Heterogeneity of ornithine decarboxylase expression in 12-O-tetradecanoylphorbol-13-acetate-treated mouse skin and in epidermal tumors

One of the earliest events after treatment of mouse skin with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) is the induction of ornithine decarboxylase (ODC). Using an immunoperoxidase technique with a rabbit antiserum specific for ODC, the localization of cells containing high levels of ODC following TPA treatment was determined. CD-1 female mice treated with multiple topical applications of TPA and killed 4.5 h after the last TPA treatment exhibited a heterogeneous localization of ODC in this hyperplastic epidermis. The cells which exhibited intense immunostaining were found predominantly in the suprabasal cells lining the hair follicles. This specific ODC staining in cells surrounding hair follicles was inhibited by pretreatment of mice with either retinoic acid or cycloheximide 1 h before TPA treatment. The induction of ODC-specific staining after TPA treatment in hyperplastic mouse skin was transient, since no staining was observed 16 or 24 h after TPA treatment. In contrast, benign papillomas produced by two-stage tumorigenesis contained some cells demonstrating high levels of ODC a week after the last TPA application. These results indicate that both normal mouse epidermal cells as well as tumor tissue display cellular heterogeneity of ODC expression.     

Effects of combined treatments with selenium, glutathione, and vitamin E on glutathione peroxidase activity, ornithine decarboxylase induction, and complete and multistage carcinogenesis in mouse skin

Several structurally different tumor promoters altered to various degrees both glutathione (GSH) peroxidase (EC 1.11.1.9) and ornithine decarboxylase (ODC, L-ornithine carboxy-lyase, EC 4.1.1.17) activities in mouse epidermis in vivo. At 5 h after their application to the skin, the complete tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and the stage 2 promoter mezerein were the most potent in inhibiting GSH peroxidase activity and inducing ODC activity. In comparison, the effects of anthralin, phorbol-12,13-didecanoate, benzoyl peroxide, H2O2, and phorbol-12,13-dibenzoate were much smaller, whereas the nontumor promoter phorbol, the hyperplastic agent ethyl phenylpropiolate, and the stage 1 promoter 4-O-methyl TPA did not alter GSH peroxidase and ODC activities. Various treatments including i.p. injections of 40 micrograms of Na2SeO3 and 100 mumol of GSH and/or topical applications of 40 mumol of D-alpha-tocopherol (vitamin E) 20 or 15 min, respectively, before tumor promoter treatment inhibited in an additive manner the effects of either TPA or mezerein on both GSH peroxidase activity and ODC induction. Moreover, these Na2SeO3, GSH, and/or vitamin E treatments inhibited in the same additive manner the tumor-promoting activity of TPA in the initiation-promotion protocol. However, when tested in the 2-stage promotion protocol with 4 doses of TPA followed by twice weekly applications of mezerein, Na2SeO3 plus vitamin E and GSH plus vitamin E treatments inhibited remarkably the tumor-promoting activity of mezerein but were ineffective in the first stage of promotion. The sequence and magnitude for the effects of 7,12-dimethylbenz[alpha]anthracene (DMBA) on GSH peroxidase and ODC activities were very different from those of the tumor promoters. In contrast with their antitumor-promoting activity, the treatments with Na2SeO3 plus vitamin E and GSH plus vitamin E failed to inhibit the carcinogenicity of a single large dose of DMBA and even enhanced the induction of skin tumors by repeated applications of subcarcinogenic doses of DMBA. These results suggest that the promoting component of DMBA carcinogenesis may be different from that of TPA. Moreover, the anticarcinogenicity of Na2SeO3, GSH, and vitamin E may be linked to their ability to facilitate or enhance the activity of the natural GSH-dependent antioxidant protective system of the epidermal cells during the later stages of skin tumor promotion.     

Relation between induction of rat hepatic ornithine decarboxylase activity by tumor promoters 12-O-tetradecanoylphorbol-13-acetate and phenobarbital and levels of the polyamines putrescine, spermidine and spermine, in vivo; differential effects of retinyl-acetate

A single i.p. injection of 12-O-tetradecanoylphorbol-13-acetate(TPA) induced a transient increase in the levels of rat liverputrescine, spermidine and spermine. These polyamine concentrationscontinuously increased until 6 h, immediately following administrationof the tumor promoter. Phenobarbital (PB) induced an increaseof the putrescine and spermidine concentrations during the 8h post administration studied, while spermine reached a plateauafter 4 h. When retinyl-acetate (RA) was injected one hour priorto TPA, the increases of the polyamines were considerably inhibited.This treatment with RA partially inhibited further increaseof putrescine and spermine by PB. These findings are discussedin respect to the concomitant ornithine decarboxylase (ODC)activities, we have previously reported. I.p. injection of RAalone caused an elevation of ODC activity as well as putrescineand spermidine concentration within 2 h of exposure.     

