Inhibition of Cyclophosphamide and Mitomycin C-Induced Sister Chromatid Exchanges in Mice by Vitamin C

Home , Cyclophosphamide

ICANCERRESEARCH46,2670-2674, June 19861 Inhibition of Cyclophosphamide and Mitomycin C-induced Sister Chromatid Exchanges in Mice by Vitamin C

G. Krishna,' J. Nath, and T. Ong Natio,wilnstitutefor OccupationaiSafety and Health, Division ofRespiratory Disease Studies, Morgansown, West Virginia 26505-2888 fG. K., T. 0.], andDivision of PlantandSoil Sciences,West VirginiaUniversity,Morgantown,West Visijnia 265O6fJ.N.J

ABSTRACF The research reported here was performed to determine the effect of ascorbic acid on SCEs induced by CPA and MMC in Ascorbic acid (vitamin C) is known to act as an antimutagen and in vivo and in vivo/in vitro conditions in bone marrow and anticarcinogen in several test systems. However, there is no report of its spleen cells of mice. Analysis of SCEs is a sensitive cytogenetic effect on carcinogen-induced chromosomal damage in vii'o in animals. technique for detecting cellular chromosomal damage (11). The present study was performed to determine whether or not ascorbic SCEs are visualized as reciprocal exchangesof staining inten acid affects sister Ã©bromatidexchanges (SCEs) induced by cyclophos sities between sister chromatid arms in metaphase cells that phamide (CPA) and mitomycin C (MMC) in bone marrow and spleen cells in mice. The results indicate that ascorbic acid per se did not cause have replicated twice in the presenceof BrdUrd. a significantincreaseinSCEsin mice.However,increasingconcentra tions of ascorbic acid caused decreasing levels of CPA- and MMC induced SCEs in both cell types in rho. At the highest concentration of MATERIALS AND METHODS ascorbic acid, 6.68 gfkg, approximately 75 and 40% SCE inhibition in Animals. Male CD-I mice were purchased from Charles River Breed both cell types was noted for CPA and MMC, respectively. likewise, under in ris'o/in ritro conditions (exposure of *nimals to experimental ing Laboratories, Wilmington, MA. All animals were 2-3 mo old and chemicals followed by culturing of cells), ascorbic acid caused a dose weighed 25â€”30g.Mice were housed separately in groups of 4 in cages related decrease in CPA- and MMC-induced SCEs, up to a dose of 3.34 containing hardwood chip bedding and excelsior nesting material. g/kg. At this concentration, approximately 50% CPA- and MMC-induced Water and Purina laboratory rodent chow were provided ad libitum SCE inhibition was observed in both cell types studied. Thus, ascorbic throughout the period of animal holding and experimentation. acid acts as an anti-SCE agent in both in rivo and in rho/in ritro Drug Treatment. CPA and MMC (Sigma Chemical Co., St. Louis, conditions in mice. MO) were dissolved in PBS and immediately injected i.p. in a volume equivalent to 10 ml/kg of body weight (40 mg of CPA and 2.5 mg of MMC per kg of body weight). These concentrations were chosen INTRODUCI1ON because they caused a significant increase in SCE levels in mice in earlier studies (12â€”14).L-AscorbiCacid sodium salt (Calbiochem-Behr Ascorbic acid (vitamin C) is known to act as an antimutagen ing Corp., La Jolla, CA), was dissolved in sterile distilled water and and anticarcinogen in various test systems (1). Under in vitro injected (i.p.) in different dosages (1.67, 3.34, and 6.68 g/kg of body conditions, it decreasescarcinogen-induced gene mutations (2, weight) immediately after drug treatment. These ascorbic acid concen 3), SCEs2 (4), and chromosomal breakages (5, 6). In humans, trations were chosen based on our preliminary studies and the studies ascorbic acid prophylaxis has been reported to decreasechro reported by others (15). Negative control animals received an equivalent volume of PBS. For the ascorbic