changes in adipogenic potential, suggesting the requirement for a more stable cell system to study FAM46A in vitro. To further characterise the role of Fam46a, metabolic phenotyping of a Fam46a knockout (KO) mouse model under hyper-caloric stress was performed in collaboration with the IMG in Prague. This model revealed a significantly smaller mouse, and increased plasma alkaline phosphatase, as previously observed. When fed a high-fat diet, KO mice had reduced overall fat mass, improved glucose clearance and reduced adipocyte size in both subcutaneous and visceral white adipose tissue (subWAT/visWAT) depots. Transcriptomic analysis of these two depots revealed an up-regulation of genes expressed in ribosome related pathways in the subWAT, whereas the visWAT showed the up-regulation of fatty acid metabolism pathways, suggesting that the KO adipose tissue is more metabolically active. Other down-regulated pathways involved stem cell differentiation and BMP/TGFÎ²-signalling. In conclusion, this study found the possible regulation of the FAM46A promoter by TFs involved in adipogenesis regulation, such as SMAD proteins, as well as strengthening its possible functional involvement in TGFÎ² signalling by enrichment of this pathway in KO studies.

EPFL2021

Metabolic and microenvironmental strategies to accelerate hematopoietic recovery post-aplasia

Vasco Ledebur de Antas de Campos

The procedure-associated mortality rate of bone marrow transplantations (BMT) remains close to 25%. The BMT is therefore only indicated for life-threatening diseases, with no replacement of this technology to be expected in the foreseeable future. Modest improvements such as the use of G-CSF to accelerate hematopoietic recovery significantly reduce mortality. Hematopoietic stem cells (HSC) reside in the bone marrow (BM) and are mainly quiescent, dividing only to self-renew. Via extracellular signals and interactions with other cells of the BM (microenvironment), a stable HSC pool is maintained, while hematopoietic progenitor cells (HPC) proliferate and differentiate to mature blood cell types. Marrow adipocytes (BMA) have recently been recognized as a hematopoietic regulatory entity, although the mechanisms by which it interacts with HSCs and HPCs (HSPC) in vivo, remain unknown. During the hematopoietic recovery period post-BMT, HSPCs increase their metabolism as a response to hematopoietic stress, partly modulated by the microenvironment. In this work we identified novel pharmacological approaches to accelerate hematopoietic recovery in the post-BMT period, addressed via two strategies: (i) by directly modulating HSC metabolism, and (ii) by regulating the differentiation of BMA. Increasing oxidative phosphorylation, combined with an impaired mitochondrial stress response, can severely compromise the regenerative capacity of HSCs. Here we demonstrate that the NAD+-boosting agent Nicotinamide Riboside (NR) reduces mitochondrial activity within HSCs through increased mitophagy, the recycling mechanism of damaged mitochondria. We observed in vitro NR treatment of HSCs to lead to increased asymmetric HSC divisions, which resulted in a significantly enlarged pool of progenitor cells, without HSC exhaustion. Dietary supplementation of NR to mice undergoing BMT accelerated blood recovery and improved survival. We therefore established a novel link between HSC mitochondrial stress, mitophagy and stem cell fate decision. Recent studies have suggested BMA maintain HSCs quiescent, which, may be detrimental to the increased expansion of HPCs in the recovery phase post-BMT. BM stromal cells (BMSC) are the cellular precursors of both adipocytes and osteoblasts, and they have been strongly implicated in supporting hematopoiesis by secreting cytokines such as SCF and Cxcl12. We observed that adipocytes quickly accumulate the marrow space of mice undergoing BMT, followed by an expansion hematopoiesis-supportive multipotent Sca1+/CD24+ stromal cells, coinciding with hematopoietic recovery. We developed a novel screening platform using Digital Holographic Microscopy (DHM), quantifying in vitro adipocytic differentiation non-invasively, allowing for follow-up co-cultures with HSPCs. Using the OP9 BMSC-line, we modulated adipocyte differentiation prior to co-culture using a pharmacological approach and observed that high adipocytic content was associated to a decrease in HPC expansion. We perfomed a screening of over 4000 FDA-approved drugs and natural compounds and identified one to efficiently prevent adipocytic differentiation and maintain the hematopoietic supportive capacity of OP9 stroma. Altogether, this work opens the door to alternative strategies accelerating hematopoietic reconstitution and unveils the potential to improve recovery of patients undergoing BMT.

EPFL2019

The role of the transcriptional regulator BCL6 in adipose tissue development and remodeling

Teresa Didonna

The adipose tissue is increasingly recognized for the important role it plays in various diseases and pathological conditions, such as obesity, diabetes, cardiovascular diseases, osteoporosis, cancer, and aging. Despite its long-held reputation for being a simple fat storage tissue, we now know the adipose tissue is a singular organ playing a crucial role in whole-body metabolism and energy homeostasis. Understanding the development and functioning of adipose tissue and, in particular, of its structural and functional fat depots, is crucial to understanding human health and disease, and can pave the way for personalized therapeutic and preventive approaches. Transcription factors are key regulators of adipose tissue biogenesis, development, and responses to changes in environmental stimuli such as feeding, fasting, and exercise. The transcription repressor BCL6 - well known in immunology and cancer - has been recently linked to adipogenesis and metabolism. In the context of this thesis, in order to better understand the role of BCL6 in adipose tissue development and remodeling, we sought to further investigate the physiological function of BCL6 in white and brown adipose tissue. We developed and characterized a fat-specific conditional knockout mouse model for Bcl6 (Bcl6 FKO), and a Pdgfra-Cre driver was used to obtain in vivo ablation of Bcl6 in adipogenic precursors and stem cells (ASPCs). We discovered that ablation of Bcl6 in adipocyte precursors has a major impact on white fat mass and its distribution in the bodies of mice. Moreover, being fed a standard and a high-fat diet has a prominent effect on visceral adipose tissue, the mass of which was significantly reduced compared to that of control mice. Additionally, Bcl6 FKO mice did not demonstrate sensitivity to high-fat diet-induced mass gain and related comorbidities, such as hepatic steatosis and glucose-homeostasis alterations. The increased number of adipocytes observed in the gonadal adipose tissue occurred without increasing overall adiposity and body weight compared to control mice under a high-caloric diet regime. In conclusion, Bcl6 FKO mice represent a lean mouse model with a healthy balance between subcutaneous and visceral white adipose tissue depots. BCL6 could be an interesting adipose-specific target for therapeutic interventions in the context of obesity and related metabolic dysfunctions.

EPFL2019

Metabolic and microenvironmental strategies to accelerate hematopoietic recovery post-aplasia

Vasco Ledebur de Antas de Campos

EPFL2019

The role and transcriptional regulation of FAM46A in adipocyte differentiation

Ann-Kristin Hov

EPFL2021

The role of the transcriptional regulator BCL6 in adipose tissue development and remodeling

Teresa Didonna

EPFL2019