Human periodontal ligament fibroblasts (PdLFs) were treated with 1-nonadecene, a metabolite from radicular cysts, in comparison to the untreated and positive controls
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ABSTRACT: Human PdLFs (iCell Bioscience, China) were treated with 1µM, 10µM, and 100µM 1-nonadecene (TCI, US-Japan). PdLFs were treated daily with fresh media containing 1-nonadecene for 2- and 6-days. Phorbol 12-myristate 13-acetate (PMA) (ab120297, Abcam, UK) at 1µM was used as an inflammatory inducer, a positive control for the expression and synthesis of MMP-1, N-cadherin, and as a negative control for the expression of E-cadherin. Untreated PdLFs were regarded as control.
Project description:Human PdLFs (iCell Bioscience, China) were treated with 1µM, 10µM, and 100µM L-lactic acid (Sigma-Aldrich, Germany). Phorbol 12-myristate 13-acetate (PMA) (ab120297, Abcam, UK) at 1µM was used as an inflammatory inducer, a positive control for the expression and synthesis of collagen and MMP-1. Untreated PdLFs were regarded as control. PdLFs were incubated at 37°C and 5% CO2 for 2 days and 6 days.
Project description:Recently, L-lactic acid was identified as a unique metabolite in periapical granulomas. However, the biological roles of this metabolite in this lesion were unknown. Therefore, we aimed to investigate the inflammatory effect of L-lactic acid on peripheral blood mononuclear cells (PBMCs).
Project description:Recently, 1-nonadecene was identified as a unique metabolite in radicular cysts. However, the biological roles of this metabolite were unknown. Therefore, we aimed to investigate the inflammatory effect of 1-nonadecene on peripheral blood mononuclear cells (PBMCs).
Project description:We investigated the effects of the TLR8 agonist 3M-002 on latently HIV infected U1 cells. We found a prominent upregulation of TLR-dependent genes. Notably, the TLR8 agonist resulted in a marked activation of HIV. We stimulated U1 cells for 6 or 10 hours with 3M-002 and subsequently harvested the cells, extracted total RNA. RNA was then used for quantifiying TLR-dependent gene upregulation using a commercial PCR array pathway
Project description:Adult human ependymal and ventral horn regions were obtained from postmortem frozen samples by Laser Capture Microdissection. Briefly, Cryostat 25 micron sections from were stained with toluidin blue and both regions microdissected and collected on eppendorf (n=4 for each region). Samples mRNA concentration and purity was assessed by electrophoresis (BioRad Experion HighSensitivity kit, USA). RQI values were lower than 6,5 in every case, so that purification was followed by 2 cycle amplification with a kit designed for highly degraded samples (ExpressArt® TRinucleotide mRNA Amplification Kit; #6299-A15, AmpTec, AMSBIO, UK). After amplification, mRNA concentration and purity was assessed both by electrophoresis (BioRad Experion StSens kit, USA) and by spectrophotometry (Nanodrop, Thermo Scientific, USA). We amplified 3.7-37 ng of total RNA, obtaining between 6 and 21 µg of mRNA after 2 rounds. After collecting samples and studying the RNA integrity and quantity, cDNA of samples was selected for gene expression assays using 384 wells Custom Taqman Low Density Arrays. We built arrays with genes belonging to a profile of stemness or ependymoma (see Garcia-Ovejero et al., 2015, BRAIN). Taqman based qPCR gene expression profiling. Ependymal and ventral horn regions obtained by LCMD from four different individuals each were used to establish genes involved in stem cell niches or in ependymoma phenotype that are enriched in control human ependyma using ventral horn as a non-ependymary, non-neurogenic region. Samples were treated as stated in the summary. Equal amount of amplified RNA (aRNA; 25ng, corresponding approximately to 500ng total RNA) from each donor was used in Custom Designed Taqman Low Density Arrays. Every value is the resultant of duplicates at least, but most of them have been assayed 4 times.
Project description:In order to identify new markers of vascular cell senescence with potential in vivo implications, primary cultured Human Umbilical Vein Endothelial Cells (HUVECs), were analysed for microRNA (miR) expression. QRT-PCR microRNA expression profiling in 3 senescent (XIII passage) vs. 3 young HUVECs (II passage).
Project description:Goal was to identify N-recognins for Arginine and Leucine by comparison of biotinylated proteins from 1 week old whole Arabidopsis seedlings constitutively expressing the bait constructs R-degron-Turbo or L-degron-Turbo. M-degron-Turbo served as a negative control. The plants were treated with 50µM biotin and either the proteasome inhibitor Bortezomib (10µM) or the inhibitor of vacuolar acidification Concanamycin A (1µM) or DMSO one hour before harvest.
Project description:We were interested in the effects of TLR8 triggering of human monocyte-derived macrophages. We observed a striking upregulation of those genes. This work was done in the context of examining the effects of TLR8 triggering for the activation of HIV. We generated monocyte-derived macrophages (MDM) by initially isolating monocytes and subsequently culturing them in RPMI supplemented with human AB serum. MDM were stimulated for 18 h with the TLR8 agonist 3M-002, and subsequently harvested for extraction of total RNA. Total RNA was then used to quantify the TLR-dependent genes using a PCR array pathway.