Ameliorative Effects of Vitamins A, C, and E on Sperm Parameters, Testis Histopathology, and Oxidative Stress Status in Zinc Oxide Nanoparticle-Treated Rats

Abstract

One of the most often utilized nanoparticles (NPs) in several technologies is zinc oxide (ZnO) NPs. However, these NPs are said to have harmful effects on the reproductive system. Thus, we designed this study to specify the potential preventive activity of vitamins (Vits) A, C, and E, as antioxidants, against toxicity of ZnO NPs in the testes of rats. A total of 54 Wistar rats were arranged in 9 groups of 6 and then orally received water (control 1), olive oil (control 2), Vit A (1000 IU/kg), Vit C (200 mg/kg), Vit E (100 IU/kg), ZnO (200 mg/kg), ZnO+Vit A, ZnO+Vit C, and ZnO+Vit E. To determine the amount of testicular injury, sperm analysis and histological evaluation were performed. In addition, oxidative stress status was examined using colorimetric and qRT-PCR methods. Our findings suggest that ZnO NPs cause adverse effects on sperm parameters and testicular histology. Furthermore, oxidative biomarkers (malondialdehyde and total oxidant capacity) were enhanced in the ZnO group. By contrast, the gene expression and activities of antioxidant enzymes (SOD, GPx, and CAT) noted a remarkable decrease in the ZnO group regarding control (p < 0.05). However, oxidative markers were remarkably mitigated after combined treatment of ZnO NPs and Vits A, C, or E compared to the rats given ZnO NPs (p < 0.05). Additionally, compared to the ZnO NP group, the rats receiving Vits+ZnO NPs exhibit increased antioxidant enzyme activity and mRNA expression (p < 0.05). The findings demonstrate the abovementioned Vits' ameliorative effects on toxicity incurred by ZnO NPs.

Figure 1

Micrographs of testicular tissue from control and treated rats. (a) Control 1 (Bi-distilled water), (b) control 2 (olive oil), (c) Vit A, (d) Vit C, and (e) Vit E show normal histology of the organ showing regular interstitial connective tissue (large arrow) and seminiferous tubules (asterisk) with typical arrangement of spermatogenic cells in various stages of maturation (ellipse). (f) ZnO NP-treated rats (arrowheads, large arrow and small arrow show atrophy of tubular epithelium, sloughing of Sertoli cells, and intratubular empty spaces, respectively.) (g), (h) and (i), respectively, indicate Vit A+ZnO, Vit C+ZnO, and Vit E+ZnO-treated rats so that the disorganized germinal epithelium of seminiferous tubules (arrow) has partly been improved (×100 magnification (inset ×1000), hematoxylin and eosin).

Figure 2

(a) Effects of zinc oxide nanoparticle (ZnO NP) exposure and administration of vitamins A, C, and E on the area of seminiferous tubules (μm²). (b) The amount of total protein in tissue lysates is adopted from the Bradford assay. ^∗p < 0.05 shows significant differences compared to the control group. Control 1: distilled water; control 2: olive oil; Vit: vitamin.

Figure 3

Effects of zinc oxide nanoparticle (ZnO NP) exposure and administration of vitamins A, C, and E on oxidative stress biomarkers in the testis of Wistar rats. (a) Malondialdehyde (MDA), (b) total oxidant status (TOS), (c) total antioxidant capacity (TAC), and (d) oxidative stress index (OSI). The results are expressed as mean ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001 show significant differences compared to the control group, and ^#p < 0.05 and ^###p < 0.001 remarkably altered from the ZnO NP group. Control 1: distilled water; control 2: olive oil; Vit: vitamin.

Figure 4

Impacts of zinc oxide nanoparticles (ZnO NPs) and vitamin A, C, and E treatment on antioxidant enzyme activities in the testis of Wistar rats. (a) Glutathione peroxidase (GPx), (b) superoxide dismutase (SOD), and (c) catalase (CAT). The results are expressed as mean ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001 show significant differences compared to the control groups, and ^#p < 0.05 and ^##p < 0.01 significantly changed from the ZnO NP-treated group. Control 1: distilled water; control 2: olive oil; Vit: vitamin.

Figure 5

Changes in the mRNA expression of antioxidant-related genes including (a) glutathione peroxidase (GPx), (b) superoxide dismutase (SOD), and (c) catalase (CAT) in the testis of Wistar rats, obtained from reverse transcription-polymerase chain reaction (qRT-PCR) analysis. Method of 2^−ΔΔCt was used to analyze the data in which the β-actin gene expression was selected as an endogenous control. The results are reported as fold change compared to the control and expressed as mean ± SEM. ^∗p < 0.05 and ^∗∗∗p < 0.001 show significant differences compared to the control group, and ^#p < 0.05, ^##p < 0.01, and ^###p < 0.001 indicate meaningful differences relative to the ZnO NP-treated group. Control 1: distilled water; control 2: olive oil; Vit: vitamin.