Cerebellar dysgenesis in rats by diaplacental effects of 7,12-dimethylbenz[a]anthracene

7,12-Dimethylbenz[a]anthracene (DMBA) administered iv to pregnant Sparague-Dawley rats produced cerebellar malfunction in at least 10% of the offspring. The underlying morphologic basis of the cerebellar symptomatology was found in maldevelopment of cerebellar cortex, ranging from focal loss of granule and Purkinje's cell layers to extensive areas of cortical disorganization with losses of granule neurons. An interference with the proliferation of the granule cell-layer primordium was suggested as a mechanism of this cerebellar dysgenesis. After a 200-day observation, no tumors were found in the offspring of the DMBA-treated, pregnant rats.     

Metabolism of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine by human cytochrome P4501A1, P4501A2 and P4501B1

While the metabolic activation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) by N-hydroxylation has been well documented, the relative roles of the human cytochrome P450 (CYP) enzymes that catalyze this reaction have not been established. Previous studies indicated that the mutagenic activation product, 2-hydroxyamino-PhIP (N2-OH-PhIP), is produced primarily by CYP1A2, and to a lesser extent by CYP1A1. We recently reported that human CYP1B1 also produces N2-OH-PhIP (Carcinogenesis, 18, 1793-1798, 1997). In the present study, we examined PhIP metabolism by microsomes containing recombinant human CYP1A1, 1A2 or 1B1 expressed in Sf9 insect cells and compared the kinetic values for PhIP metabolite formation. PhIP metabolites were analyzed by high pressure liquid chromatography with fluorescence and absorbance detection. Vmax values for N2-OH-PhIP formation were 90, 16 and 0.2 nmol/min/nmol P450, and the apparent Km values were 79, 5.1 and 4.5 microM for human CYP1A2, 1A1 and 1B1, respectively. The non-mutagenic metabolite, 4'-hydroxy-PhIP, was also formed by all three CYP enzymes with Vmax values of 1.5, 7.8 and 0.3 nmol/ min/nmol P450 and apparent Km values of 43, 8.2 and 2.2 microM for human CYP1A2, 1A1 and 1B1, respectively. Although the Vmax for N2-OH-PhIP production was highest for CYP1A2, the catalytic efficiency (Vmax/Km) of CYP1A1 was greater than that of CYP1A2. These results suggest that, for humans, extrahepatic CYP1A1 may be more important than previously thought for the metabolic activation of the dietary carcinogen PhIP.     

Black tea and mammary gland carcinogenesis by 7,12-dimethylbenz[a]anthracene in rats fed control or high fat diets

Epidemiological studies suggest that tea may reduce cancer risk, and in laboratory rodents, chemopreventive effects of tea or purified extracts of tea have been demonstrated in lung, gastrointestinal tract and skin. There is some evidence of chemoprevention by tea in the mammary gland, but the data are not conclusive. In order to evaluate more fully the possible influence of black tea on 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary gland tumors in the female S-D (Sprague-Dawley) rat, three large studies were performed: experiment 1, tumorigenesis in rats fed AIN-76A diet and given 25 mg/kg DMBA and 1.25 or 2.5% whole tea extract or water to drink; experiment 2, tumorigenesis in rats given 15 mg/kg DMBA and the same diet and fluids as in experiment 1; experiment 3, tumorigenesis in rats fed control or HF (high fat, corn oil) diet and given 15 mg/kg DMBA and 2% tea or water to drink. Tea was given throughout the experiment; DMBA was given by gastric gavage at 8 weeks of age. There was no consistent effect of tea on tumorigenesis in rats fed AIN-76A diet; there was, however, evidence in experiment 3 of a reduction of tumorigenesis by tea in rats fed the HF diet. In experiment 3, rats fed the HF diet and given water showed the expected increase in tumor burden (number and weight) compared with rats fed control diet. However, rats fed the HF diet and given 2% tea showed no increase in tumor burden; their tumor burden was significantly lower than in rats fed the HF diet and given water (P < 0.01) and was not different from rats fed control diet and given water or tea. In addition, in experiment 3, the number of malignant tumors per tumor-bearing rat was increased by the HF diet in water-drinking rats (P < 0.01) but not in tea-drinking rats. Therefore, it appears that tea partially blocked the promotion of DMBA-induced mammary tumorigenesis by the HF diet.     

