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Biomedical subjects

D G Kaufman

Publications and source records attributed to D G Kaufman.

At least 91 records · Page 5Linked to original sources
PubMed1984

Biochemical studies of the tracheobronchial epithelium.

Tracheobronchial epithelium has been a focus of intense investigation in the field of chemical carcinogenesis. We have reviewed some biochemical investigations that have evolved through linkage with carcinogenesis research. These areas of investigation have included kinetics of carcinogen metabolism, identification of carcinogen metabolites, levels of carcinogen binding to DNA, and analysis of carcinogen-DNA adducts. Such studies appear to have provided a reasonable explanation for the susceptibilities of the respiratory tracts of rats and hamsters to carcinogenesis by benzo(a)pyrene. Coinciding with the attempts to understand the initiation of carcinogenesis in the respiratory tract has also been a major thrust aimed at effecting its prevention both in humans and in animal models for human bronchogenic carcinoma. These studies have concerned the effects of derivatives of vitamin A (retinoids) and their influence on normal cell biology and biochemistry of this tissue. Recent investigations have included the effects of retinoid deficiency on the synthesis of RNA and the identification of RNA species associated with this biological state, and also have included the effects of retinoids on the synthesis of mucus-related glycoproteins. Tracheal organ cultures from retinoid-deficient hamsters have been used successfully to indicate the potency of synthetic retinoids by monitoring the reversal of squamous metaplasia. Techniques applied to this tissue have also served to elucidate features of the metabolism of retinoic acid using high pressure liquid chromatography. In brief, formidable strides have been made in biochemistry specific to this important target tissue, despite the inability to acquire tracheobronchial epithelium in large quantities.

Animals↗
PubMed1984

The heterogeneity of LDH isoenzyme patterns of human uterine sarcomas and cultured sarcoma cell lines.

Both the tissue lactate dehydrogenase (LDH) isoenzyme pattern and total LDH activity were investigated in a series of human uterine sarcomas and cell lines derived from these tumors. Abnormal LDH isoenzyme patterns characterized by shifts toward the muscle isoenzyme forms (LDH4 and 5) were found in some, but not all, of the sarcomas. The tumors containing rhabdomyosarcomatous elements usually had normal or only slightly altered LDH isoenzyme patterns. The LDH isoenzyme patterns expressed by cultured cell lines established from the uterine sarcomas, in general, resembled those found in the primary tumors. These results suggest that the normal or near normal LDH isoenzyme distributions found in uterine sarcomas with rhabdomyosarcomatous differentiation may be a potential characteristic which may be used for distinguishing these tumors from other uterine sarcomas.

Adult↗
PubMed1983

Enhancement by TPA of phenotypes associated with transformation in carcinogen-treated human cells: evidence for a selective mechanism.

The tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA) was found to increase the incidence of phenotypic alterations induced by the direct-acting carcinogen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in human endometrial stromal cells. Following carcinogen treatment, changes in saturation density, gamma-glutamyltranspeptidase expression, morphology, and growth in selective media were enhanced in cell cultures subjected to continuous TPA exposure as compared to cultures receiving ethanol vehicle. This enhancement may have resulted, at least in part, from selection of cells altered by carcinogen, as evidenced by differences in the colony-forming abilities of MNNG-treated and control cultures after prolonged TPA exposure, and by differences in morphologic response to TPA challenge in these two populations.

Cell Aggregation↗
PubMed1983

Fractionation and characterization of DNA at sites of replication from rat liver nuclei.

Regenerating rat liver nuclei when sonicated and centrifuged in a Cs2SO4 gradient were fractionated into three distinct bands. These bands were designated as light band (LB), middle band (MB), and heavy band (HB) according to their density. LB and MB were shown to consist of large granular particles with varying electron densities, but LB also contained remnants of nuclear membrane. When analysed by gel electrophoresis, LB and MB displayed more than 35 bands of proteins. The third fraction, HB, consisted largely of small chromatin fibers and its proteins were predominantly the four histones of the nucleosomal core particle. Following short pulses with [3H]thymidine in vivo, the specific activity of DNA in LB and MB was significantly higher than that of bulk DNA contained in HB. DNA in all three fractions became equally labelled as the duration of the labelling interval increased beyond 30 min. Newly synthesized DNA was characterized by electrophoresis on analytical 1.7% acrylamide -0.5% agarose composite gels. After a 1-min labelling interval in vivo, 17% of the rapidly labelled DNA from LB and MB was stationary at the gel origin like replication forks from E. coli, but only 3% of HB DNA had zero mobility. Electron microscopy confirmed the presence of DNA replication forks in LB, MB, and HB. With increasing time of synthesis the proportion of labelled DNA exhibiting zero mobility decreased in all three fractions. Denaturation of DNA or digestion of single-stranded DNA with S1 nuclease released newly synthesized DNA from the gel origin. Ribonuclease was without effect. DNA recovered from LB and MB also had a higher molecular weight than the HB DNA. Together these results indicate (1) that LB and MB are enriched in newly replicated DNA; (2) that an increased proportion of newly replicated DNA in LB and MB is associated with DNA replication forks; and (3) that the replicating DNA recovered in LB and MB may be associated with other nuclear constituents in situ because this DNA appears to be protected from the more frequent chain breaks introduced into the bulk of chromatin (HB) by sonication.

