<Emphasis Type="Italic">Trans</Emphasis>-cinnamaldehyde Modulates Hippocampal Nrf2 Factor and Inhibits Amyloid Beta Aggregation in LPS-Induced Neuroinflammation Mouse Model

Trans-cinnamaldehyde (CNM) has recently drawn attention due to its potent anti-inflammatory and antioxidant properties. The current study explored the memory enhancing effects of CNM against lipopolysaccharide (LPS)-induced neuroinflammation in mice. CNM and curcumin (a reference antioxidant) were administered at a dose of 50 mg/kg i.p. 3 h after a single LPS injection (0.8 mg/kg, i.p.) and continued daily for 7 days. Our results displayed that CNM and curcumin significantly ameliorated the LPS-induced impairment of learning and memory, neuroinflammation, oxidative stress and neuronal apoptosis. Memory functions and locomotor activity were assessed by Morris water maze, object recognition test and open field test. Both CNM and curcumin activated the nuclear factor erythroid 2 related factor 2 (Nrf2) and restored levels of downstream antioxidant enzymes superoxide dismutase and glutathione-S-transferase (GST) in the hippocampus. They also attenuated LPS-induced increase in hippocampal contents of interleukin-1β (IL-1β), malondialdehyde and caspase-3. Immunohistochemistry results showed that both CNM and curcumin reduced Aβ_1–42 protein accumulation in brain of mice. Remarkably CNM’s effect on IL-1β was less pronounced than curcumin; however it showed higher GST activity and more potent anti-apoptotic and anti-amylodogenic effect. We conclude that, CNM produces its memory enhancing effects through modulation of Nrf2 antioxidant defense in hippocampus, inhibition of neuroinflammation, apoptosis and amyloid protein burden.     

Comparative Modeling and Evaluation of Leukotriene B4 Receptors for Selective Drug Discovery Towards the Treatment of Inflammatory Diseases

Leukotriene B4 (LTB4) exerts its biological effects through stimulation of specific G protein-coupled receptors (GPCRs)—namely BLT1 and BLT2. Due to the absence of human BLT1 and BLT2 crystal structures, the current study was set to predict the 3D structures of these two receptors for structure-based anti-inflammatory drug discovery. Homology modeling of the BLT1 receptor was first constructed, based on various X-ray and NMR GPCR templates, followed by molecular dynamics (MD) refinement. Using a single-template approach, nine well-established alignment methods and ten secondary structure prediction methods during the backbone generation were implemented and assessed. The binding sites of the BLT1 receptor were then mapped using fifteen chemical probes with the help of FTMAP and AutoDock Vina 4.2 software. Model validation was performed through the docking of eight specific antagonists that have experimental inhibition constants (k_i) towards BLT1. The antagonists-BLT1 docked structures were then subjected to AMBER-based molecular mechanical minimization and the corresponding binding energies were calculated using molecular mechanics–generalized Born surface area (MM/GBSA) approach. According to the results, the most energetically stable models were constructed using SAlign method for the alignment process and PSIPRED for secondary structure prediction. In comparison, the refined BLT1 model built on 2KS9 as an NMR template has the lowest DOPE energy compared to those built on 4EA3 and 4XT1 as X-ray templates. According to the mapping results, two main binding sites were identified: one was among TMs II, III and VII and the other was among TMs III, IV and V. For the antagonists, correlation between binding energies and experimental data was in a good agreement, with a correlation coefficient (R² value) of 0.91. Due to the great amino acid sequence similarity between BLT1 and BLT2 receptors (calculated as 45.2%), BLT2 model was constructed based on the predicted BLT1 model.     

