Journal Detail Information

1

Cite

Share

Fibrin glue increases the cell survival and the transduced gene product secretion of the ceiling culture-derived adipocytes transplanted in mice.

Yasuyuki AOYAGI ; Masayuki KURODA ; Sakiyo ASADA ; Hideaki BUJO ; Shigeaki TANAKA ; Shunichi KONNO ; Masami TANIO ; Itsuko ISHII ; Masayuki ASO ; Yasushi SAITO

Experimental & Molecular Medicine.2011;43(3):161-167. doi:10.3858/emm.2011.43.3.021

The development of clinically applicable scaffolds is important for the application of cell transplantation in various human diseases. The aims of this study are to evaluate fibrin glue in a novel protein replacement therapy using proliferative adipocytes and to develop a mouse model system to monitor the delivery of the transgene product into the blood and the fate of the transduced cells after transplantation. Proliferative adipocytes from mouse adipose tissue were transduced by a retroviral vector harboring the human lecithin-cholesterol acyltransferase (lcat) gene, and were subcutaneously transplanted into mice combined with fibrin glue. The lcat gene transduction efficiency and the subsequent secretion of the product in mouse adipocytes were enhanced using a protamine concentration of 500 microg/ml. Adipogenesis induction did not significantly affect the lcat gene-transduced cell survival after transplantation. Immunohistochemistry showed the ectopic enzyme production to persist for 28 days in the subcutaneously transplanted gene-transduced adipocytes. The increased viability of transplanted cells with fibrin glue was accompanied with the decrease in apoptotic cell death. The immunodetectable serum LCAT levels in mice implanted with the fibrin glue were comparable with those observed in mice implanted with Matrigel, indicating that the transplanted lcat gene-transduced adipocytes survived and functioned in the transplanted spaces with fibrin glue as well as with Matrigel for 28 days. Thus, this in vivo system using fibrin is expected to serve as a good model to further improve the transplanted cell/scaffold conditions for the stable and durable cell-based replacement of defective proteins in patients with LCAT deficiency.
Adipocytes/*cytology/transplantation ; Animals ; Blotting, Western ; Cell Differentiation ; Cell Survival/drug effects ; Cells, Cultured ; Collagen/metabolism ; Drug Combinations ; Drug Delivery Systems ; Fibrin Tissue Adhesive/*administration & dosage ; Genetic Vectors/administration & dosage ; Humans ; Laminin/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Nude ; Phosphatidylcholine-Sterol O-Acyltransferase/*genetics/*metabolism ; Proteoglycans/metabolism ; RNA, Messenger/genetics ; Reverse Transcriptase Polymerase Chain Reaction ; *Tissue Engineering

2

Cite

Share

Neuronal activation increases the density of eukaryotic translation initiation factor 4E mRNA clusters in dendrites of cultured hippocampal neurons.

Il Soo MOON ; Sun Jung CHO ; Dae Hyun SEOG ; Randall WALIKONIS

Experimental & Molecular Medicine.2009;41(8):601-610. doi:10.3858/emm.2009.41.8.066

Activity-dependent dendritic translation in CNS neurons is important for the synapse-specific provision of proteins that may be necessary for strengthening of synaptic connections. A major rate-limiting factor during protein synthesis is the availability of eukaryotic translation initiation factor 4E (eIF4E), an mRNA 5'-cap-binding protein. In this study we show by fluorescence in situ hybridization (FISH) that the mRNA for eIF4E is present in the dendrites of cultured rat hippocampal neurons. Under basal culture conditions, 58.7 +/- 11.6% of the eIF4E mRNA clusters localize with or immediately adjacent to PSD-95 clusters. Neuronal activation with KCl (60 mM, 10 min) very significantly increases the number of eIF4E mRNA clusters in dendrites by 50.1 and 74.5% at 2 and 6 h after treatment, respectively. In addition, the proportion of eIF4E mRNA clusters that localize with PSD-95 increases to 74.4 +/- 7.7% and 77.8 +/- 7.6% of the eIF4E clusters at 2 and 6 h after KCl treatment, respectively. Our results demonstrate the presence of eIF4E mRNA in dendrites and an activity-dependent increase of these clusters at synaptic sites. This provides a potential mechanism by which protein translation at synapses may be enhanced in response to synaptic stimulation.
Animals ; Cells, Cultured ; Dendrites/*metabolism ; Eukaryotic Initiation Factor-4E/genetics/*metabolism ; Hippocampus/*cytology ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Microscopy, Confocal ; Neurons/cytology/*physiology ; Potassium Chloride/pharmacology ; Protein Biosynthesis ; RNA, Messenger/genetics/*metabolism ; Rats ; Rats, Sprague-Dawley ; Synapses ; *Up-Regulation

3

Cite

Share

Stimulatory heterotrimeric G protein augments gamma ray-induced apoptosis by up-regulation of Bak expression via CREB and AP-1 in H1299 human lung cancer cells.