Inhibitory effects of naturally occurring coumarins on the metabolic activation of benzo[a]pyrene and 7,12-dimethylbenz[a]anthracene in cultured mouse keratinocytes

Several naturally occurring coumarins to which humans are routinely exposed have been previously found to be potent inhibitors and inactivators of cytochrome P450 (P450) 1A1-mediated monooxygenase in both murine hepatic microsomes and in a reconstituted system using purified human P450 1A1 [Cai et al. (1993) Chem. Res. Toxicol., 6, 872-879 and Cai et al. (1996) Chem. Res. Toxicol., 9, 729-736]. In the present study, several of these coumarins were investigated for their inhibitory effects on the metabolism and metabolic activation of benzo[a]pyrene (B[a]P) and 7,12-dimethylbenz[a]anthracene (DMBA) in cultured mouse keratinocytes. Initial analysis of B[a]P metabolism in cultured keratinocytes showed that imperatorin, isoimperatorin, coriandrin, and bergamottin, at concentrations of 2 nM equal with B[a]P, reduced the formation of water-soluble metabolites of B[a]P by 33% to 57%. Bergamottin and coriandrin were the most potent inhibitors of the compounds examined. HPLC analysis of organic solvent-soluble metabolites of B[a]P indicated that all the coumarins tested significantly reduced the formation of individual B[a]P metabolites (including phenols, diols and tetraols). However, the greatest effect was on the formation of B[a]P tetraols. Additional experiments determined the ability of selected coumarins to block covalent binding of B[a]P and DMBA to DNA in keratinocytes. Bergamottin preferentially inhibited the binding of B[a]P to DNA by 56%, while coriandrin preferentially inhibited the binding of DMBA to DNA by 48%. Notably, analysis of individual DNA adducts formed from B[a]P and DMBA indicated that both bergamottin and coriandrin specifically inhibited the formation of anti diol-epoxide DNA adducts derived from both hydrocarbons. The preferential inhibitory effect of bergamottin and coriandrin on the formation of anti diol-epoxide adducts derived from DMBA was further confirmed by separation of anti- and syn-diol-epoxide-DNA adducts using immobilized boronate chromatography. The current study demonstrates that certain naturally occurring coumarins inhibited metabolic activation of B[a]P and DMBA in cultured mouse keratinocytes and specifically inhibited the formation of DNA adducts derived from the anti diol-epoxide diastereomers from either hydrocarbon. The current data also suggest that certain naturally occurring coumarins may possess anticarcinogenic activity toward polycyclic aromatic hydrocarbons.     

Metabolism of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) by human cytochrome P4501B1

Cytochrome P4501B1 (CYP1B1) is the most recently identified member of the dioxin-inducible CYP1 family. CYP1B1 is constitutively expressed in most human tissues, including colon and breast, and can activate numerous chemically diverse carcinogens. We evaluated the metabolism of the dietary heterocyclic amine carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) by microsomes from yeast expressing the human CYP1B1 protein. PhIP metabolites were analysed by HPLC with fluorescence and absorbance detection. We found that human CYP1B1 metabolizes PhIP to three products: N2-OH-PhIP, a mutagenic activation product; 4'-OH-PhIP, a detoxification product; and 2-OH-PhIP, the mutagenic potential of which is unknown. Metabolite identity was confirmed by co-elution with authentic standards and synchronous fluorescence spectroscopy. The identity of the 2-OH-PhIP standard was additionally confirmed by mass spectrometry. Kinetic studies of the formation of N2-OH-PhIP, 4'-OH-PhIP and 2-OH-PhIP by CYP1B1 indicated apparent Km values of 5.7 +/- 1.3, 2.2 +/- 0.5 and 1.3 +/- 0.2 microM, respectively. Apparent turnover rates were 0.40 +/- 0.03, 0.93 +/- 0.02 and 0.04 +/- 0.00 nmol product/min nmol P450, respectively. At saturating levels of substrate, CYP1B1-mediated formation of the non-mutagenic metabolite 4'-OH-PhIP was favored two-fold over that of the mutagenic metabolite, N2-OH-PhIP and >10-fold over that of 2-OH-PhIP. The formation of N2-OH-PhIP, a potent mutagen implicated in the etiology of human colon and breast cancer, indicates that CYP1B1 may play an important role in PhIP-mediated carcinogenesis.     

