Reversal of obesity and insulin resistance by a non-peptidic glucagon-like peptide-1 receptor agonist in diet-induced obese mice

Background

Glucagon-like peptide-1 (GLP-1) is recognized as an important regulator of glucose homeostasis. Efforts to utilize GLP-1 mimetics in the treatment of diabetes have yielded clinical benefits. A major hurdle for an effective oral therapy has been the difficulty of finding a non-peptidic GLP-1 receptor (GLP-1R) agonist. While its oral bioavailability still poses significant challenges, Boc5, one of the first such compounds, has demonstrated the attainment of GLP-1R agonism in diabetic mice. The present work was to investigate whether subchronic Boc5 treatment can restore glycemic control and induce sustainable weight loss in diet-induced obese (DIO) mice, an animal model of human obesity and insulin resistance.

Methodology/Principal Findings

DIO mice were treated three times a week with Boc5 (0.3, 1 and 3 mg) for 12 weeks. Body weight, body mass index (BMI), food intake, fasting glucose, intraperitoneal glucose tolerance and insulin induced glucose clearance were monitored regularly throughout the treatment. Glucose-stimulated insulin secretion, β-cell mass, islet size, body composition, serum metabolic profiles, lipogenesis, lipolysis, adipose hypertrophy and lipid deposition in the liver and muscle were also measured after 12 weeks of dosing. Boc5 dose-dependently reduced body weight, BMI and food intake in DIO mice. These changes were associated with significant decreases in fat mass, adipocyte hypertrophy and peripheral tissue lipid accumulation. Boc5 treatment also restored glycemic control through marked improvement of insulin sensitivity and normalization of β-cell mass. Administration of Boc5 (3 mg) reduced basal but enhanced insulin-mediated glucose incorporation and noradrenaline-stimulated lipolysis in isolated adipocytes from obese mice. Furthermore, circulating leptin, adiponectin, triglyceride, total cholesterol, nonesterified fatty acid and high-density lipoprotein/low-density lipoprotein ratio were normalized to various extents by Boc5 treatment.

Conclusions/Significance

Boc5 may produce metabolic benefits via multiple synergistic mechanisms and may represent an attractive tool for therapeutic intervention of obesity and diabetes, by means of non-peptidic GLP-1R agonism.     

The involvement of cytosolic lipases in converting phosphatidyl choline to substrate for galactolipid synthesis in the chloroplast envelope

Here we report that cytosolic phospholipases are involved in the utilization of phosphatidylcholine (PC) as substrate for chloroplast-localized synthesis of monogalactosyldiacylglycerol (MGDG). Isolated chloroplasts were pre-incubated with lysoPC and [14C]18:0-CoA to form [14C]PC. When soluble plant proteins (cytosol) and UDP-galactose were added, [14C] MGDG was formed. An inhibitor of phospholipase D markedly lowered the formation of [14C]MGDG, whereas thermolysin pretreatment of the chloroplasts was without effect. The cytosolic activity resided in the >100-kDa fraction. In a second approach, [14C]PC-containing lipid mixtures were incubated with cytosol. Degradation of [14C]PC to [14C]diacylglycerol was highest when the lipid composition of the mixture mimicked that of the outer chloroplast envelope. We also investigated whether PC of extraplastidic origin could function as substrate for MGDG synthesis. Isolated chloroplasts were incubated with enriched endoplasmic reticulum containing radiolabelled acyl lipids. In the presence of cytosol and UDP-galactose, there was a time-dependent transfer of [14C]PC from this fraction to chloroplasts, where [14C]MGDG was formed. We conclude that chloroplasts recruit cytosolic phospholipase D and phosphatidic acid phosphatase to convert PC to diacylglycerol. Apparently, these lipases do not interact with chloroplast surface proteins, but rather with outer membrane lipids, either for association to the envelope or for substrate presentation.     

Differences in the domain forming properties of N-palmitoylated neutral glycosphingolipids in bilayer membranes

