Cellular mechanisms by which proinsulin C-peptide prevents insulin-induced neointima formation in human saphenous vein

AIMS/HYPOTHESIS: Endothelial cells (ECs) and smooth muscle cells (SMCs) play key roles in the development of intimal hyperplasia in saphenous vein (SV) bypass grafts. In diabetic patients, insulin administration controls hyperglycaemia but cardiovascular complications remain. Insulin is synthesised as a pro-peptide, from which C-peptide is cleaved and released into the circulation with insulin; exogenous insulin lacks C-peptide. Here we investigate modulation of human SV neointima formation and SV-EC and SV-SMC function by insulin and C-peptide. METHODS: Effects of insulin and C-peptide on neointima formation (organ cultures), EC and SMC proliferation (cell counting), EC migration (scratch wound), SMC migration (Boyden chamber) and signalling (immunoblotting) were examined. A real-time RT-PCR array identified insulin-responsive genes, and results were confirmed by real-time RT-PCR. Targeted gene silencing (siRNA) was used to assess functional relevance. RESULTS: Insulin (100 nmol/l) augmented SV neointimal thickening (70% increase, 14 days), SMC proliferation (55% increase, 7 days) and migration (150% increase, 6 h); effects were abrogated by 10 nmol/l C-peptide. C-peptide did not affect insulin-induced Akt or extracellular signal-regulated kinase signalling (15 min), but array data and gene silencing implicated sterol regulatory element binding transcription factor 1 (SREBF1). Insulin (1-100 nmol/l) did not modify EC proliferation or migration, whereas 10 nmol/l C-peptide stimulated EC proliferation by 40% (5 days). CONCLUSIONS/INTERPRETATION: Our data support a causative role for insulin in human SV neointima formation with a novel counter-regulatory effect of proinsulin C-peptide. Thus, C-peptide can limit the detrimental effects of insulin on SMC function. Co-supplementing insulin therapy with C-peptide could improve therapy in insulin-treated patients

Proinsulin C-peptide elicits disaggregation of insulin resulting in enhanced physiological insulin effects

Using surface plasmon resonance (SPR) and electrospray mass spectrometry (ESI-MS), proinsulin C-peptide was found to influence insulin-insulin interactions. In SPR with chip-bound insulin, C-peptide mixed with analyte insulin increased the binding, while alone C-peptide did not. A control peptide with the same residues in random sequence had little effect. In ESI-MS, C-peptide lowered the presence of insulin hexamer. The data suggest that C-peptide promotes insulin disaggregation. Insulin/insulin oligomer μM dissociation constants were determined. Compatible with these findings, type 1 diabetic patients receiving insulin and C-peptide developed 66% more stimulation of glucose metabolism than when given insulin alone. A role of C-peptide in promoting insulin disaggregation may be important physiologically during exocytosis of pancreatic β-cell secretory granulae and pharmacologically at insulin injection sites. It is compatible with the normal co-release of C-peptide and insulin and may contribute to the beneficial effect of C-peptide and insulin replacement in type 1 diabetics

Time series analysis of the long-term hydrologic impacts of afforestation in the Agueda watershed of north-central Portugal

The north-central region of Portugal has undergone significant land cover change since the early 1900s, with large-scale replacement of natural vegetation types with plantation forests. This transition consisted of an initial conversion primarily to Pinus pinaster, followed by a secondary transition to Eucalyptus globulus. This land cover change is likely to have altered the hydrologic functioning of this region; however, these potential impacts are not fully understood. To contribute to a better understanding of the potential hydrologic impacts of this land cover change, this study examines the temporal trends in 75 years of data from the Águeda watershed (part of the Vouga Basin) over the period of 1936–2010. A number of hydrometeorological variables were analyzed using a combined Thiel–Sen/Mann–Kendall trend-testing approach, to assess the magnitude and significance of patterns in the observed data. These trend tests indicated that there have been no significant reductions in streamflow over either the entire test period, or during sub-record periods, despite the large-scale afforestation which has occurred. This lack of change in streamflow is attributed to the specific characteristics of the watershed and land cover change. By contrast, a number of significant trends were found for baseflow index, with positive trends in the early data record (primarily during Pinus pinaster afforestation), followed by negative trends later in the data record (primarily during Eucalyptus globulus afforestation). These trends are attributed to land use and vegetation impacts on streamflow generating processes, both due to species differences and to alterations in soil properties (i.e., infiltration capacity, soil water repellency). These results highlight the importance of considering both vegetation types/dynamics and watershed characteristic when assessing hydrologic impacts, in particular with respect to soil properties

