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F F Becker

Publications and source records attributed to F F Becker.

At least 37 records · Page 2Linked to original sources
PubMed1989

In vitro DNA cytosine methylation of cis-regulatory elements modulates c-Ha-ras promoter activity in vivo.

The effect of DNA cytosine methylation on promoter activity was assessed using a transient expression system employing pHrasCAT. This 551 bp Ha-ras-1 gene promoter region is enriched with 84 CpG dinucleotides, six functional GC boxes, and is prototypic of many genes possessing CpG islands in their promoter regions. Bacterial modification enzymes HhaI methyl transferase (MTase) and HpaII MTase, alone or in combination with a human placental DNA methyltransferase (HP MTase) that methylates CpG sites in a generalized manner, including asymmetric elements such as GC box CpG's, were used to methylate at different types of sites in the promoter. Methylation of HhaI and HpaII sites reduced CAT expression by approximately 70%-80%, whereas methylation at generalized CpG sites with HP MTase inactivated the promoter by greater than 95%. The inhibition of H-ras promoter activity was not attributable to methylation-induced differences in DNA uptake or stability in the cell, topological form of the plasmid, or methylation effects in non-promoter regions.

Animals↗
PubMed1989

Altered expression of acute-phase reactants in mouse liver tumors.

We studied the expression of eight liver acute-phase genes in spontaneous and diethylnitrosamine-induced mouse liver tumors (MLTs) under basal and induced conditions. Primary spontaneous and chemically induced MLTs were used for RNA isolation and histopathologic analysis. In the noninduced state, all MLTs showed similar levels of mRNA for albumin, serum amyloid A, alpha 1-acid glycoprotein, haptoglobin, and alpha-, beta-, and gamma-fibrinogens compared with control or background livers. The mRNA for the alpha 1-major acute-phase protein, however, was consistently elevated in both spontaneous and diethylnitrosamine-induced MLTs. The expression of these acute-phase reactants in 11 MLTs was examined following exposure of the mice to turpentine. The relative expression of these mRNAs in these MLTs varied widely compared with mRNA expression in controls. Though all MLTs expressed the same three species of fibrinogen mRNAs as did the controls, no MLT demonstrated downregulation of albumin mRNA levels, and only 1 of 11 MLTs showed a marginal increase in serum amyloid A mRNA levels. Synthesis of mRNA for alpha 1-acid glycoprotein and haptoglobin was intermediate. The study of the response of MLTs to specific acute-phase stimuli may make possible a better understanding of the basis for coordinated expression of acute-phase reactants and of the variable phenotypes associated with cancers.

Acute-Phase Proteins↗
PubMed1989

The modulation of astrocytic differentiation in cells derived from a medulloblastoma surgical specimen.

Medulloblastomas are cerebellar tumors which are primarily composed of sheets of uniform, small malignant cells and may have astrocytic, neuronal or no features typical of these cell types. The assessment of astrocytic differentiation in medulloblastoma rests largely on the detection in malignant cells of glial fibrillary acidic protein (GFAP), a marker present in the later stages of normal astrocyte differentiation. It is still not known whether cells that do not contain GFAP in medulloblastomas with astrocytic differentiation correspond to highly proliferative astrocyte progenitors in maturation arrest at earlier stages of differentiation. The purpose of the current study was to examine whether cells in short term culture derived from a medulloblastoma tumor specimen with astrocytic differentiation were of the astrocytic lineage and if so, whether they represented proliferative astrocyte progenitors which would morphologically and antigenically mature in response to differentiating agents. A portion of tumor specimen from a 10-month-old child with recurrent posterior fossa medulloblastoma (RB2) that contained GFAP focally in tumor cells was grown in monolayer culture. We examined cellular structure and appearance of western immunoblotting and immunohistochemical studies for GFAP and neuron-specific enolase (NSE) in RB2 cells before and after treatment with retinoic acid (RA) and dibutyryl cyclic AMP (dBcAMP). RB2 in culture consisted of small polygonal cells (93%), large flat cells (3%), and polygonal cells with cytoplasmic processes (4%). In untreated RB2, 30% of cells expressed GFAP and staining for NSE was negative. RA treatment produced flattened cells and decreased GFAP. DBcAMP reversibly induced fine cytoplasmic processes containing GFAP in 85% of cells within 96 h. Neither agent induced NSE. The results suggest that cultured cells which are derived from a medulloblastoma with astrocytic differentiation do not spontaneously differentiate but that treatment with dBcAMP suppresses proliferation, enhances cytoplasmic process formation and increases cytoplasmic GFAP. Cells in culture and in medulloblastoma tumor specimens which do not contain GFAP may represent astrocyte progenitors in maturation arrest.

