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J D Ahn,
R Morishita,
Y Kaneda,
H J Kim,
Y D Kim,
H J Lee,
K U Lee,
J Y Park, Y H Kim,
K K Park,
Y C Chang,
K H Yoon,
H S Kwon,
K G Park,
I K Lee
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ABSTRACT: Diabetic nephropathy is characterized by an expansion of glomerular mesangium, caused by mesangial cell proliferation and excessive accumulation of extracellular matrix (ECM) proteins, which eventually leads to glomerulosclerosis and renal failure. Activator protein-1 (AP-1), a transcription factor, is implicated in the transcriptional regulation of a wide range of genes participating in cell proliferation and ECM production. This investigation was undertaken to test the hypothesis that AP-1 plays an important role in ECM gene expression, and to develop a molecular therapeutic strategy based on decoy oligodeoxynucleotides (ODN). In this report, we show that transfection with AP-1 decoy ODN strongly inhibits high glucose- and angiotensin II-induced cell proliferation and expression of ECM genes in cultured mesangial cells in vitro. Administration of AP-1 decoy ODN into streptozotocin-induced diabetic rat kidney in vivo using the hemagglutinating virus of Japan (HVJ)-liposome method virtually abolished TGF-beta1 and plasminogen activator inhibitor-1 expression. Our results collectively indicate that AP-1 activation is crucial for mesangial cell proliferation and ECM production in response to high glucose and angiotensin II. Moreover, use of stable AP-1 decoy ODN combined with the highly effective HVJ-liposome method provides a novel potential molecular therapeutic strategy for the prevention of diabetic nephropathy.
Gene Therapy 07/2004; 11(11):916-23. · 3.71 Impact Factor
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ABSTRACT: The transcription factor, E2F, plays a critical role in the trans-activation of several genes involved in cell cycle regulation. Previous studies showed that the transfection of cis element double-stranded decoy oligodeoxynucleotides (ODNs) corresponding to E2F binding sites inhibited the proliferation of vascular smooth muscle cells (VSMCs) and neointimal hyperplasia in injured vessels. We have developed a novel E2F decoy ODN with a circular dumbbell structure (CD-E2F) and compared its effects with those of the conventional phosphorothioated E2F decoy (PS-E2F) ODN. CD-E2F ODN was more stable than PS-E2F ODN, largely preserving its structural integrity after incubation in the presence of nucleases and sera. Moreover, CD-E2F ODN inhibited high glucose- and serum-induced transcriptional expression of cell cycle regulatory genes more strongly than PS-E2F ODN. Transfection of CD-E2F ODN resulted in more effective inhibition of VSMC proliferation in vitro and neointimal formation in vivo, compared with PS-E2F ODN. An approximately 40-50% lower dose of CD-E2F ODN than PS-E2F ODN was sufficient to attain similar effects. In conclusion, our results indicate that CD-E2F ODN may be a valuable tool in gene therapy protocols for inhibiting VSMC proliferation and studying transcriptional regulation.
Gene Therapy 01/2003; 9(24):1682-92. · 3.71 Impact Factor
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ABSTRACT: The nucleotide sequence of the ZF5128 gene, encoding a novel Kruppel type zinc finger protein, has been determined. The ZF5128 gene has a predicted 553-amino acid open reading frame, encoding a putative 61 kDa zinc finger protein. The N-terminus of the ZF5128 coding region has a well-conserved Kruppel-associated box (KRAB) domain that consists of KRAB box A and B, whereas the C-terminus contains a Kruppel type C2H2 zinc finger domain possessing nine C2H2 zinc finger motifs in tandem arrays with the highly conserved space region of the H/C-link. Each C2H2 zinc finger motif has a typical consensus sequence of CX2CX3FX5LX2HX3H. A 3.2 kb transcript specific for ZF5128 was expressed at high levels in the spleen, thymus, and peripheral blood leukocyte, and weakly expressed in the prostate, ovary, small intestine, colon (mucosal lining), placenta, lung, and pancreas. Although there was no detectable ZF5128 mRNA in unstimulated human peripheral T cells, it was first detectable 1.5 h after activation by anti-CD3 plus anti-CD28, and reached a maximum in 25-30 h. During the cell cycle progression of Jurkat T cells, the expression of ZF5128 mRNA appeared to be induced in G1 and reached a maximum in the S phase, but declined as the cells entered the G2/M phase. The 12-O-tetradecanoylphorbol 13-acetate-induced monocytic differentiation of U937, which also resulted in growth arrest, down-regulated the expression of ZF5128 mRNA. Taken together, these results indicate that ZF5128 is a novel gene encoding a Kruppel type C2H2 zinc finger protein and is regulated at the transcriptional level depending on tissue type and the cell cycle status to support cell proliferation.
