SIAH2 is Expressed in Adipocyte Precursor Cells and Interacts with EBF1 and ZFP521 to Promote Adipogenesis.

Objective: Expression of zinc finger protein 423 (ZFP423), a key proadipogenic transcription factor in adipocyte precursor cells, is regulated by interaction of the proadipogenic early B-cell factor 1 (EBF1) and antiadipogenic ZFP521. The ubiquitin ligase seven-in-absentia homolog 2 (SIAH2) targets ZFP521 for degradation. This study asked whether SIAH2 is expressed in adipocyte precursor cells and whether SIAH2 interacts with ZFP521 and EBF1 to regulate ZFP521 protein levels during adipogenesis.

Correction to: Siah2 modulates sex-dependent metabolic and inflammatory responses in adipose tissue to a high-fat diet challenge.

Following publication of the original article [1], the authors reported that additional file 1 was incorrect. The corrected additional file 1 is given below.

Siah2 modulates sex-dependent metabolic and inflammatory responses in adipose tissue to a high-fat diet challenge.

Background: The obesity-related risk of developing metabolic syndrome is higher in males than in females of reproductive age, likely due to estrogen-mediated reduced adipose tissue inflammation and fibrosis with hypertrophied adipocytes. Depletion of the ubiquitin ligase Siah2 reduced white adipose tissue inflammation and improved glucose metabolism in obese male mice. Siah2 is a transcriptional target of estrogen, but data is lacking about the effect of Siah2 on adipose tissue of females.

Isolation of Murine Adipose-Derived Stromal/Stem Cells for Adipogenic Differentiation or Flow Cytometry-Based Analysis.

Murine models of obesity or reduced adiposity are a valuable resource for understanding the role of adipocyte dysfunction in metabolic disorders. Adipose tissue stromal vascular cells or primary adipocytes derived from murine adipose tissue and grown in culture are essential tools for studying the mechanisms underlying adipocyte development and function. Herein, we describe methods for the isolation, expansion, and long-term storage of murine adipose-derived stromal/stem cells along with protocols for inducing adipogenesis in this cell population or isolating the adipose stromal vascular fraction cells for flow cytometric analysis.

Siah2 Protein Mediates Early Events in Commitment to an Adipogenic Pathway.

Adipose tissue expansion occurs by increasing the size of existing adipocytes or by increasing the number of adipocytes via adipogenesis. Adipose tissue dysfunction in obesity is associated with adipocyte hypertrophy and impaired adipogenesis. We recently demonstrated that deletion of the ubiquitin ligase Siah2 is associated with enlarged adipocytes in lean or obese mice.

The ubiquitin ligase Siah2 regulates obesity-induced adipose tissue inflammation.

Objective: Chronic, low-grade adipose tissue inflammation associated with adipocyte hypertrophy is an important link in the relationship between obesity and insulin resistance. Although ubiquitin ligases regulate inflammatory processes, the role of these enzymes in metabolically driven adipose tissue inflammation is relatively unexplored. Herein, the effect of the ubiquitin ligase Siah2 on obesity-related adipose tissue inflammation was examined.

Prolonged proteasome inhibition cyclically upregulates Oct3/4 and Nanog gene expression, but reduces induced pluripotent stem cell colony formation.

There is ample evidence that the ubiquitin-proteasome system is an important regulator of transcription and its activity is necessary for maintaining pluripotency and promoting cellular reprogramming. Moreover, proteasome activity contributes to maintaining the open chromatin structure found in pluripotent stem cells, acting as a transcriptional inhibitor at specific gene loci generally associated with differentiation. The current study was designed to understand further the role of proteasome inhibition in reprogramming and its ability to modulate endogenous expression of pluripotency-related genes and induced pluripotent stem cells (iPSCs) colony formation.

An ethanolic extract of Artemisia dracunculus L. regulates gene expression of ubiquitin-proteasome system enzymes in skeletal muscle: potential role in the treatment of sarcopenic obesity.

Objective: Obesity is linked to insulin resistance, a primary component of metabolic syndrome and type 2 diabetes. The problem of obesity-related insulin resistance is compounded when age-related skeletal muscle loss, called sarcopenia, occurs with obesity. Skeletal muscle loss results from elevated levels of protein degradation and prevention of obesity-related sarcopenic muscle loss will depend on strategies that target pathways involved in protein degradation.

An extract of Artemisia dracunculus L. inhibits ubiquitin-proteasome activity and preserves skeletal muscle mass in a murine model of diabetes.