acid control, only the highest dose mosomal damagein peripheral lymphocytes ofcoal-tar workers (6.68 g/kg of body weight) was used. occupationallyexposed to polycyclic aromatic hydrocarbons In Vi,'o Sister Chromatid Exchange Assay. Paraffin-coated BrdUrd and benzene (7). It has been shown to be anticarcinogenic in tablets(50 mg;BoehringerMannheinBiochemicals,Indianapolis,IN) rodents treated with UV radiation, benzo(a)pyrene,and nitroso were inserted under the skin on the flank (16). At 16 h after BrdUrd compounds(8). In terminal humancancerpatients,supplemen implantation,experimentalchemicalswereinjected.Fiveh afterchem tal ascorbateis known to prolongsurvivaltimes (9). A signifi ical treatment, animals were given injections of colchicine (4 mg/kg cant dose-related decrease in the appearance of spontaneous GIBCO) and were sacrificed by cervical dislocation 3 h later. For bone mammary tumors following dietary ascorbicacid in 11111mice marrow preparations, both femora and tibia were isolated, and the has been reported (10). adherent muscle was removed. Each bone was cleaned with 70% ethyl To date, there is no report of the in vivoanticlastogenicity alcohol, and the head was cut offwith scissors. The marrow was flushed studies of ascorbic acid in animals. Likewise there is a relative out with physiological saline into a centrifuge tube, followed by cen paucity of data on metabolicactivation, detoxification, trans trifugation at 285 x g for 6 mm. The supernatant was removed, and portation, and distribution of test compoundsin the intact the pellet was resuspended with hypotonic (0.075 M KC1)solution for animal. Also, cytogenetic studies in humans would involve 20 mm at 37'C and recentrifuged. The cells were fixed in 2 changes of fixative, 10 mm each, in methanol:acetic acid (3:1). The cells were collection of peripheral lymphocytes and/or bone marrow and resuspended in approximately 0.5 ml of fixative and dropped onto culturing such cells for 1â€”2cellcyclesto assessclastogenic/ precleaned, chilled wet slides and air dried for 24 h. anticlastogenic effects of agents to which humans are exposed. For spleencellpreparations,spleensfromthesameanimalsusedfor However, the resultsfrom suchin vivo/in vitromonitoringmay bone marrow removal were removed by opening the abdominal cavity. not be comparable to the in vivo situations. Thus, to better The spleens were transferred into 15-ml centrifuge tubes, each contain assessthe effect of vitamin C, the genotoxicity studies in both ing 2 ml of Hanks' balanced salt solution (GIBCO). Spleens were in vivo/in vitro and in vivo situations in animals should prove mashed with a spatula, and the debris was removed. Cells were treated valuable. with hypotonic solution and fixed, and slides were prepared for SCE analysis as described for bone marrow cells. Received11/19/85; revised 1/15/86; accepted2/24/86. In Vipo/in Vitro Sister Chromatid Exchange Assay. Animals were The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in given injections of experimental chemicals and sacrificed 8 h later by accordance with 18 U.S.C. Section 1734 solely to indicate this fact. cervical dislocation. In these experiments, animals were not implanted 1 To whom requests for reprints should be addressed. with BrdUrd tablets. Bone marrow cells were obtained as described in 2 The abbreviations used are: SCEs, sister chromatid exchanges; CPA, cyclo the previous section except that bone marrow was flushed out with phosphamide; MMC, mitomycin C; BrdUrd, 5-bromo-2'-deoxyuridine;PBS, phosphate-bufferedsaline; GIBCO, Grand Island BiologicalCo., Grand Island, Ham's F-12 medium (Flow Laboratories, McLean, VA) into a 15-mi NY; RI, replicativeindex. centrifuge tube. After centrifugation, bone marrow cells (approximately 2670