Astrocytes↗
PubMed1989

[The removability of pesticides during the production of dialysis water (2)].

In many cases it can be demonstrated that the amount of plant protectives and plant treatments (pesticides) in drinking-water exceeds the permitted levels of the drinking-water decree which will be effective on October 1st, 1989. These components are in parts toxicologically important. Therefore, an examination was made on how far pesticides are removed during the conventional purification of dialysis water, but especially during the reverse osmosis. Retention rates of a reverse osmosis plant for 14 different pesticides were discovered which were used in different concentrations and compositions. In part 2 of this contribution the results of the investigation are presented. The figures demonstrate that almost all of the examined components were retained with an effectiveness of 92-98%. The elimination efficiency did not depend on the basic concentration of the pesticides. After an initial phase of 50 h duration, the permeat concentration reached a constant value which did not alter even after more than 700 h.

Dialysis↗
PubMed1989

[Reducing chloramines in drinking water].

Monochloramine is produced when drinking water containing ammonium is chlorinated. It has long been known that activated charcoal destroys monochloramine. However, exact data for the dimensioning of a dechloramination plant were lacking. Four different commercially available activated charcoals were characterized (size of particle, iodine number) and examined for their effectiveness in removing monochloramines. The degradation of monochloramine by active charcoal is based on a chemical reaction of first order between carbon and monochloramine. The types of activated charcoal considerably differ in terms of reaction velocity, due in part to the mean granular size. The final concentration of the monochloramines is influenced only by their length of stay in the activated carbon filter, the temperature and the inflow concentration. A mathematical model describes the dependence of the degradation rate of the monochloramines on various factors. With its aid a nomogram can be established with which, simply and quickly, the activated charcoal needed in a concrete case can be determined.

Charcoal↗
PubMed1989

[Ability to remove pesticides in the production of dialysis water (1)].

In many cases it can be demonstrated that the amount of plant protective and plant treatment substances (pesticides) in drinking water exceeds the permitted levels of the drinking water ordinance which will come into effect on October 1st, 1989. Since some of these components are of toxicological relevance, an investigation was done on how far pesticides are removed during conventional purification of dialysis water, and especially during reverse osmosis. The retention rates of a reverse osmosis plant for 14 different pesticides applied in different concentrations and compositions were determined. Almost all of the substances examined were retained with an effectiveness of 92-98%. The elimination efficiency did not depend on the initial concentration of the pesticides. After an initial phase of 50 h duration, the concentration in the treated water reached a constant value which no longer changed even after more than 700 h. In part 1 of this contribution at first the fundamentals of dyalisis water purification are reviewed and a selection of the pesticides to be investigated is carried out. In addition experimental set up and procedure are described.

Filtration↗
PubMed1988

Defective DNA ligase I in Bloom's syndrome cells. Simultaneous analysis using immunoblotting and the ligase-[32P]AMP adduct assay.

Cells from patients with Bloom's syndrome (BS), an autosomal recessive disorder associated with an increased risk of cancer, exhibit genomic instability. Increased numbers of sister-chromatid exchanges (SCE) and delayed DNA chain maturation are typically observed in BS cells. To elucidate the basis for the previously reported decreased DNA ligase I activity in BS cells, simultaneous immunoblot and activity assays for ligase-[32P]AMP adduct formation were performed on extracts from BS and normal lymphoblastoid cell lines. Immunoblot analysis using antibody to DNA ligase I indicate that the amount of the major reactive protein (98 kDa) in normal and BS cells is similar. However, a 50-90% decrease was observed in the ligase activity of the 98-kDa polypeptide in high-SCE BS cells (HG1514 and GM3403c). In contrast, the activity in low-SCE BS cells (HG1554) did not differ significantly from that in normal cells. The data, together with mixing experiments, indicate that the defect in BS ligase I is due at least in part to the loss of ATP binding and/or hydrolytic activity and not to differences in numbers of protein molecules or inhibitory substances. These results suggest that mutation of the DNA ligase I gene may account for the primary metabolic defect in BS.

Adenosine Monophosphate↗
PubMed1988

Analysis of DNA adducts in putative premalignant hepatic nodules and nontarget tissues of rats during 2-acetylaminofluorene carcinogenesis.

