Biomedical subjects

D Medina

Publications and source records attributed to D Medina.

At least 109 records · Page 6Linked to original sources

PubMed1992

Nuclear type II sites and malignant cell proliferation: inhibition by 2,6-bis-benzylidenecyclohexanones.

Methyl-p-hydroxyphenyllactate (MeHPLA) is a bioflavonoid and/or tyrosine metabolite which may regulate cellular growth and proliferation through interactions with nuclear type II sites. Our current studies suggest that type II sites may function as MeHPLA receptors which are localized on the nuclear matrix, and occupancy of this binding site by MeHPLA directly correlates with the inhibition of normal and malignant cell proliferation. This ligand is inactivated by MeHPLA esterase in mammary tumors, resulting in a deficiency in MeHPLA, high quantities of unoccupied type II sites, and uncontrolled cellular proliferation. For these reasons we synthesized 2,6-bis((3,4-dihydroxyphenyl)methylene)-cyclohexanone (BDHPC) and 2,6-bis((3-methoxy-4-hydroxyphenyl)-methylene)cyclohexanone (BMHPC) for assessment as nuclear type II site and cell growth antagonists. These two esterase stable cyclohexanone derivatives, which bind to nuclear type II sites with high affinity (Kd 1-7 nM), mimic MeHPLA as cell growth-regulating agents. Dose-dependent occupancy of type II sites in MCF-7 human cells by BDHPC and BMHPC directly correlated with the inhibition of cell proliferation, and administration of BDHPC by silastic implant inhibited mouse mammary tumor growth in vivo. These findings demonstrate that esterase-stable type II antagonists such as BDHPC and BMHPC inhibit mammary cancer cell proliferation in vitro and in vivo and support earlier studies demonstrating that MeHPLA and functionally related compounds may regulate malignant cell proliferation at the level of this binding site.

Animals↗

PubMed1992

Development of mammary preneoplasias in vivo from mouse mammary epithelial cell lines in vitro.

A series of mouse mammary epithelial cell lines has been established by a protocol that gives highly reproducible results. The mammary epithelial cell lines, designated as FSK lines, were judged to be epithelial based on positive immunostaining for keratin-intermediate filaments, negative immunostaining for vimentin-intermediate filaments, hormonal induction of casein, and the ability to exhibit ductal and alveolar mammary morphogenesis in vivo. The FSK cell lines are dependent on epidermal growth factor and insulin in a low serum (1%) medium. Conditioned medium from spindle cell cultures replaced the requirement for serum and increased the growth of FSK3 and FSK4 4-5 times in collagen gels and 12-14 times in monolayer culture, respectively. Following injection into the mammary fat pad at passages 2-11, the FSK cell lines generated stable transplantable hyperplastic alveolar outgrowth lines. The in vivo outgrowth lines were judged as preneoplastic based on their stable alveolar morphology in vivo and an increased susceptibility for tumorigenesis. The FSK cell lines and their derivative in vivo outgrowth lines provide a new and potentially productive system to examine critical molecular alterations involved in the development of mammary preneoplasias. Furthermore, the reproducibility of the in vitro culture system provides the assurance that stable cell lines of mouse mammary epithelial cells can be generated easily and at will.

Adenocarcinoma, Bronchiolo-Alveolar↗

PubMed1992

The effects of deprenyl on methamphetamine-induced dopamine depletions.

The effects of deprenyl on methamphetamine-induced dopamine depletions were studied in mice. Four SC injections of 12.5 mg/kg of methamphetamine at two-hour intervals caused substantial (72-82%) and long-lasting depletions of striatal dopamine. Pretreatment with either 25 or 40 mg/kg of deprenyl did not significantly alter the magnitude of this depletion. These results indicate that, unlike what is observed following MPTP, there is no protection afforded dopaminergic cells by deprenyl pretreatment in the methamphetamine model of parkinsonism.

Animals↗

PubMed1991

Levels and 75Se-labeling of specific proteins as a consequence of dietary selenium concentration in mice and rats.

