INT. J. RADIAT. BIOL ., 1983, VOL . 44, NO . 4, 3 77-386

Oncogenic transformation of C3H/10T2 cells by X-rays,fast-fission neutrons, and cyclotron-produced neutrons

ELIZABETH K. BALCER-KUBICZEKand GEORGE H. HARRISONDivision of Radiation Research, Department of Radiation Oncology,University of Maryland School of Medicine, Baltimore, MD 21201, U .S .A .

(Received 24 February 1983; revision received 1 June 1983;accepted 7 June 1983)

Lethality and oncogenic transformation were measured in C3H/10T2 murinefibroblasts exposed to neutrons and X-rays at doses between 0 .5 and 11 Gy .Transformation results with X-rays and low-energy, reactor-producedneutrons were used as a baseline to compare and evaluate the results obtainedwith high-energy, cyclotron-produced neutrons . The radiations were 100-kVpX-rays at 0 .49 Gy min 1, reactor fission neutrons at 0. 10 to 0. 31 Gy min-1 withan 8 to 20 per cent gamma dose component, and cyclotron-produced neutrons at0.51 Gymin-1 with mean energy 38 MeV and an 8 per cent gamma dosecomponent. The radiobiological effectiveness (r .b .e .) for cell lethality was 2 .4±0 . 2 for fission neutrons and 1 .7±-0 . 1 for high-energy neutrons . The maximumproportions of transformants per thousand surviving cells were, respectively, 3 . 7±0.8, 6 .5±0 .7, and 2 .3±0 .6 for X-rays, fission neutrons, and cyclotron-produced neutrons . The maximum observed r .b .e . for transformation inductionwas 3 . 8 for fission neutrons and 1 . 2 for cyclotron neutrons . Thus, high-energyneutrons exhibit a higher r .b .e . for cell killing capacity than for oncogenictransformation in C3H/lOT+ cells .

Indexing terms : oncogenic transformation, neutrons .

1 . IntroductionCompared to lower energy neutrons, high-energy neutrons are desirable in

clinical applications because of the improved beam penetration in tissue (Grant et al.1978, Harrison et al . 1978) and the possible reduction of the oxygen effect (Harrisonand Balcer-Kubiczek 1980) . Evaluation of neutron beams for cancer therapy shouldalso include the oncogenic risk from high-energy neutrons since the exposure ofnormal cells in or near the primary beam is unavoidable during therapy . In addition,such data are necessary for assessing the health hazards for exposed personnel tohigh-energy neutrons in man-made or extra-terrestrial environments .

In previous studies, the oncogenic effects of neutrons have not been assessed overa wide range of energy due, in part, to the limited availability of intense neutronsources with mean energy above 14 MeV . For neutrons below 14 MeV, thecumulative data from observations on atomic bomb survivors (Rossi and Kellerer1974), investigations with experimental animals (Fry 1981, Upton et al . 1970,Darden et al. 1967) and studies of oncogenic effects in vitro using cell transformationassays (Borek et al . 1978, Han and Elkind 1979) have indicated an r .b .e . significantlygreater than 1 . The high quality factors assigned to MeV-range neutrons do not takeexplicitly into account the influence of neutron energy on biological effectiveness(ICRU 1971) . Some changes in the r.b .e. value for neutrons in the high-MeV range

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E. K. Balcer-Kubiczek and G . H. Harrison

could be anticipated on the basis of the associated energy deposition processes inorganic media: at lower energies the densely ionizing quality of neutron radiation isdue to recoil protons, while at higher energies, this quality is due almost entirely toalpha particles and ion fragments resulting from neutron reactions with carbon,nitrogen and oxygen (Harrison and Balcer-Kubiczek 1980) .

We report here results of experiments on in vitro oncogenic transformation andcell lethality following exposure to fission-spectrum and cyclotron-producedneutrons ; 100 kv X-rays were used as the reference low-LET beam . The respectivemean energies of the fission neutrons and the cyclotron neutrons were 0 .5 and38 MeV, so that the physical radiation qualities of the two neutron beams differedappreciably . Experiments were carried out with the established cell line ofC3H/10T2 mouse embryo-derived fibroblasts (Reznikoff et al . 1973 a) . This systemis regarded as a valuable adjunct to studies of carcinogenesis with many agentsincluding different radiation qualities, oncogenic viruses, and chemicals (Han andElkind 1979, 1980, 1982, Yang et al . 1980, 1981, Lloyd et al . 1978, Mondal et al.1976, Reznikoff et al. 1973 b, Terzaghi and Little 1976) .

