Direct cytochemical localization of the free catalytic subunit of cAMP-dependent protein kinase.
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Aminophylline↗
Biomedical subjects
C V Byus
Publications and source records attributed to C V Byus.
Receptor-mediated action without receptor occupancy: a function for cell-cell communication in ovarian follicles.
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Animals↗
The effects of low-energy 60-Hz environmental electromagnetic fields upon the growth-related enzyme ornithine decarboxylase.
People living in the industrial society of today are unavoidably exposed to low-energy electromagnetic (EM) radiation. The potential risk to human health of such exposure has received much study. In this regard, numerous epidemiological studies have linked exposure to low-energy EM fields to increased cancer risk. We investigated the ability of low-energy 60-Hz EM fields to alter the activity of ornithine decarboxylase (ODC) in a number of established cell lines. The activity of ODC, the controlling enzyme in polyamine biosynthesis, has been shown to be elevated in growing cells or tissues and during the process of tumor promotion. A 1-h exposure to a 60-Hz EM field of an intensity of 10 mV/cm produced a 5-fold increase in ODC activity in human lymphoma CEM cells and a 2- to 3-fold increase in mouse myeloma cells (P3) relative to the unexposed cultures. Depending upon the cell type, ODC activity increased during the 1-h exposure period and remained elevated for several hours after the field exposure ended. In another series of experiments, fields of an intensity as low as 0.1 mV/cm for a 1-h period produced a 30% increase in the activity of ODC in Reuber H35 hepatoma cells grown in monolayer culture. In the H35 cells, continuous exposure to the 60-Hz EM field (10 mV/cm) for periods of 2 and 3 h resulted in either no increase in ODC activity (2 h) or a decrease in enzyme activity (3 h) compared to the unexposed control cultures. The data is discussed in relation to possible molecular mechanisms of field-cell interaction, the importance of the exposure intervals altering cellular ODC activity and the potential ability of 60-Hz EM fields to serve as a tumor promoting stimulus.
Animals↗
A rat monoclonal antibody which interacts with mammalian ornithine decarboxylase at an epitope involved in phosphorylation.
Ornithine decarboxylase was purified from androgen-treated mouse kidney to homogeneity and high specific activity. The purified enzyme was utilized for production and screening of rat monoclonal and polyclonal antibodies. A rat monoclonal antibody was isolated which was capable of immunoprecipitation of native mouse kidney ornithine decarboxylase activity or the [3H]difluoromethylornithine-inactivated enzyme. Phosphorylation of mouse ornithine decarboxylase by casein kinase-II prior to immunoprecipitation led to complete loss of the epitope recognized by the monoclonal antibody but did not alter recognition by polyclonal antibody. Mammalian ornithine decarboxylase activity obtained from several species, in crude or partially purified extracts, was subjected to quantitative immunoprecipitation with monoclonal and polyclonal antibody. Polyclonal antibody immunoprecipitated all of the ornithine decarboxylase activity from every extract tested, while monoclonal antibody was capable of only limited immunoprecipitation (60-80%). Due to the inability of the monoclonal antibody to recognize ornithine decarboxylase phosphorylated in vitro by casein kinase-II and the partial immunoprecipitation of ornithine decarboxylase activity from cell extracts, a portion of the ornithine decarboxylase molecule population must exist in a phosphorylated state. This immunological evidence further confirms existing data that the enzyme exists in at least two distinct forms.
Androgens↗
Phosphorylation of histones is stimulated by phorbol esters in quiescent Reuber H35 hepatoma cells.
Histones isolated from Reuber H35 rat hepatoma cells treated with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) were examined for possible alterations in phosphorylation. Incorporation of 32P orthophosphate into individual acid-extracted histones was monitored by autoradiography and scintillation counting of polyacrylamide gels or by reverse-phase high performance liquid chromatography. Treatment of quiescent H35 cells (arrested by serum starvation) with submicromolar doses of TPA resulted in a rapid and specific increase in phosphorylation of histones H2B and H1(0). Smaller increases in phosphorylation were observed for H4. No significant change in phosphorylation of the major H1 histones or H2A were observed after 1 h of treatment. The phosphorylation was TPA dose-dependent, with a maximum increase of approximately 14-fold for H2B, 11-fold for H1(0), and 2-fold for H4 achieved at 0.8 M TPA. The nonpromoting parent compound phorbol did not induce any of these changes. Furthermore, the mitogenic hormone insulin did not cause a similar pattern of histone phosphorylation, suggesting that the effect observed was not due to a general mitogenic response in the H35 hepatoma cells. Addition of 8-Br-cAMP also failed to reproduce the effect of TPA on histone phosphorylation, suggesting that cAMP-dependent protein kinases are not likely to be involved in mediating this response to TPA.
