Assessment of Proliferation and Apoptosis in Testes of Rats after Experimental Localized Electron Irradiation

Abstract

Background and purpose: With the emergence of linear accelerators in radiotherapy, it becomes necessary to accurately select new dosing regimens. The purpose of this study was to assess the morphological changes of spermatogenesis after radiation exposure.

Materials and methods: Male Wistar rats (n = 40) were subjected to targeted ionizing radiation on a pulsed electron accelerator "NOVAC-11" with doses of 2, 8 and 12 Gy. Spermatogenesis was assessed a week later using light microscopy and immunohistochemical method (antibodies to Ki-67, Bcl-2, p53, Caspase 3).

Results: A decrease in the number of normal germ cells was seen in all experimental groups, while radioresistant Sertoli and Leydig cells were barely affected. The most serious damage to the tubules and germ cells was observed in 8 and 12 Gy irradiation groups. IHC analysis of testes after irradiation showed a shift in the proliferative-apoptotic balance toward apoptosis of germ cells: a decrease in the expression levels of Ki-67 and Bcl-2, an increase in p53-positive and caspase 3-positive cells by the end of the experiment.

Conclusion: Dose-dependent progressive pathomorphological changes in histoarchitectonics of the testes are traced, and a decrease in the number of germ cells is seen on the seventh day after irradiation with a pulsed electron accelerator "NOVAC-11".

Keywords: electron irradiation; male infertility; seminal balls; spermatogenesis.

Figure 1

Testes of rats of experimental groups at different radiation doses; stain—hematoxylin and eosin, magn. ×200 (■—the proportion of damaged seminiferous tubules (area—28,079.84 μm²; diameter—189.083 μm; p < 0.01) with disorganization of the epithelium and loss of cell polarity accounted for up to 1/8 of the testis (1–2 transverse sections of tubules in sight); ●—1/3 of the seminiferous tubules showed the appearance of highly degenerated spermatids and spermatocytes combined into seminal balls (area—0.7 µm²; diameter—0.9 µm; p < 0.01). The area of single primary spermatocytes was 0.8 µm² at p < 0.01, and the diameter was 0.9 µm at p < 0.01, which is 1.1 times more than normal; ▲—the largest vacuoles were found in the seminiferous tubules of the irradiation group with a dose of 12 Gy. The number of seminal balls doubled, and their size was 5.0–10.0 times larger than the area of the primary spermatocyte. The dimensions of the aplastic seminiferous tubules are significantly smaller (area—14,123.72 µm²; diameter—76.142 µm; height of the spermatogenic epithelium—14.2 ± 12.1 at p < 0.01; an increase in the number of pathological mitoses) than in the previous groups, and therefore, an increase in the interstitial component was observed testes with edema. Damage to the seminiferous tubules was 3/4).

Figure 2

Testes of rats of experimental groups at different radiation doses, IHC staining, magn. ×200.