Impaired insulin signaling is a key feature of type 2 diabetes and is associated with increased ubiquitin-proteasome-dependent protein degradation in skeletal muscle. An extract of Artemisia dracunculus L. (termed PMI5011) improves insulin action by increasing insulin signaling in skeletal muscle.

The ubiquitin ligase Siah2 regulates PPARγ activity in adipocytes.

Moderate reductions in peroxisome proliferator-activated receptor (PPAR)γ levels control insulin sensitivity as effectively as activation of PPARγ in adipocytes by the thiazolidinediones. That observation suggests that PPARγ activity can be regulated by modulating the amount of PPARγ protein in adipocytes. Activation of PPARγ in adipocytes is linked to changes in PPARγ protein levels via increased degradation of PPARγ proteins by the ubiquitin proteasome system.

An improved method for isolation of RNA from bone.

Background: Bone physiology is increasingly appreciated as an important contributor to metabolic disorders such as type 2 diabetes. However, progress in understanding the role of bone in determining metabolic health is hampered by the well-described difficulty of obtaining high quality RNA from bone for gene expression analysis using the currently available approaches.

Results: We developed a simple approach to isolate bone RNA that combines pulverizing the bone and the phenol-guanidinium based RNA extraction in a single step while maintaining near-freezing temperatures.

Isolation of murine adipose-derived stem cells.

Murine models of obesity or reduced adiposity are a valuable resource for understanding the role of adipocyte dysfunction in metabolic disorders. Primary adipocytes grown in culture and derived from murine adipose tissue are essential for studying the mechanisms underlying adipocyte development and function. Herein, we describe methods for the isolation, expansion, and long-term storage of murine adipose-derived stem cells along with a protocol for inducing adipogenesis in this cell population.

High efficiency lipid-based siRNA transfection of adipocytes in suspension.

Background: Fully differentiated adipocytes are considered to be refractory to introduction of siRNA via lipid-based transfection. However, large scale siRNA-based loss-of-function screening of adipocytes using either electroporation or virally-mediated transfection approaches can be prohibitively complex and expensive.

Methodology/principal Findings: We present a method for introducing small interfering RNA (siRNA) into differentiated 3T3-L1 adipocytes and primary human adipocytes using an approach based on forming the siRNA/cell complex with the adipocytes in suspension rather than as an adherent monolayer, a variation of "reverse transfection".

PPAR-gamma AF-2 domain functions as a component of a ubiquitin-dependent degradation signal.

The nuclear hormone receptor peroxisome proliferator-activated receptor-gamma (PPAR-gamma) functions as the "master switch" in adipocyte development and is important in regulating glucose metabolism. PPAR-gamma is rapidly degraded in adipocytes by the ubiquitin proteasome pathway under basal and ligand-activated conditions. Proteasome inhibition increases PPAR-gamma activity, indicating disposal of PPAR-gamma by the ubiquitin proteasome system regulates PPAR-gamma activity.

Modulation of peroxisome proliferator-activated receptor gamma stability and transcriptional activity in adipocytes by resveratrol.

The peroxisome proliferator-activated receptor (PPAR) gamma is essential for the formation and function of adipocytes. It is also involved in regulating insulin sensitivity and is the functional target of the thiazolidinedione class of insulin-sensitizing drugs. Whereas thiazolidinediones activate PPARgamma and decrease PPARgamma protein levels, genetic models indicate that decreased expression of PPARgamma is also associated with increased insulin sensitivity.

Isolation of human adipose-derived stem cells from biopsies and liposuction specimens.

Adipose tissue has proven to serve as an abundant, accessible, and rich source of adult stem cells with multipotent properties suitable for tissue engineering and regenerative medical applications. Here, we describe a detailed method for the isolation and expansion of adipose-derived stem cells (ASCs). We present a large scale procedure suitable for processing >100 mL volumes of lipoaspirate tissue specimens and a small scale procedure suitable for processing adipose tissue biopsy specimens of < 0.

Adipogenic human adenovirus Ad-36 induces commitment, differentiation, and lipid accumulation in human adipose-derived stem cells.

Human adenovirus Ad-36 is causatively and correlatively linked with animal and human obesity, respectively. Ad-36 enhances differentiation of rodent preadipocytes, but its effect on adipogenesis in humans is unknown. To indirectly assess the role of Ad-36-induced adipogenesis in human obesity, the effect of the virus on commitment, differentiation, and lipid accumulation was investigated in vitro in primary human adipose-derived stem/stromal cells (hASC).