Biochimica et Biophysica Acta 01/2002; 1522(3):230-7. · 4.66 Impact Factor
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ABSTRACT: Apolipoprotein C-II (apoC-II), which is known to activate lipoprotein lipase (LPL), was identified by ordered differential display (ODD)-polymerase chain reaction (PCR) as a cDNA fragment exhibiting a distinct increase in expression during 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced differentiation of promonocytic U937 cells into monocytes and macrophages. The amount of apoC-II mRNA expression detectable in U937 cells significantly increased and reached a maximum 24-48 h after treatment with 32 nM TPA. apoC-II mRNA was also detected in monocytic THP-1 cells but was not detected in promyelocytic HL-60 cells. In healthy human tissues, the most significant expression of apoC-II mRNA was in the liver. Although apoC-II mRNA expression was markedly up-regulated during the induced differentiation of HL-60 cells into monocytes and macrophages with 32 nM TPA, such expression was not induced during the differentiation of HL-60 cells into granulocytes with 1.25% dimethyl sulfoxide. These results suggest that human apoC-II expression is induced at the transcription level during myelomonocytic differentiation and may confer an important role to macrophages involved in normal lipid metabolism and atherosclerosis.
Journal of Leukocyte Biology 05/2001; 69(4):645-50. · 4.99 Impact Factor
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ABSTRACT: The nucleotide sequence of Hs 3-PGDH gene, encoding human 3-phosphoglycerate dehydrogenase that catalyzes the initiating step in the phosphorylated pathway of serine biosynthesis, has been determined. The 3-PGDH gene has a predicted 533 amino acid open reading frame, encoding a 56.8kDa protein that shares 94.0% similarity with rat-liver 3-PGDH. Two different transcripts corresponding to 3-PGDH mRNA were detected in human normal tissues. A dominant 2.1kb transcript was expressed at high levels in prostate, testis, ovary, brain, liver, kidney, and pancreas, and weakly expressed in thymus, colon, and heart. A 710bp transcript also appeared as a weaker band where the 2.1kb mRNA was expressed, and it was more significant than the 2.1kb mRNA in heart and skeletal muscle. The TPA-induced monocytic differentiation of U937, which also resulted in growth arrest, abruptly downregulated the expression of 3-PGDH. Removal of TPA restored cell growth through the retrodifferentiation process and subsequent expression of 3-PGDH. The 3-PGDH mRNA was markedly expressed in human leukemias, lymphoma Sup-T1, colon adenocarcinoma COLO 320DM, epitheloid carcinoma HeLa S3, and murine lymphoma BW5147.G.1.4, but not in human leukemia K562. This report demonstrates that the human 3-PGDH gene is regulated at the transcriptional level depending on tissue specificty and cellular proliferative status, and its transcriptional regulation mechanism may be a useful target for diagnosis and therapy of cancer.
Gene 04/2000; 245(1):193-201. · 2.34 Impact Factor
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Cytogenetics and cell genetics 02/2000; 89(1-2):6-7.