1.5 x 10@cells) were cultured as described in an earlier study (17). At tested in both bone marrow and spleen cells in vivo. At the 30 h after incubation, colchicine (33 @iMfinalconcentration) was added, highest concentration, 6.68 g/kg, it causedapproximately 75% and cells were harvested 3 h later. SCE inhibition in both cell types. Spleens were obtained aseptically by opening the abdominal cavity Similar in vivo data following exposure ofanimals to ascorbic of the same mice used for bone marrow isolation. They were then acid and MMC are shown in Table 2. MMC (2.5 mg/kg) also processed and cultured as described by Neft et a!. (18) with modifica induced about a 7-fold increase in SCEs. Ascorbic acid, as in tion. The cell suspension (approximately 1.5 x 10@cells) was cultured in 5 ml of medium consisting of: 3.70 ml of RPM! 1640 with L CPA experiments, caused a dose-related decrease in MMC glutamine and 4-(2-hydroxyethyl)-l-piperazineethanesulfonic acid induced SCEs. At the highest concentration (6.68 g/kg), it buffer (GIBCO); 1 ml of heat-inactivated fetal bovine serum (GIBCO); causedapproximately a 40% decreasein SCE level in both cell 0.05 ml of penicillin and streptomycin (Flow Laboratories); 0.05 ml of types. These differences were statistically significant (P < 0.01). 200 m@iL-glutaminesolution (GIBCO); 1 x l0@ M2-mercaptoethanol The results of the inhibitory effect of ascorbic acid on CPA (Sigma); 20 gzMBrdUrd (Sigma); and 0.2 ml of lipopolysaccharide and MMC-induced SCEs, in in vivo/in vitro conditions, are (Escherichia coli serotype 01l1:B4; Sigma stock of600 zg/ml in PBS). presented in Tables 3 and 4, respectively. Ascorbic acid caused The cell suspension with the complete medium was dispensed into 25- a dose-related decreasein SCEs induced by both drugs in both cm2 Falcon tissue culture flasks, covered with aluminum foil, and then cell types up to a concentration of 3.34 g/kg. At this concentra incubated at 37'C with 98% relative humidity. At 40 h after incubation, colchicine (33 @iMfinalconcentration) was added, and cells were har tion, it caused 61 and 41% CPA-induced SCE inhibition in vested 3 h later. bone marrow and spleen cells, respectively. Under the same in Harvesting, Staining, and Scoring. For harvesting cultures, the con vivo/in vitro conditions, ascorbic acid (3.34 g/kg) caused51and tents of the flask were decanted into 15-mI Falcon centrifuge tubes. 38% decreases of MMC-induced SCEs in bone marrow and Each flask was rinsed with 3 ml ofHanks' balanced salt solution which spleen cells, respectively. These decreases were statistically was then transferred to the centrifuge tube. The tubes were centrifuged significant (P < 0.01). However, at the highest dosetested (6.68 at 285 x g for 6 mm, and cells were processed for slide preparation as g/kg), ascorbic acid acted as a co-SCE inducer under these described previously. conditions. Staining for SCE analysis was performed according to a modified The data on cell replication kinetics are shown in Tables 1 technique of Perry and Wolff (19) and Goto et a!. (20). Slides were to 4 for respective experiments. The RI of cells from treated stained for 15 mm with Hoechst 33258 (5 @g/ml)and exposed to â€œblackâ€•lightat 55â€”60C for 15 mm at a distance of 1 cm while animals was almost similar to the respective control values in immersed in Sorenson's buffer (phosphate buffer, pH 6.8). The slides in vivo conditions. Similar results were noted under in vivo/in were then rinsed with distilled water and stained with 5% Giemsa (in vitro conditions up to a concentration of 3.34 g of ascorbic acid Sorenson's buffer) for 10â€”15mm. All slides were coded, and cells with per kg. However, at the highest dose (6.68 g/kg), along with at least 38 chromosomes were analyzed for SCEs. carcinogens, ascorbic acid caused significant cell cycle delay To determine Ris, the frequencies of the first, second, and third and and toxicity. subsequentmetaphasesweredetermined in 100consecutivemetaphase cells from each of the 4 animals or cultures. Those cells whose DNA had replicated exclusively before the addition of BrdUrd could not be DISCUSSION distinguished from cells at first metaphase, and those that had gone through 3 or more cell cycles were included as third mitoses. The RI Ascorbic acid in the present study acted asan anti-SCE agent was calculated as follows under in vivo conditions in 2 cell types, against 2 known mutagens and carcinogens. CPA is an alkylating agent which RI â€”1M3+ 2M2 + 3M3 produces highly reactive carbonium ions that react with the 100 electron-rich areas of susceptible molecules, such as nucleic acidsand proteins. However, it must bebiochemically activated. where M@,M2, and M3 represent percentages of first, second, and third MMC is a natural antibiotic which is believed to be reduced in metaphases, respectively (21, 22). Statistical Analysis.The data from SCE assays ofboth in vivoand in vivo to a bifunctional alkylating agent capable of cross-linking vivo/in vitro conditionswere comparedto respectivecontrol values complementary copies of DNA and thus affects the synthesis using Student's I test. The RI data from different experiments were and function of nucleic acids. analyzed using the same test. Percentage inhibition was calculated by Shamberger (1) has recently reviewed in detail a series of using the formula reports on antimutagenic action of ascorbic acid against nitro and nitroso compounds, metal ions, and dietary components in @ 100 â€”No of SCEs/cell in the presence of ascorbic acid 100 bacteria. A decreasein fecal mutagenicity of normal humans No. of SCEs/cell in the absence of ascorbic acid has also been noted following dietary application of vitamin C The number of spontaneous SCEs per cell was subtracted from the (23, 24). Ascorbic acid is known to decrease N-methyl-N'-nitro numerator and the denominator. N-nitrosoguanidine-induced SCEs in mammalian cell cultures (4). Anticlastogenic action ofascorbic acid against 7,12-dimeth ylbenz(a) anthracene (6), CPA, and trenimon (5) in human cell RESULTS cultures has also been reported. The data on the in vivo induction of SCEs in bone marrow Ascorbic acid per se did not act as an SCE-inducing agent and spleencells following animal exposure to ascorbic acid and either under in vivo or in vivo/in vitro conditions. It did not CPA are given in Table 1. The data represent 2 separate induce any significant cell cycle delay and, thus, was not toxic. experiments with at least 2 animals in each experiment. Both Similar results have been reported by Speit et a!. (15) under in experiments were performed under identical conditions. CPA vivo conditions in Chinese hamsters with varying doses (up to (40 mg/kg) causedabout a 7-fold increase in SCEs over that in 10 g/kg of body weight) of ascorbic acid. the controls. Ascorbic acid per se at the highest concentration As both drugs used in the study are believed to interact with tested (6.68 g/kg) did not induce significant changes in SCEs chromosomes by an electrophilic attack on nucleophilic sites in over control values (Tables 1 and 2). However, it causeda dose DNA, it seems possible that an exogenous nucleophile could related decrease in CPA-induced SCEs at all concentrations compete with DNA basesfor the ultimate mutagen and thus 2671

