Therapeutic Efficacy of an ω-3-Fatty Acid-Containing 17-β Estradiol Nano-Delivery System against Experimental Atherosclerosis

Atherosclerosis and its consequences remain prevalent clinical challenges throughout the world. Initiation and progression of atherosclerosis involves a complex, dynamic interplay among inflammation, hyperlipidemia, and endothelial dysfunction. A multicomponent treatment approach targeted for delivery within diseased vessels could prove beneficial in treating atherosclerosis. This study was undertaken to evaluate the multimodal effects of a novel ω-3-fatty acid-rich, 17-β-estradiol (17-βE)-loaded, CREKA-peptide-modified nanoemulsion system on experimental atherosclerosis. In vitro treatment of cultured human aortic endothelial cells (ECs) with the 17-βE-loaded, CREKA-peptide-modified nanoemulsion system increased cellular nitrate/nitrite, indicating improved nitric oxide formation. In vivo, systemic administration of this nanoemulsion system to apolipoprotein-E knock out (ApoE^-/-) mice fed a high-fat diet significantly improved multiple parameters related to the etiology and development of occlusive atherosclerotic vasculopathy: lesion area, circulating plasma lipid levels, and expression of aortic-wall inflammatory markers. These salutary effects were attributed selectively to the 17-βE and/or ω-3 polyunsaturated fatty acid components of the nano-delivery system. At therapeutic doses, the 17-βE-loaded, CREKA-peptide modified nanoemulsion system appeared to be biocompatible in that it elicited no apparent adverse/toxic effects, as indexed by body weight, plasma alanine aminotransferase/aspartate aminotransferase levels, and liver and kidney histopathology. The study demonstrates the therapeutic potential of a novel, 17-βE-loaded, CREKA-peptide-modified nanoemulsion system against atherosclerosis in a multimodal fashion by reducing lesion size, lowering the levels of circulating plasma lipids and decreasing the gene expression of inflammatory markers associated with the disease.     

Natural Selection Shapes Maintenance of Orthologous sRNAs in Divergent Host-Restricted Bacterial Genomes

Historically it has been difficult to study the evolution of bacterial small RNAs (sRNAs) across distantly related species. For example, identifying homologs of sRNAs is often difficult in genomes that have undergone multiple structural rearrangements. Also, some types of regulatory sRNAs evolve at rapid rates. The high degree of genomic synteny among divergent host-restricted bacterial lineages, including intracellular symbionts, is conducive to sRNA maintenance and homolog identification. In turn, symbiont genomes can provide us with novel insights into sRNA evolution. Here, we examine the sRNA expression profile of the obligate symbiont of psyllids, Carsonella ruddii, which has one of the smallest cellular genomes described. Using RNA-seq, we identified 36 and 32 antisense sRNAs (asRNAs) expressed by Carsonella from the psyllids Bactericera cockerelli (Carsonella-BC) and Diaphorina citri (Carsonella-DC), respectively. The majority of these asRNAs were associated with genes that are involved in essential amino acid biosynthetic pathways. Eleven of the asRNAs were conserved in both Carsonella lineages and the majority were maintained by selection. Notably, five of the corresponding coding sequences are also the targets of conserved asRNAs in a distantly related insect symbiont, Buchnera. We detected differential expression of two asRNAs for genes involved in arginine and leucine biosynthesis occurring between two distinct Carsonella-BC life stages. Using asRNAs identified in Carsonella, Buchnera, and Profftella which are all endosymbionts, and Escherichia coli, we determined that regions upstream of these asRNAs encode unique conserved patterns of AT/GC richness, GC skew, and sequence motifs which may be involved in asRNA regulation.     

In silico model of DSF synthase RpfF protein from Xanthomonas oryzae pv. Oryzae: a novel target for bacterial blight of rice disease

BACKGROUND: Rice plant diseases play a major role as biological constraints on production. One of such rice disease is bacterial leaf blight, caused by Xanthomonas oryzae pv. Oryzae (Xoo). The diffusible signal factor (DSF) synthesized by Xoo has a major role in virulence to rice plant. The DSF synthase RpfF protein, which is related to crotonase superfamily is responsible for the maintaining concentration of DSF. DSF-dependent quorum sensing (QS) system adopts protein- protein interaction mechanism to auto regulates the production of DSF. The antibacterial activity of pesticides against Xoo has not yet been completely understood. Three dimensional structure of RpfF protein was predicted using homology modeling method by MODELLER 9V9 software, SWISS MODEL and GENO3D online tools and structures were validated by Ramachandran plot, TM-Score and RMSD. 3D structure of RpfF (accession number AAL06345) was predicted using DSF synthase of Xanthomonas campestris pv. campestris (Xcc) (PDB ID: 3M6M) as a template. The stereo chemical check reveals the structure developed from the modeller was the best one and the potential ligand binding sites were identified by CASTp Server. The predicted RpfF model provides insight into its structure, active sites and aid in the development of novel inhibitors to control bacterial leaf blight in rice plant. DSF synthase RpfF protein could be used as a novel target to control infection.     