8-Bromo Cyclic Adenosine Monophosphate↗
Insulin- and phorbol ester-stimulated phosphorylation of ribosomal protein S6.
The relative abilities of insulin and the phorbol ester tumor promoter 12-O-tetradecanoyl phorbol 13-acetate (TPA) to lead to the phosphorylation of ribosomal protein S6 in vivo were compared in a Reuber H35 hepatoma cell line shown previously to be highly responsive to these agents. In quiescent (serum-starved) cultures of H35 cells incubated with 32Pi, both insulin (10(-7) M) and TPA (1.6 X 10(-6) M) resulted in the marked phosphorylation of S6 compared to the unstimulated cultures as evidenced by an increase in radioactivity associated with S6 and by a corresponding shift in the mobility of phosphorylated S6 during two-dimensional electrophoresis. Following incubation with insulin or TPA, greater than 95% of the phosphate was in derivatives containing four to five phosphate groups. The site-specific phosphorylation of S6 in response to both optimal and suboptimal concentrations of insulin and/or TPA was examined by two-dimensional peptide mapping of the trypsin-digested ribosomal protein S6. The tryptic phosphopeptides of S6 obtained following treatment of the H35 cells with insulin and/or TPA were identical and were the same phosphopeptides as those observed previously following the phosphorylation in vitro of 40 S ribosomal subunits from reticulocytes with purified protease-activated kinase II (Perisic, O., and Traugh, J. A. (1983) J. Biol. Chem. 258, 13998-14002).
Animals↗
Intracellular kinetics of free catalytic units dissociated from adenosine 3',5'-monophosphate-dependent protein kinase in adrenocortical tumor cells (Y-1).
To define the role of cAMP in the actions of ACTH, the dissociation of cAMP-dependent protein kinase and the subsequent intracellular location of its free catalytic units were monitored after exposure of Y-1 cells to ACTH, FSH, or cyclic nucleotide analog. To accomplish this, a fluorescinated cytochemical probe was used that complexes specifically with free catalytic units from cAMP-dependent protein kinase. Also, the effects of hormone or nucleotide on secretion of fluorogenic steroids and DNA synthesis were examined. Y-1 cells dissociated protein kinase in a dose-dependent fashion when exposed to ACTH or cAMP analog, but did not respond to FSH, which was one of the control agents used. After 30 min of treatment with 1.5 X 10(-10) M ACTH, free catalytic units were observed only in the cytoplasm of Y-1 cells, whereas a similar time of exposure to 3 X 10(-10) M ACTH led to the appearance of catalytic units in nucleolus as well as in cytoplasm. ACTH (6 X 10(-10) M) caused a rise in cytoplasmic and nucleolar protein kinase dissociation proportionally greater than that seen in cultures exposed to 3 X 10(-10) M ACTH. Upon treatment with 6 X 10(-10) M ACTH, the amount of free catalytic units in cytoplasm and nucleolus was detectably greater than that in controls within 1 min of stimulation and continued to rise with increasing time of exposure to hormone. The nuclear, mostly nucleolar, content of free catalytic unit appeared to peak after 15 min of stimulation, while cytoplasmic enzyme levels continued to rise up to 60 min. Exposure of Y-1 cells to nucleotide analog caused cAMP-dependent protein kinase dissociation with temporal kinetics and a subcellular distribution similar to that seen after ACTH stimulation. We conclude that actions of ACTH are mediated by cAMP-dependent protein kinases. Further, there appear to be two intracellular pools of protein kinase, one nucleolar, the other cytoplasmic, and these may be independently regulated, with the nucleolar enzyme requiring higher concentrations of ACTH for dissociation than those needed for cytoplasm protein kinase. These observations may be relevant to the fact that more ACTH is required to inhibit DNA synthesis than is necessary to enhance steroid production.
8-Bromo Cyclic Adenosine Monophosphate↗
Intracellular localization of the catalytic subunit from cyclic AMP-dependent protein kinase in mutant Chinese hamster ovary cells deficient in this enzyme.