Selenium-labeled proteins (SLP) distinct from glutathione peroxidase (GSH-PX) recently have been purified and partially characterized. Antisera to two SLP, a 56-kDa and a 14-kDa protein, were generated in rabbits and used to examine expression of these proteins as a consequence of dietary selenium concentration (0.02, 0.2, 2.0 ppm) in mice and rats. Additionally, the kinetics of 75Se labeling in plasma, liver, kidney, and mammary gland were examined over a 40-hr time period as a function of dietary selenium concentration. A plasma 57-kDa protein was labeled by 30 min after 75Se injection and reached maximum labeling by 4 hr. The cellular 56-kDa and 14-kDa proteins, as well as GSH-Px, labeled progressively over 40 hr starting between 1 and 4 hr after injection. In general, the 56-kDa and GSH-Px followed similar labeling patterns, whereas the 14-kDa protein was labeled less and was not labeled in discernible quantities until 40 hr. The extent of labeling of all proteins was inversely proportional to the dietary selenium concentration and was probably a reflection of different endogenous selenium body pools. The most important observation was generated by the immunoblot data. The amount of 56-kDa and 14-kDa proteins as detected and measured on immunoblots was not a function of dietary selenium concentration. This result suggests that the synthesis and maintenance of the 56-kDa and 14-kDa proteins are not selenium dependent, a characteristic which distinguishes the two proteins from GSH-Px. The single exception to the above results was the 40% decrease of liver 14-kDa protein concentration in carcinogen-treated rats fed 2.0 ppm of selenium. An organic selenium compound, selenobetaine, did not lead to a decrease under similar conditions. In 15 rat mammary tumors induced by 7,12-dimethylbenzanthracene and analyzed on immunoblots, the SLP-56 was undetected in 5 cases and appeared as two bands (56,000 Da, 50,000 Da) in 10 cases. This latter result raises the possibility that the expression of SLP-56 may be altered in mammary tumors as compared with normal mammary gland.

Animals↗

PubMed1990

Methyl p-hydroxyphenyllactate and nuclear type II binding sites in malignant cells: metabolic fate and mammary tumor growth.

Previous studies in our laboratory demonstrated that methyl p-hydroxyphenyllactate (MeHPLA) is an important cell growth-regulating agent which binds to nuclear type II binding sites in normal and malignant cells. Furthermore, this compound is deficient in a variety of rat and mouse mammary tumors and human breast cancer preparations, and this deficiency correlates with the loss of regulatory control. The present studies were performed to examine the metabolic fate of [3H]MeHPLA in mouse mammary tumors. Stable analogs of this compound such as 4,4'-dihydroxy benzylidene acetophenone were also assessed for nuclear type II site binding affinity and their ability to inhibit mammary cancer cell growth and proliferation in vitro and in vivo. The results demonstrate that mouse mammary tumors contain esterase activity which hydrolyzes MeHPLA to p-hydroxyphenyllactic acid, and this was the only major metabolite detected in these tumor preparations in vitro or in vivo. 4,4'-Dihydroxy benzylidene acetophenone, an esterase-stable MeHPLA analog, was found to bind with high affinity to nuclear type II sites but not the estrogen receptor, was capable of occupying type II sites in cultured MCF-7 cells, and inhibited the proliferation of these cells in concentrations which directly correlated with type II binding site occupancy. Similarly, 4,4'-dihydroxy benzylidene acetophenone administration by silastic implant or injection resulted in a dose-dependent inhibition of the growth of transplantable mammary tumors in mice, suggesting that this stable analog mimicks MeHPLA as a cell growth-regulating agent. Taken together, these results suggest esterase hydrolysis of MeHPLA in mammary tumors may result in a deficiency in this compound which correlates with a loss of regulatory control.

Animals↗

PubMed1990

Influence of mammary cell differentiation on the expression of proteins encoded by endogenous BALB/c mouse mammary tumor virus genes.