Exposure of rats to a standard four-cycle feeding regimen of 0.06% 2-acetylaminofluorene (AAF) results in the formation of putatively premalignant hepatic nodules, but the types and magnitude of DNA adducts formed in these nodules has not been previously examined. By using a sensitive 32P-adduct assay (R. C. Gupta, Cancer Res., 45: 5656-5662, 1985), we analyzed the DNA adduct lesions in individual hepatic nodules at various times during and after exposure to AAF. Kidney, spleen, and testis were included as nontarget tissues. No qualitative difference was observed in the DNA adducts found in hepatic nodules and nontarget tissues; however, quantitative differences occurred. At least one unknown and two known (dG-C8-AF and dG-N2-AAF) DNA adducts were detected, with dG-C8-AF being predominantly (96-98%) formed, in all tissues examined. At the end of the first three weeks of AAF feeding, the concentration of the deacetylated adduct dG-C8-AF in liver (223 fmol/micrograms DNA) was found to be about 2, 6, and 5 times higher than in kidney, spleen, and testis, respectively. The concentration of the N2-acetylated adduct in liver (4.5 fmol/micrograms DNA) was 4-fold higher than in kidney and strikingly higher (51- and 42-fold, respectively) than in spleen and testis. At the end of the fourth feeding cycle, total DNA adducts measured in the hepatic nodules ranged from 30-100 fmol/micrograms DNA, while the "surrounding liver," kidney, spleen, and testis showed 235, 218, 62, and 28 fmol adducts/micrograms DNA, respectively. Sixty days following the cessation of AAF, the binding in both the persistent nodules and liver had decreased to 7% of their respective levels measured at the end of the fourth cycle, while adducts in kidney, spleen, and testis were 32%, 18% and 19%. After 88 days, the binding levels in the nontarget tissues declined further, but no additional adduct removal occurred in the nodules. Our data indicate that (a) although the metabolic apparatus for activation of AAF is diminished in the hepatic nodules, a significant level of adduct formation occurs in the cells of this putative, premalignant lesion, and (b) unlike in the nontarget tissues, repair processes in the premalignant nodules may not be operative several weeks after the cessation of AAF exposure.

2-Acetylaminofluorene↗
PubMed1988

UV-induced photoproducts of 5-methylcytosine in a DNA sequence context.

In order to detect possible m5C photoproducts, highly purified rat liver DNA-cytosine methyltransferase was used to specifically generate m5C with a radioactive methyl group. When these DNAs were subjected to a large dose (10 kJ/m2) of 254 nm or 302 nm ultraviolet light (UVB) to enhance the yield, two labeled photoproducts were detected and isolated by reverse phase HPLC after formic acid hydrolysis. Further studies using acetone as a triplet state sensitizer and UVB irradiation suggested that photoproduct II was activated via a triplet state while the more polar photoproduct I was not. Photoreversion of the purified photoproducts with 10 kJ/m2 254 nm light demonstrated the following reactions: Photoproduct I regenerated m5C, while photoproduct II is split and regenerated m5C and photoproduct I. These results suggest that photoproduct I is monomeric while photoproduct II dimeric, and from the latter's elution position possibly a cyclobutyl type dimer arising from a reaction with an adjacent cytosine. Using d[TTG] and d[Cm5CG] as models of typical sequences, irradiation with 10 kJ/m2 254 nm or 302 nm, respectively, gave rise to a small component having altered mobility in sequencing gels. The altered mobility trinucleotides were resistant to degradation by PI and micrococcal nucleases as expected from photodimerization of the pyrimidine bases. Furthermore, oligonucleotide substrates containing m5C were synthesized and shown to be susceptible to T4 endonuclease v action at locations consistent with d[Cm5C] photodimer formation when irradiated in the UVB range.

5-Methylcytosine↗
PubMed1988

Potassium inhibition of transforming protein P85gag-mos and reversal of the transformed phenotype in 6m2 cells.

K+ at high concentrations (52-72 mM hypertonic KCl) has been reported to induce reverse transformation in the 6m2 cell, which is a clone of normal rat kidney cells (NRK) infected with a temperature-sensitive transformation virus. When exposed to high K+, 6m2 cells grown at the permissive temperature (33 degrees C) exhibit normal morphology and reduced soft agar growth, characteristics of cells grown at nonpermissive temperature (39 degrees C). In the current study, flattening of cells and rearrangement of surface microvilli were demonstrated by scanning electron microscopy to occur within 6 hr of exposure to high K+, similar to the effect of temperature shift to 39 degrees C. Exposure to K+ resulted in a 90% inhibition of P85gag-mos-associated serine kinase activity within 5 min, with a subsequent reduction of up to 75% of the synthesis of this protein. These alterations in the putative transforming protein were similar to those induced by temperature shift and were considered to be the basis for retrotransformation. The cell microtubular system and F-actin cables were affected more slowly by K+ than by a temperature shift to 39 degrees C. The former did not achieve the fine reticulum network seen in NRK cells until 72 hr later, but the latter remained aberrant. The effect on the enzyme might be mediated by alteration in phosphorylation, but the mechanism by which kinase inactivation induces retrotransformation is not yet known.