Downloaded from cancerres.aacrjournals.org on September 23, 2021. © 1986 American Association for Cancer Research. VITAMIN C AND CARCINOGEN-INDUCED SCEs IN MICE Table 1 Effectofascorbic acid on cyclophosphamide-inducedsisterchromatidexchangesin mousebonemarrowand spleencellsin viva At least 4 animals were used for each treatment, and 25 second division metaphases were scored for SCEs from each animal. For replicative index, 100 consecutive metaphases were scored for M,, M3, and M3 or more cellanimal.Bone divisions from each marrow@Spleena% of in- Replicative % of in- Replicative indexNegativeTreatment SCEs/cell hibition index SCEs/cell hibition 0.05Ascorbiccontrolb 3.36 Â±0.24c 2.04 Â±0.06 3.63 Â±0.19 1.92Â± 0.08CPAacid (6.68 g/kg) 3.59 Â±0.29 2.00 Â±0.04 3.72 Â±0.36 1.98Â± 0.08CPA(40 mg/kg) 23.41 Â±0.94 1.97Â±0.03 24.12 Â±0.30 1.93Â± + ascorbicacid (40 18.40Â±1.75 24.99 2.04 Â±0.06 17.90Â±0.83 30.36 1.98Â±0.06 g/kg)CPAmg/kg + 1.67 + ascorbicacid (40 13.92Â±1.47 47.33 1.97Â±0.06 14.95Â±0.63 44.75 2.00 Â±0.09 mg/kg + 3.34 g/kg)

