Melatonin ameliorates cardiac remodelling in fructose-induced metabolic syndrome rat model by using genes encoding cardiac potassium ion channels

Molecular Biology Reports - Metabolic syndrome comprises a group of disorders, including cardiac abnormalities. Ventricular arrhythmias observed in metabolic syndrome are due to the impaired...     

Molecular Characterization of Chitinase Genes from a Local Isolate of Serratia marcescens and Their Contribution to the Insecticidal Activity of Bacillus thuringiensis Strains

The chitinase B (chiB) and C (chiC) genes and flanking regions from a local isolate of Serratia marcescens were cloned individually and sequenced. Results showed that these chiB and chiC genes have a 96 % maximum similarity with chiB and chiC from different S. marcescens species (GenBank numbers Z36295.1 and AJ630582.1, respectively). The amplified chiB fragment, including some upstream and downstream regions, is 1,689-bp long with an open reading frame of 1,500 bp. The amplified fragment of chiC is 1,844 bp with an open reading frame of 1,443 bp. These sequences were submitted to the GenBank with accession numbers JX847796 (chiB) and JX847797 (chiC). Putative promoter regions and Shine–Dalgarno sequences were identified in both genes. The genes were cloned into a shuttle vector and the constructs were designated as pHYSB and pHYSC, respectively. Both plasmids were introduced separately into kurstaki and israelensis strains of Bacillus thuringiensis and the insecticidal activities of the engineered B. thuringiensis strains were assayed in larvae of Galleria mellonella and adult of Drosophila melanogaster. Engineered B. thuringiensis strains showed higher insecticidal activity than parental strain and the parental S. marcescens. In addition, pHYSB and pHYSC were stable over 16 daily passages under non-selective conditions in transformed B. t. israelensis 5724 strain.     

Novel nanoimaging approach: Antibodious polymeric nanolabel for intracellular alpha‐fetoprotein targeted monitoring

This study describes preparation and use of novel labeled and antibodious polymeric nanolabels (anti‐alpha fetoprotein cross‐linked nanolabels) as an immunogenic and semisynthetic nanolabel with potential prognostic and therapeutic roles for hepatoma cancer. Specificity, uptake, and binding efficiencies of the nanolabel have been examined in a human hepatosarcoma cell line HepG2, a human colorectal cell line DLD‐1, and a mouse myoblast cell line C2. Labeling of the cells has been performed by treating live and fixed cells with varying concentrations of the nanolabels and then, the cells have been examined under a fluorescence microscope. In addition, all cell lines have also been labeled using FITC‐conjugated nanotrastuzumab to compare the results obtained with those of the binding of the FITC‐nanoanti‐alpha fetoprotein nanolabels. Results show that FITC‐conjugated anti‐alpha fetoprotein cross‐linked nanolabels have been taken up by both live and fixed cells and have efficiently and specifically labeled HepG2 cells at a quite low concentration. Taken all together, the results indicate that the novel targeted nanoimaging tools and technique demonstrated their ability to detect the distribution of the nanolabels as probes in hepatoma cells. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 472–479, 2013     

The advantage of laser‐capture microdissection over whole tissue analysis in proteomic profiling studies

Laser‐capture microdissection (LCM) offers a reliable cell population enrichment tool and has been successfully coupled to MS analysis. Despite this, most proteomic studies employ whole tissue lysate (WTL) analysis in the discovery of disease biomarkers and in profiling analyses. Furthermore, the influence of tissue heterogeneity in WTL analysis, nor its impact in biomarker discovery studies have been completely elucidated. In order to address this, we compared previously obtained high resolution MS data from a cohort of 38 breast cancer tissues, of which both LCM enriched tumor epithelial cells and WTL samples were analyzed. Label‐free quantification (LFQ) analysis through MaxQuant software showed a significantly higher number of identified and quantified proteins in LCM enriched samples (3404) compared to WTLs (2837). Furthermore, WTL samples displayed a higher amount of missing data compared to LCM both at peptide and protein levels (p‐value < 0.001). 2D analysis on co‐expressed proteins revealed discrepant expression of immune system and lipid metabolisms related proteins between LCM and WTL samples. We hereby show that LCM better dissected the biology of breast tumor epithelial cells, possibly due to lower interference from surrounding tissues and highly abundant proteins. All data have been deposited in the ProteomeXchange with the dataset identifier PXD002381 ( http://proteomecentral.proteomexchange.org/dataset/PXD002381 ).     

Methylation of histone H3 lysine-79 by Dot1p plays multiple roles in the response to UV damage in Saccharomyces cerevisiae

Various proteins have been found to play roles in both the repair of UV damaged DNA and heterochromatin-mediated silencing in the yeast Saccharomyces cerevisiae. In particular, factors that are involved in the methylation of lysine-79 of histone H3 by Dot1p have been implicated in both processes, suggesting a bipartite function for this modification. We find that a dot1 null mutation and a histone H3 point mutation at lysine-79 cause increased sensitivity to UV radiation, suggesting that lysine-79 methylation is important for efficient repair of UV damage. Epistasis analysis between dot1 and various UV repair genes indicates that lysine-79 methylation plays overlapping roles within the nucleotide excision, post-replication and recombination repair pathways, as well as RAD9-mediated checkpoint function. In contrast, epistasis analysis with the H3 lysine-79 point mutation indicates that the lysine-to-glutamic acid substitution exerts specific effects within the nucleotide excision repair and post-replication repair pathways, suggesting that this allele only disrupts a subset of the functions of lysine-79 methylation. The overall results indicate the existence of distinct and separable roles of histone H3 lysine-79 methylation in the response to UV damage, potentially serving to coordinate the various repair processes.     

