OBJECTIVE-Based on rodent studies, we examined the hypothesis that increased adipose tissue (AT) mass in obesity without an adequate support of vascularization might lead to hypoxia, macrophage infiltration, and inflammation.RESEARCH DESIGN AND METHODS-Oxygen partial pressure (AT pO 2 ) and AT temperature in abdominal AT (9 lean and 12 overweight/obese men and women) was measured by direct insertion of a polarographic Clark electrode. Body composition was measured by dual-energy X-ray absorptiometry, and insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp. Abdominal subcutaneous tissue was used for staining, quantitative RT-PCR, and chemokine secretion assay.RESULTS-AT pO 2 was lower in overweight/obese subjects than lean subjects (47 Ϯ 10.6 vs. 55 Ϯ 9.1 mmHg); however, this level of pO 2 did not activate the classic hypoxia targets (pyruvate dehydrogenase kinase and vascular endothelial growth factor [VEGF]). AT pO 2 was negatively correlated with percent body fat (R ϭ Ϫ0.50, P Ͻ 0.05). Compared with lean subjects, overweight/ obese subjects had 44% lower capillary density and 58% lower VEGF, suggesting AT rarefaction (capillary drop out). This might be due to lower peroxisome proliferator-activated receptor ␥1 and higher collagen VI mRNA expression, which correlated with AT pO 2 (P Ͻ 0.05). Of clinical importance, AT pO 2 negatively correlated with CD68 mRNA and macrophage inflammatory protein 1␣ secretion (R ϭ Ϫ0.58, R ϭ Ϫ0.79, P Ͻ 0.05), suggesting that lower AT pO 2 could drive AT inflammation in obesity.CONCLUSIONS-Adipose tissue rarefaction might lie upstream of both low AT pO 2 and inflammation in obesity. These results suggest novel approaches to treat the dysfunctional AT found in obesity. Diabetes 58:718-725, 2009 B oth insulin resistance and ␤-cell failure are present in individuals with type 2 diabetes. Insulin resistance is closely linked to adiposity with a central or visceral pattern, providing a greater risk of insulin resistance and metabolic dysfunction. Adipose tissue (AT) serves as an endocrine organ secreting a variety of autocrine, paracrine, and endocrine factors that can produce or prevent insulin resistance (1). The failure of AT to adequately proliferate and/or differentiate to sequester lipids away from liver, skeletal muscle, and the pancreatic ␤-cell has been proposed as a precursor to type 2 diabetes, broadening the number of potential mechanisms linking obesity to insulin resistance (2).The increase in body fat in obesity should be accompanied by an increase in vascularization, in order to provide adequate oxygen and nutrients (3). In contrast to expectations, obese mice have lower AT capillary density (rarefaction, also known as capillary drop out) and decreased vascular endothelial growth factor (VEGF), the most potent angiogenic factor (4,5). Consistent with this model, preclinical studies suggest that obese AT is hypoxic (6); however, the hypothesis that AT rarefaction might lead to hypoxia remains untested.In humans, short-term whole-body hypoxia decreases i...

The transcriptional co-activator PGC-1␣ regulates functional plasticity in adipose tissue by linking sympathetic input to the transcriptional program of adaptive thermogenesis. We report here a novel truncated form of PGC-1␣ (NT-PGC-1␣) produced by alternative 3 splicing that introduces an in-frame stop codon into PGC-1␣ mRNA. The expressed protein includes the first 267 amino acids of PGC-1␣ and 3 additional amino acids from the splicing insert. NT-PGC-1␣ contains the transactivation and nuclear receptor interaction domains but is missing key domains involved in nuclear localization, interaction with other transcription factors, and protein degradation. Expression and subcellular localization of NT-PGC-1␣ are dynamically regulated in the context of physiological signals that regulate fulllength PGC-1␣, but the truncated domain structure conveys unique properties with respect to protein-protein interactions, protein stability, and recruitment to target gene promoters. Therefore, NT-PGC-1␣ is a co-expressed, previously unrecognized form of PGC-1␣ with functions that are both unique from and complementary to PGC-1␣.

Fibers are one of the mechanical tissues that provide structural support to the plant body. To understand how the normal mechanical strength of fibers is regulated, we isolated an Arabidopsis fragile fiber (fra2) mutant defective in the mechanical strength of interfascicular fibers in the inflorescence stems. Anatomical and chemical analyses showed that the fra2 mutation caused a reduction in fiber cell length and wall thickness, a decrease in cellulose and hemicellulose contents, and an increase in lignin condensation, indicating that the fragile fiber phenotype of fra2 is a result of alterations in fiber cell elongation and cell wall biosynthesis. In addition to the effects on fibers, the fra2 mutation resulted in a remarkable reduction in cell length and an increase in cell width in all organs, which led to a global alteration in plant morphology. The FRA2 gene was shown to encode a protein with high similarity to katanin (hence FRA2 was renamed AtKTN1), a protein shown to be involved in regulating microtubule disassembly by severing microtubules. Consistent with the putative function of AtKTN1 as a microtubule-severing protein, immunolocalization demonstrated that the fra2 mutation caused delays in the disappearance of perinuclear microtubule array and in the establishment of transverse cortical microtubule array in interphase and elongating cells. Together, these results suggest that AtKTN1, a katanin-like protein, is essential not only for normal cell wall biosynthesis and cell elongation in fiber cells but also for cell expansion in all organs.

