Biomedical subjects

J F Chiu

Publications and source records attributed to J F Chiu.

At least 73 records · Page 4Linked to original sources

PubMed1989

The mechanism of the bidirectional regulation of the rat alpha-fetoprotein gene by glucocorticoid hormone.

pAFP-CAT, a recombinant plasmid containing 5'-flanking sequence from -7 kb to +7 bp of rat alpha-fetoprotein (AFP) gene can drive the expression of the bacterial chloramphenicol acetyltransferase gene in McA-RH7777 and McA-RH8994 rat hepatoma cell lines. Dexamethasone treatment suppresses pAFP-CAT expression in McA-RH7777 cells but increases its expression in McA-RH8994 cells, which mimics the dexamethasone responses of the endogenous AFP gene in both cell lines. However, dexamethasone treatment enhanced pMMTV-CAT expression in both cell lines. These data suggest that the effects of dexamethasone on AFP gene expression may be mediated by different trans-acting factors binding to the specific cis-elements of the 5'-flanking region of the rat AFP gene.

Animals↗

PubMed1989

The enhanced association of keratin with hepatoma cell nuclei.

A monoclonal antibody to rat hepatoma keratin demonstrates a close association of intermediate filaments with the nucleus in hepatoma cells. Immunoblot analysis of nuclear fractions and immunofluorescence of nuclei both prepared by standard procedures, indicate that intermediate filament proteins are consistently present. Sodium citrate extraction of these preparations diminishes the amount of intermediate filament proteins but does not totally remove the antigenic moieties, suggesting a tight association of intermediate filaments with nuclei. The results from both immunoblot analysis and immunofluorescent localization demonstrate the increased amount of keratins associated with hepatoma cell nuclei.

Animals↗

PubMed1988

Differential expression of cellular oncogenes during rat liver development.

The expression of a number of proto-oncogenes (myc, erb B, Ha-ras, bas, rel, mos, sis, myb, ki-ras, fms, src and fos) was studied in developing rat liver. Northern blot hybridization shows that cellular counterpart of erb B, Ha-ras, and fos oncogenes were in an early stage of liver development, and the expressions of these proto-oncogenes gradually decreased as the liver developed, while c-myc transcript was found only in the rat fetal liver. The transcripts of these oncogenes were found in high level in Morris hepatoma 7777. Bas proto-oncogene was found in high expression at early stages of rat liver development but was not in hepatoma 7777. The expression of other proto-oncogenes studied (src, fm, rel, mos, sis, myb and ki-ras) did not change significantly during liver development and was almost the same in hepatoma and normal adult liver. Southern blot analysis demonstrates that gene amplification and apparent gene rearrangement were not responsible for the change in expression of erb B, Ha-ras, myc and fos proto-oncogenes. Our study gives further evidence that erb B, myc, Ha-ras and fos proto-oncogenes are involved in the control of cell growth and in the process of rat hepatocarcinogenesis.

Aging↗

PubMed1987

Methylation of the alpha-fetoprotein gene in isogenic rat hepatoma and liver cell lines.

We have found that DNA methylation is inversely correlated with alpha-fetoprotein (AFP) gene expression in a series of isogenic rat hepatoma cell lines. The 5' end of the gene is extensively demethylated in AFP-producing cells and is highly methylated in cell lines which do not produce AFP. Glucocorticoid affects markedly the synthesis of AFP in the hepatoma cells. However, methylation patterns of cell lines which were treated with dexamethasone were not different from those of control cells, indicating that glucocorticoid action on AFP gene expression does not alter DNA methylation in this region of the gene.

Animals↗

PubMed1987

The expressions of oncogenes and liver-specific genes in Morris hepatomas.

The expression of three liver-specific genes and four oncogenes was studied in the Morris hepatomas 8994, 7288c, 7777, 5123tc, and 7800. Total RNA isolated from these tumors was probed with cDNA's for alpha-fetoprotein (AFP), albumin, tyrosine aminotransferase (TAT), and the oncogenes Ha-ras, Ki-ras, myc and src. When compared to mRNA's levels expressed in normal adult liver, we found AFP levels elevated in AFP-producing tumors, albumin and TAT mRNA levels depressed in all tumors, except TAT is elevated in 5123tc and the oncogenes with the exception of src elevated in all tumors. These results argue against a coordinated expression of these genes as a result of transformation, but suggest that oncogene expression is related to tumorigenesis or proliferation.

