STAT5 proteins play a role in adipocyte development and function, but their specific functions are largely unknown. To this end, we used an unbiased MS-based approach to identify novel STAT5-interacting proteins. We observed that STAT5A bound the E1β and E2 subunits of the pyruvate dehydrogenase complex (PDC). Whereas STAT5A typically localizes to the cytosol or nucleus, PDC normally resides within the mitochondrial matrix where it converts pyruvate to acetyl-CoA. We employed affinity purification and immunoblotting to validate the interaction between STAT5A and PDC subunits in murine and human cultured adipocytes, as well as in adipose tissue. We found that multiple PDC subunits interact with hormone-activated STAT5A in a dose- and time-dependent manner that coincides with tyrosine phosphorylation of STAT5. Using subcellular fractionation and immunofluorescence microscopy, we observed that PDC-E2 is present within the adipocyte nucleus where it associates with STAT5A. Because STAT5A is a transcription factor, we used chromatin immunoprecipitation (ChIP) to assess PDC's ability to interact with STAT5 DNA-binding sites. These analyses revealed that PDC-E2 is bound to a STAT5-binding site in the promoter of the STAT5 target gene cytokine-inducible SH2-containing protein (cish). We have demonstrated a compelling interaction between STAT5A and PDC subunits in adipocytes under physiological conditions. There is previous evidence that PDC localizes to cancer cell nuclei where it plays a role in histone acetylation. On the basis of our ChIP data and these previous findings, we hypothesize that PDC may modulate STAT5's ability to regulate gene expression by controlling histone or STAT5 acetylation.

Herbal remedies are increasing in popularity as treatments for metabolic conditions such as obesity and Type 2 Diabetes. One potential therapeutic option is fenugreek seeds (Trigonella foenum-graecum), which have been used for treating high cholesterol and Type 2 diabetes. A proposed mechanism for these benefits is through alterations in the microbiome, which impact mammalian host metabolic function. This study used untargeted metabolomics to investigate the fenugreek-induced alterations in the intestinal, liver, and serum profiles of mice fed either a 60% high-fat or low-fat control diet each with or without fenugreek supplementation (2% w/w) for 14 weeks. Metagenomic analyses of intestinal contents found significant alterations in the relative composition of the gut microbiome resulting from fenugreek supplementation. Specifically, Verrucomicrobia, a phylum containing beneficial bacteria which are correlated with health benefits, increased in relative abundance with fenugreek. Metabolomics partial least squares discriminant analysis revealed substantial fenugreek-induced changes in the large intestines. However, it was observed that while the magnitude of changes was less, significant modifications were present in the liver tissues resulting from fenugreek supplementation. Further analyses revealed metabolic processes affected by fenugreek and showed broad ranging impacts in multiple pathways, including carnitine biosynthesis, cholesterol and bile acid metabolism, and arginine biosynthesis. These pathways may play important roles in the beneficial effects of fenugreek.

An ethanolic extract of Artemisia scoparia (SCO) has metabolically favorable effects on adipocyte development and function in vitro and in vivo. In diet-induced obese mice, SCO supplementation significantly reduced fasting glucose and insulin levels. Given the importance of adipocyte lipolysis in metabolic health, we tested hypothesized that SCO modulates lipolysis in vitro and in vivo. Free fatty acids and glycerol were measured in the sera of mice fed a high-fat diet with or without SCO supplementation. In cultured 3T3-L1 adipocytes, the effects of SCO on lipolysis were assessed by measuring glycerol and free fatty acid release. Microarray analysis, qPCR, and immunoblotting were used to assess gene expression and protein abundance. We found that SCO supplementation of a high-fat diet in mice substantially reduces circulating glycerol and free fatty acid levels, and observed a cell-autonomous effect of SCO to significantly attenuate TNFα (tumor necrosis factor alpha)-induced lipolysis in cultured adipocytes. Although several pro-lipolytic and anti-lipolytic genes were identified by microarray analysis of subcutaneous and visceral adipose tissue from SCO-fed mice, regulation of these genes did not consistently correlate with SCO's ability to reduce lipolytic metabolites in sera or cell culture media. However, in the presence of TNFα in cultured adipocytes, SCO induced anti-lipolytic changes in phosphorylation of hormone sensitive lipase and perilipin. Together, these data suggest that the anti-lipolytic effects of SCO on adipose tissue play a role in the ability of this botanical extract to improve whole-body metabolic parameters, and support its use as a dietary supplement to promote metabolic resiliency.