The interactions between differentiation-associated cellular events in the intact mammary gland or in cultured mammary cells and the post-transcriptional activity of the endogenous mouse mammary tumor virus (MMTV) loci were investigated. The transcriptional activities of the endogenous MMTV proviruses of the BALB/c mouse strain (Mtv-6, Mtv-8 and Mtv-9) appear to be regulated differentially during pregnancy-induced mammary gland development (J.E. Knepper, D. Medina and J.S. Butel, J. Virol. 59, 518-521, 1986). Analysis of MMTV-specific proteins at various stages of mammary gland development (virgin, midpregnant, lactating, regressing) established the presence of steady-state levels of a 67,000-Mr env precursor-type polypeptide at all physiological stages. However, processing to lower-molecular-weight env-specific proteins, including a predominant 50,000-Mr species, was detected only with the transition to the functional mammary gland phenotype. The contributions of cell proliferation, cell-matrix interactions, and modulation of functional activity to the pattern of endogenous MMTV protein expression were investigated using a 3-dimensional collagen type I culture system. Growth and cell-matrix interactions (cell polarization, lumen formation) leading to formation of 3-dimensional duct-like structures were permissive for the synthesis and processing of MMTV-specific proteins; accumulation of high levels of the 50,000-Mr env-specific polypeptide was associated with the onset of the fully functional mammary cell phenotype. Expression of MMTV-specific proteins was not due to amplification of a specific cell subpopulation. The potential of the full-length Mtv-8 and Mtv-9 proviruses to be transcribed, as indicated by their methylation status, was not dramatically different between differentiated and undifferentiated mammary cells in culture. This study indicates that MMTV transcriptional activity is reflected at the protein level in mammary tissue of BALB/c mice and that viral protein synthesis and processing may serve as important markers of different physiological stages of mammary epithelial cells. These observations also suggest a general approach to the examination of potential modulatory effects of cellular interactions (cell-cell, cell-matrix or both) known to be important in various differentiated epithelial cell systems for the expression of viral genes.

Aldosterone↗

PubMed1990

DNA sequencing of a mouse liver protein that binds selenium: implications for selenium's mechanism of action in cancer prevention.

Complementary DNA clones for liver protein 56K (SLP-56) were isolated by screening a lambda Zap mouse liver library. The cloned cDNAs represented the complete message. The correct reading frame was verified by alignment of the deduced amino acid sequence with that of peptides sequenced from the purified protein. The primary sequence has not been reported previously since homologous DNA sequences were not found in GenBank. Most importantly, the DNA sequence did not contain an in-frame TGA codon that would code for seleno-cysteine, as occurs in the prototypic selenoprotein, glutathione peroxidase. Hydropathy analysis suggested the protein was not a membrane-spanning protein. SLP-56 was previously localized as a cytosolic-soluble protein on the basis of cell fractionation experiments. The results suggest that SLP-56 is different from proteins whose synthesis and concentration are dependent upon selenium and require TGA to encode for selenocysteine. In this respect, SLP-56 appears to be similar to liver fatty acid binding protein (SLP-14) for which selenium is a ligand. Our working hypothesis is that selenium exerts its inhibitory effects on cell growth by modulating the properties of existing growth regulatory proteins. The two proteins that are readily labeled by selenium in many rodent tissues, SLP-56 and SLP-14, would fit into this category.

Amino Acid Sequence↗

PubMed1990

Activation of H-ras oncogenes in preneoplastic mouse mammary tissues.

The mammary hyperplastic outgrowth (HOG) line C4, resulted from serial transplantation of a hyperplastic alveolar nodule which arose in a dimethylbenz(a)anthracene (DMBA) treated mouse. The immortalized C4 outgrowth line, on transplantation into syngeneic mice, develops as preneoplastic, hyperplastic outgrowths and subsequently into malignant carcinomas after a long latent period (greater than 6 months). Treatment of mice carrying C4 HOG transplants with DMBA resulted in a reduced latent period for tumor development (less than 3 months) and an increased tumor incidence. DNA's from C4 HOGs and mammary carcinomas of untreated as well as DMBA-treated mice were analyzed for the presence of oncogenes by the NIH3T3 focus forming assay. Transforming H-ras genes were detected in two of 6 preneoplastic HOGs and 10 of 12 carcinomas from DMBA-treated mice. DNAs from neither the HOGs nor the tumors from untreated mice were positive in this assay. The H-ras locus was then directly examined in the 61st codon by in vitro amplification of each of the tissue DNAs using PCR. The location of the activating mutation was determined by hybridization of amplified DNA to mixed sequence oligonucleotide probes. The specific nature of the mutation was defined by RFLPs using XbaI, TaqI and Sau96I restriction enzymes. Six of the H-ras oncogenes in DMBA-promoted tumors were activated by commonly observed A to T transversions at the 61st codon, while five (including an additional tumor with H-ras oncogene revealed by PCR analysis) contained novel A to G transitions. The H-ras oncogene in one DMBA-treated HOG sample was activated by A to T while the second contained an A to G mutations, representative of both modes of mutational activation involved in this model of mammary tumorigenesis. In summary, DMBA-induced point mutated H-ras oncogenes appear to potentiate the progression of hyperplastic outgrowths (HOG) to mammary carcinomas.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1989

A 14-kilodalton selenium-binding protein in mouse liver is fatty acid-binding protein.

