BACKGROUND AND OBJECTIVES: Recent reports have demonstrated that perturbation of the balance between myo-sin light chain (MLC) phosphorylation and the dephosphorylation status is associated with the development of cardiac hypertrophy. Myosin light chain phosphatase (MLCP) is a key enzyme that regulates the phosphorylation status of the MLC, but its functional roles in cardiac muscle have not been well investigated. Especially, the functions of the small-subunit of MLCP in cardiac muscles are not well elucidated. Here, whether the human heart-specific small-subunit (M21) of MLCP is associated with hypertrophic responses in a transgenic mice model were assessed. MATERIALS AND METHODS: The transgenic mice, overexpressing the human M21, were generated from a cardiac-specific transgenic construct. Cardiac tissues from the transgenic mice were subjected to histology for their morphological examination. The echocardiographic parameters of the murine heart were examined with transgenic mice aged 1, 2 and 3 months, and compared with their non-transgenic littermates. To determine whether the transgenic heart was sensitive to stress, the echocardiographic examination was also performed at the baseline, both before and after the administration of isoproterenol, at a dosage of 30 microgram/g/day, for 2 weeks. RESULTS: The histological analysis of the transgenic heart revealed myocyte disarray and nuclear hypertrophy. No significant differences were observed between the transgenic and non-transgenic mice in relation to the echocardiographic determinants, such as the left ventricular dimensions and the wall thickness. Chronic cardiac stress, induced by isoproterenol infusion, also failed to show any significant differences in relation to the same determinants. CONCLUSION: Overexpression of the human M21 in the murine heart induced myocyte hypertrophy. However, the overall cardiac functions were not affected under normal and stressed conditions.
Animals ; Cardiomegaly* ; Cardiomyopathy, Hypertrophic ; Echocardiography ; Heart ; Humans* ; Hypertrophy ; Isoproterenol ; Mice ; Mice, Transgenic* ; Muscle Cells ; Myocardium ; Myosin Light Chains* ; Myosin-Light-Chain Phosphatase* ; Myosins* ; Phosphorylation ; Protein Subunits

Failure of mitotic checkpoint machinery leads to the chromosomal missegregation and nuclear endoreduplication, thereby driving the emergence of aneuploidy and tetraploidy population. Although abnormal nuclear ploidy and the resulting impairment of mitotic checkpoint function are typical physiological event leading to human hepatocellular carcinoma, any mutational change of mitotic checkpoint regulators has not yet been discovered. Therefore, we investigated the mutation of p31(comet), a recently identified mitotic checkpoint regulator, in human hepatocellular carcinoma. Of 51 human hepatocellular carcinoma tissue and 6 cell lines tested, five samples exhibited nucleotide sequence variations dispersed on four sites within the entire coding sequence. Among these sites with sequence substitutions, three were found to be missense mutation accompanied with amino acid change but one was a silent mutation. Of these sequence substitutions, two were present in both tumor and non-tumor liver tissues, suggesting the possibility of polymorphism. The present findings indicate that p31(comet) does not have an impact on the formation of aneuploidy and tetraploidy found in human hepatocellular carcinoma.
Adaptor Proteins, Signal Transducing ; Calcium-Binding Proteins/*metabolism ; Carcinoma, Hepatocellular/genetics/*metabolism ; Carrier Proteins/*genetics/metabolism ; Cell Cycle Proteins/*genetics/*metabolism ; Cell Line, Tumor ; Humans ; Liver Neoplasms/genetics/*metabolism ; *Mutation ; Nuclear Proteins ; Polyploidy ; Repressor Proteins/*metabolism

