Search results for "transfection"

Noninvasive Monitoring of Lesion Size in a Heterologous Mouse Model of Endometriosis

2019

Here, we describe a protocol for the implementation of a heterologous mouse model in which progression of endometriosis can be assessed in real time through noninvasive monitoring of fluorescence emitted by implanted ectopic human endometrial tissue. For this purpose, biopsies of human endometrium are obtained from donor women ongoing oocyte donation. Human endometrial fragments are cultured in the presence of adenoviruses engineered to express cDNA for the reporter fluorescent protein mCherry. Upon visualization, labeled tissues with an optimal rate of fluorescence after infection are subsequently chosen for the implantation in recipient mice. One week prior to the implantation surgery, re…

Agonist‐induced desensitisation of β 3 ‐adrenoceptors: Where, when, and how?

2019

β3 -Adrenoceptor agonists have proven useful in the treatment of overactive bladder syndrome, but it is not known whether their efficacy during chronic administration may be limited by receptor-induced desensitisation. Whereas the β2 -adrenoceptor has phosphorylation sites that are important for desensitisation, the β3 -adrenoceptor lacks these; therefore, it had been assumed that β3 -adrenoceptors are largely resistant to agonist-induced desensitisation. While all direct comparative studies demonstrate that β3 -adrenoceptors are less susceptible to desensitisation than β2 -adrenoceptors, desensitisation of β3 -adrenoceptors has been observed in many models and treatment settings. Chimeric …

2019

Parthenolide (PT) is a sesquiterpene lactone isolated from Tanacetum parthenium. In this study, PT showed varying cytotoxic effects against different solid tumor cell lines. HCT116 (p53+/+) colon carcinoma cells and their parental HCT116 knockout p53 (p53-/-) cell lines showed a resistance degree of 2.36. On the other hand, wild-type U87.MG cells or cells transfected with a deletion-activated EGFR cDNA (U87.MGΔEGFR) exhibited slight sensitivity toward PT. Multidrug-resistant MDA-MB-231-BCRP cells were even more sensitive toward PT than sensitive MDA-MB-231-pcDNA cells with a resistance degree of 0.07 (collateral sensitivity). To the best of our knowledge, hypersensitivity (collateral sensit…

MicroRNA Intercellular Transfer and Bioelectrical Regulation of Model Multicellular Ensembles by the Gap Junction Connectivity.

2017

We have studied theoretically the microRNA (miRNA) intercellular transfer through voltage-gated gap junctions in terms of a biophysically grounded system of coupled differential equations. Instead of modeling a specific system, we use a general approach describing the interplay between the genetic mechanisms and the single-cell electric potentials. The dynamics of the multicellular ensemble are simulated under different conditions including spatially inhomogeneous transcription rates and local intercellular transfer of miRNAs. These processes result in spatiotemporal changes of miRNA, mRNA, and ion channel protein concentrations that eventually modify the bioelectrical states of small multi…

Triclosan-Evoked Neurotoxicity Involves NMDAR Subunits with the Specific Role of GluN2A in Caspase-3-Dependent Apoptosis

2018

Triclosan (TCS) is an antimicrobial agent that is used extensively in personal care and in sanitising products. A number of studies have shown the presence of TCS in different human tissues such as blood, adipose tissue, the liver, brain as well as in breast milk and urine. N-Methyl-D-aspartate receptors (NMDARs) are glutamate-gated ion channels that are widely expressed in the central nervous system and which play key roles in excitatory synaptic transmission. There is, however, no data on the involvement of NMDAR subunits in the apoptotic and neurotoxic effects of TCS. Our experiments are the first to show that TCS used at environmentally relevant concentrations evoked NMDA-dependent effe…

Metabolic Imaging in Multicellular Spheroids of Oncogene-transfected Fibroblasts

2000

Four rat embryo fibroblast (REF) cell lines with defined oncogenic transformation were used to study the relationship between tumorigenic conversion, metabolism, and development of cell death in a 3D spheroid system. Rat1 (spontaneously immortalized) and M1 ( myc-transfected) fibroblasts represent early nontumorigenic transformation stages, whereas Rat1-T1 (T24Ha- ras-transfected Rat1) and MR1 ( myc/T24Ha- ras-co-transfected REF) cells express a highly tumorigenic phenotype. Localized ATP, glucose, and lactate concentrations in spheroid median sections were determined by imaging bioluminescence. ATP concentrations were low in the nonproliferating Rat1 aggregates despite sufficient oxygen an…

SWATH-MS based quantitative proteomics analysis reveals that curcumin alters the metabolic enzyme profile of CML cells by affecting the activity of m…

2018

Background Chronic myelogenous leukemia (CML) is a myeloproliferative disorder caused by expression of the chimeric BCR-ABL tyrosine kinase oncogene, resulting from the t(9;22) chromosomal translocation. Imatinib (gleevec, STI-571) is a selective inhibitor of BCR-ABL activity highly effective in the treatment of CML. However, even though almost all CML patients respond to treatment with imatinib or third generation inhibitors, these drugs are not curative and need to be taken indefinitely or until patients become resistant. Therefore, to get a definitive eradication of leukemic cells, it is necessary to find novel therapeutic combinations, for achieving greater efficacy and fewer side effec…

The actin remodeling protein cofilin is crucial for thymic αβ but not γδ T-cell development

2018

Cofilin is an essential actin remodeling protein promoting depolymerization and severing of actin filaments. To address the relevance of cofilin for the development and function of T cells in vivo, we generated knock-in mice in which T-cell–specific nonfunctional (nf) cofilin was expressed instead of wild-type (WT) cofilin. Nf cofilin mice lacked peripheral αβ T cells and showed a severe thymus atrophy. This was caused by an early developmental arrest of thymocytes at the double negative (DN) stage. Importantly, even though DN thymocytes expressed the TCRβ chain intracellularly, they completely lacked TCRβ surface expression. In contrast, nf cofilin mice possessed normal numbers of γδ T cel…

The membrane-associated MUC1 improves adhesion of salivary MUC5B on buccal cells. Application to development of an in vitro cellular model of oral ep…

2015

Objectives: The mucosal pellicle is a thin layer of salivary proteins, mostly MUC5B mucins, anchored to epithelial oral cells. This pellicle is involved in protection of oral mucosae against abrasion, pathogenic microorganisms or chemical xenobiotics. The present study aimed at studying the involvement of MUC1 in mucosal pellicle formation and more specifically in salivary MUC5B binding using a cell-based model of oral epithelium. DESIGN: MUC1 mRNAs were not detected in TR146 cells, and therefore a stable cell line named TR146/MUC1 expressing this protein was developed by transfection. TR146 and TR146/MUC1 were incubated with human saliva in order to evaluate retention of MUC5B by epithelia…

Characterization of a mutant form of human apolipoprotein B (Thr26_Tyr27del) associated with familial hypobetalipoproteinemia

2016

We have previously identified a deletion mutant of human apoB [apoB (Thr26_Tyr27del)] in a subject with primary hypobetalipoproteinemia. The present study determined the effect of Thr26_Tyr27del mutation on apoB secretion using transfected McA-RH7777 cells. Transient or stable transfection of apoB-48 containing the Thr26_Tyr27del mutation showed drastically reduced secretion of the mutant as compared to wild-type apoB-48. No lipoproteins containing the mutant apoB-48 were secreted into the medium. Incubation of transfected cells in a lipid-rich medium in the presence of cycloheximide showed rapid turnover of cell-associated mutant apoB-48 as compared to that of wild-type apoB-48. Immunofluo…