Yoon Jung CHOI ; So Young KIM ; Jung Min OH ; Yong Sung JUHNN

Experimental & Molecular Medicine.2009;41(8):592-600. doi:10.3858/emm.2009.41.8.065

Stimulatory heterotrimeric GTP-binding proteins (Gs protein) stimulate cAMP generation in response to various signals, and modulate various cellular phenomena such as proliferation and apoptosis. This study aimed to investigate the effect of Gs proteins on gamma ray-induced apoptosis of lung cancer cells and its molecular mechanism, as an attempt to develop a new strategy to improve the therapeutic efficacy of gamma radiation. Expression of constitutively active mutant of the alpha subunit of Gs (GalphasQL) augmented gamma ray-induced apoptosis via mitochondrial dependent pathway when assessed by clonogenic assay, FACS analysis of PI stained cells, and western blot analysis of the cytoplasmic translocation of cytochrome C and the cleavage of caspase-3 and ploy(ADP-ribose) polymerase (PARP) in H1299 human lung cancer cells. GalphasQL up-regulated the Bak expression at the levels of protein and mRNA. Treatment with inhibitors of PKA (H89), SP600125 (JNK inhibitor), and a CRE-decoy blocked GalphasQL-stimulated Bak reporter luciferase activity. Expression of GalphasQL increased basal and gamma ray-induced luciferase activity of cAMP response element binding protein (CREB) and AP-1, and the binding of CREB and AP-1 to Bak promoter. Furthermore, prostaglandin E2, a Galphas activating signal, was found to augment gamma ray-induced apoptosis, which was abolished by treatment with a prostanoid receptor antagonist. These results indicate that Galphas augments gamma ray-induced apoptosis by up-regulation of Bak expression via CREB and AP-1 in H1299 lung cancer cells, suggesting that the efficacy of radiotherapy of lung cancer may be improved by modulating Gs signaling pathway.
Apoptosis/*radiation effects ; Cell Line, Tumor ; Cyclic AMP Response Element-Binding Protein/metabolism ; GTP-Binding Protein alpha Subunits, Gs/*metabolism ; *Gamma Rays ; Heterotrimeric GTP-Binding Proteins/metabolism ; Humans ; Lung/*cytology/physiology/radiation effects ; Lung Neoplasms ; Transcription Factor AP-1/metabolism ; *Up-Regulation ; bcl-2 Homologous Antagonist-Killer Protein/*metabolism

4

Cite

Share

Lysophosphatidylglycerol inhibits formyl peptide receptor like-1-stimulated chemotactic migration and IL-1beta production from human phagocytes.

Jae Woong SHIM ; Seong Ho JO ; Sang Doo KIM ; Ha Young LEE ; Jeanho YUN ; Yoe Sik BAE

Experimental & Molecular Medicine.2009;41(8):584-591. doi:10.3858/emm.2009.41.8.064

In this study, we observed that lysophosphatidylglycerol (LPG) completely inhibited a formyl peptide receptor like-1 (FPRL1) agonist (MMK-1)-stimulated chemotactic migration in human phagocytes, such as neutrophils and monocytes. LPG also dramatically inhibited IL-1beta production by another FPRL1 agonist serum amyloid A (SAA) in human phagocytes. However, LPG itself induced intracellular calcium increase and superoxide anion production in human phagocytes. Keeping in mind that phagocytes migration and IL-1beta production by FPRL1 are important for the induction of inflammatory response, our data suggest that LPG can be regarded as a useful material for the modulation of inflammatory response induced by FPRL1 activation.
Chemotaxis, Leukocyte/*drug effects ; Humans ; Interleukin-1beta/*biosynthesis ; Lysophospholipids/*pharmacology ; Monocytes/drug effects/immunology/metabolism/physiology ; Neutrophils/drug effects/immunology/metabolism/physiology ; Peptides/metabolism/pharmacology ; *Phagocytes/drug effects/immunology/metabolism/physiology ; Receptors, Formyl Peptide/*metabolism ; Receptors, Lipoxin/*metabolism ; Serum Amyloid A Protein/metabolism/pharmacology

5

Cite

Share

Adiponectin and thiazolidinedione targets CRTC2 to regulate hepatic gluconeogenesis.

