GROWING evidence suggests that nicotine, the addictive component of cigarettes, plays a direct role in testicular injury and infertility. The present study was intended to investigate the therapeutic effect of epigallocatechin-3-gallate (EGCG) and rutin (RUT), either alone or in combination with a low-dose radiation (LDR), against testicular injury evoked by nicotine in rats. For the induction of testicular injury, nicotine (1mg/kg) was administered orally for 30 days. Following that, rats were administered EGCG (100 mg/kg), RUT (30 mg/kg) orally, either alone or in combination with LDR (2 x 0.25 Gy) for an additional 14 days. Rats were sacrificed on day 45, the testes were then dissected for histopathological analysis, and several biochemical parameters in serum and testicular tissue were also evaluated. The results showed that nicotine administration significantly increased the testicular thiobarbituric acid reactive substances and decreased the reduced glutathione contents. Besides, the activities of testicular androgenic enzymes (3 beta-hydroxysteroid dehydrogenases and 17 beta-hydroxysteroid dehydrogenases) were reduced, whereas serum lactate dehydrogenase activity was considerably raised. In addition, the follicle-stimulating hormone, luteinizing hormone, and testosterone serum levels were reduced, indicating hormonal alterations. The testicular seminiferous tubules structure was also deformed after histological examination. On the other hand, treatment with LDR combined with either EGCG or RUT dramatically reduced the deleterious effects of nicotine compared to their individual effects, as evidenced by biochemical and histological findings. Accordingly, exposure to LDR combined with natural antioxidants, either EGCG or RUT, may be a promising candidate for treating testicular injury caused by nicotine.