Actins↗
PubMed1988

Failure of the viable yellow (Avy) and lethal yellow (Ay) genes to enhance chemical induction of thymic lymphomas.

Two dominant mutations in the agouti locus of the mouse, viable yellow, Avy, and lethal yellow, Ay, are associated with increased susceptibility to spontaneous as well as virally or chemically induced tumors. Regardless of the mechanism of initiation, this effect is dependent upon the presence of initiated cells. Although complex, the numerous physiologic alterations associated with these mutations offer an interesting model for the study of endogenous promotion. To date, it has been reported that these genes enhance the growth and early appearance of tumors in all organ sites wherein initiated cells exist, suggesting that the system was pan-promoting. In our current study, however, neither Avy nor Ay enhanced the appearance of chemically induced thymic lymphomas in two mouse strains of differing susceptibility. Thus, the concept of a pan-promoting effect is not valid. However, the presence of Ay did enhance the appearance of chemically induced hepatocellular tumors as has been reported previously for Avy. This striking difference of thymomas from other carcinogenic sequences may make possible a better understanding of the nature of susceptibility to endogenous promoting factors.

Animals↗
PubMed1987

Changes in cell surface charge and transmembrane potential accompanying neoplastic transformation of rat kidney cells.

Free flow electrophoresis measurements have been used to determine the surface charge density of normal rat kidney (NRK) cells and a clone of NRK, designated as 6m2, that exhibit a transformed phenotype at 33 degrees C and a non-transformed phenotype at 39 degrees C. A clone of 6m2, designated 54-5A4, which is transformed at both 33 degrees C and 39 degrees C was also studied. A surface charge density of -1.42 microC/cm2 was obtained for the NRK and non-transformed 6m2 cells at 39 degrees C, whereas at 33 degrees C values of -1.85 and -1.78 microC/cm2 were determined for the transformed 6m2 and 54-5A4 cells, respectively. It was found that 72% of the increased charge that appeared on the transformed 6m2 cells compared with the non-transformed 6m2 cells was RNAase sensitive. The time-dependent decrease in surface charge that accompanied the shift of the 6m2 cells from their transformed to non-transformed state was found to mirror the increase in transmembrane potential previously reported using a fluorescent dye technique, and was also comparable to the reported temporal changes in their morphology and virally-coded protein content.

Animals↗
PubMed1987

Heme enzyme patterns in rat liver nodules and tumors.

Chemically induced rat hepatocyte nodules and carcinomas have a reduced capacity to oxidize drugs. The reduction in monoxygenase activity results largely from the partial loss of cytochrome P-450, a heme-containing terminal electron acceptor. To determine whether the cytochrome P-450 deficit was indicative of an altered heme metabolism, we quantitated four heme-containing proteins in normal rat liver and in rat liver nodules and cancers induced by 2-acetylaminofluorene or diethyl-nitrosamine: cytochrome P-450; cytochrome bs; catalase (EC 1.11.1.6); and tryptophan 2,3-dioxygenase (EC 1.13.11.11). The amounts of these components in nodules were 45%, 88%, 50%, and 59% of normal liver, respectively; in 2-acetylaminofluorene-induced cancers, 65%, 74%, 64%, and 65%, respectively; and in diethylnitrosamine-induced cancers, 40%, 69%, 56%, and 52%. delta-Aminolevulinic acid synthase (EC 2.3.1.37), the rate-limiting enzyme in the heme synthetic pathway, and heme oxygenase (EC 1.14.99.3), a degradative enzyme, were also quantitated. The amounts of these enzymes in nodules were 95% and 138% of normal liver, respectively, whereas in 2-acetylaminofluorene-induced cancers, they were 47% and 233%, and in diethylnitrosamine-induced cancers, they were 50% and 175%. These data indicate that four nonmitochondrial liver hemoproteins were diminished to about the same extent in hepatic nodules and cancers. Nodules and cancers also demonstrated an increased capacity for heme degradation, while cancers also demonstrated a decreased capacity for heme synthesis. Thus, the resistance of nodules and tumors to P-450-activated cytotoxic agents may ultimately result from a disturbance in heme metabolism.

2-Acetylaminofluorene↗
PubMed1986

Protective role of thiols in carcinogen-induced DNA damage in rat liver.

