Proteomic characterization of thiazolidinedione regulation of obese adipose secretome in Zucker obese rats

Signaling molecules released by adipose tissue have been implicated in inflammation, adipocyte dysfunction and systemic insulin resistance. In this study, we used 2-D LC-MS/MS and quantitative proteomics approaches to characterize the obese adipose secretory proteins that are responsive to the thiazolidinediones class of PPAR-γ agonizts. We first showed the differential secretion profiling between obese and lean adipose tissue; 87 proteins were detected from the conditioned medium of adipose tissue of Zucker obese rats compared with 31 from lean rats. A total of 57 proteins comprising immune factors, inflammatory molecules, collagens, proteases, and extracellular matrix proteins were detected from obese, but not lean adipose tissue. More importantly, a quantitative proteomics approach using ^(18)O proteolytic labeling allowed quantification of the difference in the secretion levels of 77 proteins, and thiazolidinediones treatment suppressed the secretion of most of the obese adipose tissue secretome, thus resembling a lean tissue. We have demonstrated an application of identifying the obese adipose secretome and characterizing the regulation of adipose secretion in obesity and insulin resistance. Our data provide the first evidence of changes in adipose secretion in obesity at a global level and show that such changes are correlated with systemic insulin resistance

Waves of Adipose Tissue Growth in the Genetically Obese Zucker Fatty Rat

In mammals, calories ingested in excess of those used are stored primarily as fat in adipose tissue; consistent ingestion of excess calories requires an enlargement of the adipose tissue mass. Thus, a dysfunction in adipose tissue growth may be a key factor in insulin resistance due to imbalanced fat storage and disrupted insulin action. Adipose tissue growth requires the recruitment and then the development of adipose precursor cells, but little is known about these processes in vivo.In this study, adipose cell-size probability distributions were measured in two Zucker fa/fa rats over a period of 151 and 163 days, from four weeks of age, using micro-biopsies to obtain subcutaneous (inguinal) fat tissue from the animals. These longitudinal probability distributions were analyzed to assess the probability of periodic phenomena.Adipose tissue growth in this strain of rat exhibits a striking temporal periodicity of approximately days. A simple model is proposed for the periodicity, with PPAR signaling driven by a deficit in lipid uptake capacity leading to the periodic recruitment of new adipocytes. This model predicts that the observed period will be diet-dependent

Differential Effects of Thiazolidinediones on Adipocyte Growth and Recruitment in Zucker Fatty Rats

Background: Adipose tissue grows by two mechanisms: hyperplasia (cell number increase) and hypertrophy (cell size increase). Thiazolidinediones are insulin-sensitizing peroxisome proliferator-activated receptor gamma agonists that are known to affect the morphology of adipose tissue. Methodology: In this study, adipose cell-size probability distributions were measured in six Zucker fa/fa rats over a period of 24 days, from four weeks of age, using micro-biopsies to obtain subcutaneous (inguinal) fat tissue from the animals. Three of the rats were gavaged daily with rosiglitazone, a thiazolidinedione, and three served as controls. These longitudinal probability distributions were analyzed to obtain the rate of increase in cell-size diameter in rosiglitazone-treated animals, and the hyperplasia induced by treatment quantitatively. Conclusions: We found that treatment leads to hypertrophy that leads to an approximately linear rate of cell diameter increase (2 mm/day), and that the hyperplasia evident in treated animals occurs largely within the first eight days of treatment. The availability of additional lipid storage due to treatment may alleviate lipotoxicity and thereby promote insulin sensitivity. The hypothesis that a TZD regimen involving repeated treatments of limited duration may suffice for improvements in insulin sensitivity merits further investigation

Analysis of GLUT4 Distribution in Whole Skeletal Muscle Fibers: Identification of Distinct Storage Compartments That Are Recruited by Insulin and Muscle Contractions

The effects of insulin stimulation and muscle contractions on the subcellular distribution of GLUT4 in skeletal muscle have been studied on a preparation of single whole fibers from the rat soleus. The fibers were labeled for GLUT4 by a preembedding technique and observed as whole mounts by immunofluorescence microscopy, or after sectioning, by immunogold electron microscopy. The advantage of this preparation for cells of the size of muscle fibers is that it provides global views of the staining from one end of a fiber to the other and from one side to the other through the core of the fiber. In addition, the labeling efficiency is much higher than can be obtained with ultracryosections. In nonstimulated fibers, GLUT4 is excluded from the plasma membrane and T tubules. It is distributed throughout the muscle fibers with ∼23% associated with large structures including multivesicular endosomes located in the TGN region, and 77% with small tubulovesicular structures. The two stimuli cause translocation of GLUT4 to both plasma membrane and T tubules. Quantitation of the immunogold electron microscopy shows that the effects of insulin and contraction are additive and that each stimulus recruits GLUT4 from both large and small depots. Immunofluorescence double labeling for GLUT4 and transferrin receptor (TfR) shows that the small depots can be further subdivided into TfR-positive and TfR-negative elements. Interestingly, we observe that colocalization of TfR and GLUT4 is increased by insulin and decreased by contractions. These results, supported by subcellular fractionation experiments, suggest that TfR-positive depots are only recruited by contractions. We do not find evidence for stimulation-induced unmasking of resident surface membrane GLUT4 transporters or for dilation of the T tubule system (Wang, W., P.A. Hansen, B.A. Marshall, J.O. Holloszy, and M. Mueckler. 1996. J. Cell Biol. 135:415–430)

Insulin stimulates the halting, tethering, and fusion of mobile GLUT4 vesicles in rat adipose cells

Glucose transport in adipose cells is regulated by changing the distribution of glucose transporter 4 (GLUT4) between the cell interior and the plasma membrane (PM). Insulin shifts this distribution by augmenting the rate of exocytosis of specialized GLUT4 vesicles. We applied time-lapse total internal reflection fluorescence microscopy to dissect intermediates of this GLUT4 translocation in rat adipose cells in primary culture. Without insulin, GLUT4 vesicles rapidly moved along a microtubule network covering the entire PM, periodically stopping, most often just briefly, by loosely tethering to the PM. Insulin halted this traffic by tightly tethering vesicles to the PM where they formed clusters and slowly fused to the PM. This slow release of GLUT4 determined the overall increase of the PM GLUT4. Thus, insulin initially recruits GLUT4 sequestered in mobile vesicles near the PM. It is likely that the primary mechanism of insulin action in GLUT4 translocation is to stimulate tethering and fusion of trafficking vesicles to specific fusion sites in the PM

Spatio-temporal ecology of sympatric felids on Borneo. Evidence for resource partitioning?

Niche differentiation, the partitioning of resources along one or more axes of a species’ niche hyper-volume, is widely recognised as an important mechanism for sympatric species to reduce interspecific competition and predation risk, and thus facilitate co-existence. Resource partitioning may be facilitated by behavioural differentiation along three main niche dimensions: habitat, food and time. In this study, we investigate the extent to which these mechanisms can explain the coexistence of an assemblage of five sympatric felids in Borneo. Using multi-scale logistic regression, we show that Bornean felids exhibit differences in both their broad and fine-scale habitat use. We calculate temporal activity patterns and overlap between these species, and present evidence for temporal separation within this felid guild. Lastly, we conducted an all-subsets logistic regression to predict the occurrence of each felid species as a function of the co-occurrence of a large number of other species and showed that Bornean felids co-occurred with a range of other species, some of which could be candidate prey. Our study reveals apparent resource partitioning within the Bornean felid assemblage, operating along all three niche dimension axes. These results provide new insights into the ecology of these species and the broader community in which they live and also provide important information for conservation planning for this guild of predators