2. Materials and methods2 .1 . Biological material

The initial culture of the C3H/10T2 mouse embryo fibroblasts was generouslysupplied by Dr. S. Mondal of the University of California at Los Angeles .Subpopulations of these cells, in passages between 6 to 9, were used in this study .Cultures were grown in Basal Medium Eagle's (BME) buffered at pH =7 . 3, with 15per cent foetal bovine serum (FBS) and gentamycin at 5 ltg/ml in a humidified 5 percent CO 2 atmosphere maintained at 37°C . Stock cultures were maintained inexponential growth phase and never allowed to attain confluency. Other details ofcell storage and handling prior to irradiation followed published procedures(Terzaghi and Little 1976) . Our approach to transformation and survival studies wassimilar to that previously described by others (Han and Elkind 1979, 1980) . Briefly,aliquots containing 5 x 10 5 cells were transferred to several 25 cm 2 plastic flasks andincubated for 36 hours prior to irradiation . For experimental treatment, flasks werefilled with medium and randomly assigned to treatment and control groups allowing1 flask per group. Three hours after treatment, the medium was removed and cellswere subcultured . The cell population in each flask was determined separately usinga cell counter, and appropriate dilutions were made to yield 250 to 300 viable cells per100-mm Petri dish .

The culture medium was replaced following a refeeding protocol similar to thatdescribed by Bertram (1977), but serum content was always lowered to 5 per cent aweek after initial plating and was maintained at that level throughout the incubationperiod. The densely stained colonies were classified into 3 phenotypes using criteriaoriginally established by Reznikoff et al . (1973 b) . Only clonally-derived cell linesfrom Type 2 and Type 3 colonies have been shown to have tumorigenic potentialin vivo (Reznikoff et al . 1973 b, Terzaghi and Little 1976, Han and Elkind 1979),and we scored them as malignantly transformed .

2 .2 . Transformation assayThe transformation frequency per colony was calculated as a ratio of the

parametric mean of the Poisson distribution of the transformants induced by a given

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Oncogenic transformation of C3H/1OT 1-2 cells

379

dose to the average number of cells surviving this radiation dose (Han and Elkind1979, Kennedy et al. 1980). Standard error (SE) of the mean number of colonies perdish was calculated from formula SE =(u/N) 112 , where y is the mean number ofcolonies per dish and N is the number of dishes used to determine the mean (Boag1975). Least-square fitting procedures were applied to establish dose-responserelationships (Alper et al . 1962) . The r .b.e. of neutrons for cell survival wascalculated from the ratio of X-ray D o to that of neutrons (fission or high-energy) .Unless stated differently, the r .b.e. for transformation induction was estimated fromiso-effect dose ratios (ICRU 1963) .

2.3 . Irradiation sourcesThe relevant dosimetric data on the 3 radiation sources used in this study are

summarized in table 1 . The classical formulation of microdosimetric concepts andtheir applications is given elsewhere (Kellerer and Rossi 1972, Caswell and Coyne1978, Rossi 1978) .

The d(80) + (Be + Ta) neutron beam generated at the University of Marylandcyclotron has been previously described (Harrison et al . 1978, Harrison and Balcer-Kubiczek 1980) . The source of fission-spectrum neutrons was a General AtomicTRIGA Mark-F pool-type thermal research reactor, operated by the Armed ForceRadiobiological Research Institute (AFRRI) . Dosimetric information on the fissionneutron beam used (given in table 1) was obtained from the AFRRI staff (privatecommunication) . The X-ray source was a General Electric MaximaR-100 unit . Itgenerates X-ray beams with a half-value layer of 1 .4 mm of aluminium whenoperated at 100 kV and 5 mA with 1 mm of aluminium added filtration .

Source

Parameter

Mean energy (MeV)tDose rate (Gy min -1 )Photon component (%)YD (keV/um)$yD (keV/um)§

t Calculation of mean energies are based on experimental spectra describing similarsources (Harrison and Balcer-Kubiczek 1980, Lavigne et al . 1977, Leroux et al. 1978) .