Young Sil YOON ; Dongryeol RYU ; Min Woo LEE ; Sungpyo HONG ; Seung Hoi KOO

Experimental & Molecular Medicine.2009;41(8):577-583. doi:10.3858/emm.2009.41.8.063

During fasting periods, hepatic glucose production is enhanced by glucagon to provide fuels for other organs. This process is mediated via cAMP-dependent induction of the CREB regulated transcriptional coactivator (CRTC) 2, a critical transcriptional activator for hepatic gluconeogenesis. We have previously shown that CRTC2 activity is regulated by AMP activated protein kinase (AMPK) family members. Here we show that adiponectin and thiazolidinedione directly regulate AMPK to modulate CRTC2 activity in hepatocytes. Adiponectin or thiazolidinedione lowered glucose production from primary hepatocytes. Treatment of both reagents reduced gluconeogenic gene expression as well as cAMP-mediated induction of CRE reporter, suggesting that these reagents directly affect CREB/CRTC2- dependent transcription. Furthermore, adiponectin or thiazolidinedione mediated repression of CRE activity is largely blunted by co-expression of phosphorylation defective mutant CRTC2, underscoring the importance of serine 171 residue of this factor. Taken together, we propose that adiponectin and thiazolidinedione promote the modulation of AMPK-dependent CRTC2 activity to influence hepatic gluconeogenesis.
Adiponectin/*pharmacology ; Animals ; Cells, Cultured ; *Gene Expression Regulation ; Gluconeogenesis/*drug effects ; Glucose/metabolism ; Hepatocytes/drug effects/*metabolism ; Humans ; Liver/cytology/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Protein Kinases/genetics/metabolism ; Rats ; Rats, Sprague-Dawley ; Thiazolidinediones/*pharmacology ; Transcription Factors/genetics/*metabolism

6

Cite

Share

Angiotensin II-induced aortic ring constriction is mediated by phosphatidylinositol 3-kinase/L-type calcium channel signaling pathway.

Kee Hun DO ; Min Sung KIM ; Jae Ho KIM ; Byung Yong RHIM ; Won Suk LEE ; Chi Dae KIM ; Sun Sik BAE

Experimental & Molecular Medicine.2009;41(8):569-576. doi:10.3858/emm.2009.41.8.062

Angiotensin II (AngII) is a crucial hormone that affects vasoconstriction and exerts hypertrophic effects on vascular smooth muscle cells. Here, we showed that phosphatidylinositol 3-kinase-dependent calcium mobilization plays pivotal roles in AngII-induced vascular constriction. Stimulation of rat aortic vascular smooth muscle cell (RASMC)-embedded collagen gel with AngII rapidly induced contraction. AngII-induced collagen gel contraction was blocked by pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002) whereas ERK inhibitor (PD98059) was not effective. AngII-induced collagen gel contraction was significantly blocked by extracellular calcium depletion by EGTA or by nifedipine which is an L-type calcium channel blocker. In addition, AngII-induced calcium mobilization was also blocked by nifedipine and EGTA, whereas intracellular calcium store-depletion by thapsigargin was not effective. Finally, pretreatment of rat aortic ring with LY294002 and nifedipine significantly reduced AngII-induced constriction. Given these results, we suggest that PI3K-dependent activation of L-type calcium channels might be involved in AngII-induced vascular constriction.
1-Phosphatidylinositol 3-Kinase/*metabolism/pharmacology ; Angiotensin II/metabolism/*pharmacology ; Animals ; Aorta, Thoracic/*drug effects/physiology ; Calcium Channels, L-Type/drug effects/*metabolism ; Muscle Contraction/drug effects ; Muscle, Smooth, Vascular/drug effects/enzymology ; Rats ; Rats, Sprague-Dawley ; Signal Transduction/*drug effects ; Specific Pathogen-Free Organisms ; Vasoconstriction/*drug effects

7

Cite

Share

Ubiquitylation of Fe65 adaptor protein by neuronal precursor cell expressed developmentally down regulated 4-2 (Nedd4-2) via the WW domain interaction with Fe65.

