A Study of the Mechanisms of Polyploid Nuclei Formation Induced in Tadpoles by Carcinogen-Protein Conjugates
Authors: E. Marie Hearne Creech
Journal: Cancer Research (1958)

1.  Following immersion of frog embryos in an aqueous solution of 9,10-dimethyl-1,2-benzanthryl-3-carbamido bovine serum albumin-D42 (4 μg/ml), 78 per cent of the dividing nuclei in the tail tips were polyploid at the end of a week. The percentage increased to 90 at the end of the 2d week, declined to 58 at the end of the 5th week, and increased again to 80 at the end of the 6th week.

2.  From a careful cytological study it is concluded that this compound caused an incomplete inhibition of the spindle, resulting in the formation of defective spindles rather than preventing spindle formation. Typical C-metaphases were rare and, even in the absence of a visible spindle, the chromosomes were oriented on a metaphase plate. Polyploid nuclei resulted from this spindle inhibition.

3.  Polytene chromosomes were found frequently in tail tips treated with the protein conjugates of carcinogenic hydrocarbons, and, in some instances, it was possible to detect four chromatids in each chromosome with relational spiraling between pairs of chromatids. From this evidence, it is concluded that the chromosomes had undergone an extra division during interphase (endoreduplication). This mechanism is thought to account for some of the polyploid nuclei.

4.  Prophases similar to Geitler's endomitosis were found frequently; they increased in number at higher concentrations of compound. These could have been owing to endoreduplication during the preceding interphase, to actual prophase reduplication or to the delayed separation of C-pairs from a preceding mitosis.

5.  The unoxidized protein conjugate of 9,10-dimethyl-1,2-benzanthracene was more effective than the partially oxidized form in producing polyploid nuclei and prophases showing a wide range in the degree of chromatid separation; it also produced more polyploids in the light than in the dark. The partially oxidized conjugate was more effective in producing defective spindles. This is explained on the assumption that the unoxidized conjugate, in the dark, produced polyploid nuclei primarily by endoreduplication, whereas in the light it produced polyploids by both spindle inhibition and by endoreduplication; the partially oxidized conjugate produced polyploid nuclei primarily by action on the spindle.

6.  Although the protein conjugate of a noncarcinogen (β-anthryl carbamido horse serum albumin) induced complete spindle inhibition, thus resulting in typical C-metaphases, it produced only a low percentage of polyploid nuclei, and these did not persist. The same explanation could be applied here.

7.  The chromosome behavior induced by the protein conjugates of the carcinogens and of the noncarcinogens paralleled that reported previously for the choleic acid derivatives of carcinogens and noncarcinogens. These choleic acids possess known carcinogenic or noncarcinogenic activity.

8.  There was a disappearance of the highly polyploid nuclei with a selection towards the tetraploid and diploid level over a period of 4 weeks. Aneuploid and lower polyploid nuclei were formed by multipolar mitoses and by lagging and aberrant chromosomes. A detailed analysis of a few tetraploid nuclei revealed that the number of chromosomes was 4 n ± 1 as well as 4 n .

9.  Similar chromosome behavior was found with the protein conjugates of several carcinogenic hydrocarbons. Chromosome bridges and fragments were found in all cases but were more frequent following treatment with 3,4-benzpyrenyl-5-carbamido bovine serum albumin.

10. A much broader spectrum of effects on the chromosomes, on other cell components, and on the timing of the mitotic cycle was produced by derivatives of the carcinogens than by derivatives of the noncarcinogens or by colchicine. The numerous mechanisms for chromosome structural and numerical changes due to the former compounds would tend to create a greater chromosome or gene imbalance. This is mentioned in relation to the theory of gene imbalance in the production of malignancy.