$ Dose average lineal energy without saturation effect ; yo = 1 0'y . D(y) dy .§ Dose average lineal energy with saturation effect; yD= fo ysa, • D(y) dy where

ysai=y{(yo/y 2 )[1 -exp (-y2lyo)]} . The saturation parameter was taken to be equal to125 keV/um (Kellerer and Rossi 1972) .

Table 1 . Dosimetric characterization of radiation sources .

2.4 . Irradiation conditionsFor X-ray exposures, cellular monolayers attached to the bottom of 25 cm 2 flasks

were positioned at a source-to-surface distance (s .s .d .) of 25 cm and irradiatedthrough a 2 .5 cm layer of nutrient medium . Flasks were irradiated individually using

X-raysFissionneutrons

d(80) + (Be + Ta)neutrons

0.033 0 . 5 380.49 0.10-0. 31 0 . 51100 8-20 84. 3 75 974. 3 60 25

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3 80

E. K. Balcer-Kubiczek and G. H. Harrison

a randomized dose sequence . In fission neutron experiments, flasks were irradiatedsimultaneously at different s .s .d. values and hence, different dose rates (table 1) .Distances from the central line of the reactor core were selected to obtain the desireddose rates at the locations the cellular monolayers would occupy during theexposure. The exposure time was approximately 5 min for all doses used in thisexperiment. In experiments with the d(80+(Be+Ta) neutrons, flasks wereirradiated individually in a water phantom positioned at a s .s .d. of 43 cm . Cells wereexposed through 0.5 cm of plastic (walls of the water-phantom and flask) and 2 .5 cmof nutrient medium, at the position of maximum dose rate (Harrison et al . 1978) .With a deuteron beam current of 0 .23 µA, the dose rate nearly matched the dose ratein the control X-ray experiments) .

Experiments with X-rays were alternated with the fast-neutron experiments ; X-ray and fission neutron experiments were performed within 2 hours of each other,and cells were obtained from the same population . In all experiments, the cells werekept at ambient temperature for about 4 hours before irradiation, and subcultured 3hours after the end of irradiation . In X-ray experiments, this time-course wasimposed artificially in order to match conditions of the neutron exposures .

3. ResultsThe mean values from several experiments, in which both transformation

frequency and survival were measured, are combined in table 2 . For doses at whichmore than one experiment was performed, the data were pooled . This procedure wasjustified because experiment-to-experiment fluctuations at any dose were within theSE range of variability of individual data points . The observed number of coloniesper dish never exceeded 300, so that cell density modulation of radiation-inducedtransformation was avoided . No transformed foci were observed in all 173 controldishes, which indicates an upper limit for spontaneous transformation of 3 . 5 x 10- sper colony. Plating efficiency ranged from 7 .5 to 17 per cent .

The colony-forming ability of C3H/10T2 cells as a function of dose along withthe parameters of the dose-response curves are shown in figure 1 . Similar results forX-rays (Terzaghi and Little 1976, Han and Elkind 1979, Little et al . 1979) and forfission neutrons (Han and Elkind 1979) have been reported . The intermediate valuesof Do and Dq we found for d(80) + (Be + Ta) neutrons are expected on theoreticalgrounds (Kellerer and Rossi 1972) . The r .b .e . for cell killing was (2.4±0 .2) for fissionneutrons, and (1 . 7±0 .1) for cyclotron neutrons . The general trend of the decreasingr.b .e. with increasing neutron energy for cell inactivation has also been establishedfrom theoretical models (Kellerer and Rossi 1972, Goodhead 1980) and confirmedexperimentally in different mammalian cell systems by others (Ngo et al . 1979, Hallet al . 1972) .

We plotted in figure 2 the transformation induction per cell surviving a givendose . A logarithmic increase of transformation frequency with the d(80) + (Be + Ta)neutron dose, up to 3 .5 Gy, was observed. The r .b .e . values for transformation atdifferent transformation levels, measured at doses below 3 . 5 Gy, are given in table 3 .The decreased effectiveness of the cyclotron neutrons in the plateau region is evidentfrom the right-hand panel in figure 2. It is interesting to note quantitative differencesbetween the three radiation sources when cell lethality is the endpoint scored .In contrast, when the transformation is the endpoint scored, X-rays andd(80) + (Be + Ta) neutrons appear to have a similar potential for transformation

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Oncogenic transformation of C3H/10Tz cells

381

Total number of colonies classified as type 2 and type 3 .Parametric mean of Poisson distribution . A is derived from fraction of dishes without

transformed foci (type 2 and type 3) .§ TR/colony = transformation frequency per viable cell; it is equal to ratio of mean number

of transformants per dish (A) and mean number of colonies per plate .

