The Role of Differentially Expressed miRNAs and Potential miRNA-mRNA Regulatory Network in Prostate Cancer Progression and Metastasis

Purpose: Aberrant expression of microRNAs (miRNAs) has been discovered in prostate cancer progression however their function is not well understood, thereby further investigation is required to understand the importance of underlying mechanisms and their involvement in multiple signaling pathways, as well as their potential as therapeutic targets. In this study the role and expression levels of three miRNAs were evaluated: miR-21, miR-221 and miR-200c in different prostate cancer cell lines. In addition, based on the latest studies on miRNAs function, their association with other target genes and molecules were analyzed using bioinformatic tools. Methods: Three PCa cell lines PC3, LNCaP and VCaP and normal prostate epithelial cell line PNT1A were screened for miRNA expression levels using qPCR. miRNA target genes and their association with signaling pathways were analyzed through several Network and pathway analysis online tools. Findings: Upregulation of miR-21 and miR-221 was observed in PC3 and VCaP prostate cancer cells, respectively. According to KEGG analysis, we found that Hippo signaling pathway and cytokine-cytokine receptor interactions were affected by miR-21 while miR-221 would interfere with ECM-receptor interaction, Fatty acid elongation and Huntington disease molecular networks. Exposure of PC3 cells to TGF-β (10 µM) caused upregulation of miR-21 with the evidence with increased invasion potential. Discussion and Conclusion: miRNAs could regulate several genes in multiple signaling pathways. Here, we demonstrated that in a panel of PCa cell lines, both mir-21 and miR-221 expressions were upregulated. miR-21 may be a dignostic and prognosticbiomarker for PCa

Expression of Trichoderma reesei β-Mannanase in Tobacco Chloroplasts and Its Utilization in Lignocellulosic Woody Biomass Hydrolysis

Lignocellulosic ethanol offers a promising alternative to conventional fossil fuels. One among the major limitations in the lignocellulosic biomass hydrolysis is unavailability of efficient and environmentally biomass degrading technologies. Plant-based production of these enzymes on large scale offers a cost-effective solution. Cellulases, hemicellulases including mannanases and other accessory enzymes are required for conversion of lignocellulosic biomass into fermentable sugars. β-mannanase catalyzes endo-hydrolysis of the mannan backbone, a major constituent of woody biomass. In this study, the man1 gene encoding β-mannanase was isolated from Trichoderma reesei and expressed via the chloroplast genome. PCR and Southern hybridization analysis confirmed site-specific transgene integration into the tobacco chloroplast genomes and homoplasmy. Transplastomic plants were fertile and set viable seeds. Germination of seeds in the selection medium showed inheritance of transgenes into the progeny without any Mendelian segregation. Expression of endo-β-mannanase for the first time in plants facilitated its characterization for use in enhanced lignocellulosic biomass hydrolysis. Gel diffusion assay for endo-β-mannanase showed the zone of clearance confirming functionality of chloroplast-derived mannanase. Endo-β-mannanase expression levels reached up to 25 units per gram of leaf (fresh weight). Chloroplast-derived mannanase had higher temperature stability (40°C to 70°C) and wider pH optima (pH 3.0 to 7.0) than E.coli enzyme extracts. Plant crude extracts showed 6–7 fold higher enzyme activity than E.coli extracts due to the formation of disulfide bonds in chloroplasts, thereby facilitating their direct utilization in enzyme cocktails without any purification. Chloroplast-derived mannanase when added to the enzyme cocktail containing a combination of different plant-derived enzymes yielded 20% more glucose equivalents from pinewood than the cocktail without mannanase. Our results demonstrate that chloroplast-derived mannanase is an important component of enzymatic cocktail for woody biomass hydrolysis and should provide a cost-effective solution for its diverse applications in the biofuel, paper, oil, pharmaceutical, coffee and detergent industries

Withania somnifera Root Extract Enhances Chemotherapy through ‘Priming’

Withania somnifera extracts are known for their anti-cancerous, anti-inflammatory and antioxidative properties. One of their mechanisms of actions is to modulate mitochondrial function through increasing oxidative stress. Recently ‘priming’ has been suggested as a potential mechanism for enhancing cancer cell death. In this study we demonstrate that ‘priming’, in HT-29 colon cells, with W. somnifera root extract increased the potency of the chemotherapeutic agent cisplatin. We have also showed the W. somnifera root extract enhanced mitochondrial dysfunction and that the underlying mechanism of ‘priming’ was selectively through increased ROS. Moreover, we showed that this effect was not seen in non-cancerous cells