Regulation of ornithine decarboxylase gene expression in mouse epidermis and epidermal tumors during two-stage tumorigenesis

Topical treatment of mouse skin with the potent tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) results in an array of biochemical alterations, one of the earliest being a more than 200-fold transient induction of epidermal ornithine decarboxylase (ODC) activity. There is an excellent correlation between the induction of epidermal ODC activity and changes in the level of immunoreactive ODC protein following a single TPA treatment to skin. Both ODC activity and protein levels peak at 4.5 h after TPA treatment and rapidly fall to basal levels by 24 h. Cycloheximide treatment of mice in which ODC had been previously induced by TPA indicated a similar rapid turnover of both ODC catalytic activity and protein levels. Northern blot analysis of polyadenylated RNA isolated from mouse epidermis after a single TPA treatment revealed the stimulation of one species of ODC mRNA of 2.0 kilobases with a maximum at 3.5 h declining by 16 h. The same-sized species of ODC mRNA was detected 4.5 h after multiple biweekly treatments with TPA as well as in mouse papillomas and carcinomas not treated with TPA for at least 1 week. Southern blot analysis of EcoRI or BamHI digests of DNA derived from mouse liver, papillomas, or carcinomas revealed no ODC gene amplification or rearrangement during neoplastic progression. These observations indicate that the induction of epidermal ODC activity following TPA treatment results in a transient increase in the steady state levels of ODC mRNA and in the rate of synthesis of ODC protein, in contrast to epidermal tumors where the levels of ODC mRNA and protein are constitutively elevated.     

Inhibition by indomethacin and 5,8,11,14-eicosatetraynoic acid of the induction of rat hepatic ornithine decarboxylase by the tumor promoters phenobarbital and 12-O-tetradecanoylphorbol-13-acetate in vivo

The involvement of arachidonate metabolism in the inductionof rat hepatic ornithine decarboxylase (ODC) activity by thetumor promoters 12-O-tetradecanoylphorbol-13-acetate (TPA) andphenobarbital (PB) was investigated. Pretreatment of the ratswith indomethacin or 5,8,11,14-eicosateraynoic acid dose dependentlyinhibited the induction of ODC by both tumor promoters. Bothinhibitors were more potent inhibitors of PB induction thanTPA induction of ODC. The data are consistent with an involvementof arachidonate cyclooxygenase products in the induction ofrat hepatic ODC by the tumor promoters.     

Characterization of the tumor-promoting activity of thapsigargin in cf-1 mouse skin

The ornithine decarboxylase (ODC), hydroperoxide (HPx) and DNA responses to 12-O-tetradecanoylphorbol-13-acetate (TPA) and mezerein (MEZ) are similar in vivo. Thapsigargin (TG) respectively mimics about 15, 75 and 75% of the ODC, HPx and DNA responses to TPA and these differences persist after chronic treatments. The peak of ODC induction 5 h after 2 TG treatments at 12 to 48-h intervals is twice that produced 16 h after a single TG treatment or 5 h after 2 TG treatments at a 72-h interval. The ODC-inducing activity of TG is dose dependent from 1 to 15 nmol and its magnitude is maximal after 2 applications. The biochemical effects of TG and TPA are neither synergistic nor additive. But the ODC response to TG is greater when this compound is applied 48 h after TPA than after another TG treatment. In initiated skins promoted 2x/week with 5 nmol of TPA, MEZ, or TG, the Ist papillomas (PAs) appear at 7, 12 and 17 weeks, the tumor incidences are 100, 40 and 24%, and there are 17, 1 and 0.6 PAs/mouse, respectively. The tumor-promoting activity of TG is increased at a higher dose (15 nmol), slightly accelerated at a higher frequency (1x/2 days) and decreased at a lower frequency (1x/week). TPA applied 1x/3 weeks is insufficient to promote tumors but slightly enhances the tumor-promoting activity of TG. MEZ applied 1x/4 weeks in stage 2 also accelerates the tumor-promoting activity of TG. TG is effective as a stage 1 promoter but its promoting activity is not enhanced by stage 1 treatments with TPA. In contrast to TPA or even MEZ, TG may be a very weak tumor promoter because it is a very weak ODC inducer and mobilizes enough intracellular free Ca2+ to impair tumor cell proliferation.     