Table 2 . Survival and transformation of C3H/10TZ cells by X-rays, fission-spectrumor d(80) + (Be + Ta) neutrons .

Dose (Gy) SFTotaldishes

Totalfocit

Total dishesw/foci 1$

TR/colony§x 10 -4

X-ray0 . 0 1 . 0 90 0 0 -1 . 5 0. 9 98 3 2 0.0206 1 . 52 . 0 0-85 63 6 4 0-0656 4. 52-5 0-76 20 5 3 0. 163 6 . 93-0 0-72 41 11 8 0.217 11 . 03 . 5 0-56 28 9 6 0. 241 14. 44-0 0 . 51 41 20 16 0. 495 25 . 44 . 5 0 . 35 21 12 8 0-480 31 . 25-0 0 . 33 39 14 13 0.406 30-15 . 5 0 . 18 20 17 13 1-050 37 . 06-0 0 . 16 37 20 15 0-520 38 . 07 . 0 0 .065 33 17 14 0-552 37 . 68 . 0 0-042 14 10 7 0.693 35 . 39 . 0 0 . 019 20 14 9 0.597 38 . 3

10 . 0 0-013 36 16 12 0-405 33-211 . 0 0 .0056 20 12 8 0. 511 36-5

Fission neutrons0.0 1 . 0 39 0 0 -0. 50 0 . 91 46 3 2 0-0445 4 . 21 .00 0 . 89 39 10 6 0-167 12 . 11-50 0 . 52 20 6 4 0-223 23 . 72. 00 0 .44 18 17 10 0. 811 38 . 82.50 0-15 17 18 10 0. 887 50. 43 .00 0-071 19 20 12 0.999 55-83 . 50 0-049 19 28 14 1 . 335 57 . 54.00 0 .011 20 17 11 0 . 799 65-05-00 0-0031 20 22 12 0-916 64-1

d(80) + (Be + Ta)0. 0

neutrons1 .0 44 0 0 -

1 . 50 0-86 45 4 3 0-069 2-52-00 0-59 33 6 6 0-129 5-92-50 0 . 26 31 10 3 0-215 9-63-00 0-17 39 15 11 0-331 17 . 43 . 50 0-13 42 16 12 0-336 21-84.00 0 .070 35 16 13 0-464 23-14. 50 0-050 30 14 11 0-457 23-25 . 00 0-018 37 17 10 0-392 22 . 36. 00 0-0091 32 19 13 0.521 23-4

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38 2

E. K. Balcer-Kubiczek and G . H. Harrison

10.

z0U 1 .0QW

z

> 0.1

0.01

>-1(52

zOo~ UU

on- 10-3UZ Za W

HaWXU_ 410

4

6

8DOSE (Gy)

Figure 1 . The survival of C3H/10TZ cells after exposure to different doses of X-rays, fission-spectrum and d(80) + (Be + Ta) neutrons . Survival curve parameters : Do, D q , and nare listed with their standard errors (SE) following symbols representing differentradiation sources. 0 X-rays : D0 =(1 .62±0.04)Gy, Dq =(2 .77±0 .07)Gy, n=5 .6 ;• d(80)+(Be+Ta) neutrons : D0 =(0 .98±0.04)Gy, Dq =(1 . 37±0.06)Gy, n=4 .0; Ofission neutrons : D0 =(0.68±0 .04)Gy, Dq =(1 .15±0 .07)Gy, n=5 . 4. SEs of survivingfractions are smaller than the symbols we used to plot the colony-forming ability at agiven dose .

0 2 10 12

d

d

I i i

I i i i0

4

8

0

4DOSE (Gy)

Figure 2 . The transformation frequency per surviving cell (colony) in C3H/I0T2 fibroblastsafter exposure to different doses of X-rays, fission-spectrum neutrons (left panel) and tod(80)+(Be+Ta) neutrons (right panel). In the right panel, solid lines trace the curvesfor fission neutrons and X-rays from the left panel . Bars are SEs of pooled data .0 X-rays, • d(80) + (Be + Te) neutrons, O fission neutrons .