Evidences show that eukaryotic mRNAs can perform protein translation through internal ribosome entry sites (IRES). 5'-Untranslated region of the mRNA encoding apoptotic protease-activating factor 1 (Apaf-1) contains IRES, and, thus, can be translated in a cap-independent manner. Effects of changes in protein translation pattern through rapamycin pretreatment on 4-(methylnitrosamino)-1-(3-pyridyl)-butanone(NNK, tobacco-specific lung carcinogen)-induced apoptosis in human bronchial epithelial cells were examined by caspase assay, FACS analysis, Western blotting, and transient transfection. Results showed that NNK induced apoptosis in concentration- and time-dependent manners. NNK-induced apoptosis occurred initially through cap-independent protein translation, which during later stage was replaced by cap-dependent protein translation. Our data may be pplicable as the mechanical basis of lung cancer treatment.
Antibiotics, Antineoplastic/pharmacology ; Apoptosis/*drug effects ; Apoptotic Protease-Activating Factor 1 ; BH3 Interacting Domain Death Agonist Protein ; Blotting, Western ; Bronchi/metabolism/*pathology ; Carcinogens/*pharmacology ; Carrier Proteins/metabolism ; Caspases/metabolism ; Cytochromes c/metabolism ; Dose-Response Relationship, Drug ; Epithelial Cells/metabolism/*pathology ; Eukaryotic Initiation Factor-4E/metabolism ; Flow Cytometry ; Humans ; Nitrosamines/*pharmacology ; Protein Biosynthesis ; Proteins/metabolism ; Proto-Oncogene Proteins c-bcl-2/metabolism ; RNA Cap-Binding Proteins/*physiology ; Sirolimus/pharmacology ; Time Factors ; bcl-2-Associated X Protein

Biocompatible silica-overcoated magnetic nanoparticles containing an organic fluorescence dye, rhodamine B isothiocyanate (RITC), within a silica shell [50 nm size, MNP@SiO2(RITC)s] were synthesized. For future application of the MNP@SiO2(RITC)s into diverse areas of research such as drug or gene delivery, bioimaging, and biosensors, detailed information of the cellular uptake process of the nanoparticles is essential. Thus, this study was performed to elucidate the precise mechanism by which the lung cancer cells uptake the magnetic nanoparticles. Lung cells were chosen for this study because inhalation is the most likely route of exposure and lung cancer cells were also found to uptake magnetic nanoparticles rapidly in preliminary experiments. The lung cells were pretreated with different metabolic inhibitors. Our results revealed that low temperature disturbed the uptake of magnetic nanoparticles into the cells. Metabolic inhibitors also prevented the delivery of the materials into cells. Use of TEM clearly demonstrated that uptake of the nanoparticles was mediated through endosomes. Taken together, our results demonstrate that magnetic nanoparticles can be internalized into the cells through an energy-dependent endosomal-lysosomal mechanism.
Biocompatible Materials/*pharmacokinetics ; Cell Line, Tumor ; Drug Delivery Systems/methods ; Endocytosis/*physiology ; Endosomes/physiology ; Humans ; Lung Neoplasms/drug therapy/*metabolism ; Macrolides/pharmacology ; Microscopy, Confocal ; Microscopy, Electron, Transmission ; Nanoparticles/*administration & dosage ; Sodium Azide/pharmacology ; Sucrose/pharmacology ; Temperature

Inorganic phosphate (Pi) plays a critical role in diverse cellular functions, and regulating the Pi balance is accomplished by sodium-dependent Pi co-transporter (NPT). Pulmonary NPT has recently been identified in mammalian lungs. However, to date, many of the studies that have involved Pi have mainly focused on its effect on bone and kidney. Therefore, current study was performed to discover the potential effects of low Pi on the lung of developing transgenic mice expressing the renilla/firefly luciferase dual reporter gene. Two-weeks old male mice divided into 2 groups and these groups were fed either a low PI diet or a normal control diet (normal: 0.5% Pi, low: 0.1% Pi) for 4 weeks. After 4 weeks of the diet, all the mice were sacrificed. Their lungs were harvested and analyzed by performing luciferase assay, Western blotting, kinase assay and immunohistochemistry. Our results demonstrate that low Pi affects the lungs of developing mice by disturbing protein translation, the cell cycle and the expression of fibroblast growth factor-2. These results suggest that optimally regulating Pi consumption may be important to maintain health.
Animals ; Blotting, Western ; Carrier Proteins/metabolism ; Immunohistochemistry ; Lung/drug effects/enzymology/*growth & development/metabolism ; Male ; Mice ; Mice, Transgenic ; Phosphoproteins/metabolism ; Phosphorus, Dietary/*administration & dosage ; Phosphorylation ; Phosphotransferases (Alcohol Group Acceptor)/metabolism ; Proto-Oncogene Proteins c-akt/metabolism ; Sodium-Phosphate Cotransporter Proteins, Type IIa/*metabolism