Inhibition of the induction of ornithine decarboxylase activity by 12-O-tetradecanoylphorbol-13-acetate in mouse skin by sphingosine sulfate

We investigated the effect of sphingosine sulfate on the inductionof ODC (ornithine decarboxylase) activity by TPA (12-O-tetradecanoylphorbol-13-acetate)in mouse skin. When applied topically to the shaved skin ofSENCAR mice at dosages of 10–40 µunol per animal,30 min before the superficial application of 8.5 nmol of TPA,sphingosine sulfate dramatically inhibited the induction ofODC activity by the tumor promoter. Significant inhibition ofTPA-induced ODC activity was observed at 4, 6 and 8 h afterTPA treatment in separate studies. The results indicate thatsphingosine sulfate is an effective inhibitor of ODC inductionby TPA in mouse skin.     

Induction of mouse epidermal ornithine decarboxylase activity and skin tumors by 7,12-dimethylben

Application of a single large dose (3.6 micromol) or smaller weekly repeated doses (0.2 micromol) of 7,12-dimethylbenz[a]anthracene (DMBA) to the skin of CD-1 mice led to a 20 to 50-fold increase in epidermal ornithine decarboxylase (ODC) (EC 4.1.1.17) activity as well as tumor formation. Retinoic acid (0.17-68 nmol), a potent inhibitor of both the induction of ODC activity and tumor formation by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), failed to inhibit both the induction of ODC activity and tumor formation by DMBA. In contrast, 7,8-benzoflavone (367 nmol), which did not inhibit the induction of ODC activity by TPA, effectively inhibited the induction of ODC activity as well as the formation of skin tumors caused by DMBA. These results indicate that (a) the mechanism of the induction of ODC activity and tumor formation by a complete carcinogen appears to be different from that of the tumor promoter TPA, (b) DMBA-induced ODC activity may be an important component of the mechanism of DMBA carcinogenesis, and (c) the protective effect of retinoic acid on skin carcinogenesis is not universal; it inhibits skin tumor formation by some agents and not by others.     

Effect of polycyclic aromatic compounds and phorbol esters on ornithine decarboxylase and aryl hydrocarbon hydroxylase activities in mouse liver

Single i.p. injections of 3-methylcholanthrene (MC; 50 mg/kg) administered to inbred C57BL/6 mice or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 100 micrograms/kg) to DBA/2 mice gave an increase in the hepatic activities of ornithine decarboxylase (ODC) and aryl hydrocarbon hydroxylase (AHH) with peaks occurring by 12 and 48 hr, respectively. A single i.p. dose of the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA; 100 micrograms/kg) enhanced the activity of ODC about 70-fold within 12 hr in C57BL/6 mice and 18-fold within 24 hr in DBA/2 mice without affecting AHH activity markedly. 4-O-Methyl-12-O-tetradecanoylphorbol-13-acetate (100 micrograms/kg) raised ODC activity to about 25% of the TPA-treated value in C57BL/6 mice; in DBA/2 mice, TPA and 4-O-methyl-12-O-tetradecanoylphorbol-13-acetate induced ODC activity to roughly the same level. Benzo(e)pyrene (50 mg/kg) failed to affect ODC and AHH activities significantly in either strain. The inducing effect of TPA on ODC activity was potentiated by a simultaneous administration of MC to C57BL/6 mice; combined TPA and TCDD to DBA/2 mice exerted an additive effect on hepatic ODC activity. Difluoromethylornithine administered i.p. effectively inhibited the induction of ODC activity elicited by TPA, MC, or TCDD either alone or in various combinations but did not interfere with AHH induction. These data indicate that different regulatory factors are involved in the ODC induction process elicited by TPA and polycyclic aromatic compounds and that MC and TCDD may induce ODC activity by different mechanisms. The results also confirm our earlier findings in rat skin and cells in culture which suggest that the ODC and AHH induction processes can occur independently of each other. Additionally, there is a strain-related difference in sensitivity with regard to ODC-inducing activity of TPA in the livers of C57BL/6 and DBA/2 mice.     

