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Radioprotective potential of melatonin against ⁶⁰Co γ-ray-induced testicular injury in male C57BL/6 mice.

Khan S, Adhikari JS, Rizvi MA, Chaudhury NK - J. Biomed. Sci. (2015)

Bottom Line: In addition, radiation-induced sperm abnormalities, motility and viability in cauda-epididymes were significantly reduced by melatonin.The detailed study will benefit in understanding the role of melatonin in modulation of radiation-induced ATM-dependent p53-mediated pro-vs.-anti apoptotic proteins in testicular injury.These results can be further exploited for use of melatonin for protection of male reproductive system in radiotherapy applications involving hemibody abdominal exposures.

View Article: PubMed Central - PubMed

Affiliation: Chemical Radioprotector and Radiation Dosimetry Research Group, Division of Radiation Biosciences, Institute of Nuclear Medicine and Allied Sciences, Defence Research & Development Organization, Brig. S. K. Mazumdar Road, New Delhi, Delhi, 110054, India. shahanshah88@hotmail.com.

ABSTRACT

Background: Melatonin, the chief secretary product of pineal gland, is a strong free radical scavenger and antioxidant molecule. The radioprotective efficacy and underlying mechanisms refer to its antioxidant role in somatic cells. The purpose of this study, therefore, was to investigate the prophylactic implications of melatonin against γ-ray-induced injury in germinal cells (testes). C57BL/6 male mice were administered melatonin (100 mg/kg) intra-peritoneally 30 min prior to a single dose of whole-body γ-irradiation (5 Gy, 1 Gy/minute) using (60)Co teletherapy unit. Animals were sacrificed at 2h, 4h and 8h post-irradiation and their testes along with its spermatozoa taken out and used for total antioxidant capacity (TAC), lipid peroxidation, comet assay, western blotting and sperm motility and viability. In another set of experiment, animals were similarly treated were sacrificed on 1(st), 3(rd), 7(th), 15(th) and 30(th) day post-irradiation and evaluated for sperm abnormalities and histopathological analysis.

Results: Whole-body γ-radiation exposure (5 Gy) drastically depleted the populations of spermatogenic cells in seminiferous tubules on day three, which were significantly protected by melatonin. In addition, radiation-induced sperm abnormalities, motility and viability in cauda-epididymes were significantly reduced by melatonin. Melatonin pre-treatment significantly inhibited radiation-induced DNA strands breaks and lipid peroxidation. At this time, radiation-induces activation of ATM-dependent p53 apoptotic proteins-ATM, p53, p21, Bax, cytochrome C, active caspase-3 and caspases-9 expression, which were significantly reversed in melatonin pre-treated mice. This reduced apoptotic proteins by melatonin pre-treatment was associated with the increase of anti-apoptotic-Bcl-x and DNA repair-PCNA proteins in irradiated mice. Further, radiation-induced decline in the TAC was significantly reversed in melatonin pre-treated mice.

Conclusions: The present results indicated that melatonin as prophylactic agent protected male reproductive system against radiation-induced injury in mice. The detailed study will benefit in understanding the role of melatonin in modulation of radiation-induced ATM-dependent p53-mediated pro-vs.-anti apoptotic proteins in testicular injury. These results can be further exploited for use of melatonin for protection of male reproductive system in radiotherapy applications involving hemibody abdominal exposures.

No MeSH data available.


Related in: MedlinePlus

Effect of melatonin pre-treatment on spermatogenic cell in mice exposed to whole-body 60Co γ-irradiation. Animals were sacrificed through cervical dislocation and testes were collected on 1st, 3rd, 7th, 15th and 30th days post-irradiation. After fixation and processing, cross sections of testes (5 μm) were stained with H & E and spermatogenic cells were analyzed and represented. *p < 0.001
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Fig2: Effect of melatonin pre-treatment on spermatogenic cell in mice exposed to whole-body 60Co γ-irradiation. Animals were sacrificed through cervical dislocation and testes were collected on 1st, 3rd, 7th, 15th and 30th days post-irradiation. After fixation and processing, cross sections of testes (5 μm) were stained with H & E and spermatogenic cells were analyzed and represented. *p < 0.001

Mentions: Radiation cause morphological changes in the testes histological architecture mainly due to the killing of spermatogonial cells [42], therefore, counting of spermatogonia is considered as gold standard to measure radiation-induced effects in testes. Spermatogenic cells were counted in seminiferous tubules to assess the γ-ray-induced testicular injury in mouse. A statistically significant (p < 0.001) reduction in spermatogenic cells (excluding sertoli cell) was observed on 3rd day post-irradiation in irradiated mice (Fig. 2). Melatonin pre-treatment increased (p < 0.001) spermatogenic cells on 7th day in comparison to radiation alone treated mice (Fig. 2). Sertoli cells appeared to be more radio resistant, therefore, did not show changes (p > 0.05) till 30th day of observation (Additional file 1: Figure S1). Thus, the sertoli cells were considered as reference standard cells, only if they had nucleolus in the plane of section. The total number of spermatogonia was divided by total number of sertoli cells and expressed as ratio of spermatogonia/sertoli cells. The results showed that more number of spermatogonia per sertoli cell was present in melatonin pre-treated mice (p < 0.001) on 7th day following irradiation (Fig. 2d). On the other hand, irradiation resulted a significant decrease in the number of spermatogonia per sertoli cell in irradiated mice (p < 0.001) as observed on 3rd day (Fig. 2d). The present results suggest that a single prophylactic dose of melatonin recover spermatogenic cells in irradiated testes of mice as a function of post-irradiation days. Melatonin treatment alone did not show any change in the histological artchitechture and spermatogenic population of testes till 30th day of observation (Figs. 1 and 2).Fig. 2


Radioprotective potential of melatonin against ⁶⁰Co γ-ray-induced testicular injury in male C57BL/6 mice.