A direct cytochemical procedure that specifically locates free catalytic subunits (C) from cAMP-dependent protein kinase has been used to follow the kinetics of kinase dissociation in parental Chinese hamster ovary cells (CHO 10001) and 4 mutant CHO cell lines variously deficient in this enzyme (CHO 10215, 10248, 10260, 10265) (1-5). When cultures of wild-type cells were stimulated with 8BrcAMP a time- and dose-dependent dissociation of kinase was observed. The catalytic unit appeared first in the cytoplasm and nucleolus and with time in the nucleoplasm as well. At peak protein kinase activation (30 min) more than 80% of the cells possessed abundant C in all of these subcellular compartments. The data indicate that both the type I and type II isozymes of the cAMP-dependent protein kinase are localized in similar areas of the cell. Stimulation of the mutant cell lines with 8-BrcAMP revealed that they each contained activatable kinase, determined cytochemically, that paralleled in amount the total assayable protein kinase determined biochemically (1-6). There were differences in the basal (unstimulated) levels of free C in the mutants relative to each other and to wild-type cells. For example, wild-type and mutants 10248, and 10260 had barely detectable cytoplasmic catalytic-subunit in unstimulated cultures whereas mutant 10215 possessed a significant amount of free C. Upon stimulation with 8-BrcAMP, the subcellular distribution of C was in all cases similar; although there were significant quantitative disparities between the wild type and various mutant cell lines. All cell lines examined had roughly equivalent amounts of nucleolar C but differed predominantly in the quantity of cytoplasmic kinase. Specifically, two of the mutant lines (10260 and 10248) contained barely detectable amounts of nucleoplasmic C whereas the other mutants had moderate (10265) to near normal (10215) amounts of nucleoplasmic enzyme. Mutants having only type I isozyme (10248) and those with only type II isozyme (10265) gave similar patterns of free C localization after 8BrcAMP stimulation.
Animals↗
A role for ornithine in the regulation of putrescine accumulation and ornithine decarboxylase activity in Reuber H35 hepatoma cells.
We investigated the ability of intracellular ornithine to alter both the biosynthesis of putrescine and the activity of ornithine decarboxylase in Reuber H35 hepatoma cells in culture incubated with 12-O- tetrade - canoylphorbol 13-acetate (TPA). In confluent cultures of H35 cells, the addition of TPA (1.6 microM) caused the activity of ornithine decarboxylase to increase by more than 100-fold within 4 h. When exogenous ornithine (0.1-1.0 mM) was added to the culture medium with TPA, a marked dose-dependent increase in the production of putrescine was observed. The activity of ornithine decarboxylase in the same cultures incubated with ornithine decreased in a similar dose-dependent manner. The addition of arginine (0.1-1.0 mM) (but not lysine or histidine) to the H35 cells in culture concomitant with TPA also led to a relative increase in putrescine biosynthesis and a decrease in ornithine decarboxylase activity compared to cultures not receiving the amino acids. A similar response to exogenous ornithine and TPA was observed in a series of less confluent rapidly growing cultures which were in culture for a shorter period of time. The confluent cultures possessed a basal level of arginase (55 units/mg protein) which increased approx. 2-fold upon treatment with TPA. The intracellular concentration of ornithine in the unstimulated cells was in the order of 0.02-0.03 mM. Upon incubation of the cells with exogenous ornithine or arginine, the intracellular pools of these amino acids increased 4- to 8-fold.
Animals↗
Tumor-promoting phorbol esters stimulate the phosphorylation of ribosomal protein S6 in quiescent Reuber H35 hepatoma cells.
