A NEUROGENIC GENE EXPRESSION SIGNATURE SUPPORTS HUMAN THERMOGENIC ADIPOSE TISSUE DEVELOPMENT IN VIVO.

Human beige/brite thermogenic adipose tissue exerts beneficial metabolic effects and may be harnessed to improve metabolic health. To uncover mechanisms by which thermogenic adipose tissue is generated and maintained we developed a species-hybrid model in which human mesenchymal progenitor cells are induced in vitro to differentiate into white or thermogenic adipocytes and are then implanted into immuno-compromised mice. Upon implantation, thermogenic adipocytes form a more densely vascularized and innervated adipose tissue compared to non-thermogenic adipocytes. Mouse endothelial and stem/progenitor cells recruited by implanted human thermogenic adipocytes are also qualitatively different, with differentially expressed genes mapping predominantly to circadian rhythm pathways. We trace the formation of this enhanced neurovascular architecture to higher expression of a distinct set of genes directly associated with neurogenesis (THBS4, TNC, NTRK3 and SPARCL1), and to lower expression of genes associated with neurotransmitter degradation (MAOA, ACHE) by adipocytes in the developed tissue. Further analysis reveals that MAOA is abundant in human adipocytes but absent in mouse adipocytes, revealing species-specific mechanisms of neurotransmitter tone regulation. In summary, our work discovers specific neurogenic genes associated with development and maintenance of human thermogenic adipose tissue, reveals species-specific mechanisms of control of neurotransmitter tone, and suggests that targeting adipocyte MAOA may be a strategy for enhancing thermogenic adipose tissue activity in humans.

Abstract P158: Endothelin Inhibits Adiponectin Production Via ETB Receptor Activation On Adipocytes

Endothelin-1 has been implicated in obesity related insulin resistance. Our lab recently demonstrated that ET-1 is elevated in adipose tissue of obese mice, and blockade of ET-1 receptors improves insulin sensitivity in a mouse model of diet induced obesity. One potential mechanism by which ET-1 promotes insulin resistance is through activation of the ET-1 type B receptor (ET B ). Blockade of ET B receptors improves insulin sensitivity and increases circulating adiponectin, an adipokine only released by adipose tissue. Therefore, the current hypothesis is that ET-1 causes insulin resistance and inhibits adiponectin production by adipocytes. Primary mouse adipocytes were cultured and chronically treated with ET-1 for 3 days. ET-1 treated adipocytes had significantly lower peroxisome proliferator activator gamma, a transcription factor that drive adiponectin production, and adiponectin mRNA expression and release into media. This response was attenuated by co-treatment with an ET B receptor antagonist (BQ-788; 57.2±2.0 vehicle, 42.5±4.5 ET-1, 59.8±1.5 ET-1+BQ788, ng/ml; p<0.05) and in adipocytes from adipocyte ET B receptor knockout mice. Further, expression of several genes in the insulin signaling pathway, including Glut4 and insulin receptor substrates 1 and 2 were significantly reduced in adipocytes treated with ET-1, a response that was attenuated with ET B receptor blockade or knockout of the ET B receptor. These data suggest that increased ET-1 production in adipose tissue promotes insulin resistance on adipocytes and inhibits the release of insulin sensitizing adipokines such as adiponectin, a potential mechanism by which ET-1 receptor blockade improves insulin sensitivity in obese mice.

Long Non-coding RNA 332443 Inhibits Preadipocyte Differentiation by Targeting Runx1 and p38-MAPK and ERK1/2-MAPK Signaling Pathways

Long non-coding RNAs (lncRNAs) have emerged as integral regulators of pathophysiological processes, but their specific roles and mechanisms in adipose tissue development remain largely unknown. Here, through microarray analysis, co-expression, and tissue specific analysis of adipocyte tissues after fasting for 72 h, we found that Lnc-FR332443 expression was dramatically decreased, as well as the expression of Runx1. The UCSC database and Ensembl database indicated that Lnc-FR332443 is the antisense lncRNA of Runx1. Lnc-FR332443 and Runx1 are highly enriched in adipose tissue and downregulated during adipogenic differentiation. Adipose tissue-specific knockdown of Lnc-FR332443 increased fat mass in vivo, and specific knockdown of Lnc-FR332443 in 3T3-L1 preadipocytes promoted adipogenic differentiation. In this process, Runx1 expression was decreased when Lnc-FR332443 was downregulated in adipocytes or 3T3-L1 preadipocytes, and vice versa, when Lnc-FR332443 was upregulated, the expression of Runx1 was increased. However, overexpression of Runx1 decreased the expression of the adipocyte cell marker genes PPARγ, C/EBPα and FABP4 significantly, while not affected the expression of Lnc-FR332443. Mechanistically, Lnc-FR332443 positively regulates Runx1 expression in mouse adipocytes and suppresses adipocyte differentiation by attenuating the phosphorylation of MAPK-p38 and MAPK-ERK1/2 expression. Thus, this study indicated that Lnc-FR332443 inhibits adipogenesis and which might be a drug target for the prevention and treatment of obesity.