Sequence analysis of the conserved protamine gene cluster shows that it contains a fourth expressed gene

Structural data are presented on the protamine gene cluster (PGC) of human, mouse, rat, and bull. By restriction mapping we demonstrate that the organization of the protamine cluster is conserved throughout all four species, i.e., the genes are situated in a head to tail arrangement in the order: protamine l-protamine 2-transition protein 2. Further, we established the nucleotide sequence of the entire human PGC (25 kb in total) and the 3′ portion of the rat protamine cluster (PRM2 and TNP2 genes and intergenic region). In addition, a 1 kb fragment of the bovine and murine protamine cluster, situated between PRM2 and TNP2, was sequenced. This fragment is conserved regarding sequence, position, and orientation in all species examined, and was classified as likely coding region by gene recognition program GRAIL. Using the rat fragment as a probe in RNA blots, we detected a testis-specific signal of about 0.5 kb. Finally, we demonstrate a high density of Alu elements, both full and fragmented copies, in the human PGC and discuss their localization with respect to evolutionary and functional aspects. © 1996 Wiley-Liss, Inc.     

Paraoxonase 192 gene polymorphism and serum paraoxonase activity in high grade gliomas and meningiomas

The purpose of this study was to demonstrate the relationship between serum PON1 activity and PON 192 polymorphism in brain tumours. The distribution of PON 192 polymorphism in 42 high grade gliomas and 42 meningiomas were determined by polymerase chain reaction--based restriction fragment length polymorphism analysis and compared with 50 healthy control subjects. Serum paraoxonase1 activities were also measured and compared in the same population. We found that in both tumour groups serum PON1 activity was significantly lower than the control group (p < 0.001), but did not differ between meningiomas and high grade gliomas. There was no significant difference either in distribution of the AA, AB and BB genotypes or in the allelic frequencies, between the patient group and control subjects (p > 0.05). Our results suggest that serum PON1 as a part of the lipid peroxidation scavenging systems might be involved in the tumourigenesis of brain tumours.     

Do MCF7 cells cope with metformin treatment under energetic stress in low glucose conditions?

There is a growing body of evidence about metformin being effective in cancer therapy. Despite controversies about the ways of its effectiveness, several ongoing clinical trials are evaluating the drug when used as an adjuvant or a neo-adjuvant agent. We aimed to investigate metformin’s effects on proliferation, metastasis, and hormone receptor expressions in breast cancer cell line MCF-7 incubated in two different glucose conditions. MCF-7 cells were incubated in high or low glucose media and treated with various doses of metformin. The cell viability was studied using MTT test. The Ki-67, estrogen and progesterone receptor expression were evaluated by ICC and galectin-3 expression was evaluated by ELISA or spectrophotometrically. The cell viability following consecutive metformin doses in either glucose condition for 24 and 48 h represented a significant decrease when compared to control. The proliferation detected in low glucose medium following metformin at doses < 20 mM was found significantly decreased when compared to high glucose medium at 48 h. In terms of galectin-3 levels, the increase in high glucose medium treated with metformin and the decrease in low glucose medium were found statistically significant when compared to control. Progesterone receptor staining demonstrated a significant increase in low glucose medium. Our findings represent better outcomes for cancer lines incubated in low glucose medium treated with metformin in terms of viability, receptor expression and metastatic activity, and highlight the potential benefit of metformin especially in restraining the cancer cell’s ability to cope energetic stress in low glucose conditions.     

Co-culture of rat luteal cells with islet cells enhances islet viability and revascularization

Islet cell transplantation is a major treatment strategy for type I diabetes, and has proven to be effective for maintaining glucose homeostasis. However, this treatment requires an extended period of immunosuppression to prevent rejection and recurrent transplantation to maintain function. Thus, to enhance the properties of transplanted islet cells, we examined the effect of the co-culture of luteal cells, which secrete progesterone, on islet cell viability, functionality, and revascularization. It was found that islet viability and functionality were higher in the co-cultured group than in single cultures of islets at 48 and 96 h, in parallel with increased progesterone and vascular endothelial growth factor (VEGF) secretion from luteal cells. In the co-culture groups, VEGF levels at 48 and 96 h and CD31 levels at 48 h were significantly higher than those in the islet groups (p?<?0.001 and p?<?0.05, respectively), and basic fibroblast growth factor (bFGF) levels were increased at 96 h (p?<?0.001). Thus, co-culture with luteal cells may increase islet vascularity by enhancing VEGF and bFGF levels for up to 96 h, which could help to markedly increase the pre-transplantation time to allow for effective immunosuppression therapy. This method may also promote islet cell viability and functionality. Progesterone and angiogenic factors secreted from luteal cells may be responsible for these positive effects.