Khan S, Adhikari JS, Rizvi MA, Chaudhury NK - J. Biomed. Sci. (2015)

Effect of melatonin pre-treatment on spermatogenic cell in mice exposed to whole-body 60Co γ-irradiation. Animals were sacrificed through cervical dislocation and testes were collected on 1st, 3rd, 7th, 15th and 30th days post-irradiation. After fixation and processing, cross sections of testes (5 μm) were stained with H & E and spermatogenic cells were analyzed and represented. *p < 0.001
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4514449&req=5

Fig2: Effect of melatonin pre-treatment on spermatogenic cell in mice exposed to whole-body 60Co γ-irradiation. Animals were sacrificed through cervical dislocation and testes were collected on 1st, 3rd, 7th, 15th and 30th days post-irradiation. After fixation and processing, cross sections of testes (5 μm) were stained with H & E and spermatogenic cells were analyzed and represented. *p < 0.001
Mentions: Radiation cause morphological changes in the testes histological architecture mainly due to the killing of spermatogonial cells [42], therefore, counting of spermatogonia is considered as gold standard to measure radiation-induced effects in testes. Spermatogenic cells were counted in seminiferous tubules to assess the γ-ray-induced testicular injury in mouse. A statistically significant (p < 0.001) reduction in spermatogenic cells (excluding sertoli cell) was observed on 3rd day post-irradiation in irradiated mice (Fig. 2). Melatonin pre-treatment increased (p < 0.001) spermatogenic cells on 7th day in comparison to radiation alone treated mice (Fig. 2). Sertoli cells appeared to be more radio resistant, therefore, did not show changes (p > 0.05) till 30th day of observation (Additional file 1: Figure S1). Thus, the sertoli cells were considered as reference standard cells, only if they had nucleolus in the plane of section. The total number of spermatogonia was divided by total number of sertoli cells and expressed as ratio of spermatogonia/sertoli cells. The results showed that more number of spermatogonia per sertoli cell was present in melatonin pre-treated mice (p < 0.001) on 7th day following irradiation (Fig. 2d). On the other hand, irradiation resulted a significant decrease in the number of spermatogonia per sertoli cell in irradiated mice (p < 0.001) as observed on 3rd day (Fig. 2d). The present results suggest that a single prophylactic dose of melatonin recover spermatogenic cells in irradiated testes of mice as a function of post-irradiation days. Melatonin treatment alone did not show any change in the histological artchitechture and spermatogenic population of testes till 30th day of observation (Figs. 1 and 2).Fig. 2

Bottom Line: In addition, radiation-induced sperm abnormalities, motility and viability in cauda-epididymes were significantly reduced by melatonin.The detailed study will benefit in understanding the role of melatonin in modulation of radiation-induced ATM-dependent p53-mediated pro-vs.-anti apoptotic proteins in testicular injury.These results can be further exploited for use of melatonin for protection of male reproductive system in radiotherapy applications involving hemibody abdominal exposures.

View Article: PubMed Central - PubMed

Affiliation: Chemical Radioprotector and Radiation Dosimetry Research Group, Division of Radiation Biosciences, Institute of Nuclear Medicine and Allied Sciences, Defence Research & Development Organization, Brig. S. K. Mazumdar Road, New Delhi, Delhi, 110054, India. shahanshah88@hotmail.com.

ABSTRACT

Background: Melatonin, the chief secretary product of pineal gland, is a strong free radical scavenger and antioxidant molecule. The radioprotective efficacy and underlying mechanisms refer to its antioxidant role in somatic cells. The purpose of this study, therefore, was to investigate the prophylactic implications of melatonin against γ-ray-induced injury in germinal cells (testes). C57BL/6 male mice were administered melatonin (100 mg/kg) intra-peritoneally 30 min prior to a single dose of whole-body γ-irradiation (5 Gy, 1 Gy/minute) using (60)Co teletherapy unit. Animals were sacrificed at 2h, 4h and 8h post-irradiation and their testes along with its spermatozoa taken out and used for total antioxidant capacity (TAC), lipid peroxidation, comet assay, western blotting and sperm motility and viability. In another set of experiment, animals were similarly treated were sacrificed on 1(st), 3(rd), 7(th), 15(th) and 30(th) day post-irradiation and evaluated for sperm abnormalities and histopathological analysis.

Results: Whole-body γ-radiation exposure (5 Gy) drastically depleted the populations of spermatogenic cells in seminiferous tubules on day three, which were significantly protected by melatonin. In addition, radiation-induced sperm abnormalities, motility and viability in cauda-epididymes were significantly reduced by melatonin. Melatonin pre-treatment significantly inhibited radiation-induced DNA strands breaks and lipid peroxidation. At this time, radiation-induces activation of ATM-dependent p53 apoptotic proteins-ATM, p53, p21, Bax, cytochrome C, active caspase-3 and caspases-9 expression, which were significantly reversed in melatonin pre-treated mice. This reduced apoptotic proteins by melatonin pre-treatment was associated with the increase of anti-apoptotic-Bcl-x and DNA repair-PCNA proteins in irradiated mice. Further, radiation-induced decline in the TAC was significantly reversed in melatonin pre-treated mice.

Conclusions: The present results indicated that melatonin as prophylactic agent protected male reproductive system against radiation-induced injury in mice. The detailed study will benefit in understanding the role of melatonin in modulation of radiation-induced ATM-dependent p53-mediated pro-vs.-anti apoptotic proteins in testicular injury. These results can be further exploited for use of melatonin for protection of male reproductive system in radiotherapy applications involving hemibody abdominal exposures.

No MeSH data available.


Related in: MedlinePlus