In a previous study, we purified three selenium-binding proteins (molecular masses 56, 14, and 12 kDa) from mouse liver using column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The aim of the present study was to determine the amino acid sequence of the 14-kDa protein thereby establishing any relationship with known proteins. Although the amino terminus of the 14-kDa protein was blocked, separate in situ digestions of the protein with endoproteinases Glu-c and Lys-c gave overlapping peptides that provided a continuous sequence of 93 amino acids. This sequence exhibited a 92.5% sequence homology with rat liver fatty acid-binding protein. In situ enzymatic digestion and partial sequencing of a 12-kDa selenium-binding protein revealed identical homology to the 14-kDa protein. The 14-kDa protein bound specifically to an oleate-affinity column from which the protein and 75Se coeluted. Delipidation or sodium dodecyl sulfate treatment failed to remove 75Se from the protein, indicating that the selenium moiety was tightly bound to the protein. These observations confirm that the mouse liver selenium-binding 14-kDa protein is a fatty acid-binding protein. The nature of the selenium linkage to the protein still needs to be explored.

Amino Acid Sequence↗

PubMed1989

Spontaneous progression of hyperplastic outgrowths of the D1 lineage to mammary tumors: expression of mouse mammary tumor virus and cellular proto-oncogenes.

Mammary cancer in mice is characterized by progression through defined stages of preneoplasia, with the most common preneoplastic stage being the hyperplastic alveolar nodule (HAN). We determined the relative levels of RNA expression of various cellular proto-oncogenes and endogenous mouse mammary tumor virus genes in outgrowths and tumors of three sublines of the transplantable D1 HAN preneoplastic outgrowth line. The three sublines differed in relative tumor-producing capabilities. Subline D1B produced a high incidence of tumors with short latency periods, whereas sublines D1C and D1D produced low incidences of tumors with long latency periods. No consistent alteration in proto-oncogene expression correlated with relative tumorigenicity, although tumors frequently contained higher levels of one or more proto-oncogene transcripts as compared with preneoplastic tissue. Slightly elevated (2- to 6-fold) levels of different oncogene transcripts were detected in 13 of 17 tumors as compared with outgrowth tissue, including abl (2 tumors), fps (5 tumors), Ha-ras (6 tumors), and Ki-ras (8 tumors). One tumor contained 45 times more Ki-ras-specific RNA than outgrowth tissue because of a comparable amplification of Ki-ras DNA sequences. Elevated levels of Ha-ras occurred more frequently in tumors of a high-incidence subline than in a less-aggressive subline (5/10 vs 1/7), but this difference was not statistically significant. However, consistent changes in MMTV expression accompanied progression from preneoplastic tissues to mammary tumors. All 17 tumors displayed reduced levels of the MMTV-specific long terminal repeat (LTR) transcript (1.6 kb) as compared with HAN tissue; tumors with moderate levels of LTR transcript expressed the 3.8-kb envelope message as well, one not detected in HANs. Expression of the LTR transcript is apparently influenced by factors in addition to the methylation status of endogenous mouse mammary tumor virus genes, which was similar in outgrowths and tumors. As the survey of representative proto-oncogenes failed to identify a uniform change between HAN and tumors, it is likely that other genes are involved in tumor progression in the mammary gland.

Animals↗

PubMed1989

Clonal populations of the mouse mammary cell line, COMMA-D, which retain capability of morphogenesis in vivo.

Clonal populations were isolated from the mouse mammary cell line, COMMA-D, by transfection with a dominant-selectable gene, pSV2Neo, which confers resistance to the antibiotic, G418. Seven of twenty-four clones isolated retained the ability of the parental line to repopulate cleared mammary fat pads in vivo as ductal-alveolar hyperplasias. Two sublines designated CDNR2 and CDNR4 retained hyperplastic growth potential after multiple passages in vitro with low incidence of tumor formation. A third subpopulation, CDNR1, contained a single integration site for the pSV2Neo plasmid indicating a bonafide clonal origin for this subline. CDNR1 cells displayed heterogeneous growth phenotypes in vivo including hyperplasia, adenocarcinoma, and bone formation. Functional differentiation of CDNR1 cells organized as alveolarlike structures in vivo or on floating collagen gels in vitro was observed as determined by immunoperoxidase staining for the milk-specific protein, casein. Overall, the results indicate that a subset of cells from the COMMA-D cell line may be functionally analogous to stem cells existing in the mammary gland.

Adipose Tissue↗

PubMed1989

Time course of selenite metabolism in confluent cultures of mouse mammary epithelial cells.