Tissue distribution of DNA adducts in rats treated by intramammillary injection with dibenzo[a,l]pyrene, 7,12-dimethylbenz[a]anthracene and benzo[a]pyrene

Recombinant human bone morphogenetic protein (rhBMP-2) was examined for its in vitro effects on biochemical markers representing osteoblast phenotype. Primary cultures of fetal rat calvarial osteoblasts were used in this study. The results indicated that rhBMP-2 stimulated alkaline phosphatase activity, parathyroid hormone (PTH)-induced cyclic AMP production, and collagen biosynthesis in a dose-dependent manner in confluent cultures. The percent collagen synthesis also increased in a dose-dependent manner. Alkaline phosphatase activity was stimulated in a time-dependent manner by rhBMP-2 that reached its maximum 5 days after initiation. Cycloheximide (2 micrograms/ml) inhibited rhBMP-2-stimulated alkaline phosphatase indicating de novo protein synthesis of the enzyme. Transforming growth factor-beta 1 (TGF-beta 1)-induced inhibition of alkaline phosphatase activity observed in confluent primary cultures was completely abolished by rhBMP-2 at a concentration that was 43 times greater than the TGF-beta 1 concentration. Also, rhBMP-2 produced a small stimulation of alkaline phosphatase activity in cells grown in the absence of ascorbic acid; however, the effect was greatly enhanced in cells cultivated in the presence of ascorbic acid (50 micrograms/ml). In view of the potentiating effect of ascorbic acid on rhBMP-2-induced stimulation of alkaline phosphatase, we speculate that ascorbic acid could amplify the osteoinductive effects of rhBMP-2 and thereby augment the efficacy of the BMP when used as bone repair material in vivo. rhBMP-2 (4.3-86 ng/ml) did not exhibit mitogenic effects on cultured osteoblasts. These data suggest that rhBMP-2 has the ability to induce expression of various markers associated with the osteoblast phenotype in primary cultures of fetal rat calvarial osteoblasts. In addition, we speculate that TGF-beta 1 may play a regulatory role in BMP-induced bone formation and ascorbic acid may potentiate the effects of rhBMP-2 in vivo.     

Induction of mammary tumors in virgin female BALB/c mice by single low doses of 7,12-dimethylbenz[a]anthracene

The induction of mammary tumors in virgin female inbred BALB/c mice after administration of 7,12-dimethylbenz[a]anthracene (DMBA) over a wide range of doses was studied. Mice were exposed at 12 weeks of age to single or multiple doses of DMBA ranging from 0.0025 to 12.0 mg by gastric intubation and were checked regularly for mammary tumors. The experiment was terminated when the mice were 800 days of age. In the dose range of 0.0025--0.125 mg DMBA, the incidence of mammary tumors was dose-dependent. At higher doses, the mammary tumor incidence became less dose-dependent and was nearly independent of doses above the 0.25-mg level. Analysis of the data for the rate of appearance of mammary tumors with age of the animals and for the age at death of non-mammary tumor-bearing animals indicated that in the low dose range induction of mammary tumors was the predominant effect of DMBA exposure, whereas at moderate to high doses the toxic and carcinogenic effects of DMBA on other tissues significantly influenced the final incidence of mammary tumors. Greater than 90% of the tumors that resulted from administration of low doses of DMBA were adenocarcinomas. In contrast, adenocarcinomas and adenoacanthomas were found in approximately equal proportions following administration of high doses of DMBA.     