CPA + ascorbicacid (40 9.02Â± 1.19 72.27 2.02Â±0.11 8.15Â±0.36 77.94 1.97Â±0.07 mg/kg + 6.68 g/kg) a The decrease in SCEs with increased concentrations of ascorbic acid with CPA was significant (P <0.01). b Phosphate-buffered saline.

C Mean Â± SD.

Table 2 Effect ofascorbic acid on mitomycin C-induced sister chromatid exchanges in mouse bone marrow and spleen cells in vivo At least 4 animals were used for each treatment, and 25 second division metaphases were scored for SCEs from each animal. For replicative index, 100 consecutive metaphaseswerescored for M,, M2,and hi3 or more cellanimal.Bone divisionsfrom each marrow@Splecna% of in- Replicative % of in- Replicative indexNegativeTreatment SCEs/cell hibition index SCEs/cell hibition 0.14Ascorbiccontrolâ€• 3.23 Â±0.16c 1.96Â±0.11 3.43 Â±0.42 1.93Â± 0.08MMCacid(6.68 g/kg) 3.16 Â±0.13 1.98Â±0.13 3.47 Â±0.09 1.97Â± 0.15MMC(2.5 mg/kg) 24.69 Â±0.67 1.97Â±0.05 24.84 Â±0.61 1.95Â± + ascorbicacid (2.5 21.53 Â±0.58 14.72 2.00 Â±0.04 21.12 Â±0.89 27.53 1.94Â±0.07 g/kg)MMCmg/kg + 1.67

+ ascorbic acid (2.5 18.55 Â±0.57 28.61 1.93 Â±0.05 18.36 Â±0.56 38.84 1.99 Â±0.05 mg/kg + 3.34 g/kg)

MMC + ascorbic acid (2.5 16.20 Â±0.24 39.56 1.97 Â±0.06 16.64 Â±0.64 45.85 1.91 Â±0.07 mg/kg + 6.68 g/kg)

a The decrease in SCEs with increased concentrations ofascorbic acid with MMC was significant (P < 0.01). b Phosphate-buffered saline.

C Mean Â± SD.

Table 3 Effect ofascorbic acid on cyclophosphamide-induced sister chromatid exchanges in mouse bone marrow and spleen cells in vivo/in vitro At least 4 animals were used for each treatment, and 25 second division metaphases were scored for SCEs from each animal culture. For replicative index, 100 consecutivemetaphaseswerescoredfor M,, M2,and Al3oranimal.Bone more celldivisionsfrom each Spleen@%marrowâ€• of in- Replicative % of in- Replicative indexNegativeTreatment SCEs/cell hibition index SCEs/cell hibition 0.06Ascorbiccontrolâ€• 6.35 Â±0.4lc 1.95Â±0.09 6.72 Â±0.78 1.97Â± 0.03CPAacid (6.68 g/kg) 6.91 Â±0.31 1.99Â±0.13 6.89 Â±0.70 1.90Â±

0.05CPA(40 mg/kg) 43.64 Â±1.76 1.96 Â±0.01 125.47 Â±2.12 1.98 Â± + ascorbicacid (40 31.47 Â±3.07 32.64 1.98Â±0.08 101.34Â±3.18 20.32 1.99Â±0.09 g/kg)CPAmg/kg + 1.67

+ ascorbic acid (40 21.03 Â±1.05 60.63 1.98 Â±0.16 76.40 Â±2.24 41.32 1.97 Â±0.06 g/kg)CPAmg/kg + 3.34 + ascorbicacid (40 52.94 Â±2.03 â€”â€˜ 1.70Â±0.28' Toxic, no M2cells mg/kg + 6.68 g/kg)

a The decrease and the increase in SCES with various concentrations ofascorbic acid with CPA were significant (P < 0.01). b Phosphate-buffered saline.