The addition of the tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA) to serum-starved quiescent Reuber H35 hepatoma cells results in a rapid 5- to 11-fold increase in the incorporation of 32Pi into a Mr = 32,000 ribosomal protein. The Mr = 32,000 protein was the major phosphorylated protein extracted from isolated 80 S ribosomes and was identified as the 40 S ribosomal protein S6 based upon its migration in two-dimensional gels. Insulin, which has been demonstrated to increase the phosphorylation of S6 in a number of cell lines, caused a 10- to 20-fold increase in the incorporation of 32Pi into this Mr = 32,000 ribosomal protein. S6 phosphorylation was dose- and time-dependent being detected as early as 5 min following the addition of 1.6 microM TPA. Maximal phosphorylation of ribosomal protein S6 was achieved by 60 min and remained elevated for at least 90 min in the presence of TPA. The 50% effective dose for TPA was estimated to be 0.14 microM. Based upon the altered migration of S6 in pH 8.5 urea-polyacrylamide gels, it was demonstrated that the increased 32Pi labeling of S6 by TPA was due to a net increase in the incorporation of phosphates into the S6 molecule. Non-tumor-promoting phorbol esters were ineffective in increasing the phosphorylation of S6. In whole cells, exogenously added 1 mM 8-bromoadenosine 3':5'-monophosphate failed to substantially increase phosphorylation of S6 suggesting that the TPA-induced phosphorylation of S6 occurs via a cyclic AMP-independent mechanism. The S6 amino acid residue phosphorylated in response to TPA was phosphoserine. A possible role for protein kinase C in the phosphorylation of ribosomal protein S6 is discussed.
Animals↗
Alterations in protein kinase activity following exposure of cultured human lymphocytes to modulated microwave fields.
Cultures of human tonsil lymphocytes were exposed in a Crawford cell to a 450-MHz field (peak envelope intensity 1.0 mW/cm2), sinusoidally amplitude modulated (depth 80%) at frequencies between 3 and 100 Hz for periods up to 60 min. The Crawford cell was housed in a temperature-controlled chamber (35 degrees C) and control cultures were placed in the same chamber. Activity of cAMP-dependent protein kinase relative to controls remained unaltered by fields modulated at 16 or 60 Hz with exposures of 15, 30, and 60 min. By contrast, total non-cAMP-dependent kinase activity fell to less than 50% of unexposed control levels after 15 and 30 min exposures, but, despite continuing field exposure, returned to control or preexposure levels by 45 and 60 min. A smaller reduction (20-25%) also occurred with 60-Hz modulation and was also restricted to exposure durations of 15 and 30 min. CW 450-MHz fields were without effect. Reduced enzyme activity occurred with 16-, 40-, and 60-Hz modulation frequencies, but not with 3-, 6-, 80-, or 100-Hz modulation. The specific identity of this kinase is unknown. This rapid but transient reduction in lymphocyte protein kinase activity restricted to modulation frequencies between 16 and 60 Hz and to less than 30 min exposure is consistent with "windowing" with respect to modulation frequency and exposure duration.
Cells, Cultured↗
Induction of murine fibrosarcomas by low dose treatment with 3-methylcholanthrene followed by promotion with 12-O-tetradecanoyl-phorbol-13-acetate.
Repeated application of the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA) to the skin of mice previously treated with an initiating dose of the carcinogen 7,12-dimethylbenz[a] anthracene has been shown to lead to an increased incidence of papilloma. The studies presented here describe a modified murine two-stage carcinogenesis model in which a single subcutaneous administration of the carcinogen 3-methylcholanthrene (3-MC) is followed by multiple applications of TPA administered subcutaneously or intraperitoneally. TPA was observed to act as a promoter under these conditions when given either subcutaneously or intraperitoneally. When a carcinogenic dose of 3-MC was administered (0.5 mg/mouse) followed by regular treatment with TPA (10 micrograms/mouse) the percent of tumor-bearing mice increased and the length of time until tumors developed significantly shortened. At a subcarcinogenic dose of 3-MC (0.025 mg/mouse), repeated treatment with TPA led to tumor development whereas no tumors were observed in mice not treated with TPA. All tumors were found to be fibrosarcomas. Thus, TPA is capable of acting as a systemic promoter of mesenchymally derived tumors.
Animals↗
Cyclic AMP and tumor promoters cause differential induction of ornithine decarboxylase and accumulation of putrescine in Chinese hamster ovary cells deficient in cyclic AMP-dependent protein kinase.
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8-Bromo Cyclic Adenosine Monophosphate↗
Activation of cyclic adenosine 3':5'-monophosphate-dependent protein kinase in H35 hepatoma and Chinese hamster ovary cells by a phorbol ester tumor promoter.