Eun Jeoung LEE ; Sunghee HYUN ; Jaesun CHUN ; Sung Hwa SHIN ; Sang Sun KANG

Experimental & Molecular Medicine.2009;41(8):555-568. doi:10.3858/emm.2009.41.8.061

Fe65 has been characterized as an adaptor protein, originally identified as an expressed sequence tag (EST) corresponding to an mRNA expressed at high levels in the rat brain. It contains one WW domain and two phosphotyrosine interaction/phosphotyrosine binding domains (PID1/PID2). As the neuronal precursor cell expressed developmentally down regulated 4-2 (Nedd4-2) has a putative WW domain binding motif (72PPLP75) in the N-terminal domain, we hypothesized that Fe65 associates with Nedd4-2 through a WW domain interaction, which has the characteristics of E3 ubiquitin-protein ligase. In this paper, we present evidence for the interaction between Fe65 WW domain and Nedd4-2 through its specific motif, using a pull down approach and co-immunoprecipitation. Additionally, the co-localization of Fe65 and Nedd4-2 were observed via confocal microscopy. Co-localization of Fe65 and Nedd4-2 was disrupted by either the mutation of Fe65 WW domain or its putative binding motif of Nedd4-2. When the ubiquitin assay was performed, the interaction of Nedd4-2 (wt) with Fe65 is required for the cell apoptosis and the ubiquitylation of Fe65. We also observed that the ubiquitylation of Fe65 (wt) was augmented depending on Nedd4-2 expression levels, whereas the Fe65 WW domain mutant (W243KP245K) or the Nedd4-2 AL mutant (72PPLP75 was changed to 72APLA75) was under-ubiquitinated significantly. Thus, our observations implicated that the protein-protein interaction between the WW domain of Fe65 and the putative binding motif of Nedd4-2 down-regulates Fe65 protein stability and subcellular localization through its ubiquitylation, to contribute cell apoptosis.
Adaptor Proteins, Signal Transducing/chemistry/genetics/*metabolism ; Cell Line ; *Down-Regulation ; Endosomal Sorting Complexes Required for Transport/genetics/*metabolism ; *Gene Expression Regulation, Developmental ; Humans ; Immunoprecipitation ; Microscopy, Confocal ; Mutation ; Protein Interaction Mapping ; Protein Structure, Tertiary/*physiology ; Transfection ; Ubiquitin-Protein Ligases/genetics/*metabolism ; Ubiquitination

8

Cite

Share

Cordycepin inhibits UVB-induced matrix metalloproteinase expression by suppressing the NF-kappa B pathway in human dermal fibroblasts.

Young Rae LEE ; Eun Mi NOH ; Eun Yong JEONG ; Seok Kweon YUN ; Young Ju JEONG ; Jong Hyeon KIM ; Kang Beom KWON ; Byeong Soo KIM ; Sung Ho LEE ; Chang Sik PARK ; Jong Suk KIM

Experimental & Molecular Medicine.2009;41(8):548-554. doi:10.3858/emm.2009.41.8.060

Cordycepin (3'-deoxyadenosine) has been shown to exhibit many pharmacological activities, including anti-cancer, anti-inflammatory, and anti-infection activities. However, the anti-skin photoaging effects of cordycepin have not yet been reported. In the present study, we investigated the inhibitory effects of cordycepin on matrix metalloproteinase-1 (MMP-1) and -3 expressions of the human dermal fibroblast cells. Western blot analysis and real-time PCR revealed cordycepin inhibited UVB-induced MMP-1 and -3 expressions in a dose-dependent manner. UVB strongly activated NF-kappa B activity, which was determined by I kappa B alpha degradation, nuclear localization of p50 and p65 subunit, and NF-kappa B binding activity. However, UVB-induced NF-kappa B activation and MMP expression were completely blocked by cordycepin pretreatment. These findings suggest that cordycepin could prevent UVB-induced MMPs expressions through inhibition of NF-kappa B activation. In conclusion, cordycepin might be used as a potential agent for the prevention and treatment of skin photoaging.
Aging/physiology ; Cells, Cultured ; Deoxyadenosines/*pharmacology ; *Dermis/cytology/drug effects/physiology/radiation effects ; Dose-Response Relationship, Drug ; Enzyme Induction/drug effects ; Fibroblasts/drug effects/metabolism/radiation effects ; Gene Expression Regulation, Enzymologic ; Humans ; Infant, Newborn ; Male ; *Matrix Metalloproteinase 1/antagonists & inhibitors/biosynthesis/genetics/radiation effects ; Matrix Metalloproteinase 3/antagonists & inhibitors/*biosynthesis/genetics/radiation effects ; NF-kappa B/*antagonists & inhibitors/genetics/metabolism ; Skin/physiopathology/radiation effects ; *Ultraviolet Rays