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ABSTRACT: In an effort to identify novel genes that are expressed differentially in an infant thymus, we constructed an oligo-d(T) primed cDNA library from a human infant thymus followed by single-run partial sequencing to generate expressed sequence tags (ESTs). Characterization of more than 1400 sequences enabled us to convert human thymus transcripts into 1223 useful ESTs. These ESTs consisted of 613 (50.1%) showing homology to known human genes, 51 (4.2%) matching to genes from other species, 289 (23.6%) matching ESTs of unknown functions, and 182 (14.9%) being novel transcripts. The expression profile of an infant thymus features a high number of genes related to cell division-DNA synthesis and gene-protein expression, indicating the active growth stage of an infant thymus. To identify the chromosomal localization of 43 thymus ESTs, PCR-based mapping was performed using a human-rodent somatic cell hybrid or radiation hybrid mapping panel. The results indicated that several novel genes were determined to be located in the vicinity of previously mapped disease loci; histidinemia loci, plasminogen Tochigi disease loci, Ehlers-Danlos syndrome, hypertriglyceridemia, thyroid resistance locus, ocular albinism, galactosemia, porphyria variegata, Charcot-Marie-tooth disease, FEOM (fibrosis of extraocular muscles), Prader-Willi syndrome.
Genome 07/1999; 42(3):457-64. · 1.65 Impact Factor
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ABSTRACT: A 1204 bp full-length cDNA encoding murine cdc2 was isolated and used as a probe to obtain four overlapping cdc2 genomic clones which span the entire cdc2 sequence. Characterization of these clones revealed that the cdc2 mRNA is distributed over 28 kb and in 8 exons ranging in size from 63 to approximately 303 bp. Since the 5'-untranslated sequence is interrupted by intron 1, the initiation codon is located in exon 2. The PSTAIRE region is in exon 3, and the stop codon is in exon 8. Primer extension analysis of cdc2 mRNA isolated from immobilized anti-CD3 activated T-cells demonstrated that the murine cdc2 gene utilizes multiple transcriptional start sites. No consensus sequence for a TATA box exists at an appropriate position within the promoter region. Instead, several putative transcription factor binding sites for PEA3, CREB, C/EBP, E box factor, YY1, ATF-like, Spl, and E2F were detected. Analysis of the promoter activity of the 5'-flanking region suggests that the sequence between -188 to -38, which possesses two Spl and one E2F motif, contains a major positive regulatory activity for the expression of murine cdc2.
Molecules and Cells 01/1999; 8(6):731-40. · 2.18 Impact Factor
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ABSTRACT: The mechanism for apoptosis induced by aburatubolactam C was investigated in human Jurkat T cells. When the cells were treated with 3 micrograms/ml of aburatubolactam C, apoptotic DNA fragmentation was first detectable in 3 hr and then increased time-dependently in accordance with upregulation in the protein level of Fas ligand (FasL). Both the DNA fragmentation and upregulation of FasL expression reached a maximal level in 7-8 hr, at which time a significant increase in the tyrosine phosphorylation of multiple cellular proteins was detected, suggesting that the enhanced tyrosine phosphorylation of cellular proteins may result from activation of Fas-mediated death signaling. However, these aburatubolactam C-induced cellular changes and accompanied apoptosis were completely blocked in the presence of genistein, a known protein tyrosine kinase inhibitor. These results indicate that upregulation of FasL expression dictated by protein tyrosine kinase activation and subsequent mediation of Fas death signaling account for aburatubolactam C-induced apoptosis in Jurkat T cells.
Biochemical and Biophysical Research Communications 06/1998; 246(1):276-81. · 2.48 Impact Factor
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ABSTRACT: The human homologue of the hamster mitotic centromere-associated kinesin (HsMCAK) gene containing a central type motor domain was isolated from a Jurkat T-cell derived cDNA library. The HsMCAK gene has a predicted 723 amino acid open reading frame, encoding a 81 kDa protein that shares 79.2% homology with hamster MCAK. Unstimulated T lymphocytes contained no detectable HsMCAK-specific mRNA. Activation of resting T-cells by immobilized anti-CD3 resulted in the expression of a 2.9-kb transcript during the S phase of the cell cycle. The TPA-induced monocytic differentiation of U937 which also results in growth-arrest abruptly downregulates the expression of HsMCAK. Removal of TPA restored the growth of the cell through the retrodifferentiation process and the subsequent expression of HsMCAK. HsMCAK is expressed in tissues containing dividing cells, such as thymus, testis, small intestine, colon (mucosal lining), and placenta. These results suggest that the expression of HsMCAK is first detected in early S phase to support the proliferative response and is strictly regulated at the transcriptional level.