Effects of 7,12-dimethylbenz[a]anthracene on the superantigen toxic shock syndrome toxin (TSST-1)-induced proliferation and antibody secretion by human lymphocytes

Toxic shock syndrome toxin (TSST-1), a 22-kDa exotoxin secreted by Staphylococcus aureus, can act as a nominal antigen and induce proliferation and immunoglobulin secretion in human B-cells. The purpose of the present studies was to examine the effect of 7,12-dimethylbenz[a]anthracene (DMBA), a well-characterized immunosuppressant of both cell-mediated and humoral immunity in murine lymphocytes, upon the mixed lymphocyte reaction (MLR) and TSST-1-induced immune responses in human lymphocytes. The MLR, using human tonsillar lymphocytes (HTL) from four different donors, was inhibited in a dose-dependent manner from 1 to 100 microM. The IC50 for the suppression of the MLR ranged from 10 to 40 microM. TSST-1 is a potent stimulator of T-cells bearing specific VB regions on the T-cell receptor (TCR). In contrast with the results from the MLR, DMBA inhibited TSST-1-induced T-cell proliferation only at 100 microM in HTL. A similar profile of activity was determined with splenic T-cells from a single donor. TSST-1 has also been demonstrated to induce specific B-cell proliferation and differentiation in the presence of irradiated T-cells. TSST-1-induced B-cell proliferation was only consistently and markedly inhibited by DMBA at 100 microM in tonsillar and splenic lymphocytes. In contrast, TSST-1-induced B-cell differentiation, as manifested by IgM and IgG secretion, was inhibited in a dose-dependent manner from 1 to 100 microM DMBA in B-cells from human tonsils and spleens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibitory effect of TMK688 on skin tumor initiation caused by 7,12-dimethylbenz[a]anthracene in relation to inhibition of aryl hydrocarbon hydroxylase activity and Cyp1a1 mRNA induction

The effects of single and repeated (9 times) administration of two dihydropyridines (DHPs), nimodipine (NIM) and nifedipine (NIF) (5 mg/kg per 12 h and 2.5 mg/kg per 12 h, IP), on the behavior of male adult rats in the holeboard and in the plus-maze, were investigated. Besides, the effects of repeated administration of the drugs on the levels of dopamine (DA), serotonin (5-HT), and their respective major metabolites in several regions of the central nervous system (CNS) were also assessed. The effects of single and repeated administration of the drugs were similar. Both DHPs caused a significant decrease in general motor activity which was evident in both tests and more marked, with the higher doses. The two exploratory parameters measured in the holeboard, i.e. head-dipping frequency and duration, were dissociated under pharmacological treatment. The drug-treated animals did not show an increased emotionality in the holeboard. However, in the plus-maze, NIF (5 mg/kg) and to a lesser extent NIM, appeared to induce some anxiety-related responses which may be secondary, at least in part, to the depressing effect on activity and exploration. Repeated administration of NIM and NIF caused an increase in striatal DA and DOPAC levels, whilst no effects were found on serotonergic system in any of the regions of the CNS analyzed.     

Demonstration and partial characterization of a 7,12-dimethylbenz[alpha]anthracene-binding protein in rat adrenal

Rat adrenal cytosol was found to contain a 7,12-dimethylbenz[a]-anthracene (DMBA)-binding protein which is characterized by a pI of 7.2, a Kd-value of 3 microM and a maximal capacity of about 47 pmol/mg protein. The binding is highly specific for DMBA and is not displaced by 3-methylcholanthrene (MC), benz[a]pyrene (BP) or other polycyclic hydrocarbons. Likewise, various androgens, estrogens or glucocorticoids have no effect on the DMBA binding. It is proposed that the DMBA-binding protein may have a role in the toxic effects of DMBA or DMBA metabolites in adrenal and possibly in other DMBA-sensitive organs as well.     

