Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe a highly efficient calcium phosphate transfection protocol capable of achieving 100% transfection efficiency of reporter genes transiently expressed in the human hepatoma cell lines HuH7 and HepG2. This procedure, a modification of that described by Chen and Okayama, is reliable, reproducible, and eliminates the requirement for the inclusion of cotransfected internal control plasmids. While Chen and Okayama described the pH of the 2x BBS (N,N-bis[2-hydroxyethyl]-2-aminoethanesulfonic acid-buffered saline) and DNA concentration as being critical factors for optimal transfection efficiency, we show that a reduced and strictly monitored standing time of the DNA/CaCl2/2x BBS cocktail prior to addition to cultured cells is essential for a particular combination of pH and DNA concentration. We also show that the inclusion of internal control plasmids can inhibit reporter gene activity in a promoter- and dose-dependent manner. The method so described is also applicable for the transfection of other mammalian cell lines including COS and HeLa, and conceivably for the generation of stable transfectants at high frequency.
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PMID:Optimization of experimental variables influencing reporter gene expression in hepatoma cells following calcium phosphate transfection. 781 89

Genetic variants of human alpha1-antitrypsin unable to fold into the native structural conformation are poorly secreted from hepatocytes. The molecular chaperone calnexin coimmunoprecipitates with secretion-incompetent variant null(Hong Kong) retained in stably transfected mouse hepatoma cells (Le, A., Steiner, J. L., Ferrell, G. A., Shaker, J. F., and Sifers, R. N. (1994) J. Biol. Chem. 269, 7514-7519). Mobilization of intracellular Ca2+ stores with metabolic poisons diminished interaction with calnexin and coincided with coimmuoprecipitation of a 150-kDa protein (p150). Mobilization of endoplasmic reticulum lumenal Ca2+ with thapsigargin, an inhibitor of the microsomal Ca2+ATPase, gave a similar result. Coimmunoprecipitation of p150 was specifically disrupted in response to incubation of the cell lysate with exogenous CaCl2. Finally, in ECL Western blotting, p150 was recognized by polyclonal antiserum against UDP-glucose:glycoprotein glucosyltransferase that likely functions in glycoprotein folding and quality control (Sousa, M. C., Ferrero-Garcia, M. A., and Parodi, A. J. (1992) Biochemistry 31, 97-105). The data are consistent with a model in which perturbation of endoplasmic reticulum Ca2+ results in a stable physical association between unfolded human alpha1-antitrypsin and UDP-glucose:glycoprotein glucosyltransferase.
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PMID:Intracellular association between UDP-glucose:glycoprotein glucosyltransferase and an incompletely folded variant of alpha1-antitrypsin. 914 70

A 12 kDa ribonuclease preferential for poly U and with much lower activity toward poly A, poly G and poly C was isolated from fresh fruiting bodies of the mushroom Pleurotus sajor-caju. A purification procedure involving ion exchange chromatography on CM-cellulose, affinity chromatography on Red-Sepharose and Heparin-Sepharose, and fast protein liquid chromatography-gel filtration on Superdex 75 was used. The ribonuclease was adsorbed on all of the first three types of chromatographic media. It exhibited some activity toward herring sperm DNA and calf thymus DNA. The ribonuclease activity was unaffected in the presence of KCl (10 and 100 mM) and NaCl (100 mM and 1 M), but was strongly inhibited by CuSO4 (0.01 and 0.1 mM) and less potently inhibited by other divalent salts including MgCl2, CaCl2, ZnCl2, ZnSO4 and FeSO4. The optimal pH was 5.5 and the ribonuclease was stable up to 60 degrees C for 1 h. The ribonuclease inhibited mycelial growth in the fungi Fusarium oxysporum and Mycosphaerella arachidicola with an IC50 value of 95 and 72 microM, respectively. Out of the 12 species of bacteria tested, only Pseudomonas aeruginosa and Staphylococcus aureus were inhibited in growth by the ribonuclease. Viability of the tumor cells HepG2 (hepatoma) and L1210 (leukemia) was reduced with an IC50 of 0.22 and 0.1 microM, respectively in the presence of the ribonuclease. The ribonuclease inhibited translation in a cell-free rabbit reticulocyte lysate system with an IC50 of 158 nM and 3H-methyl-thymidine uptake by murine splenocytes with an IC50 of 65 nM.
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PMID:A ribonuclease with antimicrobial, antimitogenic and antiproliferative activities from the edible mushroom Pleurotus sajor-caju. 1500 51


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