Biochimica et Biophysica Acta 01/1998; 1359(3):181-6. · 4.66 Impact Factor
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ABSTRACT: Cyclin D2 is normally expressed in G1 and promotes progression through G1 of the cell cycle. From a murine genomic library constructed with spleen DNA, two overlapping genomic clones of cyclin D2 were isolated. These clones contain most of the exon of cyclin D2 except exon 5. Characterization of these clones revealed that murine cyclin D2 mRNA spans over 18 kb and 5 exons ranging from 149 to approximately 462 bp in length, and suggested that exon 5 may be at least >5 kb downstream from exon 4. Primer extension analysis of cyclin D2 mRNA isolated from murine activated T cells detected 5 putative sites of transcription initiation. These are located at - 499, - 417, - 391, - 373, and - 349 relative to the translation start site, which is given as + 1. No consensus sequence for TATA box existed at an appropriate position within the promotor region. Instead, several putative transcriptional factor binding sites for C/EBP, PEA3, AP2, NF-Y, Sp1, c-Myc, GATA-1, AP1, v-Myb, and CREB were detected. The 5'-flanking region of the cyclin D2 gene up to nucleotide - 945 shared about 61% sequence homology between mouse and human. Functional analysis of promoter activity of the 5'-flanking region of cyclin D2 suggested that the region - 1,100 to - 805 including C/EBP, PEA3, AP2, NF-Y, c-Myc, and Sp1 may have a major positive regulatory activity for expression of cyclin D2.
Molecules and Cells 08/1997; 7(4):537-43. · 2.18 Impact Factor
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ABSTRACT: By using radiolabeled murine cyclin D3 cDNA as a probe, two cyclin D3 genomic clones, MCD3P-117 and MCD3P-327, were isolated from a murine genomic library constructed with murine liver DNA. Physical mapping and DNA sequence analysis revealed that these clones contain approximately 1.5 kb uninterrupted linear sequence similar to murine cyclin D3 cDNA, indicating that the 1.5 kb sequence is a processed pseudogene for cyclin D3. When the nucleotide sequence of the cyclin D3 pseudogene was compared with that of cyclin D3 cDNA at the nucleotide level, the pseudogene contained 229 bp of 5'- and 371 bp of 3'-untranslated regions, and a recognizable complete coding region that is 90% identical to murine cyclin D3. This sequence is bounded by the repeat sequence (GC/AGCTCTCC), which is common to many processed pseudogenes. However, multiple genetic lesions, including substitution, deletion and/or insertion events that result in modification of the reading frame were found in the pseudogene sequence. The pseudogene appeared to accumulate 67 random point mutations in the functional coding region composed of 879 nucleotide positions. It is thus estimated that the cyclin D3 pseudogene arose approximately 11 million years (Myr) ago. These data provide the first characterization of murine cyclin D3 pseudogene and insight into its evolutionary age.
Molecules and Cells 05/1997; 7(2):278-83. · 2.18 Impact Factor
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ABSTRACT: The taxonomic validity of morphological group III Acanthamoeba spp. is uncertain. In the present study, six type strains of group III Acanthamoeba spp., A. culbertsoni, A. healyi, A. pustulosa, A. palestinensis, A. royreba and A. lenticulata were subjected for the evaluation of their taxonomic validity by comparison of the isoenzyme patterns by isoelectic focusing on polyacrylamide gels, mitochondrial DNA (Mt DNA) restriction fragment length polymorphism (RFLP), and small subunit ribosomal DNA (ssu rDNA) PCR-RFLP patterns. The Mt DNA RFLP patterns were heterogeneous between the species. The type strains of A. palestinensis and A. pustulosa showed almost identical patterns of isoenzymes and rDNA PCR RFLP with an estimated sequence divergence of 2.6%. The other species showed heterogeneous patterns of isoenzymes and rDNA PCR-RFLP. It is likely that A. pustulosa is closely related with A. palestinensis and that the former may be regarded as a junior synonym of the latter.
The Korean Journal of Parasitology 01/1997; 34(4):259-66. · 1.04 Impact Factor
