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Abstract

Bisphenol A (BPA) is a widespread compound associated with the manufacture of many consumer products. The BPA-induced reproductive toxicity was reported to be mainly attributed to oxidative stress. However, the role of antioxidants usage to decrease the injurious effects of BPA, on male reproductive functions, remains to unveil. The present research is established to evaluate the role of selenium (Se) and its nano form (NSe) as protective agents to alleviate BPA-induced testicular toxicity. Ninety mature albino male rats were assigned into six equal groups: negative control; orally BPA 150 mg/kg; Se 3 mg/kg; NSe 2 mg/kg; both BPA 150 mg/kg and Se 3 mg/kg; and BPA 150 mg/kg + NSe 2 mg/kg. The experiment lasted for 70 consecutive days, and then serum was collected for estimation of prostatic acid phosphatase. Testicular tissues were subjected to measurement of antioxidant status, lipid peroxidation, DNA damage, and expression of some apoptotic genes. Our results reported that BPA-induced marked testicular damage evidenced by significant elevations in serum prostatic acid phosphatase activity, malondialdehyde levels, a decrease in testicular catalase activity and reduced glutathione level. Moreover, marked DNA internucleosomal fragmentation pattern as well as upregulation of cyclooxygenase-2 and estrogen receptor-2 NSe genes were detected. Coadministration of Se and NSe attenuated the reproductive toxicity induced by BPA via improvement of the antioxidant activity, genetic changes, and restoration of testicular tissue nearly as control one. These results indicated that both Se and NSe forms could be used as reproductive protective agents against the detrimental effect induced by BPA. However, the NSe surpassed the selenium in modulating the DNA laddering, and the studied gene expression levels, and offered a potent reproductive protection.

Keywords

Bisphenol nano selenium male fertility oxidative stress COX-2 ER-2

Introduction

Nanotechnology has been applied in many areas, its application in diseases diagnosis and treatment attracting much of interest. Nanominerals are minerals that have been manufactured into nanoparticles using nanotechnology. Little research work has been done to evaluate the efficiency of the nanominerals.¹ Nanoparticles exhibit novel properties, such as great specific surface area, high surface activity, a lot of surface active centers, high catalytic efficiency, high absorbed ability, and low toxicity.² Selenium (Se) is an essential trace element, which plays an important role in antioxidant defense systems. Furthermore, it is necessary for fertility, growth, and immunity in humans and animals.³ Sodium selenite is used as the commercial Se source supplement in animal. Lately, nanoparticles of elemental Se (NSe) have attracted considerable attention due to its high biological efficiency. The main reasons for its efficiency are mainly due to its high bioavailability, high surface activity, and low toxicity comparing to routine Se.^4,5 NSe as highly stable and nano-defined size in the redox state of zero has been used as nutritional supplements and in medical therapy applications.⁶ NSe could be used more effectively as dietary supplements comparing to inorganic or organic Se.¹

Bisphenol A (BPA) is an industrial chemical that has endocrine-disrupting properties. It is used as a common ingredient in the production of polycarbonate plastics and in many plastic consumer products. Detectable levels of BPA had been found in commercial food products as it can leach from their plastic containers. About 92.6% of the human population showed elevated BPA levels in their bodies.⁷ Reproductive tissues are highly sensitive biological systems to the exposure to various environmental toxicants.⁸ BPA is among the most distinguished environmental contaminants that associated with impairment of male reproductive function.⁹ BPA has been shown to have adverse male reproductive effects even at low levels in early life or adulthood.¹⁰

BPA is considered as a xenoestrogen because its chemical structure is similar to estradiol and diethylstilbestrol. BPA can bind to the estrogen receptors and triggers its estrogenic effect.¹¹ The mammalian testes produce estrogen that could be implicated in the male reproductive function.¹² Estrogen receptors are considered vital modulators of the testicular function such that, estrogen performs its physiological influences via its receptors, which were distributed throughout male genital system.¹³ In male, estrogen regulates apoptosis and differentiation of germ cells; besides, it controls the motility and transport of spermatozoa. Thus, estrogen receptors are critical for fertility and normal reproduction in male.¹⁴ Acid phosphatases represent a heterogeneous group of enzymes containing many isoenzymes; each specific for one type of tissue, prostate is particularly rich in this enzyme, and serum enzyme levels have been used as a tumor marker of prostate cancer.¹⁵

BPA affects the reproductive organs through multiple pathways, such as modulating hormone synthesis and inducing oxidative stress in the testis and epididymis. Therefore, research related to reproductive toxicity induced by BPA is of scientific significance.¹⁶ The cyclooxygenase-2 (COX-2), an inducible enzyme, contributes to the generation of prostanoids (prostaglandins, prostacyclins, and thromboxanes) from arachidonic acid. Prostaglandin is involved in gonadal somatic cells differentiation through upregulation of the major genes responsible for germ line differentiation.¹⁷ Nevertheless, few reports on the biological effects of COX-2 on Sertoli cells are available.