Peripheral adipose tissue insulin resistance alters lipid composition and function of hippocampal synapses

Compelling evidence indicates that type 2 diabetes mellitus, insulin resistance (IR), and metabolic syndrome are often accompanied by cognitive impairment. However, the mechanistic link between these metabolic abnormalities and CNS dysfunction requires further investigations. Here, we evaluated whether adipose tissue IR and related metabolic alterations resulted in CNS changes by studying synapse lipid composition and function in the adipocyte‐specific ecto‐nucleotide pyrophosphate phosphodiesterase over‐expressing transgenic (AtENPP1‐Tg) mouse, a model characterized by white adipocyte IR, systemic IR, and ectopic fat deposition. When fed a high‐fat diet, AtENPP1‐Tg mice recapitulate essential features of the human metabolic syndrome, making them an ideal model to characterize peripherally induced CNS deficits. Using a combination of gas chromatography and western blot analysis, we found evidence of altered lipid composition, including decreased phospholipids and increased triglycerides (TG) and free fatty acid in hippocampal synaptosomes isolated from high‐fat diet‐fed AtENPP1‐Tg mice. These changes were associated with impaired basal synaptic transmission at the Schaffer collaterals to hippocampal cornu ammonis 1 (CA1) synapses, decreased phosphorylation of the GluN1 glutamate receptor subunit, down‐regulation of insulin receptor expression, and up‐regulation of the free fatty acid receptor 1.

     

Modulator of Apoptosis 1 (MOAP-1) Is a Tumor Suppressor Protein Linked to the RASSF1A Protein

Modulator of apoptosis 1 (MOAP-1) is a BH3-like protein that plays key roles in cell death or apoptosis. It is an integral partner to the tumor suppressor protein, Ras association domain family 1A (RASSF1A), and functions to activate the Bcl-2 family pro-apoptotic protein Bax. Although RASSF1A is now considered a bona fide tumor suppressor protein, the role of MOAP-1 as a tumor suppressor protein has yet to be determined. In this study, we present several lines of evidence from cancer databases, immunoblotting of cancer cells, proliferation, and xenograft assays as well as DNA microarray analysis to demonstrate the role of MOAP-1 as a tumor suppressor protein. Frequent loss of MOAP-1 expression, in at least some cancers, appears to be attributed to mRNA down-regulation and the rapid proteasomal degradation of MOAP-1 that could be reversed utilizing the proteasome inhibitor MG132. Overexpression of MOAP-1 in several cancer cell lines resulted in reduced tumorigenesis and up-regulation of genes involved in cancer regulatory pathways that include apoptosis (p53, Fas, and MST1), DNA damage control (poly(ADP)-ribose polymerase and ataxia telangiectasia mutated), those within the cell metabolism (IR-α, IR-β, and AMP-activated protein kinase), and a stabilizing effect on microtubules. The loss of RASSF1A (an upstream regulator of MOAP-1) is one of the earliest detectable epigenetically silenced tumor suppressor proteins in cancer, and we speculate that the additional loss of function of MOAP-1 may be a second hit to functionally compromise the RASSF1A/MOAP-1 death receptor-dependent pathway and drive tumorigenesis.     

Sensitive determination of trimetazidine in spiked human plasma by HPLC with fluorescence detection after pre-column derivatization with 9-fluorenylmethyl chloroformate

A high-performance liquid chromatographic method for the determination of trimetazidine dihydrochloride (TMZ) in spiked human plasma is described. The method is based on the pre-column derivatization with 9-fluorenylmethyl chloroformate (FMOC-Cl) using the fluorimetric detection technique. Fluoxetine HCl (FLX) was used as internal standard. Both, TMZ and FLX were completely derivatized after heating at 50 degrees C for 20 min in borate buffer pH 8.0. Samples were analyzed by high performance liquid chromatography (HPLC) using Zorbax-TMS column (250 mm x 4.6 mm, i.d., 5 microm) and mobile phase consist of acetonitrile, methanol and 20 mM sodium acetate pH 4.7 (44:6:50; v/v/v). Fluorescence detector (FLD) was adjusted at excitation and emission wavelengths; 265 and 311 nm, respectively. The linearity of the method was in the range of 4.5-200 ng/ml. Limits of detection (LOD) and quantification (LOQ) were 1.5 and 4.5 ng/ml, respectively. Trimetazidine recovery was 96.5+/-1.3% (n=6; RSD=2.1%).     