Animals↗
PubMed1983

A comparison between the activation of benzo[a]pyrene in organ cultures and microsomes from the tracheal epithelium of rats and hamsters.

Epidermoid cancers of tracheal origin produced in Syrian golden hamsters and Fischer strain 344 rats are models for human bronchogenic carcinomas. These two species differ, however, in the sensitivity of their tracheal epithelia to tumor induction elicited by intratracheal benzo[a]pyrene (BP)-ferric oxide administration. The tracheas of hamsters are quite sensitive to the carcinogenic effects of BP-ferric oxide, but rat tracheas are apparently resistant to effects of comparable treatments by this route of administration. Rat tracheas are not completely resistant to polynuclear hydrocarbon carcinogenesis because in the heterotopic tracheal graft model, epidermoid carcinomas have been produced frequently. To determine whether differences in BP metabolism could explain this difference between species, quantitative kinetic and chromatographic studies of benzo[a]pyrene monoxygenase activity were carried out in epithelial microsomes and cells from organ cultures of rat and hamster tracheas. The Vmax was 2-fold greater in hamster tracheal cells than in rat tracheal cells whereas the Km values were identical. H.p.l.c. profiles from microsomes of rat and hamster tracheal epithelial cells incubated with BP exhibited extreme differences. Hamster tracheal microsomes produced large proportions of BP-quinones, BP-phenols, and BP-diols but rat tracheal microsomes produced mostly 3-OH BP. The total metabolic rate for BP in rat tracheal organ cultures was half that in cultures of hamster tracheas. The metabolites isolated in organ cultures of hamster and rat tracheas well reflected secondary reactions of conjugation and recycling. When evaluated with respect to the amount of tissue used, the most striking difference between rat and hamster tracheal organ cultures was in the amount of products which co-chromatographed with bay-region BP-tetrols. The amount of BP-tetrols produced by hamster tracheas was 0.22 pmol/mg tissue/24 h, and by rat, 0.012 pmol/mg tissue/24 h. The level of BP-DNA binding (in pmol/microgram DNA/24 h) catalyzed by hamster tracheal cells was 26.6 +/- 11.4, and by rat tracheas was 1.55 +/- 1.29. These interspecies differences associated with the formation of BP-diol epoxide, a presumed ultimate carcinogenic form of BP, are consistent with the differences between these two rodent species in susceptibility to carcinogenesis in this tissue.

Animals↗
PubMed1983

Characterization of a pure heterologous sarcoma of the uterus: rhabdomyosarcoma of the corpus.

A case of a pure heterologous sarcoma of the uterine corpus composed exclusively of rhabdomyosarcomatous elements has been studied by multiple morphologic and biochemical techniques. The neoplasm filled the endometrial cavity and protruded out the cervical os, but the myometrium was only superficially invaded. The tumor did not extend outside the corpus. The pathologic features are discussed in detail. Evidence of striated muscle differentiation could be identified on light microscopic and ultrastructural examination. Immunoperoxidase staining of tumor cells with antibodies to myoglobin were positive. Histochemical preparations for lactate dehydrogenase, succinic dehydrogenase, and acid phosphatase were also positive in neoplastic cells. Other stains gave equivocal or negative results. These findings are discussed in comparison with previous reports.

Acid Phosphatase↗
PubMed1983

Alterations of human endometrial stromal cells produced by N-methyl-N'-nitro-N-nitrosoguanidine.

Stromal cell cultures obtained from human endometrium were treated repetitively with N-methyl-N'-nitro-N-nitrosoguanidine in vitro at concentrations ranging from 0.5 to 4.0 micrograms/ml, and alterations in growth potential and morphology were analyzed. A single exposure to the carcinogen resulted in morphological evidence of toxicity and reductions in growth rates, plating efficiency, and saturation density as compared to solvent-treated control cells. Cytotoxicity was reduced after additional exposures to the carcinogen. Following repetitive treatments with N-methyl-N'-nitro-N-nitrosoguanidine, human endometrial stromal cells developed enhanced growth potential, the capacity to form macroscopic colonies in soft agar, and elevated gamma-glutamyltranspeptidase activity. Carcinogen-treated cells displayed atypical morphology characterized by irregularities in cell and nuclear size and shape, large bizarre nucleoli, increased nuclear:cytoplasmic ratios, and cellular crowding. Control cells did not display altered morphology or growth parameters even following multiple exposures to solvent and repetitive subculturing. These alterations in growth potential and morphology suggest that the cells are progressing towards preneoplastic and perhaps neoplastic transformation in vitro.