Comparison of extraction methods for the comprehensive analysis of mouse brain proteome using shotgun-based mass spectrometry

This study compares 16 different extraction methods for the comprehensive extraction of mouse brain proteome in combination with "shotgun"-based mass spectrometry (MS). Membrane proteins (MPs) are responsible for a large part of the regulatory functions of the cell and are therefore of great interest to extract and analyze. Sixteen protein extraction protocols were evaluated in regards to protein yield and number of identified proteins with emphasis on MPs. The extracted proteins were delipidated, on-filter digested, and analyzed by reversed phase nanoliquid chromatography (RP-nanoLC) in combination with electrospray ionization (ESI) tandem mass spectrometry (MS/MS) using a 7 T hybrid LTQ-FT mass spectrometer. Detergent-based lysis buffers showed higher efficiencies and yields in the extraction of proteins from the brain tissue compared to solubilization with organic solvents or organic acids. The detergent octyl-β-D-glucopyranoside gave the highest number of identified proteins (541) as well as numbers and percentages of identified MPs (29%). Detergent-based protocols are the best sample preparation tools for central nervous system (CNS) tissue and can readily be applied to screen for candidate biomarkers of neurological diseases.     

Elucidation of conformational diversity of druggable enzymes and classification of chemical modulators based on inhibitor-bound structures

Communicated by Ramaswamy H. Sarma     

Protective effect of ellagic acid against TCDD-induced renal oxidative stress: Modulation of CYP1A1 activity and antioxidant defense mechanisms

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) belongs to toxicologically important class of poly halogenated aromatic hydrocarbons and produce wide variety of adverse effects in humans. The present study investigated the protective effect of ellagic acid, a natural polyphenolic compound against TCDD-induced nephrotoxicity in Wistar rats. TCDD-induced nephrotoxicity was reflected in marked changes in the histology of kidney, increase in levels of kidney markers (serum urea, serum creatinine) and lipid peroxides. A significant increase in activity of phase I enzyme CYP1A1 with concomitant decline in the activities of phase II enzymes [non-enzymic antioxidant and various enzymic antioxidants such as superoxide dismutase, catalase, glutathione peroxidase, glutathione-s-transferase] was also observed. In addition, TCDD treated rats showed alterations in ATPase enzyme activities such as Na⁺ K⁺-ATPase, Mg²⁺ ATPase and Ca²⁺ ATPase. Oral pre-treatment with ellagic acid prevented TCDD-induced alterations in levels of kidney markers. Ellagic acid pre-treatment significantly counteracted TCDD-induced oxidative stress by decreasing CYP1A1 activity and enhancing the antioxidant status. Furthermore, ellagic acid restored TCDD-induced histopathological changes and alterations in ATPase enzyme activities. The results of the present study show that significant protective effect rendered by ellagic acid against TCDD-induced nephrotoxicity might be attributed to its antioxidant potential.     

Phytochemical Analysis,Estimation of Quercetin,and <i>in Vitro</i> Anti-Diabetic Potential of Stevia Leaves Samples Procured from Two Geographical Origins

The current study used RP-HPLC to compare phytochemicals and estimate the bioactive constituents found in Stevia rebaudiana Bert. (SRB) leaves collected from two different geographical sources. SRB leaves were collected from Bangalore, Karnataka, India, and Reduit, Mauritius. Extracts were prepared using ethanol and aqueous solvents. Proximate analysis was used to evaluate moisture content, ash values, crude fibers, and extractive values. Following that, preliminary phytochemical screening was done on both ethanol leaves extracts, and subsequently total flavonoid content was determined. In addition, TLC chromatograms and RP-HPLC studies were performed on both plant extracts to determine the presence of flavonoid components in both leaves extracts, followed by in vitro anti-diabetic activity was performed with alpha amylase and alpha glucosidase enzymes against Acarbose as standard. Results revealed that both the extracts from two different geographical sources varied significantly with the yield, content of chemicals, and presence of quercetin (flavonoid) content when estimated through the RP-HPLC standardized method. Glycosides, flavonoids, proteins, steroids, saponins, terpenoids, and phenols were found in various concentrations during phytochemical screening. Among both zones, the ethanol leaves extract of SRB taken from Mauritius had a greater content of phytochemicals and a higher yield than other extracts due to the soil nature. The Mauritius sample had greater total flavonoid levels as well as more quercetin (0.92 ± 0.011) than the other extracts. Following that, ethanol extract inhibited enzymes (alpha amylase, alpha glucosidases) more than aqueous extract, and this inhibition was dose dependent. Among them, the Mauritius ethanol sample showed higher anti-diabetic efficacy than the Indian sample, but this difference was not significant. Overall, SRB ethanol leaves extracts outperformed other leaves extracts in terms of yield, phytoconstituents, and total flavonoids. Overall, both SRB samples had high quercetin levels and possessed anti-diabetic potential, but they were greater in the Mauritius sample, demonstrating that plant traits are influenced by geographic location.     

Targeting insulin amyloid assembly by aminosugars and their derivatives

The use of small carbohydrates that stabilize proteins from misfolding is important from pharmaceutical point of view. We have investigated the role of small isomeric amino sugars on the in vitro aggregation of insulin amyloid. Using mass spectrometry, we screened 6 isomeric aminosugars for their role on inhibition of insulin amyloid formation and the results were compared with transmission electron microscopy imaging. We found that three N-acetylamino sugars promote insulin fibril formation. Among three isomeric aminosugars studied, only galactosamine showed few fibrils whereas other two isomers showed enhanced fibrils. The results demonstrated here may contribute to future designing of small amine derivatised galactose sugars as amyloid inhibitors and understanding their action.     

A database of six eukaryotic hypothetical genes and proteins

Assigning functions to proteins of unknown function is of considerable interest to the proteomic researchers as the genes encoding them are conserved over various species. Here, we describe HypoDB, a database of hypothetical genes and proteins in six eukaryotes. The database was collected and organized based on the number of entries in each chromosome with few annotations. Hypothetical protein database contains information related to gene and protein sequences, chromosome number and location, secondary and tertiary structure related data. AVAILABILITY: The database is available for free at http://www.trimslabs.com/database/hypodb/index.html.