Disruption of Yarrowia lipolytica TPS1 Gene Encoding Trehalose-6-P Synthase Does Not Affect Growth in Glucose but Impairs Growth at High Temperature

We have cloned the Yarrowia lipolytica TPS1 gene encoding trehalose-6-P synthase by complementation of the lack of growth in glucose of a Saccharomyces cerevisiae tps1 mutant. Disruption of YlTPS1 could only be achieved with a cassette placed in the 3′half of its coding region due to the overlap of its sequence with the promoter of the essential gene YlTFC1. The Yltps1 mutant grew in glucose although the Y. lipolytica hexokinase is extremely sensitive to inhibition by trehalose-6-P. The presence of a glucokinase, insensitive to trehalose-6-P, that constitutes about 80% of the glucose phosphorylating capacity during growth in glucose may account for the growth phenotype. Trehalose content was below 1 nmol/mg dry weight in Y. lipolytica, but it increased in strains expressing YlTPS1 under the control of the YlTEF1promoter or with a disruption of YALI0D15598 encoding a putative trehalase. mRNA levels of YlTPS1 were low and did not respond to thermal stresses, but that of YlTPS2 (YALI0D14476) and YlTPS3 (YALI0E31086) increased 4 and 6 times, repectively, by heat treatment. Disruption of YlTPS1 drastically slowed growth at 35°C. Homozygous Yltps1 diploids showed a decreased sporulation frequency that was ascribed to the low level of YALI0D20966 mRNA an homolog of the S. cerevisiae MCK1 which encodes a protein kinase that activates early meiotic gene expression

Injectable Calcium Phosphate Cement as a Bone-Graft Material Around Peri-implant Dehiscence Defects: A Dog Study Source.

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Raman micro-spectroscopic investigation of the interaction of cultured HCT116 colon cancer cells with alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase

The interaction of cultured colon cancer cells with alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, has been investigated, using Raman micro-spectroscopy, in order to investigate DFMO induced effects. Raman spectra of the cultured HCT116 colon cancer cells treated with DFMO at different concentrations (0, 1, 2.5, 5, and 7.5 mM) were recorded in the range 550-2300 cm(-1). It has been shown that second derivative profile of the raw Raman spectrum of the colon cancer cells (i.e., the original experimental spectrum without any computational corrections) discriminates the weak but sharp bands from the strong, broad fluorescence background, and gives information about the position of the peaks and their band widths. The relative integrated intensities of the 781 cm(-1) and 788 cm(-1) DNA/RNA marker bands to that of 1451 cm(-1) band are found to decrease by addition of DFMO. Up to 65% reduction in the magnitude of the 1003 cm(-1) band, the characteristic phenylalanine ring breathing mode, in comparison 10 that of 1451 band, is observed. The results indicate DFMO induced apoptosis. On the other hand the intensity ratio of the tyrosine Fermi doubled around 830 cm(-1) and 850 cm(-1), which is a marker of hydrogen-bonding state of phenolic OH, is changed. The addition of DFMO may alter the tyrosine environment in cells, and parts of tyrosine residues are exposed. We also observed some modifications in amide I band, pointing out the alterations of the secondary structure of cell proteins by the presence of DFMO. (C) 2011 Elsevier B.V. All rights reserved

Raman micro-spectroscopic analysis of cultured HCT116 colon cancer cells in the presence of roscovitine

Raman micro-spectroscopic analysis of cultured HCT116 colon cancer cells in the presence of roscovitine, [seliciclib, 2-(1-ethyl-2-hydroxy-ethylamino)-6-benzylamino-9-isopropylpurine], a promising drug candidate in cancer therapy, has been performed for the first time. The aim of this study was to investigate modulations in colon cancer cells induced by roscovitine. Raman spectra of the cultured HCT116 colon cancer cells treated with roscovitine at different concentrations (0,5,10, 25 and 50 mu M) were recorded in the range 400-1850 cm(-1). It was shown that the second derivative profile of the experimental spectrum gives valuable information about the wavenumbers and band widths of the vibrational modes of cell components, and it eliminates the appearance of false peaks arising from incorrect baseline corrections. In samples containing roscovitine, significant spectral changes were observed in the intensities of characteristic protein and DNA bands, which indicate roscovitine-induced apoptosis. Roscovitine-induced apoptosis was also assessed by flow cytometry analysis, and analysis of propidium iodide staining. We observed some modifications in amide I and III bands, which arise from alterations in the secondary structure of cell proteins caused by the presence of roscovitine. (C) 2011 Elsevier B.V. All rights reserved