Antitumour activity of coumarin and 7-hydroxycoumarin against 7,12-dimethylbenz[a]anthracene-induced rat mammary carcinomas

Female SD rats with established 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumours were treated with coumarin (20 mg/kg body weight; six times per week) or its metabolite 7-hydroxycoumarin (20 mg/kg) for 4 weeks. The anti-oestrogen tamoxifen (8.8 mg/kg) served as the reference drug. The inhibitory effect of coumarin was similar to that of tamoxifen [mean change of tumour area: 428% (coumarin) compared to 528% (tamoxifen); control 822%]. The strongest inhibition was observed with 7-hydroxycoumarin (248%); the difference compared to the control was significant (P < 0.01). Neither coumarin nor 7-hydroxycoumarin reduced the number of tumours appearing during treatment as tamoxifen did. However, the size of the tumours treated with coumarin or its metabolite was generally much smaller than those in the tamoxifen group or in the control group. From the data obtained it appears that coumarin and 7-hydroxycoumarin inhibit the growth of tumours that have reached a certain size but do not prevent the formation of tumours after exposure to the carcinogen.     

Oxidative stress and cytotoxicity induced by ferric-nitrilotriacetate in HepG2 cells that express cytochrome P450 2E1

Iron can potentiate the toxicity of ethanol. Ethanol increases the content of cytochrome P450 2E1 (CYP2E1), which generates reactive oxygen species, and transition metals such as iron are powerful catalysts of hydroxyl radical formation and lipid peroxidation. Experiments were carried out to attempt to link CYP2E1, iron, and oxidative stress as a potential mechanism by which iron increases ethanol toxicity. The addition of ferric-nitrilotriacetate (Fe-NTA) to a HepG2 cell line expressing CYP2E1 decreased cell viability, whereas little effect was observed in control cells not expressing CYP2E1. Toxicity in the CYP2E1-expressing cells was markedly enhanced after the depletion of glutathione. Lipid peroxidation was increased by Fe-NTA, especially in cell extracts and medium from the CYP2E1-expressing cells. Toxicity was completely prevented by vitamin E or by 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid, which also decreased the lipid peroxidation. Levels of ATP were lowered by Fe-NTA, and this was associated with a decreased rate of oxygen consumption by permeabilized cells with substrates donating electrons to complexes I, II, and IV of the respiratory chain. This mitochondrial damage was prevented by vitamin E. Toxicity was accompanied by DNA fragmentation, and this fragmentation was prevented by antioxidants. Overexpression of bcl-2 decreased the toxicity and DNA fragmentation produced by the combination of CYP2E1 plus Fe-NTA, as did a peptide inhibitor of caspase 3. These results suggest that elevated generation of reactive oxygen species in HepG2 cells expressing CYP2E1 leads to lipid peroxidation in the presence of iron, and the ensuing prooxidative state damages mitochondria, releasing factors that activate caspase 3, leading to a loss in cell viability and DNA fragmentation.     

Effects of estradiol and progesterone on cytochrome P4501A1 expression in Hepa 1c1c7 cells

The effects of estradiol and progesterone on the expression of cytochrome P4501A1 were investigated in Hepa 1c1c7 cells. Both steroids, at 10 microM concentration, increased P4501A1-mediated 7-ethoxyresorufin O-deethyalase activity and amounts of its immunoreactive protein and CYP1A1 mRNA. Gel shift assay revealed that the steroids could induce both AhR transformation and binding of the ligand-AhR complex to its specific DNA recognition site. Transient transfection demonstrated that 5'flanking region of CYP1A1 could respond to the steroid action. The competitive binding assay showed that the steroids bound to AhR with moderate affinity. These results suggested that steroidal structure can be AhR ligands and induce CYP1A1 expression in AhR-dependent manner.     