Regulation of ornithine decarboxylase gene expression in normal and transformed hamster embryo fibroblasts following stimulation by 12-O-tetradecanoylphorbol-13-acetate

We have compared the regulation of ornithine decarboxylase (ODC)gene expression in primary cultures of hamster embryo fibroblastsand in two independently transfonned hamster embryo cell lines.Previous studies have demonstrated that 12-O-tetradecanoylphorbol-13-acetate(TPA) can greatly potentiate the serum growth factor inductionof ODC enzyme activity in transformed cells, but not in normalhamster embryo fibroblasts. Treatment of either normal or transformedcells with both TPA and serum yielded greater accumulationsof ODC mRNA than with either treatment alone, which is consistentwith changes at the protein level. However, treatment of thetransformed cells with TPA and serum resulted in a greater increasein steady state levels of ODC mRNA than that observed usingnormal fibroblasts. The time course for the induction of ODCmRNA was similar for both normal and transformed cells withmaximal accumulations 4–8 h after treatment. Studies withactinomycin D further suggests that ODC mRNA is comparativelylong-lived in both normal and transformed cells. The accumulationof ODC mRNA after stimulation with TPA and serum is blockedby cycloheximide in normal hamster fibroblasts suggesting thatthis induction is dependent upon protein synthesis. In contrast,cydoheximide did not affect the accumulation of ODC mRNA undersimilar treatment conditions in transformed cells. This alteredregulation of ODC gene expression in transformed hamster embryofibroblasts cannot be explained by either gene rearrangementor the amplification of an ODC gene. These data suggest thattransformation of hamster embryo cells results in a loss ofcellular control over ODC gene regulation which includes analteration in the requirement for protein synthesis for ODCmRNA accumulation.     

Induction of epidermal ornithine decarboxylase activity in mouse skin exposed to biogenic silica fibers.

The present study demonstrates that biogenic silica fibers (BSF), previously shown to promote skin tumors in mice and more recently to promote the induction of mesotheliomas when injected into the pleural cavity of rats, rapidly induces epidermal ornithine decarboxylase (ODC) activity in SENCAR mice following topical application. The time course for induction of epidermal ODC by BSF was very similar to that observed following topical treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA). Maximal ODC activity was observed 4-6 h following treatment with BSF. Cycloheximide (70 mg/kg i.p.) partially inhibited (61%) the induction of ODC by BSF at 5 h. In addition, retinoic acid (RA, 5 micrograms per mouse given 30 min before BSF) effectively inhibited BSF-induced ODC by 68%, while indomethacin (100 micrograms per mouse 2 h before BSF) had little or no effect. Copper(II) bis(diisopropylsalicylate) (2 mumol 30 min before BSF), an effective inhibitor of TPA-induced ODC activity and tumor promotion, also had little or no effect on BSF-induced ODC. The work described in this paper suggests that BSF induces epidermal ODC by a very specific mechanism that exhibits both similarities and differences with that of the phorbol ester, TPA. Nevertheless, this response strongly supports the conclusion that BSF is an effective tumor promoter in mouse skin and that ODC induction is an integral part of the mechanism of action of this environmental promoter.     

Inhibition of phorbol ester--induced human epidermal ornithine decarboxylase activity by oral compounds: a possible role in human chemoprevention studies

Extensive animal data have suggested that, in some systems, the induction of ornithine decarboxylase (ODC) is an essential, although not sufficient, aspect of tumor promotion and that compounds that inhibit ODC can inhibit tumor formation. Using fasting human volunteers, we report that human epidermal and dermal ODC are consistently induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) in a manner similar to that seen in mouse skin. There is a marked intersubject variation in TPA-induced epidermal ODC activity levels. Orally administered compounds significantly inhibited TPA-caused human epidermal ODC induction. These data may be useful in the further development of drugs, doses, and dose schedules for use in human cancer chemoprevention studies.     

Comparison of epidermal protein kinase C activity, ornithine decarboxylase induction and DNA synthesis stimulated by TPA or dioctanoylglycerol in mouse strains with differing susceptibility to TPA-induced tumor promotion