These results are consistent with basic science data, suggesting that COL6A3 might contribute to adipose tissue inflammation.

It has long been hypothesized that cortical microtubules (MTs) control the orientation of cellulose microfibril deposition, but no mutants with alterations of MT orientation have been shown to affect this process. We have shown previously that in Arabidopsis, the fra2 mutation causes aberrant cortical MT orientation and reduced cell elongation, and the gene responsible for the fra2 mutation encodes a katanin-like protein. In this study, using field emission scanning electron microscopy, we found that the fra2 mutation altered the normal orientation of cellulose microfibrils in walls of expanding cells. Although cellulose microfibrils in walls of wild-type cells were oriented transversely along the elongation axis, cellulose microfibrils in walls of fra2 cells often formed bands and ran in different directions. The fra2 mutation also caused aberrant deposition of cellulose microfibrils in secondary walls of fiber cells. The aberrant orientation of cellulose microfibrils was shown to be correlated with disorganized cortical MTs in several cell types examined. In addition, the thickness of both primary and secondary cell walls was reduced significantly in the fra2 mutant. These results indicate that the katanin-like protein is essential for oriented cellulose microfibril deposition and normal cell wall biosynthesis. We further demonstrated that the Arabidopsis katanin-like protein possessed MT-severing activity in vitro; thus, it is an ortholog of animal katanin. We propose that the aberrant MT orientation caused by the mutation of katanin results in the distorted deposition of cellulose microfibrils, which in turn leads to a defect in cell elongation. These findings strongly support the hypothesis that cortical MTs regulate the oriented deposition of cellulose microfibrils that determines the direction of cell elongation.

Cortical microtubules have long been hypothesized to regulate the oriented deposition of cellulose microfibrils. However, the molecular mechanisms of how microtubules direct the orientation of cellulose microfibril deposition are not known. We have used fibers in the inflorescence stems of Arabidopsis to study secondary wall deposition and cell wall strength and found a fragile fiber ( fra1 ) mutant with a dramatic reduction in the mechanical strength of fibers. The fra1 mutation did not cause any defects in cell wall composition, secondary wall thickening, or cortical microtubule organization in fiber cells. An apparent alteration was found in the orientation of cellulose microfibrils in fra1 fiber walls, indicating that the reduced mechanical strength of fra1 fibers probably was attributable to altered cellulose microfibril deposition. The FRA1 gene was cloned and found to encode a kinesin-like protein with an N-terminal microtubule binding motor domain. The FRA1 protein was shown to be concentrated around the periphery of the cytoplasm but absent in the nucleus. Based on these findings, we propose that the FRA1 kinesin-like protein is involved in the microtubule control of cellulose microfibril order.

Dietary methionine restriction (MR) produces a rapid and persistent remodeling of white adipose tissue (WAT), an increase in energy expenditure (EE), and enhancement of insulin sensitivity. Recent work established that hepatic expression of FGF21 is robustly increased by MR. Fgf21−/− mice were used to test whether FGF21 is an essential mediator of the physiological effects of dietary MR. The MR-induced increase in energy intake and EE and activation of thermogenesis in WAT and brown adipose tissue were lost in Fgf21−/− mice. However, dietary MR produced a comparable reduction in body weight and adiposity in both genotypes because of a negative effect of MR on energy intake in Fgf21−/− mice. Despite the similar loss in weight, dietary MR produced a more significant increase in in vivo insulin sensitivity in wild-type than in Fgf21−/− mice, particularly in heart and inguinal WAT. In contrast, the ability of MR to regulate lipogenic and integrated stress response genes in liver was not compromised in Fgf21−/− mice. Collectively, these findings illustrate that FGF21 is a critical mediator of the effects of dietary MR on EE, remodeling of WAT, and increased insulin sensitivity but not of its effects on hepatic gene expression.

Achieving the vision of identifying and quantifying cancer-related events and targets for future personalized oncology is predicated on the existence of synthetically accessible and economically viable probe molecules fully able to report the presence of these events and targets in a rapid, and highly selective and sensitive fashion. Delineated here are the design and evaluation of a newly synthesized turn-on probe whose intense fluorescent reporter signature is revealed only through probe activation by a specific intracellular enzyme present in tumor cells of multiple origins. Quenching of molecular probe fluorescence is achieved through unique photo-induced electron transfer (PeT) between the naphthalimide dye reporter and a covalently attached, quinone-based enzyme substrate. Fluorescence of the reporter dye is turned on by rapid removal of the quinone quencher, an event that immediately occurs only after highly selective, two-electron reduction of the sterically and conformationally restricted quinone substrate by the cancer-associated human NAD(P)H:quinone oxidoreductase isozyme 1 (hNQO1). Successes of the approach include rapid differentiation of NQO1-expressing and non-expressing cancer cell lines via the unaided eye, flow cytometry, fluorescence imaging, and two-photon microscopy. The potential for use of the turn-on probe in longer-term cellular studies is indicated by its lack of influence on cell viability and its in vitro stability.