Discovery of novel thienoquinoline-2-carboxamide chalcone derivatives as antiproliferative EGFR tyrosine kinase inhibitors

Novel thienoquinoline carboxamide-chalcone derivatives were prepared via the cyclization of acylated chalcones and 2-mercaptoquinoline-3-carbaldehyde in DMF with K₂CO₃. Thienoquinolines 9a–f, h exhibited promising antiproliferative effect against all the tested cell lines and gave a significant activity as EGFR inhibitors, with IC₅₀ values ranging from 0.5 and 3.2?µM, and compounds 9e and 9f being the most active of the series. They also showed better activity than Erlotinib against melanoma cancer cell line A375. Moreover, compound 9f influenced pre G1 apoptosis and cell cycle arrest at G2/M phase. The binding mode of the best EGFR inhibitor 9e in the EGFR active site revealed that the thienoquinoline ring occupied the ATP-binding site while the chalcone moiety is located in the allosteric site and is responsible for the enhanced activity of these compounds.     

Effect of spdC gene expression on virulence and antibiotic resistance in clinical Staphylococcus aureus isolates

International Microbiology - Surface protein display C (SpdC) protein was described as a novel virulence factor of Staphylococcus aureus that affects biofilm formation and pathogenesis and favors...     

Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites

A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10%) were used without daily replacement of the medium. The results of a high-throughput screening assay revealed that the best HCT was 2.5% for bovine Babesia parasites and 5% for equine Babesia and Theileria parasites. The IC₅₀ values of diminazene aceturate obtained by fluorescence and microscopy did not differ significantly. Likewise, the IC₅₀ values of luteolin, pyronaridine tetraphosphate, nimbolide, gedunin, and enoxacin did not differ between the two methods. In conclusion, our fluorescence-based assay uses low HCT and does not require daily replacement of culture medium, making it highly suitable for in vitro large-scale drug screening against Babesia and Theileria parasites that infect cattle and horses.     

Multigene Panel Sequencing Reveals Cancer-Specific and Common Somatic Mutations in Colorectal Cancer Patients: An Egyptian Experience

This study aims at identifying common pathogenic somatic mutations at different stages of colorectal carcinogenesis in Egyptian patients. Our cohort included colonoscopic biopsies collected from 120 patients: 20 biopsies from patients with inflammatory bowel disease, 38 from colonic polyp patients, and 62 from patients with colorectal cancer. On top of this, the cohort included 20 biopsies from patients with non-specific mild to moderated colitis. Targeted DNA sequencing using a customized gene panel of 96 colorectal related genes running on the Ion Torrent NGS technology was used to process the samples. Our results revealed that 69% of all cases harbored at least one somatic mutation. Fifty-seven genes were found to carry 232 somatic non-synonymous variants. The most frequently pathogenic somatic mutations were localized in TP53, APC, KRAS, and PIK3CA. In total, 16 somatic mutations were detected in the CRC group and in either the IBD or CP group. In addition, our data showed that 51% of total somatic variants were CRC-specific variants. The average number of CRC-specific variants per sample is 2.4. The top genes carrying CRC-specific mutations are APC, TP53, PIK3CA, FBXW7, ATM, and SMAD4. It seems obvious that TP53 and APC genes were the most affected genes with somatic mutations in all groups. Of interest, 85% and 28% of the APC and TP53 deleterious somatic mutations were located in Exon 14 and Exon 3, respectively. Besides, 37% and 28% of the total somatic mutations identified in APC and TP53 were CRC-specific variants, respectively. Moreover, we identified that, in 29 somatic mutations in 21 genes, their association with CRC patients was unprecedented. Ten detected variants were likely to be novel: six in PIK3CA and four variants in FBXW7. The detected P53, Wnt/βcatenin, Angiogenesis, EGFR, TGF-β and Interleukin signaling pathways were the most altered pathways in 22%, 16%, 12%, 10%, 9% and 9% of the CRC patients, respectively. These results would contribute to a better understanding of the colorectal cancer and in introducing personalized therapies for Egyptian CRC patients.