Effect of dietary soy protein isolate, genistein, and 1,4-phenylenebis(methylene)selenocyanate on DNA binding of 7,12-dimethylbenz[a]anthracene in mammary glands of CD rats

We examined whether a soy protein isolate or one of its major components (genistein) influences the initiation stage of carcinogenesis via DNA binding studies of 7,12-dimethylbenz[a]anthracene (DMBA) in liver and mammary tissue of female CD rats. A semipurified high-fat diet (23.5% corn oil) containing the soy protein isolate (10%), genistein (111 ppm), or 1,4-phenylenebis(methylene)selenocyanate (p-XSC) (5 ppm as selenium) as a positive control was fed to 6-week-old virgin female CD rats for 1 week before carcinogen treatment. Neither soy nor genistein affected the extent of DMBA-DNA binding in liver. In mammary tissue, 111 ppm genistein in the diet was more effective than the soy protein isolate, although the latter contains the same amount of genistein, mainly present as a glucoside conjugate. As shown before, p-XSC inhibited DMBA-DNA binding in mammary tissue. Total binding was inhibited because of reduced formation of three major adducts: anti-diol epoxide deoxyguanosine, syn-diol epoxide deoxyadenosine, and anti-diolepoxide deoxyadenosine. Thus, an additional experiment with 111 and 222 ppm of genistein was performed; 222 ppm genistein had a weaker effect than that observed for 111 ppm. Nevertheless, 111 ppm of genistein in the diet appears to inhibit the initiation phase of DMBA-induced rat mammary tumors and may partially account for the reported inhibitory effect of soy against DMBA-induced rat mammary tumors.     

Development of Caco-2 cells expressing high levels of cDNA-derived cytochrome P4503A4

PURPOSE: To develop Caco-2 cell derivatives expressing high levels of human cytochrome P450 drug metabolizing enzymes. METHODS: The cDNAs for two cytochrome P450 forms, CYP2A6 and CYP3A4, were introduced into an extrachromosomal vector under control of the cytomegalovirus early intermediate promoter. Vector-bearing cells were selected via resistance to hygromycin B. RESULTS: Transfected cells exhibited high levels of cDNA-derived protein as measured by Western blot, spectrophotometric P450 determination and/or cytochrome P450 form-selective enzyme assay. CYP3A4 and CYP2A6 catalytic activities were about 100 fold higher than in control cells. cDNA-expressing cells were found to form tight monolayers and were suitable for study of xenobiotic transport and metabolism. The permeabilities of cephalexin, phenylalanine, mannitol and propranolol across transfected monolayers were found to be similar to those across untransfected monolayers. The appropriate transfected monolayers metabolized the CYP2A6 substrate coumarin and the CYP3A4 substrates testosterone and nifedipine. CONCLUSIONS: A Caco-2 cell system to simultaneously study drug transport and metabolism has been developed.     

Spindle disturbances induced by benzo[a]pyrene and 7, 12-dimethylbenz[a]anthracene in a Chinese hamster cell line (V79-MZ) and the stability of the numerical chromosome aberrations that follow

As was shown (1), analysis of human hair on the population level and mapping of large territories using hair elemental composition are promising approaches for estimation of both the environmental situation and the population health status. In (1,2) the map of Uzbekistan (sampling in 1981) was discussed. Ten years later (1991), samples from the territory in the vicinity of the drying out Aral Sea were taken again. Samples were analyzed for 24 elements using instrumental neutron activation analysis. Comparison of the data and maps drawn for 1981 and 1991 and their comparison with changes of the health status have shown that repeated mapping of territories using data on human hair elemental composition could be used in medical geography, especially for prediction of health status changes in ecologically unfavorable areas.     

