The minor allele of the CREBRF rs373863828 p.R457Q coding variant is associated with reduced levels of myostatin in males: Implications for body composition.

OBJECTIVE: The minor allele (A) of the rs373863828 variant (p.Arg457Gln) in CREBRF is restricted to indigenous peoples of the Pacific islands (including New Zealand Maori and peoples of Polynesia), with a frequency of up to 25% in these populations. This allele associates with a large increase in body mass index (BMI) but with significantly lower risk of type-2 diabetes (T2D). It remains unclear whether the increased BMI is driven by increased adiposity or by increased lean mass. METHODS: We undertook body composition analysis using DXA in 189 young men of Maori and Pacific descent living in Aotearoa New Zealand. Further investigation was carried out in two orthologous Arg458Gln knockin mouse models on FVB/NJ and C57BL/6j backgrounds. RESULTS: The rs373863828 A allele was associated with lower fat mass when adjusted for BMI (p < 0.05) and was associated with significantly lower circulating levels of the muscle inhibitory hormone myostatin (p < 0.05). Supporting the human data, significant reductions in adipose tissue mass were observed in the knockin mice. This was more significant in older mice in both backgrounds and appeared to be the result of reduced age-associated increases in fat mass. The older male knockin mice on C57BL/6j background also had increased grip strength (p < 0.01) and lower levels of myostatin (p < 0.05). CONCLUSION: Overall, these results prove that the rs373863828 A-allele is associated with a reduction of myostatin levels which likely contribute to an age-dependent lowering of fat mass, at least in males.

Synergistic Effects of Ad-Libitum Low-Dose Fructose Drinking and Low-Dose Streptozotocin Treatment in Wistar Rats: A Mild Model of Type 2 Diabetes.

To develop a convenient animal model of T2D by pretreatment with low-dose 10% w/v fructose (FRC) solution followed by the injection of low doses of streptozotocin (STZ) in Wistar rats. For this 8-week experimental study; rats were first fed a standard chow ad-libitum diet and either tap water (n=40) or 10% w/v FRC solution (n=40) for 4 weeks. Next, rats in each category were randomly allocated to 4 subgroups (n=10 each) of low-dose STZ (25,35, and 45 mg/kg). The final mean fasting blood sugar (FBG) of FRC+STZ45 (197±55.87 mg/dl) were significantly higher than that of the STZ45 (P=0.015) and FRC (P=0.019) groups. FRC+STZ45 showed the highest insulin resistance demonstrated by insulin tolerance test [area under the curve (AUC) of insulin tolerance test; P<0.05]. AUC was not significantly different between the STZ45 and non-STZ groups and between FRC and non-FRC fed groups. Furthermore, FBG levels did not differ between FRC and non-FRC groups. Body weight measurement showed that the FRC+STZ45 group had the lowest body weight compared to all other groups. Our data provide the evidence that FRC and STZ45 synergistically could induce hyperglycemia and insulin resistance in Wistar rats. Here we presented a feasible model for initial forms of T2D by employing pretreatment with low-dose FRC solution and treatment with low-dose STZ.

Tissue resident macrophages: Key players in the pathogenesis of type 2 diabetes and its complications.

There is increasing evidence showing that chronic inflammation is an important pathogenic mediator of the development of type 2 diabetes (T2D). It is now generally accepted that tissue-resident macrophages play a major role in regulation of tissue inflammation. T2D-associated inflammation is characterized by an increased abundance of macrophages in different tissues along with production of inflammatory cytokines. The complexity of macrophage phenotypes has been reported from different human tissues. Macrophages exhibit a phenotypic range that is intermediate between two extremes, M1 (pro-inflammatory) and M2 (anti-inflammatory). Cytokines and chemokines produced by macrophages generate local and systemic inflammation and this condition leads to pancreatic ß-cell dysfunction and insulin resistance in liver, adipose and skeletal muscle tissues. Data from human and animal studies also suggest that macrophages contribute to T2D complications such as nephropathy, neuropathy, retinopathy and cardiovascular diseases through cell-cell interactions and the release of pro-inflammatory cytokines, chemokines, and proteases to induce inflammatory cell recruitment, cell apoptosis, angiogenesis, and matrix protein remodeling. In this review we focus on the functions of macrophages and the importance of these cells in the pathogenesis of T2D. In addition, the contribution of macrophages to diabetes complications such as nephropathy, neuropathy, retinopathy and cardiovascular diseases is discussed.