The distribution of 75Se selenite in various macromolecules over time was evaluated in confluent cultures of the LMA mouse mammary epithelial cell line. 75Se label did not distribute equally to all fractions at the same time point. The label initially entered the selenonucleic acid fraction and subsequently increasing quantities were detected in the selenoprotein fraction. The time period of increasing inhibition of DNA synthesis correlated with increasing levels of selenoproteins. The major selenoprotein found in this time frame in confluent phase LMA cells was an acidic, 58K selenoprotein.

Animals↗

PubMed1989

Biochemical characteristics of mouse mammary tissues, preneoplastic lesions and tumors.

A number of metabolizing systems are measured in normal, preneoplastic and neoplastic mouse mammary tissues derived under three different conditions. These biochemical functions are considered to be important in the activation and detoxification of carcinogens and other xenobiotics and have been linked to the process of rat liver hepatocarcinogenesis. The cytochrome P450-dependent enzyme aminopyrine N-demethylase, consistently depressed in hepatocarcinogenesis models in mouse and rat, does not show a significant change among normal, preneoplastic and neoplastic mammary tissues. Glutathione and the enzymes of glutathione metabolism and utilization (e.g. glutathione-S-transferases and gamma-glutamyl transferase), active in the detoxification of xenobiotics, show no significant differences in carcinogen-induced tumors or in their homologous preneoplasias compared to control tissue. There is no increase in the anionic glutathione-S-transferase, a principal marker in rat hepatocarcinogenesis. The only observed biochemical change was a significant decrease in gamma-glutamyl cysteine synthetase the glutathione synthetic enzyme, in the carcinogen-induced preneoplastic and neoplastic line compared to control. Also inorganic glutathione peroxidase was lower in the preneoplastic, but not in the neoplastic tissues.

Aminopyrine N-Demethylase↗

PubMed1989

Evidence for two selenium-binding proteins distinct from glutathione peroxidase in mouse liver.

Labeling studies with 75selenium (75Se) have suggested the existence of selenium-binding proteins in addition to glutathione peroxidase (GSH-Px) in rodent tissues. Three selenium-binding proteins of apparent mol. wt 56, 14 and 12K on SDS-PAGE were isolated from mouse liver using Sephadex G-150 and DEAE-Sephadex chromatography. The proteins were electroeluted from SDS-PAGE gels and injected into rabbits to elicit antibodies. Western immunoblot experiments indicated that the 56K protein was distinct from the 14 and 12K proteins. The latter two proteins appeared to be immunologically related, perhaps as differentially processed variants. The 56 and 14/12K proteins appeared to be distinct from GSH-Px and the 57K plasma selenium-binding proteins. These results indicate that the mouse liver contains at least two selenium-binding proteins distinct from GSH-Px. The existence of the antibodies should permit experiments which help to examine the role of these proteins in the biological function of selenium in mammals.

Animals↗

PubMed1988

Selenite distribution in log and confluent growth phase murine mammary epithelial cells.

Two highly selenite sensitive cell lines with different growth rates were used to evaluate the effect of cell growth phase on selenite retention, selenite distribution, selenite inhibition of DNA synthesis and presence of selenoproteins. Autoradiography of log and confluent phase MOD cells revealed a uniform retention of selenite in log phase cells and a marked lack of uniformity of selenite retention in confluent phase cells. A higher total percentage of selenite was retained and covalently incorporated into proteins by confluent phase cells. Levels of the 58K selenoprotein, but not the 26K and 23K selenoproteins, were higher in confluent versus log phase cells. The results suggest that the 58K selenoprotein accumulated in cell populations where DNA synthesis was inhibited in contrast to cells actively undergoing cell proliferation. In addition, the 58K selenoprotein was the only major selenoprotein present in both log and confluent phase cells during and before inhibition of DNA synthesis. The implications of these results are discussed in terms of potential combination chemoprevention protocols in animal tumor experiments.

Animals↗

PubMed1988

A clonal derivative of mammary epithelial cell line COMMA-D retains stem cell characteristics of unique morphological and functional heterogeneity.

