Anti-inflammatory effect of SGLT-2 inhibitors via uric acid and insulin.

Sodium-glucose cotransporter 2 (SGLT-2) inhibitors (i) reduce cardiovascular and renal events in patients with and without type 2 diabetes (T2D). However, the underlying mechanisms are debated. Low-grade inflammation (LGI) is a key driver of vascular complications, suggested to be attenuated by SGLT-2i in animal models. Based on a specific working hypothesis, here we investigated the net effect of SGLT-2i on LGI in patients with T2D and the possible underlying mechanism. We enrolled patients with T2D treated either with a stable therapy with SGLT-2i or with other glucose-lowering drugs (GLD) (n = 43 per group after matching for a range of pro-inflammatory variables), and tested hs-CRP and interleukin (IL)-6 as primary variables of interest. Patients treated with SGLT-2i had lower circulating levels of IL-6, a prototypical marker of LGI, but also of uric acid and fasting insulin, compared with patients treated with other GLD. Then, to explore whether uric acid and insulin might mediate the effect of SGLT-2i on IL-6, we tested physiologically pertinent doses of these two molecules (i.e. 0.5 mM uric acid and 1 nM insulin) in two in vitro models of LGI, i.e. monocytes (THP-1) treated with LPS and endothelial cells (HUVEC) exposed to hyperglycaemia. Results from in vitro models supported a pro-inflammatory role for uric acid and its combination with insulin in monocytes and for uric acid alone in hyperglycaemia-stimulated endothelial cells. On the contrary, we observed no drug-intrinsic, anti-inflammatory effect for dapagliflozin, empagliflozin, and canagliflozin in the same models. Overall, these results suggest that SGLT-2i possess a tangible activity against LGI, an effect possibly mediated by their ability to lower uric acid and insulin concentrations and that juxtaposes other proposed mechanisms in explaining the observed benefit of this class on cardiovascular and renal endpoints.

EBP1 is a nucleolar growth-regulating protein that is part of pre-ribosomal ribonucleoprotein complexes.

EBP1 was identified as a protein that interacts with the ErbB-3 receptor and possibly contributes to transducing growth regulatory signals. The existence of EBP1 homologs across species from simple eukaryotes to humans and its wide tissue expression pattern suggest that EBP1 acts as a general signaling molecule. We provide evidence that EBP1 is localized to the cytoplasm and to the nucleolus, and that its nucleolar localization requires amino-acid sequences present at both the amino- and carboxy-terminus of the molecule. We also show that EBP1 overexpression inhibits proliferation of human fibroblasts, and that this effect is linked to its nucleolar localization. Using mass spectrometry we demonstrate that EBP1 is part of ribonucleoprotein complexes and associates with different rRNA species. It is becoming clear that cell growth and proliferation are actively coordinated with rRNA processing and ribosome assembly. Our findings indicate that EBP1 is a nucleolar growth-regulating protein, and we propose that it could represent a new link between ribosome biosynthesis and cell proliferation.

Effect of chromogranin A-derived vasostatin-1 on laser-induced choroidal neovascularization in the mouse.

PURPOSE: To verify the effect of vasostatin-1 (VS-1), an anti-angiogenic fragment of chromogranin A, in the prevention of choroidal neovascularization (CNV) in an established mouse model of laser-induced ocular neovascularization. METHODS: Bruch's membrane, the innermost layer of the choroid, was broken by laser photocoagulation in C57/Bl6 mice, to induce CNV. Mice were then treated daily for 14 days by intraperitoneal injection of VS-1 or vehicle (6 mice/group). CNV and vascular leakage were measured at three time-points (day 0, 7 and 14) in vivo by spectral domain optical coherence tomography (OCT) and fluorescein angiography (FA). Ex vivo analysis of CNV was also performed at day 14 by confocal microscopy analysis of dextran-perfused choroidal flat-mounts. RESULTS: In vivo analyses showed that VS-1 significantly reduced CNV at day 14 (p = 0.03) and vascular leakage at day 7 (p = 0.01) and 14 (p = 0.04). Ex vivo confocal microscopy analysis of CNV performed on dextran-perfused choroidal flat-mounts at day 14 confirmed the protective activity of VS-1 (p = 0.01). A significant correlation between the results of in vivo and ex vivo analyses of CNV was also observed (p = 0.001, R(2) = 0.81). CONCLUSION: The results indicate that VS-1 can prevent CNV and vascular leakage in a mouse model of ocular neovascularization, suggesting that this polypeptide might have therapeutic activity in human ocular diseases that are complicated by neovascularization or excessive vascular permeability.

Cyclic RGD-containing functionalized azabicycloalkane peptides as potent integrin antagonists for tumor targeting.

Cyclic RGD-containing functionalized azabicycloalkane peptides were synthesized with the aim of developing high-affinity selective integrin ligands as carriers for therapeutic and diagnostic purposes. Herein we describe the synthesis and in vitro screening of these RGD derivatives, as well as the determination of their conformational properties in solution by spectroscopic and computational methods. Docking studies with the X-ray crystal structure of the extracellular domain of integrin alpha(v)beta(3) were also performed to elucidate the structural binding requirements and to rationalize the biological results. One compound in particular was found to be the best alpha(v)beta(3) integrin binder (IC(50)=53.7 nM) among the new functionalized RGD cyclic peptides, thus emerging as a promising candidate for covalent bonding and selective homing of useful functional units.

Ebp1 is a dsRNA-binding protein associated with ribosomes that modulates eIF2alpha phosphorylation.

dsRNA-binding domains (dsRBDs) characterize an expanding family of proteins involved in different cellular processes, ranging from RNA editing and processing to translational control. Here we present evidence that Ebp1, a cell growth regulating protein that is part of ribonucleoprotein (RNP) complexes, contains a dsRBD and that this domain mediates its interaction with dsRNA. Deletion of Ebp1's dsRBD impairs its localization to the nucleolus and its ability to form RNP complexes. We show that in the cytoplasm, Ebp1 is associated with mature ribosomes and that it is able to inhibit the phosphorylation of serine 51 in the eukaryotic initiation factor 2 alpha (eIF2alpha). In response to various cellular stress, eIF2alpha is phosphorylated by distinct protein kinases (PKR, PERK, GCN2, and HRI), and this event results in protein translation shut-down. Ebp1 overexpression in HeLa cells is able to protect eIF2alpha from phosphorylation at steady state and also in response to various treatments. We demonstrate that Ebp1 interacts with and is phosphorylated by the PKR protein kinase. Our results demonstrate that Ebp1 is a new dsRNA-binding protein that acts as a cellular inhibitor of eIF2alpha phosphorylation suggesting that it could be involved in protein translation control.