The ability of the phorbol ester tumor promoter 12-O-tetradecanoylphorbol- 13-acetate (TPA) to lead to the activation of cyclic adenosine 3':5'-monophosphate (cAMP)-dependent protein kinase was investigated in three cell lines. In the Reuber H35 hepatoma cells, TPA (1.6 microM) led to an increase in the protein kinase activity ratio from a value of 0.13 in control cultures to 0.40 in the stimulated cells within 30 min of addition. A specific fluorescent cytochemical procedure was further utilized to study the intracellular localization of the free catalytic subunit following activation of the holoenzyme by TPA. A marked increase in cytoplasmic and nucleolar fluorescence indicative of the appearance of the free catalytic subunit (activation of cAMP-dependent protein kinase) in these cellular compartments was observed in both Reuber H35 hepatoma and Chinese hamster ovary 10001 cells incubated with TPA (1.6 microM) for 30 min. In the H35 cells monitored with the cytochemical probe for the catalytic subunit, protein kinase was activated within 5 min of the addition of TPA (1.6 microM) with 0.16 microM TPA in the culture medium resulting in the maximal degree of activation. No increase in intracellular fluorescence throughout a 40-min period was observed in a cAMP-dependent protein kinase-deficient mutant strain Chinese hamster ovary 10260 cells incubated with TPA. These studies indicate that, while TPA does lead to the marked activation of cAMP-dependent protein kinase in at least two cell types shown to be responsive to TPA, this activation may be only one of several rapid events which mediate the many specific effects of the phorbol esters.
Animals↗
Effects of insulin-glucagon interactions on glycogenolysis and protein kinase activity in rat hepatocytes.
The effects of insulin and glucagon on cAMP accumulation, protein kinase activation, and glycogenolysis were investigated in isolated rat hepatocytes. Glucagon (0.01 nM to 10 micro M) increased the activation state of protein kinase and the rate of glucose accumulation. Addition of 1.0 nM insulin to cells preincubated with 0.1 nM glucagon attenuated the rate of glucose accumulation, but did not alter the protein kinase activity ratio. Addition of 0.1 nM glucagon to cells preincubated with 1.0 nM insulin caused a rapid activation of protein kinase; however, glycogenolysis was not immediately affected. These effects were enhanced with pharmacological concentrations of glucagon and insulin. These data indicate that the degree of protein kinase activation does not always correlate temporally or quantitatively with rates of glycogenolysis in liver cells exposed to insulin and glucagon.
Animals↗
Direct cytochemical localization of catalytic subunits dissociated from cAMP-dependent protein kinase in Reuber H-35 hepatoma cells. I. Development and validation of fluorescinated inhibitor.
A specific and sensitive procedure has been developed that reliably localizes intracellular sites of free catalytic unit (C) dissociated from cAMP-dependent protein kinase. The method is based on a FITC conjugate (F:PKI) of affinity column-purified heat-stable protein inhibitor (PKI) of free C. The fidelity of this cytochemical probe was determined using cultures of Reuber H-35 hepatoma cells that had been stimulated for 2 h with 0.1 mM DBcAMP, or with diluent, then fixed with anhydrous acetone at -30 degrees C. In these preparations the F:PKI probe complexed with free C in cytoplasm, nucleolus, and, to a minor extent, in nucleoplasm. Binding of the F:PKI molecule to free C was competitively diminished by arginine analogues, guanidinium HCI and polyarginine, each used over a 2-log dose range. When the inhibitor's arginine residues were blocked by reaction with cyclohexanedione it no longer inhibited phosphotransferase activity of free C, and when fluorescinated it failed to localize C in stimulated cells. Similarly, when F:PKI was preabsorbed with excess pure C it no longer functioned as a cytochemical stain. Affinity column-purified antibody to free C also reduced significantly the ability of F:PKI to complex with C in cell cultures stimulated with 0.1 mM DBcAMP. 1 microgram of antibody reduced by approximately 10% the binding of F:PKI to all cell compartments while 5 microgram of antibody diminished binding by greater than 50%. Together, these results indicate that the F:PKI binds specifically, perhaps exclusively, to the catalytic units of cAMP-dependent protein kinase. The cytochemical procedure, unlike its biochemical counterparts, is able to locate the dissociation of cAMP-dependent protein kinase in individual cells of functionally or histologically complex cultures. Also, it reveals variations in the time- and dose-dependent activation of the kinase amongst clonal cells stimulated with cyclic nucleotide analogues or hormones.
Animals↗
Direct cytochemical localization of catalytic subunits dissociated from cAMP-dependent protein kinase in Reuber H-35 hepatoma cells. II. Temporal and spatial kinetics.