The COMMA-D cell line derived from mammary epithelial cells of midpregnant mice was shown previously to be heterogeneous as determined by phase-contrast microscopy, immunocytochemical staining, DNA content, and oncogenic potential (K.D. Danielson et al. (1984) Proc. Natl. Acad. Sci. USA 81, 3756; D. Medina et al. (1986) J. Natl. Cancer Inst. 76, 1143). Clonal subpopulations of COMMA-D cells have now been isolated by both transfection and selection using a dominant-selectable gene transfer vector and by limiting dilution. Despite their clonal origin, these subpopulations in many cases retained the heterogeneity of the parental COMMA-D line. Of 18 clonal lines assayed, only 5 were able to express beta-casein mRNA. Pooled populations of G418-resistant cells expressed substantially higher levels of beta-casein mRNA than the clonal lines. One of the expressing clonal lines, BNW-7, was characterized further, using immunocytochemical techniques. Approximately 10% of BNW-7 cells expressed casein under the appropriate hormonal and cell-substratum conditions by indirect immunofluorescent staining. Casein immunoperoxidase staining of BNW-7 cells on floating collagen gels revealed that casein-producing cells were localized in small alveolar structures, which were formed in a non-hormone-dependent fashion. The cells in these alveolar structures were cuboidal with basally located nuclei, expressed keratin intermediate filament proteins preferentially, and comprised approximately 18% of the total cells. Cells elsewhere on the surface of the gel displayed a flattened morphology, and expressed vimentin intermediate filament proteins preferentially. A proportion of COMMA-D cells, therefore, appeared to have some of the characteristics of mammary stem cells, and retained the ability to differentiate and form phenotypically heterogeneous cell populations in vitro.

Animals↗

PubMed1988

Distinguishing features of cytotoxic and pharmacological effects of selenite in murine mammary epithelial cells in vitro.

Previous studies have demonstrated that selenite exerts a reversible and non-lethal inhibition of cell growth which could be correlated with a 58 K selenoprotein. Prolonged exposure to 5 microM selenite is cytotoxic. This cytotoxic effect is heralded by the cells floating off the dish. These floating cells contained over a log more selenite than did the attached cells. The floating cells also had a higher total amount per cell of covalently incorporated selenite in the form of volatile selenides. No specific selenoprotein could be correlated with the cytotoxic effect of selenite. However, a greater aggregation of selenoproteins was observed in dead vs. viable cells. This change probably was a late manifestation of cell death, whereas the increased amount of volatile selenides marked the early stages of acute selenosis and cell death. The results suggest that the cellular response to selenite exposure contains two phases: an early reversible inhibition of cell growth, and a late irreversible cytotoxic effect. The former is characterized by an increase in a 58 K selenoprotein, whereas the latter by an increase in volatile selenides. The results suggest that experiments evaluating specific effects of selenite exposure on biochemical function need to distinguish between the two phases of cell response.

Animals↗

PubMed1988

Intracellular 58-kd selenoprotein levels correlate with inhibition of DNA synthesis in mammary epithelial cells.

Five mouse mammary epithelial cell lines (MMEC) with different growth rates were used to examine the relationship between selenoprotein levels and selenite-mediated inhibition of DNA synthesis. The inhibition of DNA synthesis preceded and was significantly greater than inhibition of protein synthesis. Cycloheximide, a protein synthesis inhibitor, caused a coordinate inhibition of both DNA and protein synthesis over a 50-fold dose range. Of the selenoproteins detected by one-dimensional SDS-PAGE, the 58-, 26- and 23-kd proteins were the only major selenoproteins observed in common among the five MMEC lines before and during inhibition of DNA synthesis. Other selenoproteins were present in some cell lines or after inhibition of DNA synthesis. The level of the 58-kd selenoprotein was most closely correlated with the degree of inhibition of DNA synthesis (r2 = 0.85), whereas the 26- and 23-kd proteins most closely correlated with selenite retention (r2 = 0.78). Upon selenite withdrawal from the growth medium, the decrease in 58-kd, but not in the 26- and 23-kd proteins correlated with resumption of DNA synthesis. Similarly, dose-response studies indicated that the 58-kd protein increased greater than 20-fold, whereas the 26- and 23-kd proteins increased only 5-fold. At high doses of selenite, other selenoproteins of mol. wts 30-101 kd were present but these proteins seemed to appear after inhibition of DNA synthesis. The possibility exists that these proteins may be product/precursors of the major selenoproteins. Since these experiments attempted to quantitate selenoproteins by densitometry of one-dimensional SDS-PAGE autoradiographs, the quality controls used for the experiments are discussed. The results are discussed in terms of the hypothesis that the 58-kd selenoproteins may mediate the effects of selenite on DNA synthesis.

Animals↗