Cell Irradiation Radiotherapy Discussion Chapter

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ionizing radiation on meiotic spindles, 34 oocytes were divided up and then exposed to 0, 74, or 222 Gy of ionizing radiation. Of the six control oocytes that were sham exposed to radiation, one degenerated (Figure, 83.3%). A total of 14 oocytes were exposed to 74 Gy of radiation and only one lacked spindles (Figure, 92.9%). Exposing 14 oocytes to 222 Gy of radiation destroyed spindles in nine oocytes (Figure, 28.6%). These results suggest radiation doses above 74 Gy are capable of degrading meiotic spindles in oocytes.

Sufficient irradiation eliminates oocyte spindles. Oocytes were either sham exposed to radiation, or exposed to 74 or 222 Gy of radiation. Percent of oocytes with intact spindles after irradiation were visualized by confocal microscopy and fluorescent staining for tubulin.

Discussion

Our results indicate that exposure of oocytes to sufficient doses of ionizing radiation can destroy meiotic spindles. The fate of these spindles can't be determined from these experiments, but prior research studies examining irradiation-induced mitotic arrest of cells indicated that spindles depolymerized (Zaremba and Irwin, 1981). The data presented here is consistent with meiotic spindle depolymerization occurring at the highest dose of ionizing radiation used.

Mechanism of Spindle Depolymerization

Spindle formation can occur in the absence of protein synthesis (Inoue et al., 1975; Inoue & Ritter, 1978) and ionizing radiation doesn't alter the rate of tubulin synthesis in cells (Noland et al., 1974).

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This suggests that mitotic spindles form from existing stores of subunits (Zaremba and Irwin, 1981). In support of this theory, cytoplasmic microtubules present during interphase disappear immediately prior to mitotic spindle formation (Brinkley et al., 1975; Fujiwara & Pollard, 1978). In addition, mitotic spindles appear to be in a state of equilibrium with the pool of subunits (Inoue, 1964). Together this research suggests that there exists a cell cycle-regulated state of equilibrium between mitotic spindles, cytoplasmic microtubules, and the tubulin-based subunits which form these structures. A similar state of equilibrium would also be expected to exist between oocytes meiotic spindles and its subunits.

The reduction in the size of the viable spindle subunit pool due to ionizing radiation has been studied in greater detail (Zaremba and Irwin, 1981; Coss, Bamburg, and Dewey, 1981) and the findings revealed two free sulfhydryl groups are lost from the 6S subunit as they form a disulfide bridge. The formation of the disulfide bridge causes a conformational change that lowers the affinity of the subunit for GTP. This covalent modification removes these subunits from the pool of subunits available for spindle formation, and shifts the stoichiometry in favour of spindle depolymerization. Based on these findings the 222 Gy of ionizing radiation probably acted in a similar manner, by depleting the pool of available subunits through covalent modification, which triggered depolymerization of.....

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