8 12

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d(80) + (Be + Ta) cyclotron-produced neutrons (d(80)n) .

induction while the reactor neutrons differ significantly from both of them . In theplateau region, the maximum proportion of transformants following X-ray orhigh-energy neutron irradiation were not statistically distinguishable . Note thatwhile high energy neutrons exhibit a higher r .b .e . for cell killing then for oncogenictransformation, conversely fission neutrons exhibit a higher r .b .e . for transformationthan for cell killing .

4 . DiscussionThe parameters of survival and transformation induction curves from our

experiments with X-rays and fission neutrons agree with other previous work usingthe C3H/10T2 system (Terzaghi and Little 1976, Han and Elkind 1979) . Qualitat-ively similar results were obtained by Borek et al . (1978), even though different cellsystems were used . This supports the view that the malignant transformation assay iscapable of providing objective measures of relative oncogenicity of a test treatmentcompared to a standard treatment, although there appears to be some dependence ofexpression of malignancy in vitro on experimental conditions and techniques . InC3H/lOT- cells, transformation frequency has been shown to vary with cell densityduring post-treatment incubation (Terzaghi and Little 1976, Han and Elkind 1979,Kennedy et al . 1980), refeedings with media supplemented with different serumconcentrations (Bertram 1977), or the use of specialized topical antibiotics (Terzaghiand Little 1976) . Other factors pertaining to the cell culture system in the course ofthe experiments, such as phase of growth (Little et al . 1979) and passage number(Lloyd et al . 1979), have also been shown to affect the efficiency of the expression oftransformation . The experiments reported here were controlled for factors known orsuspected to influence the transformation process, so that comparisons to otherpublished results could be made . Thus, while the absolute magnitude of thetransformation frequency could a priori be dependent on the particular combinationof experimental conditions and techniques employed, the r .b .e . estimates are validsince they were obtained using identical conditions and techniques for the differentradiations .

For C3H/10T2 cells, the r .b .e . for transformation at therapeutic dose levels hasbeen reported for few high LET radiation sources . Only the oncogenic effect offission neutrons has been studied in detail (Han and Elkind 1979, 1982) . Experi-ments with heavy ions (Yang et al . 1980, 1981, Yang and Tobias 1981) wereperformed using virally-infected C3H/10T2 cells ; thus, the r .b.e. for transformationwith these beams, but without intervening viral influence, remains to be determined .

Oncogenic transformation of C3H/ l0T' cells 383

Radiation dose (Gy) r .b .e .

TR/colony X-ray fn d(80)n fn

d(80)n

5 x 10 -4 2. 20 0. 58 1 . 87 3 .8

1 .21 x 10 -3 2.90 0. 85 2 . 45 3 .4

1 .21 .5x10" 3 3 . 51 1 . 13 2 . 91 3 . 1

122 x 10 -3 3 . 82 1 . 37 3 . 36 2.8

1 . 1

Table 3 . The radiobiological effectiveness (r .b .e .) of fission-spectrum (fn) and

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E. K. Balcer-Kubiczek and G . H. Harrison

The precise relationship between r.b .e . and radiation quality cannot beestablished until more data are obtained . However, our results suggest that this pointmay now be tentatively addressed . The effectiveness of the densely ionizing qualityof these radiations can be correlated with a number of microdosimetric parameters .We have chosen L (Caswell and Coyne 1978, Rossi 1978) because this parameter canbe estimated for all radiation types in question, and because it takes into account thebiological overkill effect of saturation occurring for extremely densely ionizingradiation effects in tissue (Rossi 1978) . yA decreases as neutron energy increases(Caswell and Coyne 1978) . A survey of all the available data for C3H/lOT2 cells atdifferent yD values also reveals a positive correlation between r .b .e . and yD . It shouldbe noted that the r .b .e . for transformation as well as for cell killing cannot bedelineated in :PD . This suggests the possibility that lesions with similar energyrequirements might be involved in both cellular processes .

To compare our results with in vivo experimental findings, we note that Fry(1981) has demonstrated that the r.b .e . for Harderian tumour induction in RF micedecreases by approximately a factor of 2, as mean neutron energy increased from0 .85 MeV (fission-spectrum neutrons) to 25 MeV (cyclotron-produced, p(65) + Beneutrons) .