TNF-α knockdown alleviates palmitate-induced insulin resistance in C2C12 skeletal muscle cells.

Insulin resistance is a cardinal feature of Type 2 Diabetes (T2D), which accompanied by lipid accumulation and TNF-α overexpression in skeletal muscle. The role of TNF-α in palmitate-induced insulin resistance remained to be elucidated. Here, we assessed effects of TNF-α knockdown on the components of insulin signaling pathway (IRS-1 and Akt) in palmitate-induced insulin resistant C2C12 skeletal muscle cells. To reduce TNF-α expression, C2C12 cells were transduced with TNF-α-shRNA lentiviral particles. Afterwards, the protein expression of TNF-α, IRS-1, and Akt, as well as phosphorylation levels of IRS-1 and Akt were evaluated by western blot. We also measured insulin-stimulated glucose uptake in the presence and absence of palmitate. TNF-α protein expression in C2C12 cells significantly increased by treatment with 0.75 mM palmitate (P < 0.05). In TNF-α knockdown cells, the protein expression level of TNF-α was significantly decreased by almost 70% (P < 0.01) compared with the control cells. Our results also revealed that, in control cells, palmitate treatment significantly reduced the insulin-induced phosphorylations of IRS-1 (Tyr632) and Akt (Ser473) by 60% and 66% (P < 0.01), respectively. Interestingly, these phosphorylations, even in the presence of palmitate, were not significantly reduced in TNF-α knockdown cells with respect to the untreated control cells (P > 0.05). Furthermore, palmitate significantly reduced insulin-dependent glucose uptake in control cells, however, it was not able to reduce insulin-stimulated glucose uptake in TNF-α knockdown cells in comparison with the untreated control cells (P < 0.01). These findings indicated that TNF-α down-regulation maintains insulin sensitivity, even in the presence of palmitate, therefore, TNF-α inhibition could be a good strategy for the treatment of palmitate-induced insulin resistance.

Hydrodynamic-based delivery of PTP1B shRNA reduces plasma glucose levels in diabetic mice.

Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of insulin signaling which is overexpressed in the liver of diabetic animals. The aims of this study were to generate liver-specific PTP1B knockout mice using a PTP1B­short hairpin RNA (shRNA) plasmid and to investigate the effect of PTP1B inhibition on plasma glucose levels in streptozotocin-induced diabetic mice. We first validated the hydrodynamic tail vein injection in mice using a vector carrying the luciferase gene. Expression of the PTP1B gene was quantified by real-time PCR. The level of phosphorylated Akt was examined by western blot analysis. The injection of the plasmid containing firefly luciferase revealed that the highest transfer of the vector into the liver was obtained 24 h after the injection of 20 µg plasmid. The injection of PTP1B-shRNA, but not the scrambled shRNA plasmid, resulted in a reduction in PTP1B expression levels by up to 84% in the liver of the diabetic mice. Plasma glucose levels following the injection of PTP1B-shRNA remained significantly lower in the diabetic mice for 5 days. In addition, mice receiving PTP1B-shRNA in the basal and insulin-stimulated states had higher levels of Akt phosphorylation in the liver cells compared with mice that were injected with the scrambled sequence (35 and 60%, respectively; p<0.01). Furthermore, PTP1B overexpression was observed in the muscle, liver, adipose, heart and kidney tissues of the diabetic mice. The data from this study demonstrate that PTP1B inhibition may be a promising approach for lowering plasma glucose levels in diabetic patients. However, further studies using non-viral carriers are required to deliver the plasmid safely into the liver.