9

Cite

Share

Nickel (II)-induced apoptosis and G2/M enrichment.

Sang Han LEE ; Do Kwon KIM ; Young Rok SEO ; Kee Min WOO ; Chang Se KIM ; Man Hee CHO

Experimental & Molecular Medicine.1998;30(3):171-176.

Treatment with certain DNA-damaging agents induce a complex cellular response comprising pertubation of cell cycle progression and/or apoptosis on proliferating mammalian cells. Our studies were focused on the cellular effects of nickel (II) acetate, DNA-damaging agent, on Chinese hamster ovary (CHO) cells. Fragmented DNAs were examined by agarose gel electrophoresis and cell cycle was determined by DNA flow cytometry using propidium iodide fluorescence. Apparent DNA laddering was observed in cells treated with 240 microM nickel (II) and increased with a concentration-dependent manner. Treatment of nickel (II) acetate resulted in apoptosis which was accompanied by G2/M cell accumulation. Proportion of CHO cells in G2/M phase was also significantly increased in cells exposed to at least 480 microM nickel (II) from 57.7% of cells in the G0/G1 phase, 34.7% in the S phase, and 7.6% in the G2/M1 phase for 0 microM nickel (II), to 58.6%, 14.5%, and 26.9% for 640 microM nickel (II). These findings suggest that nickel (II) can modulate cellular response through some common effectors involving in both apoptotic and cell cycle regulatory pathways.
Animal ; Apoptosis/drug effects* ; CHO Cells/drug effects* ; CHO Cells/cytology ; Cell Cycle/drug effects* ; DNA Fragmentation/drug effects ; Flow Cytometry ; G2 Phase/drug effects ; Hamsters ; Mitosis/drug effects ; Nickel/pharmacology*

10

Cite

Share

Urinary N-acetyl-beta-D-glucosaminidase in rheumatoid arthritis.

M Perwaiz IQBAL ; Azra A ALI ; M Anwar WAQAR ; N MEHBOOBALI

Experimental & Molecular Medicine.1998;30(3):165-169.

Excretion of urinary N-acetyl beta-D-glucosaminidase (NAG) and its isoenzyme patterns were studied in two groups of patients with rheumatoid arthritis (RA) and in normal control subjects. Urine samples were collected from 30 seropositive RA patients, 19 seronegative RA patients, and 15 normal healthy subjects. All the patients and normal subjects were assessed to have normal liver and kidney functions. A small portion of the urine sample was dialyzed against 0.01 M phosphate buffer, pH 7.0 and NAG activity was monitored. Mean +/- SD values of urinary NAG in seropositive RA patients, in seronegative RA patients and in normal healthy subjects were found to be 4.20 +/- 3.73 U/g creatinine, 2.96 +/- 2.11 U/gm creatinine, and 1.71 +/- 0.6 U/g creatinine, respectively. The mean urinary, NAG value in RA patients was found to be significantly higher (P < 0.05) in seropositive RA compared to the mean NAG value in normal healthy subjects and patients with seronegative RA when analyzed by one way ANOVA and Tukey-HSD test. The mean proportion of isoenzyme form B to isoenzyme form A in seropositive RA patients was also found to be significantly different (P < 0.05) from the mean proportion of these forms in normal healthy subjects and seronegative RA patients. There also appears to be a correlation between the concentration of urinary NAG and severity of the disease in seropositive RA.
Acetylglucosaminidase/urine* ; Adult ; Arthritis, Rheumatoid/urine* ; Arthritis, Rheumatoid/immunology ; Arthritis, Rheumatoid/enzymology* ; Chromatography, Liquid/methods ; Comparative Study ; Female ; Human ; Isoenzymes ; Male ; Predictive Value of Tests ; Severity of Illness Index

DISPLAY OPTIONS

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share

Cite

Share