C Mean Â± SD. d Ascorbic acid acted as a co-SCE inducer.

C Statistically significant over negative control (P < 0.01). serve as a possible preventive agent against mutagenesis and fact, it has been suggested by Edgar (25) that ascorbate might carcinogenesis. Although usually known for its antioxidant protect against electrophilic attack on cellular DNA by inter property, L-ascorbate, which is the predominant form at phys- cepting reactive agents. It is also possible that vitamin C might iological pH, possessessubstantial nucleophilic character. In haveinhibited cytochrome P-450 enzymesinvolved in CPA and 2672

Table 4 Effect ofascorbic acid on mitomycin C-induced sister chromatid exchanges in mouse bone marrow and spleen cells in vivo/in vitro At least 4 animals were used for each treatment, and 25 second division metaphases were scored for SCEs from each animal culture. For replicative index, 100 consecutive metaphases were scored for M1, M2, and M3 oranimal.Bone more cell divisions from each Spleena%marrowâ€• of in- Replicative % of in- Replicative indexNegativeTreatment SCEs/cell hibition index SCEs/cell hibition 0.12Ascorbiccontrolb 6.04 Â±0.27@ 1.93Â±0.10 6.03 Â±0.20 1.95Â± 0.05MMCacid (6.68 g/kg) 6.09 Â±0.34 1.88Â±0.08 6.14 Â±0.23 1.97Â± 0.08MMC(2.5 mg/kg) 27.56 Â±1.97 2.00 Â±0.09 30.07 Â±1.03 2.00 Â± + ascorbicacid (2.5 21.27 Â±1.34 29.23 1.98Â±0.14 24.48Â±0.62 23.25 2.02 Â±0.07 gfkg)MMCmg/kg + 1.67 + ascorbicacid (2.5 16.56Â±0.92 51.12 1.93Â±0.08 20.94Â±0.80 37.98 1.94Â±0.04 g/kg)MMCmg/kg + 3.34 + ascorbicacid (2.5 31.44 Â±0.73 â€”â€˜ 1.62Â±0.10' 36.35 Â±1.64 1.57Â±0.09' mg/kg + 6.68 gJkg)

a The decrease and the increase in SCEs with various concentrations of ascorbic acid with MMC were significant (P < 0.01). b Phosphate-buffered saline.

C Mean Â± SD. d Ascorbic acid acted as a co-SCE inducer.

â€˜Statistically significant over negative control (P < 0.01).

