Search results for "Transfection"
showing 10 items of 581 documents
The Ras/Raf-1/MEK1/ERK Signaling Pathway Coupled to Integrin Expression Mediates Cholinergic Regulation of Keratinocyte Directional Migration
2005
The physiologic mechanisms that determine directionality of lateral migration are a subject of intense research. Galvanotropism in a direct current (DC) electric field represents a natural model of cell re-orientation toward the direction of future migration. Keratinocyte migration is regulated through both the nicotinic and muscarinic classes of acetylcholine (ACh) receptors. We sought to identify the signaling pathway mediating the cholinergic regulation of chemotaxis and galvanotropism. The pharmacologic and molecular modifiers of the Ras/Raf-1/MEK1/ERK signaling pathway altered both chemotaxis toward choline and galvanotropism toward the cathode in a similar way, indicating that the sam…
Pore-forming Staphylococcus aureus alpha-toxin triggers epidermal growth factor receptor-dependent proliferation.
2006
Staphylococcal alpha-toxin is an archetypal killer protein that homo-oligomerizes in target cells to create small transmembrane pores. The membrane-perforating beta-barrel motif is a conserved attack element of cytolysins of Gram-positive and Gram-negative bacteria. Following the recognition that nucleated cells can survive membrane permeabilization, a profile of abundant transcripts was obtained in transiently perforated keratinocytes. Several immediate early genes were found to be upregulated, reminiscent of the cellular response to growth factors. Cell cycle analyses revealed doubling of S + G2/M phase cells 26 h post toxin treatment. Determination of cell counts uncovered that after an …
Rottlerin induces a transformed phenotype in human keratinocytes.
2001
PKCdelta plays a fundamental role in cell cycle control. Consistent with its proposed tumour suppressor function, ras transfection of the human keratinocyte cell line HaCaT results in a loss of PKCdelta expression mediated by TGFalpha (Exp. Cell Res., 219, 299, 1995). To get more insight into the role of PKCdelta in keratinocytes, we investigated the effects of Rottlerin, a specific inhibitor of protein kinase Cdelta, in HaCaT cells. After Rottlerin treatment, HaCaT cells lost their cobble-stone morphology and displayed a spindle-shaped, fibroblastic phenotype. Additionally, the establishment of cell-cell contacts was prevented. This was caused by an internalization of E-cadherin and beta-c…
Downregulation of KLF8 expression by shRNA induces inhibition of cell proliferation in CAL27 human oral cancer cells
2013
Objectives: KLF8 is a member of KLF transcription factors which play an important tolr in oncogenesis. It is barely expressed in normal human epithelial cells but highly overexpressed in several types of human cancer cell lines. In the present study, we investigate the role of KLF8 in oral cancer and the effects of KLF8 knockdown via lentivirus mediated siRNA infection in human adenosquamos carcinoma CAL 27 cells. Study Design: �e developed a vector-based siRNA expression system that can induce RNAi in CAL 27 oral canDesign: �e developed a vector-based siRNA expression system that can induce RNAi in CAL 27 oral canesign: �e developed a vector-based siRNA expression system that can induce RN…
Isolation and subsequent analysis of murine lamina propria mononuclear cells from colonic tissue
2007
Studies on colonic cells in the lamina propria (LP) of mice are important for understanding the cellular and immune responses in the gut, especially in inflammatory bowel diseases (such as morbus crohn and colitis ulcerosa). This protocol details a method to isolate LP cells and characterize freshly isolated cells by quality control experiments to obtain cells that can be used for further investigations. After different steps of digestion of the tissue using collagenase, DNase and dispase, the resulting cells are purified using Percoll gradient. The success of the isolation can be analyzed by cell viability test (Trypan Blue exclusion test) and by flow cytometric analysis to assess apoptosi…