The purpose of this study was to examine the activity and associated kinetic parameters of epidermal protein kinase C (PKC) following stimulation by sn-1,2-dioctanoylglycerol (DIC8) or 12-O-tetradecanoylphorbol-13-acetate (TPA) and to examine the relationship between levels of epidermal PKC activity and the induction of ornithine decarboxylase by these agents, utilizing various stocks and strains of mice. Importantly, the mouse strains and stock used in this study have known differing susceptibilities to undergo TPA-induced tumor promotion: the CD-1 stock and the DBA/2 strain (both sensitive to TPA-induced tumor promotion) and the C57BL/6 strain (resistant to TPA-induced tumor promotion). TPA-stimulated protein kinase C activity was measured in the 10(5)g supernatant fraction of epidermal homogenates using lysine-rich histone as a phosphate acceptor substrate. The maximal velocities for TPA-stimulated epidermal PKC activity in CD-1, DBA/2 and C57BL/6 were 0.28, 0.29 and 0.27 nmol PO4-histone/mg 10(5)g protein/min, respectively. TPA-stimulated epidermal PKC from CD-1, DBA/2 and C57BL/6 had similar theoretical Vmax values and the apparent concentrations of TPA yielding half-maximal stimulation of PKC were also similar. DiC8-stimulated PKC activity to a greater Vmax; however, the concentration required to yield half-maximal stimulation of PKC was one thousand times greater than that of TPA. There were no strain differences in these parameters when the enzyme was stimulated with DiC8. Thus, the levels of epidermal PKC activity in CD-1, DBA/2 and C57BL/6 mice exhibit no strain differences when stimulated by TPA or DiC8 using lysine-rich histone as a phosphate acceptor substrate. Since sn-1,2-diacylglycerols are known effective inducers of epidermal ornithine decarboxylase (ODC) activity, the induction of epidermal ODC was examined in each mouse strain 5 h after topical application of 2 nmol TPA, 5 nmol TPA or 2.5 mumol DiC8. After topical treatment with TPA, C57BL/6 demonstrated an unexpected 2- and 4-fold increase in ODC activity over CD-1 and DBA/2 mice. After treatment with DiC8, C57BL/6 demonstrated a 6- and 10-fold increase in ODC activity over CD-1 and DBA/2, respectively. Thus, the resistant strain (C57BL/6) demonstrated a 'hyperinducibility' of epidermal ODC activity by TPA or DiC8. The time course for the induction of epidermal ODC was examined in each strain, and at every time point measured (3-15 h), the C57BL/6 strain exhibited this 'hyperinducibility' of ODC relative to the other strains. Epidermal DNA synthesis was stimulated to a similar extent in C57BL/6 and CD-1 mice.(ABSTRACT TRUNCATED AT 400 WORDS)     

Characterization of the hydroperoxide response observed in mouse skin treated with tumor promoters in vivo

The production of hydroperoxides is rapidly increased and remains at 200-280% of the control 1-24 h after the second daily application of 17 nmol of 12-O-tetradecanoylphorbol-13-acetate (TPA) to mouse skin in vivo. The levels of hydroperoxides are increased 1.63-, 2.64-, 4.07-, and 4.31-fold 18 h after one, two, three, or four applications of TPA at 24-h intervals, respectively. The hydroperoxide response to TPA observed in whole skin reflects almost entirely the increased hydroperoxide-producing activity of the epidermis. Such hydroperoxide responses are triggered to various degrees by the anthrone derivatives and the phorbol esters and diterpene with complete and/or stage 2 tumor-promoting activities but not by the agents with only inflammatory, hyperplastic or stage 1 tumor-promoting activities. However, the Ca2+ ionophores A23187 and ionomycin are potent inducers of hydroperoxide formation. Several discrepancies are observed between the hydroperoxide response to TPA and the known effects of the tumor promoter on ornithine decarboxylase (ODC) induction. In contrast to the refractory state against ODC induction caused by TPA treatments repeated at intervals of less than 48 h, the time interval required for recovery of the hydroperoxide response to TPA in TPA-pretreated skins is only 5 h. The stimulatory effects of A23187, ionomycin and various diacylglycerols (DAGs) on hydroperoxide production do not correlate with their ODC-inducing activities. The increasing susceptibilities of C57BL/6, CF-1, and SEN-CAR mice to skin tumor promotion correlate with their hydroperoxide responses but not with their ODC responses to TPA. alpha-Difluoromethylornithine (DFMO) and other inhibitors of TPA-induced ODC activity fail to alter hydroperoxide production whereas the compounds that inhibit the hydroperoxide response to TPA, such as fluocinolone acetonide, have no or only minimal inhibitory activity against ODC induction. This would suggest that the hydroperoxide response to TPA does not require ODC induction and may not be essential for ODC induction. The hydroperoxide response to TPA is mimicked, but to a lesser degree, by the activator of protein kinase C, 1,2-dioctanoyl-sn-glycerol, and inhibited by verapamil, trifluoperazine, and palmitoylcarnitine. Populations of TPA-treated keratinocytes, therefore, may be responsible not only for ODC activation but also for hydroperoxide production. However, these two responses, which involve, at least in part, Ca2+ mobilization and protein kinase C activation and play important roles in the mechanism of skin tumor promotion, do not appear to be correlated.     