Oxidation of benzo[a]pyrene by recombinant human cytochrome P450 enzymes

The oxidation of benzo[a]pyrene (B[a]P) was examined using reconstituted systems prepared with recombinant human cytochrome P450 (P450) enzymes 1A1, 1A2, 2C8, 2C10, 2E1, and 3A4 and with microsomes prepared from Saccharomyces cerevisiae expressing recombinant human P450s 2C8, 2C9, and 2C18. Products measured by HPLC included the 3- and 9-phenols, the 4,5-, 7,8-, and 9,10-dihydrodiols (detected in the presence of epoxide hydrolase), and products in the polar fraction eluting immediately after the void volume. The most active enzyme in all reactions was P450 1A1. P450 3A4 and P450 1A2 formed appreciable amounts of several of the products, including the 3-phenol. P450 2C enzymes and P450 2E1 formed relatively low amounts of all B[a]P products. Consideration of these patterns along with knowledge of levels of expression of the P450s in human tissues and previous results with microsomes leads to the conclusion that P450 1A1 should dominate the oxidation of B[a]P in tissues where it is present and inducible. In human liver the level of P450 1A1 is low and P450 3A4, P450 2C subfamily enzymes, and P450 1A2 probably all contribute. Of the human P450s considered here, P450 1A2 was the most active hepatic enzyme forming the 7,8-dihydrodiol. 7,8-Benzoflavone stimulated the oxidation of B[a]P by P450 3A4 and inhibited the oxidations catalyzed by P450 1A2. The extent of inhibition of P450 1A1 was less (than with P450 1A2), probably due to the rapid oxidation of 7,8-benzoflavone by P450 1A1. The major 7,8-benzoflavone product appears to be the 5,6-oxide.     

Histologic characterization of rat ovarian carcinoma induced by intraovarian insertion of a 7,12-dimethylbenz[a]anthracene-coated suture: common epithelial tumors of the ovary in rats?

The frequency of the butyrylcholinesterase K mutation was calculated on the basis of data obtained by polymerase chain reaction primer-introduced restriction analysis (PCR-PIRA). The population sample was composed of 177 Brazilians: 95 whites of predominantly European ancestry and 82 admixed individuals (European and African origin). The frequencies--18.4 +/- 2.8% for whites and 17.1 +/- 2.9% for admixed--did not differ from those previously obtained in North America, Scotland, Japan, and Denmark. The occurrence of the K mutation in Europeans, East Asians, and Africans suggests a relatively old origin for this mutation, and the similar frequencies found in these populations may suggest the operation of selective forces.     

Mutual education between hematopoietic cells and bone marrow stromal cells through direct cell-to-cell contact: factors that determine the growth of bone marrow stroma-dependent leukemic (HB-1) cells

The spermatogenic cell-specific isoform of glyceraldehyde 3-phosphate dehydrogenase (GAPD-S) may regulate glycolysis and energy production required for sperm motility. Although the steady-state level of Gapd-s mRNA is maximal at step 9 of mouse spermatogenesis, GAPD-S protein was not detected by immunohistochemistry until steps 12-13. This result suggests that Gapd-s is translationally regulated. Western blot analysis of isolated germ cells confirmed that GAPD-S is not detected in pachytene spermatocytes or round spermatids. A major immunoreactive protein migrating with a molecular weight (M(r)) of 69,200 was observed in condensing spermatids and cauda sperm. Additional minor proteins that migrated at M(r) 55,200, 32,500, and 27,500 were detected in sperm. The molecular weight of GAPD-S is higher than the predicted molecular weight of 47,445, apparently due to a proline-rich 105-amino acid domain at the N-terminus. Recombinant GAPD-S protein lacking the proline-rich region migrated at M(r) 38,250, comparably to somatic GAPD, which also lacks the proline-rich domain. Indirect immunofluorescence demonstrated that GAPD-S is restricted to the principal piece in the sperm flagellum. Western blot analysis indicated that GAPD-S is tightly associated with the fibrous sheath of the flagellum, consistent with a potential role in regulating sperm motility.