Animals↗

PubMed1987

A specific alpha-fetoprotein gene binding protein in alpha-fetoprotein producing rat hepatomas.

A specific alpha-fetoprotein (AFP) gene binding nuclear protein (Mol. Wt. 149,000) was determined in Morris hepatoma 7777 cells by the protein blotting technique. This protein is not present in normal adult rat liver and non-AFP producing Morris hepatoma 5123tc. Neoplasia induced in rats fed the hepatocarcinogen 3'-methyl-4-dimethylaminoazobenzine enhanced AFP gene activity and re-expressed specific AFP gene binding nuclear protein. The precise role of this protein in AFP gene regulation remains to be determined.

Animals↗

PubMed1987

The expression of oncogenes in human developing liver and hepatomas.

Oncogene expression was examined in the human fetal liver and human hepatomas. Erb (B), erb (A+B), Ha-ras, myc, fos and fms oncogene expression elevated in certain stages of fetal liver development and in hepatoma as compared to the normal adult human liver. In contrast, rel, src, mos, sis, myb, Ki-ras and bas oncogenes showed no apparent change of their mRNA levels during fetal liver development and in hepatoma. Further study of erb B oncogene expression in human cirrhotic liver and hepatoma demonstrated a strong correlation between erb B expression and alteration of its gene structure.

Adult↗

PubMed1987

Hepatoma-associated nonhistone chromosomal proteins are present in active chromatin.

Recently a group of nonhistone proteins with molecular weights ranging from 180-200 K were discovered which are associated with rat hepatoma chromatin specifically (Burkhardt et al., Biochim. Biophys. Acta 781, 165-172, 1984). These hepatoma-associated nonhistone proteins appeared and increased in rats treated with a hepatocarcinogen. Two approaches were used in this study to investigate whether the hepatoma-associated nonhistone chromosomal proteins are present in actively transcribed regions. We found that the limited DNase I digestion of Morris hepatoma 7777 chromatin released antigenic proteins not detected in normal liver chromatin digests. The association of antigenic nonhistone proteins with nuclear matrices was also studied. Using immunoblot analysis of nuclear matrices and total chromatin, the antigenic nonhistone chromosomal proteins were determined. Hepatoma-associated nonhistone protein antigens were extensively concentrated in the nuclear matrices. In the present study, the transcriptionally-active alpha-fetoprotein gene and the nontranscribed beta-globin gene were used as gene markers to determine the transcriptionally active chromatin region. Data presented in this paper indicate that hepatoma-associated NHPs are localized in active chromatin.

Animals↗

PubMed1987

Monoclonal antibodies to nonhistone proteins associated with human colon cancer nuclear matrix.

Hybridoma technology was applied in an effort to create highly specific probes for nonhistone proteins associated with human colon cancer nuclear matrix. Three stable monoclonal antibodies producing cloned cells No. 39, 54 and 58 are described here. All these antibodies showed high reactivity with human colon tumor nuclear matrix. Both antibodies No. 39 and 58 showed an extensive cross reactivity at high concentration of normal colon nuclear matrix. The antigens were determined to be a heterogeneous group of proteins with a major antigen of molecular weight of 140,000 for antibody No. 54 subclone 54-c-5-6 and two major antigens of molecular weight for 105,000 and 116,000 for antibody No. 39 subclone 39-d-11-12. Immunohistochemical localization of the antigens by the horseradish peroxidase bridge method demonstrated their presence in the nuclei.

Antibodies, Monoclonal↗

PubMed1986

Mechanism of the dexamethasone effect on alpha-fetoprotein gene expression in McA-RH8994 rat hepatoma cells.