Agar↗
PubMed1982

Monolayer culture of human endometrium: methods of culture and identification of cell types.

Monolayer cultures can be established from human endometrial tissue after enzymatic dispersal into isolated glands or single cells. Three cell types that have distinct morphology by light and electron microscopy are observed in the resulting primary cultures. One cell type, an elongated spindle cell, is similar in appearance to fibroblasts derived from other tissues. A second cell type forms colonies of tightly cohesive cells, ranging in shape from oval to polygonal. These cells have typical organelles and junctional complexes characteristic of epithelial cells from the endometrium. The third cell type assumes a pavement-like appearance composed of polygonal cells when viewed by phase contrast microscopy, but lacks distinctive ultrastructural features of epithelial cells. These cells in culture resemble the endometrial stromal cell, the predominant cell type of the human endometrium in vivo. The epithelial cell does not survive subculturing but the other two cell types can be passaged through several generations and can be stored in liquid nitrogen and subsequently returned to culture.

Cells, Cultured↗
PubMed1982

Morphology and growth potential of stromal cell cultures derived from human endometrium.

Propagable cell cultures derived from human endometrial tissue were determined to contain cells predominantly of stromal cell origin based on their morphologic resemblance to endometrial stromal cells. These features included nexi, solitary cilia, and predecidual cytology. In addition to morphology the cell cultures retained a normal karyotype and responded to steroid hormones as evidenced by cellular aggregation. The stromal cells were evaluated for a variety of characteristics associated with transformed cells and seemed to be biologically normal without neoplastic phenotypes. Growth potential of the stromal cell cultures was also characterized in normal maintenance medium, in nutritionally depleted medium with reduced levels of calcium or serum, and in medium with increased levels of serum. The prolonged survival of the stromal cells in vitro coupled with the retention of in vivo characteristics and an absence of neoplastic phenotype provides a human cell system that is amenable to a variety of long-term experimental analyses.

Cell Aggregation↗
PubMed1982

DNA in proximity to the site of replication is preferentially alkylated in S phase 10T1/2 cells treated with N-methyl-N-nitroso-urea.

Replicating DNA is more susceptible to modification by N-methyl-N-nitrosourea (MNU), a spontaneously active methylating agent, than bulk DNA. This conclusion is supported by results from two different experimental approaches. First, synchronized C3H 10T1/2 clone 8 cells were treated in S phase with MNU and DNA replicated during the period of treatment was separated from bulk DNA. This was done by digesting the purified DNA with restriction enzymes and retaining the replication fork-associated DNA in nitro-cellulose filters. Second, synchronized C3H 10T1/2 clone 8 cells were exposed to 5-bromodeoxyuridine and [3H]MNU and the density-labelled, replicated DNA was separated in CsCl gradients. Both methods show 2.6 to 5.0 times more [3H]methyl adducts per nucleotide residue associated with replicating DNA than that expected from random methylation. These experiments were done at low MNU concentrations (0.018-0.115 mM) that did not cause any detectable inhibition of DNA synthesis or stimulation of repair replication in 10T1/2 cells.

Animals↗
PubMed1981

Benzo[a]pyrene metabolism in organ cultures of human endometrium.

Twenty-six specimens of normal human endometrium were obtained for studies of [3H]benzo[alpha]pyrene metabolism in short-term organ culture. The tissues were capable of converting benzo[alpha]pyrene (BP) to oxygenated derivatives which co-chromatographed with dihydrodiols, quinones and monohydroxy derivatives of BP. A sulfate conjugate of a monohydroxy BP was also found among the acetone/ethyl acetate soluble products. When the acetone/ethyl acetate soluble metabolites were analyzed using high-pressure liquid chromatography (HPLC), the proportion of metabolites were found to vary greatly from specimen to specimen. Eight women smoked cigarettes but the BP metabolite profiles from organ cultures of endometrial tissue from these women did not differ from those who did not smoke. The average value for sulfate conjugates of monohydroxy BP was significantly lower in endometrial tissue from three postmenopausal women (ages 49, 57 and 58) than among premenopausal women. Specimens from premenopausal women which had been in organ culture for 2 weeks prior to the addition of [3H] BP produced metabolite profiles similar to those of specimens from postmenopausal women with a greatly reduced proportion of BP-phenol sulfate conjugates. These data suggest that the absence of hormonal stimulus (i.e., advanced age or conditions of organ culture) may affect the metabolism of BP in human endometrial tissue.