MMC metabolism in vivo.A similar link between cytochrome corbic acid in mice. The threshold levels of ascorbic acid as P-450 enzymes and inhibitory effect of estrogen-induced renal anti- or co-SCE inducer under culture conditions are notewor carcinoma in Syrian hamsters by vitamin C has been proposed thy. (26). Since the highest dose of ascorbate (6.68 g/kg) did not decreasecarcinogen-induced SCEs in in vivo/in vitro conditions, REFERENCES it appears that the possible direct inactivation of the parent mutagen/carcinogen to harmless intermediates by ascorbate 1. Shamberger, R. J. Genetic toxicoloaj@of ascorbic acid. Mutat. Rca., 133: does not occur under present conditions. 135â€”159,1984. The decreaseofCPA- and MMC-induced chromosomal dam 2. Guttenplan, J. B. Inhibition by L-ascOrbate of bacterial mutagenesis induced by two N-nitroso compounds. Nature (Lond.), 268: 368â€”370,1977. age by ascorbic acid under in vivo/in vitro conditions in the 3. Raina, V., and Gurtoo, H. L. Effect ofvitamins A, C, and E on aflatoxin B,- present study (at least with 2 concentrations) is comparable to induced mutagenesis in Salnwnella typhimurium TA-98 and TA-l00. Tera the study of Sram et a!. (7) wherein ascorbic acid prophylaxis togen.Carcinogen.Mutagen.,5: 29â€”40,1985. 4. Galloway, S. M., and Painter, R. B. Vitamin C is positive in the DNA decreasedthe number ofchromosomal aberrations in peripheral synthesis inhibition and sister-chromatid exchange tests. Mutat. Res., 60: blood lymphocyte cultures of coal-tar workers occupationally 321â€”327,1979. 5. Gebhart, E., Wagner, H., Grziwok, K., and Behnsen, H. The action of exposed to polycyclic aromatic hydrocarbons. However, vita anticlastogens in human lymphocyte cultures and their modification by rat mm C has been reported to act as a co-SCE-inducing agent in liver S9. II. Studies with vitamins C and E. Mutat. Res., 149: 83â€”94,1985. human peripheral blood lymphocyte cultures (5). In our study, 6. Shamberger, R. J., Baughman, F. F., Kalchert, S. L, Willis, C. E., and Hoffman, G. C. Carcinogen-induced chromosomal breakage decreased by the highest concentration of ascorbic acid tested (6.68 g/kg) antioxidants. Proc. Nati. Acad. Sci. USA, 70: 1461â€”1463,1973. also acted as a co-SCE-inducing agent in bone marrow and was 7. Sram, R. J., Dobias, L, Pasterkova, A., Rossner, P., and Janca, L Effect of toxic to spleen cells (as evidenced by the absenceof dividing ascorbic acid prophylaxis on the frequency ofchromosome aberrations in the peripheral lymphocytesofcoal-tarworkers. Mutat. Res., 120: 181â€”186,1983. cells in spleen following CPA treatment) under in vivo/in vitro 8. Ames, B. N. Dietary carcinogens and anticarcinogens. Science (Wash. DC), conditions. However, the same dose in in vivoconditions caused 221: 1256â€”1264,1983. 9. Cameron, E., and Pauling, L. Supplemental ascorbate in the supportive SCE inhibition. At this time, there is no suitable explanation treatmentofcancer prolongationofsurvivaltimesin terminalhumancancer. for this inconsistency. This phenomenon may be related to the Proc. NatI. Acad. Sci. USA, 73: 3685â€”3689,1976. selection of cells, differences in cell cycle length, and/or over 10. Pauling, L., Nixon, J. C., Stiu, F., Marcuson, R., Dunham, W. B., Barth, R., Bensch, K., Herman, Z. S., Blaisdell, B. E., Tsao, C., Prender, M., whelming of repair mechanisms in cells in the presence of Andrews, V., Willoughby, R., and Zuckerkandl, E. Effect ofdietary ascorbic excessiveamounts ofascorbic acid. Also, it appearsthat vitamin acid on the incidence of spontaneous mammary tumors in RIII mice. Proc. NatI. Acad. Sci. USA, 82: 5185â€”5189,1985. C hasa threshold level for its action as anti- or co-SCE inducer 11. Tice, R., Lambert, A., Morimoto, K., and Hollaender, A. A review of the under culture conditions. Weitberg and Weitzman (27) have international symposium on sister chromatid exchanges: twenty-five years of suggested that ascorbic acid, depending upon whether it is in experimental research. Environ. Mutagen., 6: 737â€”752,1984. 12. Kram, D., Bynum, G. D., Dean, R., Schneider, E. L, Farland, W. H., and the oxidized or reduced state, can function biologically aseither Williams, J. R. Effects of acute and chronic administration of mitomycin C a prooxidant or oxygen radical scavenger.Whether it functions on the induction of sister chromatid exchanges in vivo. Environ. Mutagen., 3:489â€”495, 1981. as a pro- or antioxidant in a particular situation will also depend 13. Krishna, G., Nath, J., and Ong, T. Murine bone marrow culture system for on the concentrations and availability of other reactants. The cytogenetic analysis. Mutat. Res., in press, 1986. prooxidant effect is probably mediated by the generation of 14. Nakanishi, Y., and Schneider, E. L. In viva sister-chromatid exchange: a sensitive measure of DNA damage. Mutat. Rca., 60: 329â€”337,1979. oxygen radicals, which in turn can react with ascorbic and 15. Speit, G., Wolf, M., and Vogel, W. The SCE-inducing capacity of vitamin dehydroascorbic acids, thus accounting for the chemical com C: investigationsin vitroand in vivo.Mutat. Res., 78:273â€”278,1980. plexities of these systems. 16. McFee, A. F., Lowe, K. W., and San Sebastian, J. R. Improved sister chromatid differentiation using paraffin-coatedbromodeoxyuÃ±dinetablets The major significance of this study, which involved 2 known in mice. Mutat. Res., 119: 83-88, 1983. mutagenic carcinogens and 2 tissue/cell types under in vivoand 17. Krishna, G., Nath, J., and Ong, 1. Preparation of mouse bone marrow primaryculturesfor sisterchromatidexchangeand chromosomalaberration in vivo/in vitro conditions, is that the level of carcinogen studies.J. TissueCult. Methods, 9: 193â€”198,1985. induced SCEs decreaseswith increasing concentrations of as 18. Neft, R. E., Conner, M. K., and Takeshita, T. Long-term persistence of ethyl 2673