Biological thiols are known to play an important role in the detoxification of xenobiotics, including chemical carcinogens. To determine the influence of cellular thiols on carcinogen induction of hepatic DNA damage in the rat, diethylmaleate (DEM) administration was used to deplete intracellular glutathione (GSH). The effects of administration of the synthetic thiols, N-acetylcysteine (NAC) and alpha-mercaptopropionylglycine (alpha MPG), on the induction of DNA lesions were also examined. Pretreatment with DEM reduced liver GSH levels by greater than 70%. As assessed by the technique of alkaline elution, subsequent administration of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) resulted in DNA damage 4 h post MNNG treatment which was 4- to 8-fold greater than that induced in the livers of rats treated with MNNG alone. However, DEM pretreatment had little effect on the extent of DNA damage induced by methylnitrosourea (MNU). DEM alone did not cause any measurable DNA damage. Pretreatment with alpha MPG or NAC reduced MNNG-induced DNA damage by as much as 77%. In contrast, MNU-induced DNA damage was increased by alpha MPG treatment whereas NAC treatment was without effect. These results indicated that in the rat liver, the activity of some DNA alkylating agents may be modulated in varying degree by the concentration of intracellular thiols. These data support the notion that thiols play an important role in protection against carcinogen damage, and that synthetic thiols such as alpha MPG and NAC may be useful as anti-carcinogenic agents against certain carcinogens.

Acetylcysteine↗
PubMed1986

Progression of tumor histiotype during mouse hepatocarcinogenesis associated with the viable yellow (Avy) gene.

Increasing attention has been focused recently upon those factors in carcinogenesis that are responsible for the proliferation of initiated cells and the increasingly aberrant phenotype that they progressively manifest. The agouti locus allele Avy (viable yellow) has been shown to be associated with conditions which favor promotion of cells that have been initiated by a wide variety of causes, in many organs, but has not been previously associated with tumor progression in those systems. In the current study, the presence of the Avy gene in a strain of mice not normally predisposed to hepatocarcinogenesis, C57BL/6N was, for the first time, associated not only with much earlier appearance, but with progression of the histiotype of hepatic tumors, following neonatal administration of diethylnitrosamine. At 52 weeks, 28 C57BL/6N mice demonstrated 7 mouse liver tumors 0.5 cm or greater in diameter, all of more benign histiotype, without associated metastasis. The 31 C57BL/6N-Avy demonstrated 194 mouse liver tumors at that time, 22% of which were of malignant histiotype, 19% of which were associated with metastasis. This system would appear to offer the possibility of identifying the underlying mechanisms for components of the carcinogenic process. In addition, the C57BL/6N-Avy mouse appears to offer advantages as a test animal in bioassay procedures that use the liver as a target organ. Thus, it represents a mouse with little or no spontaneous predisposition to hepatocarcinogenesis, with a predicted short lag period toward response to hepatocarcinogens.

Animals↗
PubMed1986

Heme enzyme patterns in genetically and chemically induced mouse liver tumors.

Chemically induced rat hepatocyte nodules and hepatomas have repeatedly been shown to be deficient in Phase I drug-metabolizing enzymes. Some of these reduced activities are attributable to a diminution of the heme-containing terminal electron acceptor, cytochrome P-450. We recently demonstrated that spontaneous mouse liver tumors exhibit the same deficiency. Therefore, chemically induced and spontaneous liver tumors share common metabolic alterations which are likely to represent intrinsic characteristics of the tumorigenic process and are independent of its etiology. To determine whether the cytochrome P-450 deficit was the result of an altered heme metabolism, we quantitated four heme-containing proteins in normal mouse liver, spontaneous mouse liver tumors, and those induced by a single injection of diethylnitrosamine: cytochrome P-450; cytochrome b5; tryptophan 2,3-dioxygenase (EC 1.13.11.11); and catalase (EC 1.11.1.6). The amounts of these components in spontaneous tumors relative to normal liver were 0.35, 0.68, 0.76, and 0.51, respectively. Similar values were obtained with chemically induced tumors. The enzymes delta-aminolevulinic acid synthase (EC 2.3.1.37), the rate-limiting enzyme in the heme synthetic pathway, and heme oxygenase (EC 1.14.99.3), a degradative enzyme, were also quantitated. The amounts of these enzymes in spontaneous tumor relative to liver were 0.49 and 1.51, respectively. Again, similar values were observed for the chemically induced tumors. Alteration of the latter two enzyme activities may be sufficient for the altered hemoprotein patterns seen in mouse liver tumors. Further, this pattern of metabolic alteration is common to both chemically induced and spontaneous tumors. Thus, tumor resistance to cytotoxic agents activated by the monooxygenase system is not necessarily induced by exposure to these agents, nor as a result of selection.

5-Aminolevulinate Synthetase↗