Diet-dependent Natriuretic Peptide Receptor C expression in adipose tissue is mediated by PPARγ via long-range distal enhancers

In addition to their established role to maintain blood pressure and fluid volume, the cardiac natriuretic peptides (NPs) can stimulate adipocyte lipolysis and control the brown fat gene program of nonshivering thermogenesis. The NP “clearance” receptor C (NPRC) functions to clear NPs from the circulation via peptide internalization and degradation and thus is an important regulator of NP signaling and adipocyte metabolism. It is well appreciated that the Nprc gene is highly expressed in adipose tissue and is dynamically regulated with nutrition and environmental changes. However, the molecular basis for how Nprc gene expression is regulated is still poorly understood. Here we identified Peroxisome Proliferator-Activated Receptor gamma (PPARγ) as a transcriptional regulator of Nprc expression in mouse adipocytes. During 3T3-L1 adipocyte differentiation, levels of Nprc expression increase in parallel with PPARγ induction. Rosiglitazone, a classic PPARγ agonist, increases, while siRNA knockdown of PPARγ reduces, Nprc expression in 3T3-L1 adipocytes. We demonstrate that PPARγ controls Nprc gene expression in adipocytes through its long-range distal enhancers. Furthermore, the induction of Nprc expression in adipose tissue during high-fat diet feeding is associated with increased PPARγ enhancer activity. Our findings define PPARγ as a mediator of adipocyte Nprc gene expression and establish a new connection between PPARγ and the control of adipocyte NP signaling in obesity.

Dermal fibroblast-like cells reprogrammed directly from adipocytes in mouse

In deep burns, early wound closure is important for healing, and skin grafting is mainly used for wound closure. However, it is difficult to achieve early wound closure in extensive total body surface area deep burns due to the lack of donor sites. Dermal fibroblasts, responsible for dermis formation, may be lost in deep burns. However, fat layers composed of adipocytes, lying underneath the dermis, are retained even in such cases. Direct reprogramming is a novel method for directly reprograming some cells into other types by introducing specific master regulators; it has exhibited appreciable success in various fields. In this study, we aimed to assess whether the transfection of master regulators (ELF4, FOXC2, FOXO1, IRF1, PRRX1, and ZEB1) could reprogram mouse adipocytes into dermal fibroblast-like cells. Our results indicated the shrinkage of fat droplets in reprogrammed mouse adipocytes and their transformation into spindle-shaped dermal fibroblasts. Reduced expression of PPAR-2, c/EBP, aP2, and leptin, the known markers of adipocytes, in RT-PCR, and enhanced expression of anti-ER-TR7, the known anti-fibroblast marker, in immunocytochemistry, were confirmed in the reprogrammed mouse adipocytes. The dermal fibroblast-like cells, reported here, may open up a new treatment mode for enabling early closure of deep burn wounds.

YAP and TAZ protect against white adipocyte cell death during obesity

The expansion of the white adipose tissue (WAT) in obesity goes along with increased mechanical, metabolic and inflammatory stress. How adipocytes resist this stress is still poorly understood. Both in human and mouse adipocytes, the transcriptional co-activators YAP/TAZ and YAP/TAZ target genes become activated during obesity. When fed a high-fat diet (HFD), mice lacking YAP/TAZ in white adipocytes develop severe lipodystrophy with adipocyte cell death. The pro-apoptotic factor BIM, which is downregulated in adipocytes of obese mice and humans, is strongly upregulated in YAP/TAZ-deficient adipocytes under HFD, and suppression of BIM expression reduces adipocyte apoptosis. In differentiated adipocytes, TNFα and IL-1β promote YAP/TAZ nuclear translocation via activation of RhoA-mediated actomyosin contractility and increase YAP/TAZ-mediated transcriptional regulation by activation of c-Jun N-terminal kinase (JNK) and AP-1. Our data indicate that the YAP/TAZ signaling pathway may be a target to control adipocyte cell death and compensatory adipogenesis during obesity.