Brain Insulin Dysregulation: Implication for Neurological and Neuropsychiatric Disorders

Arduous efforts have been made in the last three decades to elucidate the role of insulin in the brain. A growing number of evidences show that insulin is involved in several physiological function of the brain such as food intake and weight control, reproduction, learning and memory, neuromodulation and neuroprotection. In addition, it is now clear that insulin and insulin disturbances particularly diabetes mellitus may contribute or in some cases play the main role in development and progression of neurodegenerative and neuropsychiatric disorders. Focusing on the molecular mechanisms, this review summarizes the recent findings on the involvement of insulin dysfunction in neurological disorders like Alzheimer’s disease, Parkinson’s disease and Huntington’s disease and also mental disorders like depression and psychosis sharing features of neuroinflammation and neurodegeneration.     

Cannabidiol inhibits the skeletal muscle Nav1.4 by blocking its pore and by altering membrane elasticity

Cannabidiol (CBD) is the primary nonpsychotropic phytocannabinoid found in Cannabis sativa, which has been proposed to be therapeutic against many conditions, including muscle spasms. Among its putative targets are voltage-gated sodium channels (Navs), which have been implicated in many conditions. We investigated the effects of CBD on Nav1.4, the skeletal muscle Nav subtype. We explored direct effects, involving physical block of the Nav pore, as well as indirect effects, involving modulation of membrane elasticity that contributes to Nav inhibition. MD simulations revealed CBD’s localization inside the membrane and effects on bilayer properties. Nuclear magnetic resonance (NMR) confirmed these results, showing CBD localizing below membrane headgroups. To determine the functional implications of these findings, we used a gramicidin-based fluorescence assay to show that CBD alters membrane elasticity or thickness, which could alter Nav function through bilayer-mediated regulation. Site-directed mutagenesis in the vicinity of the Nav1.4 pore revealed that removing the local anesthetic binding site with F1586A reduces the block of I_Na by CBD. Altering the fenestrations in the bilayer-spanning domain with Nav1.4-WWWW blocked CBD access from the membrane into the Nav1.4 pore (as judged by MD). The stabilization of inactivation, however, persisted in WWWW, which we ascribe to CBD-induced changes in membrane elasticity. To investigate the potential therapeutic value of CBD against Nav1.4 channelopathies, we used a pathogenic Nav1.4 variant, P1158S, which causes myotonia and periodic paralysis. CBD reduces excitability in both wild-type and the P1158S variant. Our in vitro and in silico results suggest that CBD may have therapeutic value against Nav1.4 hyperexcitability.     

The Effect of Antimicrobial Peptide Temporin-Ra on Cell Viability and Gene Expression of Pro-inflammatory Factors in A549 Cell Line

Antimicrobial peptide Temporin-Ra was isolated from the skin secretions of marsh frog Rana ridibunda. Temporin-Ra was chemically synthesized and purified using RP-HPLC technique. The cytotoxicity of peptide on lung airway epithelial cell line (A549) and peripheral blood mononuclear cells (PBMC) was studied by MTT assay. Furthermore, the effect of Temporin-Ra on the expression of pro-inflammatory factors such as IL-1β and IL-8 in A549 cell line was evaluated at peptide concentrations of 15, 30 and 50 μg/mL for 6, 12 and 24 h using semi-quantitative RT-PCR and real-time PCR methods. The result of our experiments revealed that Temporin-Ra decreased cell viability about 3–13 % in A549 cells and 4–6 % in PBMC cells. Moreover, Temporin-Ra induced the mRNA expression of IL-1β and IL-8 genes in a dose- and time-dependent manner. According to our results, maximum IL-8 mRNA expression was observed after a 24-h treatment of cancer cells with 50 μg/mL peptide with approximately 18-fold increase in expression. The least expression level of IL-1β was observed after 6-h of incubation in the presence of 15 μg/mL peptide with 2.5-fold increase in expression whereas the most expression level was obtained following 24 h-treatment with 50 μg/mL peptide with 26-fold increase in mRNA expression. Eventually, the present study highlights the role of Temporin-Ra as a potent antimicrobial peptide in the activation and maintenance of inflammatory processes.     

A hidden aggregation‐prone structure in the heart of hypoxia inducible factor prolyl hydroxylase

Prolyl hydroxylase domain‐containing protein 2 (PHD2), as one of the most important regulators of angiogenesis and metastasis of cancer cells, is a promising target for cancer therapy drug design. Progressive studies imply that abnormality in PHD2 function may be due to misfolding. Therefore, study of the PHD2 unfolding pathway paves the way for a better understanding of the influence of PHD2 mutations and cancer cell metabolites on the protein folding pathway. We study the unfolding of the PHD2 catalytic domain using differential scanning calorimetry (DSC), fluorescence spectroscopy, and discrete molecular dynamics simulations (DMD). Using computational and experimental techniques, we find that PHD2 undergoes four transitions along the thermal unfolding pathway. To illustrate PHD2 unfolding events in atomic detail, we utilize DMD simulations. Analysis of computational results indicates an intermediate species in the PHD2 unfolding pathway that may enhance aggregation propensity, explaining mutation‐independent PHD2 malfunction. Proteins 2016; 84:611–623. © 2016 Wiley Periodicals, Inc.     