The Peptide Hemopressin Acts through CB1Cannabinoid Receptors to Reduce Food Intake in Rats and Mice
2010
Hemopressin is a short, nine amino acid peptide (H-Pro-Val-Asn-Phe-Lys-Leu-Leu-Ser-His-OH) isolated from rat brain that behaves as an inverse agonist at the cannabinoid receptor CB1, and is shown here to inhibit agonist-induced receptor internalization in a heterologous cell model. Since this peptide occurs naturally in the rodent brain, we determined its effect on appetite, an established central target of cannabinoid signaling. Hemopressin dose-dependently decreases night-time food intake in normal male rats and mice, as well as in obeseob/obmale mice, when administered centrally or systemically, without causing any obvious adverse side effects. The normal, behavioral satiety sequence is …
Selective uptake and degradation of c-Fos and v-Fos by rat liver lysosomes
1996
AbstractThe transcription factor c-Fos is a short-lived protein and calpains and ubiquitin-dependent systems have been proposed to be involved in its degradation. In this report, we consider a lysosomal degradation pathway for c-Fos. Using a cell-free assay, we have found that freshly isolated lysosomes can take up and degrade c-Fos with high efficiency. v-Fos, the oncogenic counterpart of c-Fos, can also be taken up by lysosomes, yet the amount of incorporated protein is much lower. c-Fos uptake is independent of its phosphorylation state but it appears to be regulated by dimerization with differentially phosphorylated forms of c-Jun, while v-Fos escapes this regulation. Moreover, we show …
Inhibition of NF-κB Activation and iNOS Induction by ent-Kaurane Diterpenoids in LPS-Stimulated RAW264.7 Murine Macrophages
2009
Xerophilusin A (1), xerophilusin B (2), longikaurin B (3), and xerophilusin F (4) from Isodon xerophylus inhibit LPS-induced NO production in RAW 264.7 macrophages, with IC(50) values of 0.60, 0.23, 0.44, and 0.67 muM, respectively, and they all inhibited mRNA production in these same cells. They decreased the luciferase activity in RAW 264.7 cells transiently transfected with the NF-kappaB-dependent luciferase reporter, with IC(50) values of 1.8, 0.7, 1.2, and 1.6 muM, respectively. Compounds 1-3 reduced NF-kappaB activation, with compound 4 showing no effect, but p65 translocation from the cytoplasm to the nucleus and the LPS-induced degradation of IkappaB were inhibited by all four test …
Highly efficient liposome-mediated gene transfer of inducible nitric oxide synthase in vivo and in vitro in vascular smooth muscle cells.
2000
Objective: The efficient introduction of regulatory genes into vascular smooth muscle cells (SMCs) is one of the most promising options for gene therapy of cardiovascular diseases. Cationic liposome-mediated gene transfer may become a favorable transfection technique with regard to patient’s safety for in vivo administration. However, this method until now has its limitation in a low transfection efficiency. Therefore, the present study was designed to improve cationic liposome-mediated transfection of rabbit vascular SMCs in vitro and in vivo, in order to enhance transfection efficiency and present an optimized system which may offer a potential therapeutic benefit for in vivo application.…
Bcl-2 is a negative regulator of interleukin-1β secretion in murine macrophages in pharmacological-induced apoptosis
2010
BACKGROUND AND PURPOSE Cucurbitacin R, a natural anti-inflammatory product, has been shown to exhibit activity against both adjuvant-induced arthritis and delayed-type hypersensitivity reactions induced by various agents. Previous studies have demonstrated that the effects of cucurbitacin R stem from its inhibition of both cytokine production and lymphocyte proliferation. EXPERIMENTAL APPROACHES Effects of cucurbitacin R were investigated on lipopolysaccharide-stimulated RAW 264.7 cells. Cell cycle evolution was analysed by flow cytometry, detection of apoptosis by DNA ladder, Bcl-2, p21, p53, Bax, cleaved caspase-1 (p10), caspase-9, and caspase-3, cleaved caspase (p17) and interleukin-1β d…