We have investigated the mechanism by which dexamethasone (DEX) increases alpha-fetoprotein (AFP) secretion by the McA-RH8994 rat hepatoma cell line. In contrast to the 7777 cell line, DEX was found to increase the amount of hybridizable cytoplasmic AFP mRNA in McA-RH8994 cells. This effect was dose-dependent, reversible after removal of DEX, and was partially blocked by progesterone in excess, indicating a receptor-mediated response. In vitro nuclear transcription experiments showed that DEX increased AFP gene transcription in McA-RH8994 cells. Cycloheximide blocked the effect of DEX on AFP mRNA levels in both the McA-RH8994 and 7777 cell lines, showing that ongoing protein synthesis is required for glucocorticoids either to enhance or to suppress AFP gene expression in these cell lines.

Animals↗

PubMed1986

Keratin species in type II pneumocytes in culture and during lung injury.

A detailed understanding of alveolar epithelial cell transitions during remodeling after lung injury requires the identification of specific markers. We have developed a panel of monoclonal antibodies against species of the intermediate filament protein, keratin. These individual species are recognized markers of the state of differentiation of various epithelial cells. These and complementary protein analytic methods have been applied to studies of isolated, enriched Type II pneumocyte preparations as well as to normal and injured lung tissues. Monoclonal antibody 24A3, initially raised against Morris hepatoma 7777 keratins, decorated a filament network in isolated cultured rat Type II pneumocytes by indirect immunofluorescence; it reacts by 2-dimensional polyacrylamide gel immunoblot procedures with an acidic, 46,000-dalton keratin. Monoclonal antikeratin antibodies AE1 and AE3, raised against human epidermal keratins, reacted poorly with isolated Type II cells; however, AE3 reacted by immunoblot technique with the 55,000-dalton keratin subclass. The bronchial epithelium reacted intensely with 24A3 as well as with a mix of AE1 plus AE3 in ethanol-fixed, paraffin-embedded sections of normal and injured rat lung. Alveolar regions of normal lung reacted poorly with all 3 antibodies, however, as visualized by light microscopy. At the same time, very large, presumptive epithelial cells in the alveolar regions stained intensely with 24A3 3 days after intratracheal instillation of bleomycin, whereas thin cells lining the alveoli in injured regions were intensely reactive 14 days after bleomycin treatment. These elongated cells may represent Type II pneumocytes in the process of converting to Type I cells.

Animals↗

PubMed1985

alpha-Fetoprotein gene DNA-binding proteins.

The technique of protein blotting was used to study nuclear protein interaction with the alpha-fetoprotein (AFP) gene. The DNA-binding specificity was optimized by varying the amount of competitor DNA and the ionic strength. The specific binding of AFP gene DNA was observed for a set of Morris hepatoma 7777 nuclear proteins. Similar specificity was not seen for these same proteins in liver. In normal rat livers, however, two unique proteins were observed which displayed specific binding. The speculated involvement of these proteins in AFP gene regulation is discussed.

Animals↗

PubMed1985

Dexamethasone inhibits alpha-fetoprotein gene transcription in neonatal rat liver and isolated nuclei.

The effect of dexamethasone on rat alpha-fetoprotein (AFP) expression has been further examined. Quantitation of serum AFP levels from newborns treated with dexamethasone showed a dose-response relationship between the quantity of dexamethasone administered and the reduction in AFP serum level. RNA blots, utilizing cloned AFP cDNA as probe, showed a marked reduction in AFP mRNA in dexamethasone treated livers. The extent of AFP mRNA depletion was correlated with dexamethasone dosage. The effect of dexamethasone on AFP mRNA concentration was relatively rapid; a substantial reduction occurred 12 hours after a single injection. The effect of dexamethasone appeared to be irreversible as hormone withdrawal did not cause AFP mRNA levels to rise. One putative AFP nuclear RNA precursor was identified which rapidly disappeared following dexamethasone treatment. AFP mRNA synthesis was also diminished in nuclei transcribed in vitro. The direct inhibitory effect of glucocorticoid hormone on AFP gene transcription was demonstrated in a reconstituted cell-free nuclear system.

Animals↗

PubMed1985

Oncogene expression in rat hepatomas and during hepatocarcinogenesis.

In this study we examined oncogene expression in several Morris hepatomas. We found elevated transcripts for Ha-ras, Ki-ras, and the myc oncogenes, while the src oncogene was not increased. Northern analysis revealed transcript sizes of 1.4 kb for Ha-ras and 2.5 kb for myc. Analysis of primary tumors induced by the carcinogen 3'-methyl-4-dimethylaminoazobenzene (3'-MDAB) showed increasing oncogene transcript production with increasing length of diet. The results show enhanced ras and myc expression in rat hepatocellular carcinomas and during hepatocarcinogenesis.