Rational design of HIV vaccine and microbicides: report of the EUROPRISE annual conference

EUROPRISE is a Network of Excellence sponsored from 2007 to 2011 by the European Commission within the 6th Framework Program. The Network encompasses a wide portfolio of activities ranging from an integrated research program in the field of HIV vaccines and microbicides to training, dissemination and advocacy. The research program covers the whole pipeline of vaccine and microbicide development from discovery to early clinical trials. The Network is composed of 58 partners representing more than 65 institutions from 13 European countries; it also includes three major pharmaceutical companies (GlaxoSmithKline, Novartis and Sanofi-Pasteur) involved in HIV microbicide and vaccine research. The Network displays a dedicated and informative web page: http://www.europrise.org. Finally, a distinguishing trait of EUROPRISE is its PhD School of students from across Europe, a unique example in the world of science aimed at spreading excellence through training

Liver Fat Content in Type 2 Diabetes: Relationship With Hepatic Perfusion and Substrate Metabolism

OBJECTIVE - Hepatic steatosis is common in type 2 diabetes. It is causally linked to the features of the metabolic syndrome, liver cirrhosis, and cardiovascular disease. Experimental data have indicated that increased liver fat may impair hepatic perfusion and metabolism. The aim of the current study was to assess hepatic parenchymal perfusion, together with glucose and fatty acid metabolism, in relation to hepatic triglyceride content. RESEARCH DESIGN AND METHODS - Fifty-nine men with well controlled type 2 diabetes and 18 age-matched healthy normoglycemic men were studied using positron emission tomography to assess hepatic tissue perfusion, insulin-stimulated glucose, and fasting fatty acid metabolism, respectively, in relation to hepatic triglyceride content, quantified by proton magnetic resonance spectroscopy. Patients were divided into two groups with hepatic triglyceride content below (type 2 diabeteslow) or above (type 2 diabetes-high) the median of 8.6%. RESULTS - Type 2 diabetes-high patients had the highest BMI and A1C and lowest whole-body insulin sensitivity (ANOVA, all P < 0.001). Compared with control subjects and type 2 diabeteslow patients, type 2 diabetes-high patients had the lowest hepatic parenchymal perfusion (P = 0.004) and insulin-stimulated hepatic glucose uptake (P = 0.013). The observed decrease in hepatic fatty acid influx rate constant, however, only reached borderline significance (P = 0.088). In type 2 diabetic patients, hepatic parenchymal perfusion (r = -0.360, P = 0.007) and hepatic fatty acid influx rate constant (r = -0.407, P = 0.007) correlated inversely with hepatic triglyceride content. In a pooled analysis, hepatic fat correlated with hepatic glucose uptake (r = -0.329, P = 0.004). CONCLUSIONS - In conclusion, type 2 diabetic patients with increased hepatic triglyceride content showed decreased hepatic parenchymal perfusion and hepatic insulin mediated glucose uptake, suggesting a potential modulating effect of hepatic fat on hepatic physiology. © 2010 by the American Diabetes Association

Self Protection from Anti-Viral Responses – Ro52 Promotes Degradation of the Transcription Factor IRF7 Downstream of the Viral Toll-Like Receptors

Ro52 is a member of the TRIM family of single-protein E3 ligases and is also a target for autoantibody production in systemic lupus erythematosus and Sjögren's syndrome. We previously demonstrated a novel function of Ro52 in the ubiquitination and proteasomal degradation of IRF3 following TLR3/4 stimulation. We now present evidence that Ro52 has a similar role in regulating the stability and activity of IRF7. Endogenous immunoprecipitation of Ro52-bound proteins revealed that IRF7 associates with Ro52, an effect which increases following TLR7 and TLR9 stimulation, suggesting that Ro52 interacts with IRF7 post-pathogen recognition. Furthermore, we show that Ro52 ubiquitinates IRF7 in a dose-dependent manner, resulting in a decrease in total IRF7 expression and a subsequent decrease in IFN-α production. IRF7 stability was increased in bone marrow-derived macrophages from Ro52-deficient mice stimulated with imiquimod or CpG-B, consistent with a role for Ro52 in the negative regulation of IRF7 signalling. Taken together, these results suggest that Ro52-mediated ubiquitination promotes the degradation of IRF7 following TLR7 and TLR9 stimulation. As Ro52 is known to be IFN-inducible, this system constitutes a negative-feedback loop that acts to protect the host from the prolonged activation of the immune response