Adult↗
PubMed1981

Benzo(a)pyrene binding to DNA in organ cultures of human endometrium.

Benzo(a)pyrene was found to bind to DNA in human endometrial tissue in vitro. Among specimens from 41 individuals examined, there was a 70-fold range in the observed specific activities of carcinogen binding to DNA. To determine whether this interindividual variability was correlated with the hormonally determined state of differentiation of the endometrial tissue, this population was subdivided to separate postmenopausal patients from premenopausal patients; among premenopausal patients, further division was made according to location within the menstrual cycle. Tissue obtained late in the proliferative phase or early in the secretory phase of the menstrual cycle had the highest mean specific activity of benzo(a)pyrene binding. In spite of the relatively small group sizes, the observed difference between this and the level of benzo(a)pyrene binding in the mid- and late secretory phases was statistically significant. The average binding level among the small number of patients studied who had entered a natural menopause was lower than the average binding for any of the subgroups of premenopausal patients and significantly lower than the mean for the whole population of premenopausal patients.

Adult↗
PubMed1981

Cycle-dependent removal of certain methylated bases from DNA of 10T1/2 cells treated with N-methyl-N'-nitro-N-nitrosoguanidine.

The loss of N-methyl-N-nitro-N-nitrosoguanidine (MNNG)-induced adducts from DNA was quantitated during the G1 and S phases of synchronously proliferating mouse 10T1/2 cells exposed to MNNG (2 microgram/ml) when they were in either confluence-induced arrest of proliferation or at the G2-S border. When treated at either time, N-7-methylguanine and O6-methylguanine were not excised from the template DNA during the subsequent S phase. However, both lesions were efficiently removed during the G1 phase immediately following exposure to MNNG, as well as during the second G2 phase after an S phase during which no loss occurrred. In contrast, N-3-methyladenine was lost rapidly during both the G1 and S phases following MNNG treatment. N-7 Methylguanine and O6-methylguanine were removed from logarithmically growing cell populations more slowly than from cells passing synchronously through the G1 phase. However, when the observed rate of loss in logarithmic cultures was corrected for the fraction of the logarithmic population located in the S phase, the rates of loss of the two methylated bases were the same as those observed in the G1 phase in synchronous cultures.

Animals↗
PubMed1981

Reversible inhibition of rat hepatocyte proliferation by hydrocortisone and its effect on cell cycle-dependent hepatocarcinogenesis by N-methyl-N-nitrosourea.

The susceptibility of hepatocytes to carcinogenesis in vivo may be influenced by the phase of the cell cycle at which carcinogen-induced damage is incurred. In order to better understand this relationship, hepatic cell proliferation in juvenile male Fischer 344 rats was charted following a two-thirds partial hepatectomy. For hepatocytes, two distinct waves of DNA synthesis occurred which were followed after 6 to 8 hr by waves of mitotic cell division. In contrast to this kinetic pattern, when hydrocortisone was given after the partial hepatectomy, the initial waves of DNA synthesis and mitosis by hepatocytes were each delayed by about 15 hr. In rats not given hydrocortisone, susceptibility to heptocarcinogenesis by N-methyl-N-nitrosourea was greatest at 20 hr after partial hepatectomy when the peak fraction of proliferating hepatocytes was in the S phase. By shifting the time of onset of DNA synthesis, the hydrocortisone treatments also shifted the time with greatest sensitivity to N-methyl-N-nitrosourea, with hepatocytes in late G1 or S again the most susceptible. Numerous tumors were also induced by N-methyl-N-nitrosourea in extrahepatic tissues, including intestine, Zymbal's gland, nervous system, kidneys, odontogenic tissues, and peritesticular mesothelium. The results illustrate the importance of cell proliferation in carcinogenesis and further point to the specific sensitivity of certain cell cycle phases.

Animals↗
PubMed1981

Risks of cervical intraepithelial neoplasia among DES-exposed women.

The increased occurrence of cervical intraepithelial neoplasia (CIN) in women with in utero exposure to diethylstilbestrol (DES) is described. Among 335 self-referred women exposed to DES, 14.9% (50) had a least 1 cytologic smear interpreted as CIN, and 16.2% (54) had at least 1 biopsy interpreted as CIN. Excluding those with grade 1 CIN, 8.7% (29) had at least grade 2 CIN on biopsy and 4.2% (14) had biopsies interpreted as grade 3 CIN. No invasive lesions were detected. Because the problem of differentiating true intraepithelial neoplasia from morphologically indistinguishable but benign lesions in DES-exposed women remains unresolved, the authors believe a conservative approach to treatment is still indicated. This study supports the recommendation that DES-exposed women merit continued thorough gynecologic evaluation, including regular cytologic evaluation and colposcopy, with biopsies as indicated.

Adult↗