Induction of circadian gene expression in human subcutaneous adipose-derived stem cells.

Objective: Genes encoding the circadian transcriptional apparatus exhibit robust oscillatory expression in murine adipose tissues. This study tests the hypothesis that human subcutaneous adipose-derived stem cells (ASCs) provide an in vitro model in which to monitor the activity of the core circadian transcriptional apparatus.

Research Methods And Procedures: Primary cultures of undifferentiated or adipocyte-differentiated ASCs were treated with dexamethasone, rosiglitazone, or 30% fetal bovine serum.

Cryopreservation characteristics of adipose-derived stem cells: maintenance of differentiation potential and viability.

With the emergence of regenerative medicine, many researchers have turned to fat tissue as a source of adipose-derived stem cells (ASCs). Because freshly collected adipose tissue is not always readily available, there will be a need for improved cryopreservation methods to reproducibly maintain ASC viablility and multipotentiality in long-term storage. This study examines the efficiency of conventional dimethyl sulphoxide cryopreservation methods by measuring the maintenance of differentiation potential after one freeze cycle.

Characterization of equine adipose tissue-derived stromal cells: adipogenic and osteogenic capacity and comparison with bone marrow-derived mesenchymal stromal cells.

Objective: To characterize equine adipose tissue-derived stromal cell (ASC) frequency and growth characteristics and assess of their adipogenic and osteogenic differentiation potential.

Study Design: In vitro experimental study.

Animals: Horses (n=5; aged, 9 months to 5 years).

Cytokine profile of human adipose-derived stem cells: expression of angiogenic, hematopoietic, and pro-inflammatory factors.

Adipose tissue serves as a source of adipokines and cytokines with both local and systemic actions in health and disease. In this study, we examine the hypothesis that multipotent human adipose-derived stem cells (ASCs), capable of differentiating along the adipocyte, chondrocyte, and osteoblast pathways, contribute to adipose tissue-derived cytokine secretion. Following exposure to basic fibroblast growth factor (bFGF) or epidermal growth factor (EGF), the ASCs significantly increase their secretion of hepatocyte growth factor (HGF), a cytokine implicated in hematopoiesis, vasculogenesis, and mammary epithelial duct formation.

Effects of prolyl hydroxylase inhibitors on adipogenesis and hypoxia inducible factor 1 alpha levels under normoxic conditions.

Adipocyte function is highly regulated in response to changing oxygen levels and physiological regulation of adipocyte formation involves factors originally identified as hypoxia-responsive proteins. Inhibition of adipogenesis at low oxygen tension is associated with activation of hypoxia inducible factor-1 alpha (HIF-1alpha), a transcription factor essential for cellular responses to decreased oxygen levels whose activity is regulated by prolyl hydroxylase (PHD) enzymes. However, HIF-1alpha RNA expression has been detected during the initial stages of adipocyte formation under aerobic conditions, suggesting a physiological role for HIF-1alpha during adipogenesis under a range of oxygen levels.

Circadian oscillation of gene expression in murine calvarial bone.

Unlabelled: The genes encoding the core circadian transcription factors display an oscillating expression profile in murine calvarial bone. More than 26% of the calvarial bone transcriptome exhibits a circadian rhythm, comparable with that observed in brown and white adipose tissues and liver. Thus, circadian mechanisms may directly modulate oxidative phosphorylation and multiple metabolic pathways in bone homeostasis.

Cell growth characteristics and differentiation frequency of adherent equine bone marrow-derived mesenchymal stromal cells: adipogenic and osteogenic capacity.

Objectives: To characterize equine bone marrow (BM)-derived mesenchymal stem cell (MSC) growth characteristics and frequency as well as their adipogenic and osteogenic differentiation potential.

Study Design: In vitro experimental study.

Animals: Foals (n=3, age range, 17-51 days) and young horses (n=5, age range, 9 months to 5 years).

Secretome of primary cultures of human adipose-derived stem cells: modulation of serpins by adipogenesis.

Studies of adipogenic protein induction have led to a new appreciation of the role of adipose tissue as an endocrine organ. Adipocyte-derived "adipokines" such as adiponectin, leptin, and visceral adipose tissue-derived serine protease inhibitor (vaspin) exert hormone-like activities at the systemic level. In this study, we examined the secretome of primary cultures of human subcutaneous adipose-derived stem cells as an in vitro model of adipogenesis.