The activation of cyclic AMP-dependent protein kinase has been found to be the predominant mode by which cyclic AMP (cAMP) leads to alterations of a large variety of cellular functions. The activation of the kinase results in the release of the catalytic subunit which as the free enzyme possesses phosphotransferase activity for a variety of specific protein substrates. Using a sensitive and specific cytofluorometric technique we monitored the appearance of free catalytic subunit in Reuber H35 hepatoma cells in culture after incubation with N6-1'-O-dibutyryl-cyclic AMP (DBcAMP), 8-bromoadenosine-3':5'-cyclic monophosphate (8-BrcAMP), and glucagon. The cytochemical method employs the heat-stable inhibitor of the free catalytic subunit which has been conjugated to fluorescein isothiocyanate (F:PKI) and was validated as described in the companion paper (Fletcher and Byus. 1982. J. Cell Biol. 93:719-726). Here we studied the temporal and spatial kinetics of the free catalytic subunit following activation of cAMP-dependent protein kinase by increasing concentrations of DBcAMP,8-BrcAMP, and glucagon. Under similar conditions protein kinase activation was also assessed biochemically in H35 cell supernatants by assaying the protein kinase activity ratio. Incubation of the hepatoma cells with DBcAMP (0.1 mM) led to an increase in the activity ratio from 0.2 in control cultures to a value of nearly 1.0 within a 1- to 2-h period. During this same period using the F:PKI probe, a significant increase in cytoplasmic and nucleolar fluorescence indicative of the release of the free catalytic subunit was coincidentally observed. In contrast to the rapid appearance of catalytic subunit in the cytoplasm and nucleolus of the cell within 5-15 min of the addition of DBcAMP, discernible nucleoplasmic fluorescence did not occur until after 1 h. H35 cell cultures incubated with 8-BrcAMP (0.01-1.0 mM) exhibited a more rapid activation of the protein kinase measured cytochemically compared to the cells treated with DBcAMP. Cultures incubated with 8-BrcAMP had significantly increased cytoplasmic and nucleolar fluorescence compared to unstimulated cells within 1 min of the addition of the analogue and reached a maximal level within 15 min. By employing a microspectrophotometer a distinct dose-dependent increase in cellular fluorescence (i.e., free catalytic subunit) was observed as the concentration of 8-BrcAMP was increased from 0.01 to 1.0 mM at 1, 5, 15, and 60 min following stimulation. The addition of glucagon (10(-6) M) to the culture also led to the activation of cAMP-dependent protein kinase as determined by an increase in the activity ratio. This increase was paralleled throughout the incubation period by a marked elevation in cytoplasmic and nucleolar fluorescence. The results reported herein suggest that both cyclic nucleotide analogues and a polypeptide hormone lead to the activation of cAMP-dependent protein kinase in similar intracellular compartments in Reuber H35 hepatoma cells...
Adenosine Monophosphate↗
Tumor promoting phorbol-ester derivatives increase ornithine decarboxylase activity and polyamine biosynthesis in the liver of the rat and mouse.
The ability of the phorbol-ester tumor promoters to alter ornithine decarboxylase (ODC) activity in the liver of the rat and mouse was determined. The injection of 12-O-tetra-decanoyl phorbol-13-acetate (TPA, 100 microgram, i.p.) led to a 250-fold increase in hepatic ODC activity within 4 h of administration. This increase in ODC activity required both RNA and protein synthesis and did not occur when a variety of the non-tumor promoting phorbol-ester derivatives were administered to the rat. A distinct dose-dependent increase in hepatic ODC activity could be observed at 4 h following the injection of increasing amounts of TPA (0-100 microgram, i.p.). As little as 1.0 microgram TPA (i.p.) administered to a rat resulted in a significant stimulation in the activity of ODC in the liver compared to the control unstimulated values. Both 200 micrograms and 500 micrograms TPA produced less of an elevation in hepatic ODC activity than did the optimal dose of 100 micrograms. In the mouse, the administration of 1 microgram and 20 micrograms of TPA (i.p.) both led to a marked increase in hepatic ODC activity at 7 h and 4 h, respectively, following injection. A 4-5-fold increase in putrescine levels occurred in the rat liver in a biphasic manner between 4-8 h and 16-24 h following the injection of TPA (100 micrograms). No alterations in either spermidine or spermine were observed during this period. The administration of 100 micrograms of TPA to the rat did not alter the incorporated control animals. Under these identical conditions partial hepatectomy led to a large increase in DNA synthesis.
Animals↗