We have compared the domain forming properties of three neutral acyl chain defined glycosphingolipids differing in their head group structures. The aim of the study was to explore if glycosphingolipids and sterols exist in the same lateral domains in bilayer membranes and how the structure of the head group influences the capacity of the glycosphingolipids to colocalize with cholesterol. The glycosphingolipids used in the study were galactosyl-, glucosyl- and lactosylceramides with a palmitic acid in the N-linked position. Domain formation in mixed bilayer vesicles was examined using fluorescent reporter molecules associating with ordered domains, together with a fluorescence quencher lipid in the disordered membrane phase. Our results show that the glycosphingolipids studied were poor in forming sterol-enriched domains compared to palmitoyl-sphingomyelin as detected by cholestatrienol quenching. However, the tendency to associate with cholesterol was clearly dependent on the carbohydrate structure of the glycosphingolipids, also when two glycosphingolipids with different head groups were mixed in the bilayer. All palmitoylated glycosphingolipids associated with palmitoyl-sphingomyelin/cholesterol domains. Our results show that the head group structures of neutral glycosphingolipids markedly affect their domain forming properties in bilayers both with and without cholesterol. The most striking observation being that large differences in domain forming properties were seen even between glucosylceramide and galactosylceramide, which differ only in the stereochemistry of one hydroxyl group in the carbohydrate head group.     

TGF-beta induced G(1) cell cycle arrest requires the activity of the proteasome pathway. Transforming growth factor

Transforming growth factor-beta (TGF-beta) induces a potent G(1)/S-phase cell cycle arrest of epithelial cells by inhibiting the activities of cyclin D- and cyclin E-associated kinase complexes. Downregulation of the kinase activities is mediated by induction of cyclin dependent kinase (CDK) inhibitor p15(Ink4b) which blocks CDK4 and CDK6 kinases and leads to binding of p27(Kip1) to CDK2-cyclin E complex. Levels of several of these factors are controlled by the ubiquitin-proteasome pathway. We demonstrate here that proteasomal inhibitors release the cells from TGF-beta imposed G(1)-phase arrest and instigate the entry of the cells into S-phase. Proteasomal inhibitors are shown to specifically increase the activity of the cyclin D-kinase complex by increasing the levels of p27(Kip1) and cyclin D and by maintaining CDK4/6 protein levels leading to phosphorylation of the retinoblastoma protein without increasing cyclin E-associated kinase activity. The results indicate caution in the potential therapeutic use of the proteasome inhibitors due to unscheduled initiation of DNA replication in the presence of a physiological growth inhibitor.     

ErbB2 Receptor Immunoreactivity in Prostate Cancer: Relationship to the Androgen Receptor,Disease Severity at Diagnosis and Disease Outcome

Background

ErbB2 is a member of the epidermal growth factor family of tyrosine kinases that is centrally involved in the pathogenesis of prostate cancer and several studies have reported that a high expression of this protein has prognostic value. In the present study, we have investigated whether tumour ErbB2 immunoreactivity (ErbB2-IR) has clinically useful prognostic value, i.e. that it provides additional prognostic information to that provided by routine clinical tests (Gleason score, tumour stage).

Methodology/Principal Findings

ErbB2-IR was measured in a well-characterised tissue microarray of tumour and non-malignant samples obtained at diagnosis. Additionally, mRNA levels of ErbB2-IR in the prostate were determined in the rat following manipulation of circulating androgen levels. Tumour ErbB2-IR was significantly associated with the downstream signalling molecule phosphorylated-Akt and with the cell proliferation marker Ki-67. The significant association of tumour ErbB2-IR with the Gleason score at diagnosis was lost when controlled for the association of both parameters with Ki-67. In the rat prostate, mRNA for ErbB2 was inversely associated with circulating androgen levels. There was no association between ErbB2-IR and the androgen receptor (AR)-IR in the tumours, but an interaction between the two parameters was seen with respect to their association with the tumour stage. Tumour ErbB2-IR was confirmed to be a prognostic marker for disease-specific survival, but it did not provide significant additive information to the Gleason score or to Ki-67.

Conclusions/Significance

It is concluded that tumour ErbB2-IR is of limited clinical value as a prognostic marker to aid treatment decisions, but could be of pathophysiological importance in prostate cancer.     

Multispectral imaging for determination of astaxanthin concentration in salmonids

Multispectral imaging has been evaluated for characterization of the concentration of a specific cartenoid pigment; astaxanthin. 59 fillets of rainbow trout, Oncorhynchus mykiss, were filleted and imaged using a rapid multispectral imaging device for quantitative analysis. The multispectral imaging device captures reflection properties in 19 distinct wavelength bands, prior to determination of the true concentration of astaxanthin. The samples ranged from 0.20 to 4.34 g per g fish. A PLSR model was calibrated to predict astaxanthin concentration from novel images, and showed good results with a RMSEP of 0.27. For comparison a similar model were built for normal color images, which yielded a RMSEP of 0.45. The acquisition speed of the multispectral imaging system and the accuracy of the PLSR model obtained suggest this method as a promising technique for rapid in-line estimation of astaxanthin concentration in rainbow trout fillets.