Male fertility and the process of spermatogenesis are dependent upon an adequate Sertoli cell function, which is directly related to prostaglandin signaling, which in turn controlled by COX-2 gene.¹⁸ Therefore, using antioxidants may be an effective strategy to mitigate the disturbances induced by BPA.⁹ Consequently, this study attempts to compare the protective effect of NSe and Se against BPA-induced testicular toxicity in rat through evaluating their impacts on the DNA fragmentation level and the expression of COX-2 as inflammation associated gene and ER-2 gene as a modulator in spermatogenesis, prostatic acid phosphatase activity, and oxidative stress biomarkers in the exposed rats.

Materials and methods

Animals

Ninety mature male albino rats (weight 150–180 g) were maintained under controlled temperature (25 ± 2 °C), humidity (55 ± 7%), and a 12-h light/12-h dark cycle. Animals acquired free access to water and standard rodent diet throughout the whole experimental period (70 days + 1 week acclimatization). All experimental and euthanasia procedures were carried out according to the protocol approved by (Institutional Animal Care and Use Committee, Beni-Suef University).

Chemicals

BPA and sodium selenite were obtained from Sigma Chemical Co. (St. Louis, Missouri, USA). NSe in the size range of 20–60 nm was prepared as described previously by Zhang et al.² Commercial kits for estimation of prostatic acid phosphatase activity (PAP), catalase (CAT), reduced glutathione (GSH), and lipid peroxidation were obtained from Biodiagnostic Company (Egypt).

Experimental treatment and sample collection

Rats were randomly assigned to six groups (n = 15). Group I (negative control group) orally received corn oil (2 ml/kg/day) as a vehicle for BPA. Group II (BPA group) orally administrated BPA in corn oil only at a dose of 150 mg/kg/day equal to 1/20 median lethal dose.¹⁹ Group III (Se group) was orally given sodium selenite in a dose of 3 mg/kg.²⁰ Group IV (NSe group) orally received a dose of 2 mg/kg.²¹ Group V (BPA + Se group) was administered both treatments together as described in groups II and III. Group VI BPA + NSe group orally received both treatments in the same way in groups II and IV.

After 70 consecutive days of treatment, all the rats were anesthetized by diethyl ether, and blood samples were collected from orbital venous plexus. Sera were obtained by blood centrifugation at 3000 r/min for 10 min at 4 °C. Then rats were euthanized using capitation and the right testes were immediately removed, washed, and perfused with phosphate-buffered saline to be homogenized in the range of 5–10 ml cold buffer then centrifuged at 4000 r/min for 10 min at 4 °C. The supernatant was stored at −20 °C for biochemical analysis. The left testis specimens were stored −80 °C for RNA isolation and DNA fragmentation test.

Estimation of PAP activity

PAP in serum was estimated by the method described by Hillmann.²²

Determination of testicular antioxidant enzymes

The activity of catalase and reduced glutathione content were estimated in testicular homogenate according to Aebi²³ and Beutler et al.,²⁴ respectively.

Assessment of lipid peroxidation

Lipid peroxidation in testicular tissue was estimated by measuring of thiobarbituric acid reactive substance according to Ohkawa et al.²⁵

mRNA expression analysis

Levels of gene transcript were conducted by real-time quantitative polymerase chain reaction (qPCR) method. Total RNA was isolated from testis by GF-1 total RNA Extraction Kit according to the manufacturer’s instructions. Its yield and purity were assessed at 260 and 280 nm, respectively, using Nano drop ND-2000 spectrophotometer (Thermo Electron). A total amount of 1 μg T-RNA was used for complementary DNA synthesis by Viva 2-steps RT-PCR Kit according to the manufacturer’s protocol. Real-time PCR for the COX-2,²⁶ ER-2 (designed using primer-3 software) were performed using Luminaris Color HiGreen Low ROX qPCR Master kit (Thermo Scientific, USA; Cat. No. #K0371) in an iQ5 iCycler thermal cycler (Bio-Rad, German). The assay was done in triplicate. The primer sequences for the genes analyzed are reported in Table 1. The conditions were composed of an initial denaturation step of 5 min at 95°C, followed by 40 cycles of denaturation at 95°C for 15 s and annealing at 60°C for 30 s.

Table 1.

Primers used for real time PCR.