carbamate-induced sister chromatid exchanges in murine lymphocytes. Can 23. Bruce, W. R., Varghese,A. J., Furrer, R., and Land, P. C. A mutagen in cerRes.,45:4115â€”4121,1985. [email protected]. Hiatt, J. D. Watson,and J. A. Winsten(eds.),Origins 19. Perry, P., and Wolff, S. New Giemsa method for the differential staining of of Human Cancer, pp. 1641-1644. Cold Spring Harbor, NY: Cold Spring sister chromatids. Nature (Lond.) 251: 156â€”158,1974. Harbor Laboratory, 1977. 20. Goto, K., Maeda, S., Kano, Y., and Sugiyama, T. Factors involved in 24. Dion, P. W., Bright-See, E. B., Smith, C. C., and Bruce, W. R. The effect of differential Giemsa-stainingof sister chromatids. Chromosoma (Berl.), 66: dietary ascorbicacid and a-tocopherol on fecal mutagenicity.Mutat. Res., 351â€”359,1978. 21. Krishna,G., Xu, 3., Nath, J., Petersen, M., and Ong, T. In vivocytogenetic 102: 27â€”37,1982. studies on mice exposed to ethylene dibromide. Mutat. Res., 158: 81â€”87, 25. Edgar, J. A. Ascorbic acid and biological alkylating agents. Nature (Lond.), 248:136â€”137,1974. 1985. 22. Schneider, E. L., and Lewis, J. Aging and sister chromatid exchange. VIII. 26. Liehr, J. G., and Wheeler, W. J. Inhibition of estrogen-induced renal carci Effectof the agingenvironmenton sister chromatid exchangeinductionand noma in Syrian hamsters by vitamin C. Cancer Res., 43: 4638â€”4642,1983. cell cycle kinetics in Ehrlicb ascites tumor cells, a brief note. Mech. Ageing 27. Weitberg, A. B., and Weitzman, S. A. The effect of vitamin C on oxygen Dev., 17: 327â€”330,1981. radical-induced sister-chromatid exchanges. Mutat. Res., 144: 23â€”26,1985.

2674

Downloaded from cancerres.aacrjournals.org on September 23, 2021. © 1986 American Association for Cancer Research. Inhibition of Cyclophosphamide and Mitomycin C-induced Sister Chromatid Exchanges in Mice by Vitamin C

G. Krishna, J. Nath and T. Ong

Cancer Res 1986;46:2670-2674.

Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/46/6/2670

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/46/6/2670. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.