The central result of our studies with C3H/lOTZ cells is that the r .b.e. forneutron-induced cell lethality and transformation is less for high-energy cyclotron-produced neutrons than for fission neutrons . These results, along with the in vivoresults of Fry cited above, are consistent with a trend of lower oncogenic risk ofneutron irradiation at therapeutic dose levels with increasing neutron energy .

AcknowledgmentsThe authors wish to thank Professor J . E . Robinson of the University of

Maryland Department of Radiation Oncology for advice and helpful discussionsduring the course of this work . The authors are indebted to Dr. P. G . Roos andDr. D. Goldberg, and to the cyclotron operators, Mr . S. Shanks and Mr . J . Brettfrom the University of Maryland Department of Physics for their technical help farbeyond the call of duty ; the last experimental run performed at the University ofMaryland Cyclotron was devoted to this project . The support from Professor FrankJ. Munno of the University of Maryland Department of Nuclear Engineering, andthe technical assistance of the staff of the AFRRI reactor are also gratefullyacknowledged . We are also indebted to Dr . Antun Han who read the manuscript andmade helpful suggestions .

La destruction et la transformation oncogene ont ete mesurees sur du murin fibrocyteC314/10T-f' expose a un bombardement de neutrons et a un rayonnement X a des dosescomprises entre 0,5 et 11 Gy .

Les resultats de transformations obtenus avec des rayons X et un bombardement deneutrons basse energie produits en reacteur ont servi de base a la comparaison et al'interpretation des resultats obtenus avec des bombardements de neutrons haute energieproduits en cyclotron .

L'intensite de radiation utilisee a ete 100kVp rayons X a 0,39Gy min -1 ; neutrons defission de 0,1 a 0,31 Gymin -1 avec dose composant gamma de 8 a 20 pour cent, et neutronsproduits en cyclotron a 0,51 Gymin -1 avec energie moyenne de 38 MeV et dose composantgamma a 8 pour cent .

L'efficacite radiobiologique (e .r .b .) pour la destruction des cellules a ete 2,4±0,2 pour lesneutrons de fission, et 1,7 ±0,1 pour les neutrons haute energie . Les proportions maximales de

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385

transformation par millier de cellules ayant survecu ont ete 3,7±0,8 pour le rayonnement X,6,5±0,7 pour les neutrons de fission, et 2,3±0,6 pour les neutrons produits en cyclotron .L'efficacite radiobiologique maximum observee pour les transformations induites a ete 3,8pour les neutrons de fission et 1,2 pour les neutrons produits en cyclotron .

Ainsi, les neutrons haute energie presentent une meilleure e .r .b. dans l'aptitude a detruireque pour la transformation oncogene des cellules C3H/lOT2 .

Letalitat and onkogenische Transformation wurden in C3H/IOTZ Gewebezellen derMaus gemessen, die Neutronen and Rontgenstrahlen ausgesetzt wurden, bei Mengenzwischen 0,5 and 11 Gy . Transformationsergebnisse fur Rontgenstrahlen and im Reaktorgebildete Neutronen mit niedriger Energie wurden als Ausgangspunkt zum Vergleich and zurBewertung von Ergebnissen fur im Zyklotron gebildete Neutronen mit hoher Energieverwendet . Die Strahlungen waren lOOkVp Rontgenstrahlen bei 0,49Gymin -1 , Re-aktorspaltungsneutronen bei 0,10 bis 0,31 Gymin - 1 mit 8 bis 20 prozent Gammadosisanteil,and im Zyklotron gebildete Neutronen bei 0,51 Gy min -1 mit Durchschnittsenergie 38 MeVand 8 prozent Gammadosisanteil . Die radiobiologische Wirksamkeit (r .b .e .) der Zellen-letalitat war 2,4±0,2 fur Spaltungsneutronen and 1,7±0,1 fur Neutronen mit hoher Energie .Der maximale Anteil an Umwandlern pro Tausend uberlebende Zellen war 3,7±0,8, 6,5±0,7, bzw. 2,3±0,6 fur Rontgenstrahlen, Spaltungsneutronen, bzw . im Zyklotron gebildeteNeutronen. Die maximale beobachtete r .b .e . fur Transformationserregung war 3,8 furSpaltungsneutronen and 1,2 fur Zyklotronneutronen . Also weisen Neutronen mit hoherEnergie eine hohere r .b .e. zur Zellentotungsfahigkeit auf als zur onkogenischen Transform-ation in C3H/IOTZ Zellen .

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