Brachytherapy with gamma radiation of a coronary artery for in-stent restenosis may induce the regression of in-stent restenosis of an adjacent coronary artery without angioplasty. First case report and review of the literature

Here, we present a case of a 63-year-old male who presented with in-stent restenosis of two coronary arteries simultaneously (mid circumflex and proximal ramus). After the brachytherapy of the circumflex artery for in-stent restenosis, the patient refused the staged procedure for the ramus in-stent restenosis. After approximately 2 years, the patient underwent coronary angiography for recurrent chest pain. Surprisingly, the proximal ramus stent showed marked regression of in-stent restenosis. We hypothesized that the gamma brachytherapy of the circumflex artery could have induced the regression of in-stent restenosis of the adjacent ramus artery due to the deep tissue penetration of gamma radiation. Based on our observation, we believe that in the treatment of in-stent restenosis of a coronary artery, the initial balloon angioplasty may not be as important as the radiation itself. This observation warrants further study to evaluate the effect of external or internal radiation on in-stent restenosis without balloon angioplasty. If our hypothesis is confirmed, the treatment of in-stent restenosis with external radiation could substantially simplify the treatment of this disease. This case report follows a brief review of the literature.     

Antisense-inhibition of ADP-glucose pyrophosphorylase in Vicia narbonensis seeds increases soluble sugars and leads to higher water and nitrogen uptake

We previously reported on Vicia narbonensis seeds with largely decreased alpha- D-glucose-1-phosphate adenyltransferase (AGP; EC 2.7.7.27) due to antisense inhibition [H. Weber et al. (2000) Plant J 24:33-43]. In an extended biochemical analysis we show here that in transgenic seeds both AGP activity and ADP-glucose levels were strongly decreased but starch was only moderately reduced and contained less amylose. The flux control coefficient of AGP to starch accumulation was as low as 0.08, i.e. AGP exerts low control on starch biosynthesis in Vicia seeds. Mature cotyledons of antisense seeds had increased contents of lipids, nitrogen and sulfur. The protein content was higher due, in particular, to increased sulfur-rich albumins. Globulin fractions of storage proteins had a lower ratio of legumin to vicilin. Isolated cotyledons partitioned less [14C]sucrose into starch and more into soluble sugars with no change in the protein fraction. Respiration of isolated cotyledons and activities of the major glycolytic and carbohydrate-metabolizing enzymes were not affected. Sucrose and the hexose-phosphate pool were increased but UDP-glucose, 3-phosphoglyceric acid, phospho enolpyruvate, pyruvate, ATP and ADP were unchanged or even lower, indicating that carbon partitioning changed from starch to sucrose without affecting the glycolytic and respiratory pathways. Soluble compounds were increased but osmolality remained unchanged, indicating compensatory water influx resulting in higher water contents. Developmental patterns of water and nitrogen accumulation suggest a coupled uptake of amino acids and water into cotyledons. We conclude that, due to higher water uptake, transgenic cotyledons take up more amino acids, which become available for protein biosynthesis leading to a higher protein content. Obviously, a substantial part of amino acid uptake into Vicia seeds occurs passively and is osmotically controlled and driven by water influx.     

Transgenic tobacco plants expressing antisense ferredoxin-NADP(H) reductase transcripts display increased susceptibility to photo-oxidative damage

Ferredoxin-NADP(H) reductase (FNR) catalyses the final step of the photosynthetic electron transport in chloroplasts. Using an antisense RNA strategy to reduce expression of this flavoenzyme in transgenic tobacco plants, it has been demonstrated that FNR mediates a rate-limiting step of photosynthesis under both limiting and saturating light conditions. Here, we show that these FNR-deficient plants are abnormally prone to photo-oxidative injury. When grown under autotrophic conditions for 3 weeks, specimens with 20-40% extant reductase undergo leaf bleaching, lipid peroxidation and membrane damage. The magnitude of the effect was proportional to the light intensity and to the extent of FNR depletion, and was accompanied by morphological changes involving accumulation of aberrant plastids with defective thylakoid stacking. Damage was initially confined to chloroplast membranes, whereas Rubisco and other stromal proteins began to decline only after several weeks of autotrophic growth, paralleled by partial recovery of NADPH levels. Exposure of the transgenic plants to moderately high irradiation resulted in rapid loss of photosynthetic capacity and accumulation of singlet oxygen in leaves. The collected results suggest that the extensive photo-oxidative damage sustained by plants impaired in FNR expression was caused by singlet oxygen building up to toxic levels in these tissues, as a direct consequence of the over-reduction of the electron transport chain in FNR-deficient chloroplasts.     

Effect of oligosaccharides and their monosaccharide mixtures on the stability of proteins: a scaled particle study

Experimental results of RNase-A stabilization by sugar osmolytes show that the change in the Gibbs free energy (ΔGD) associated with the equilibrium, N (native) state ? D (denatured) state of the protein in the presence of equimolar mixture of monosaccharides is larger than that of the corresponding oligosaccharides at a given temperature and pH. However, at the molar scale, ΔGD obtained in the presence of an oligosaccharide is much higher as compared with ΔGD obtained using individual monosaccharide. We used scaled particle theory (SPT) to explain these experimental observations. The effective length, called Tolman's length that describes the curvature correlations to a surface area or surface tension and in turn contributes to the change in free energy, is discussed. Tolman's length is higher for corresponding monomer mixture than the oligosaccharide molecules. Based on SPT analysis, a geometrical model is proposed for clustering of monosaccharides in the mixture due to high particle density. The cluster is presumed to have weak interaction among them due to larger hydrodynamic radius than that of the bonded molecules of oligosaccharides.