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 ornithinedecarboxylase (ODC) activity in the liver of the rat and mousewas determined. The injection of 12-O-tetradecanoyl phorbol-13-acetate(TPA, 100 µg, i.p.) led to a 250-fold increase in hepaticODC activity within 4 h of administration. This increase inODC activity required both RNA and protein synthesis and didnot occur when a variety of the non-tumor promoting phorbol-esterderivatives were administered to the rat. A distinct dose-dependentincrease in hepatic ODC activity could be observed at 4 h followingthe injection of increasing amounts of TPA (0–100 µg,i.p.). As little as 1.0 µg TPA (i.p.) administered toa rat resulted in a significant stimulation in the activityof ODC in the liver compared to the control unstimulated values.Both 200 µg and 500 µg TPA produced less of an elevationin hepatic ODC activity than did the optimal dose of 100 µg.In the mouse, the administration of 1 µg and 20 µgof TPA (i.p.) both led to a marked increase in hepatic ODC activityat 7 h and 4 h, respectively, following injection. A 4–5-foldincrease in putrescine levels occurred in the rat liver in abiphasic manner between 4–8 h and 16–24 h followingthe injection of TPA (100 µg). No alterations in eitherspermidine or spernine were observed during this period. Theadministration of 100 µg of TPA to the rat did not alterthe incorporation of [3H]thymidine into DNA in the liver comparedto untreated control animals. Under these identical conditionspartial hepatectomy led to a large increase in DNA synthesis.     

Induction of ornithine decarboxylase and histidine decarboxylase activities in rat colon mucosa after application of 12-o-tetradecanoylphorbol-13-acetate (TPA), sodium deoxycholate and indole

Ornithine decarboxylase (ODC) and histidine decarboxylase (HDC) activities of rat colon mucosa were induced after intrarectal instillation of 12-o-tetradecanoylphorbol-13-acetate (TPA) and sodium deoxycholate. After instillation of sodium deoxycholate, ODC and HDC activities increased rapidly and reached a peak at 4 h, then decreased quickly towards control levels. Both enzymes activities also increased significantly 4 h after instillation in animals treated with TPA and sodium deoxycholate plus indole. However, there were no changes in ODC and HDC activities from 0-8 h after indole administration. These data are the first to show the induction of HDC activity in colon mucosa by TPA and sodium deoxycholate and suggest that the induction of activities in these two enzymes might be one mechanism of their action as cancer promoters.     

Ornithine decarboxylase induction by chemically complex liquids from two solvent refined coal processes

Studies with a variety of chemically purified substances havesuggested that induction of the enzyme ornithine decarboxylase(ODC) in mouse epiderinal cells may be a reliable indicatorof neoplastic transformation. In an effort to extend these observationson ODC to chemically complex materials, we examined ODC inductionby carcinogenic and non-carcinogenic mixtures and compared theseresults with tumorigenicity data for these materials. For thesestudies several boiling range fractions and several solvent-derivedsubfractions from two solvent-refined coal processes (SRC-Iand SRC-II) were evaluated for their ability to induce ODC.Single applications of heavy distillate (HD), the SRC-II high-boilingfraction and a potent mouse skin carcinogen, produced ODC inductionkinetics which were similar to that for 12-O-tetradecanoylphorbol-13-acetate(TPA). Both HD and TPA stimulated maximal ODC activity 3–5h after application, with epidermal ODC levels returning tobasal levels within 12 h. The magnitude of ODC induction aftermultiple applications of HD was not as great as that observedfor TPA. Single skin applications of TPA and HD also transientlyelevated hepatic ODC levels 27- and 7-fold, respectively; however,liver ODC activity did not increase following multiple applicationsof either chemical. Further, ODC induction by HD was also dose-dependent.Relative to controls, single applications of HD and processsolvent (boiling range >250°C) elevated ODC levels 145-to 205-fold, light distillate and light oil (boiling range <180°C)increased ODC levels 23- to 32-fold, and middle distillate andwash solvent (boiling range 180– 250°C) stimulated<2- to 8-fold increases in ODC. Single applications of threesolvent-derived subtractions of HD, which are complete carcinogens,induced 3- to 7-fold ODC elevations over background levels;multiple applications of two of these subtractions elevatedODC levels 10- to 22-fold. Of the complex mixtures evaluatedduring this study, all complete carcinogens induced ODC; however,the magnitude and temporal pattern of induction vaned with thematerial tested.