Acta Medica Okayama volume17 issue5

As has been well established, reticulocytes (RC) synthesize the species specific protein, globin, actively for about 24 hours or more till the time of their complete maturation1,2,3. This will be possible only in the presence of messenger RNA (m-RNA)4,5. Since the splendid hypothesis of m-RNA proposed by JACOB and MONOD6 for explaining the mechanism of the transfer of genetic information from nucleus to cytoplasm, it has largely been accepted through the numerous observations that followed7,8,9,10. However, the m-RNA hypothesis, which has been deduced by observing the protein synthesis in E. Coli, includes the meaning of labile RNA which is incessantly decomposed and newly synthesized to compensate the rapid degradation. As m-RNA cannot be synthesized in RC which have no detectable DNA, it has been supposed that the m-RNA of RC should be considerably stablell,12,13. Even in the denucleated cells, however, the RNA synthesis might be possible because Borsook reported the positive RNA synthesis of RC14, and this result has recently been reconfirmed by BURNY15.

Cytochemical observation of the activities of diphosphopyridine nucleotide diaphorase (DPNH-D), triphosphopyridine nucleotide diaphorase (TPNH-D), succinic dehydrogenase (SDH) and a-glycerophosphate dehydrogenase (α-GDH) of Ehrlich ascites carcinoma cells were made and following results were obtained. The smeared cells showed moderate reactions and no marked difference in the intensity among the individual cells. The free floating cells were stained relatively faint but showed the differences in the staining intensity in individual cells. In the presence of benzalkonium, the reaction intensity proved to be intermediate between the smeared cells and free floating cells without benzalkonium and the differences in the staining intensity in individual cells were more marked. Observations revealed that the reaction intensity changes closely corelated with the stage of mitotic cycle of each cell. Namely, DPNH-D activity of the tumor cells, which generally hihger than that of leucocytes, increased remarkably in the end stage of interphase and decreases abruptly in mitotic stage reaching the lowest level in metaphase. After the metaphse the activity increased slightly and it is kept at almost the same level during the first half of interphase. This enzyme is localized mainly in the granules of the eytoplasm. The activity of TPNH-D showed the similar localizations as those of DPNH-D, though the reaction intensity is lower than that of DPNH-D. The activity of SDH of the tmnor cells is lower than that of leucocytes and its diformazan granules are localized in mitochondria. Its activity decreases in the mitotic stage the lowest level in metaphase and in the followed interphase it is kept in a almost constant low level. α-GDH activity of the tumor is lower than that of SDH but show the similar localizations as the latter.

Based on our original concept, a fibroblast-inhibiting agent, chloroquine, was used against various animal tumors. Among transplanted animal tumors, the drug was most effective on relatively connective tissue-rich Bashford and Brown-Pearce tumors, as reflected by prolongation of life span, inhibition of tumor growth, inhibition of lowering of liver catalase activity, improvement of iron metabolism, increase of tumor necrosis, inhibition of connective tissue formation, and decrease of acid mucopolysaccharide. On the other hand, it was of little advantage in Ehrlich, Yoshida and MH134 tumors which contain little connective tissue, except for a decrease of the amount of ascites and ascites tumor cells in the former two tumors. These results indicate that chloroquine suppress the growth of the tumors relatively rich in connective tissue. This effect of chloroquine appears to be due to the primary attack of the stromal connective tissue of tumors being followed by the degeneration of tumor cells, though its probable anti-tumor activity by the indirect effects through its anti-inflammatory and systemic humoral activities should be taken into consideration.

The effects of chloroquine on the growth and morphology of fibroblasts in tissue culture, and in vivo granulomas were investigated. As the result, the drug was shown to have a potent action to inhibit fibroblast growth, which has led to a possibility of its clinical use to patients with malignant tumors.

1. Attempts have been made to confirm how the formazan formation is affected in the presence of oxygen gas when the cells are incubated with neotetrazolium salt and the subsrtates for the enzymes to be tested. 2. In the cases of succinoxidase formazan formation is minimized under pure O2tension, it increases with decrease in O2 tension, and reaches its maximum value under N2gas. 3. This relationship between the oxygen tension and the diformazan formmation can likewise be observed even after pretreatment of the system with KCN. 4. In measuring enzyme activity of the DPN-diaphorase system with L-glutamate and DPN as substrate and NT as hydrogen acceptor, the same relationships between the oxygen tension and the NT-reduction can be seen as in succinoxidase system. 5. In the determination of enzyme activity of the cytochrome-c-cytochrome oxidase system with p-phenylene-diamine as substrate and NT as hydrogen acceptor, likewise the diformazan formation is markedly affected by oxygen tension and increased with the reduced oxygen tension but under pure Ns gas the value is reduced. When the systen is pretreated with KCN, however, the diformazan formation reveals its maximum value under pure nitrogen gas, the values of which correspond to those values of endogenous reaction without substrate. 6. The above results show that the neotetrazolium salt can compete with O2 as hydrogen acceptor, and less values of formazan formation may be obtained under higher oxygen tension and the higher values under lower oxygen tension independently from the true activity of the enzyme.

Since the classic work of Ranvier, it is well known that the mammalian striated muscle is composed of two types of muscle fibers, i. e., the red and white muscle fibers. In the previous paper1 it has been reported that the limb muscle fibers of mammals can be divided into three types from their activities of the histochemically demonstrable oxidative enzymes. Namely, the small red muscle fibers had a higher activity of oxidative enzymes, the large white muscle fibers a lower activity and the third type of muscle fibers being called "medium fiber" or "intermediate fiber" showed an intermediate activity between those of the red and white muscle fibers.