Animals↗

PubMed1985

Colon adenocarcinoma-associated nuclear antigens.

Colon adenocarcinoma-associated chromosomal nonhistone proteins were determined in rat colon adenocarcinoma chromatin and in colon chromatin of Sprague-Dawley rats treated with 1,2-dimethylhydrazine (DMH) by the enzyme-linked immunosorbent assay method. The specific nuclear proteins are tissue-specific oncofetal antigens. They were also detected in fetal rat colon chromatin but not in adult rat colon chromatin. The specific tumor-associated nonhistone proteins were identified by immunoblot. The molecular weight ranges of these proteins were 79,000-89,000 and 40,000-56,000 daltons. Administration of DMH to Sprague-Dawley rats induced colon tumor and colon tumor-associated nonhistone proteins in the chromatin. However, DMH failed to induce colon tumor and tumor-associated nonhistone proteins in Lobund-Wistar rats.

Adenocarcinoma↗

PubMed1985

Effect of sodium butyrate on alpha-fetoprotein gene expression in rat hepatoma cells in vitro.

Sodium butyrate has been reported to induce cellular differentiation and reduce the tumorigenicity of certain tumor cells. We have examined the effects of butyrate on alpha-fetoprotein (AFP) gene expression in 7777 and McA-RH8994 rat hepatoma cells and have found that nontoxic concentrations of the drug decrease AFP mRNA levels in both cell lines. However, McA-RH8994 requires a 10-fold lower concentration (0.5 mM) of butyrate to affect a 50% reduction in AFP mRNA levels within 48 h. At 2 mM, sodium butyrate reduces AFP mRNA levels in McA-RH8994 cells by at least 90% after 48 h, while having little effect on the expression of either the 7S RNA or Harvey-ras genes. Time-course studies show that the effect of butyrate on McA-RH8994 AFP mRNA levels is immediate and is accompanied by an accumulation of cells in the G1/G0 phase of the cell cycle. Sodium butyrate was found to reduce AFP mRNA levels in both dexamethasone-treated 7777 and McA-RH8994 cells; dexamethasone decreases AFP mRNA levels in the former cell line and increases AFP mRNA levels in the latter. Therefore, it is unlikely that butyrate acts simply by reducing the dexamethasone receptor concentration in 7777 cells.

Animals↗

PubMed1984

Tissue specific control of alpha-fetoprotein gene expression.

The expression of the alpha-fetoprotein (AFP) gene was studied in rat liver and kidney. A significant level of AFP mRNA was found in neonatal liver and kidney, but not in adult tissues. Unlike liver the re-expression of AFP mRNA was not seen upon chemically induced regeneration of the kidney. Treatment of neonatal rats with dexamethasone caused a decrease in liver AFP mRNA levels, but a similar decrease was not apparent in kidney. Northern analysis revealed AFP mRNA size to be identical in neonatal liver and kidney. The results suggest different gene regulatory mechanisms in liver and kidney for the AFP gene.

Aging↗

PubMed1984

Hepatoma-associated nonhistone chromosomal proteins.

In order to compare nonhistone proteins in normal and neoplastic hepatocytes, we elicited antisera to Morris hepatoma 7777 dehistonized chromatin. By the enzyme-linked immunosorbent assay, the antisera demonstrated specificity for Morris hepatoma 7777 and little reactivity to normal rat liver chromatin. Morris hepatomas 7288c, 7800 and 5123tc shared some antigenic hepatoma nonhistone proteins. Neoplasia induced in rats fed 3'-methyl-4-dimethylaminoazobenzene changed the immunospecificity of the liver chromatin to a new type that was antigenically similar to Morris hepatoma 7777. Fetal rat liver chromatin and regenerating rat liver chromatin did not bind antibody. To further characterize the antigenic nonhistone proteins, we analyzed Morris hepatoma 7777 chromatin and normal rat liver chromatin by the immunoblot technique. Nonhistone proteins that demonstrated immunoreactivity were predominantly high molecular weight proteins.

Animals↗