Gene	PCR primers	Gene bank accession number
COX-2 F	5-AGCCTCGTCCAGAT GCTA-3	NM-017232.3
COX-2 R	5-TGGTGGCTGTCTTG GTAGG-3	NM-017232.3
ER-2 F	5-CAAACCAATGCACC ATCGATAA-3	NM_012689.1
ER-2 R	5-TTTTCGTATCCCGC CTTTCA-3	NM_012689.1
GAPDH F	5-ACCACAGTCCATG CCATCAC-3	NM_017008.4
GAPDH R	5-TCCACCACCCTGTT G CTGTA-3	NM_017008.4

The relative expression values of the target genes were normalized to values obtained for GAPDH (internal control) expression and evaluated using CT, ΔCT, and ΔΔCT by Mxpro qPCR Mx3000P software (Stratgene).²⁷

DNA fragmentation assay

Internucleosomal DNA fragmentation is a discrete evidence of apoptotic cell death. It can be detected as a DNA laddering pattern using agarose gel electrophoresis. The DNA fragmentation assay was conducted using diphenylamine (DPA) method according to Ibrahim et al.²⁸

Statistical analysis

Data obtained from our study were expressed as mean ± SE of 15 rats per group, and the statistical significances were evaluated by one-way analysis of variance using SPSS version 21 software package (SPSS, Inc, Chicago, Illinois, USA), in which the difference between any two groups were evaluated using Duncan’s test, and a p < 0.05 was considered as statistically significant.

Results

PAP activity

The present results revealed that male rats treated with BPA showed an increase (p < 0.05) in the level of serum PAP activity when compared to that of the negative control group (Table 2 and Figure 1(a)). Coadministration of BPA with either Se or NSe induced a significant decrease in PAP activity compared to BPA group and the activity was nearly similar to the negative control group.

Table 2.

Prostatic acid phosphatase activity and testicular oxidative stress parameters and DNA fragmentation.

	PAP (U/ul)	GSH (mg/g)	CAT (U/g)	LPO (nmol/g)	DNA fragmentation percentage
Control	0.691 ± 0.00^a	6.8 ± 0.47^a,b	0.56 ± 0.05^a,b,c,d	29.53 ± 0.84^a,c,d	18.66 ± 1.47^a,c,d
BPA	1.03 ± 0.01^e,f,b,c,d	4.37 ± 0.34^e,f,b,c,d	0.19 ± 0.01^e,f,b,c,d	37.63 ± 0.71^e,f,b,c,d	43.00 ± 0.70^e,f,b,c,d
Se	0.67 ± 0.02^a	7.05 ± 0.55^a	0.58 ± 0.06^a,c,d	30.70 ± 0.8^a,b	20.65 ± 0.66^a,c
NSe	0.65 ± 0.0^a	8.19 ± 0.56^e,a,c,d	0.62 ± 0.04^e,a,c,d	26.54 ± 0. 5^a,f,c,d	21.3 ± 0.44^a,c
BPA + Se	0.68 ± 0.01^a	5.96 ± 0.51^a,f,b	0.33 ± 0.03^e,a,f,b,d	33.27 ± 0.17^e,a,b	30.49 ± 1.37^e,a,f,b,d,
BPA + NSe	0.74 ± 0.00^a	6.16 ± 0.34^a,b	0.46 ± 0.03^e,a,f,b,c	33.30 ± 0.8^e,a,bd	25.70 ± 0.83^e,a,c

PAP: prostatic acid phosphatase activity; CAT: catalase GSH: glutathione; Se: selenium; BPA: bisphenol A.

Values are presented as mean ± SE (n = 15 rats/group).

^a Significantly different from BPA group at p < 0.05.

^b Significantly different from NSe group at p < 0.05.

^c Significantly different from BPA + Se at p < 0.05.

^d Significantly different from BPA + NSe group at p < 0.05.

^e Significantly different from the corresponding control negative group at p < 0.05.

^f Significantly different from Se group at p < 0.05 compared with NSe.

Figure 1.

The effects of Se and NSe on BPA-intoxicated rats. (a) Result of PAP; (b) results of CAT activity; (c) the GSH contents; and (d) the lipid peroxidation levels. Se: selenium; BPA: bisphenol A; GSH: glutathione; PAP: prostatic acid phosphatase activity; CAT: catalase. ^aSignificantly different from BPA group at p < 0.05. ^bSignificantly different from NSe group at p < 0.05. ^cSignificantly different from BPA + Se at p < 0.05. ^dSignificantly different from BPA + NSe group at p < 0.05. ^eSignificantly different from the corresponding control negative group at p < 0.05. ^fSignificantly different from Se group at p < 0.05 compared with NSe.

Activity of CAT and GSH level in rat’s testicular tissue

The current data revealed that exposure of male rats to BPA induced a significant decrease (p < 0.05) in CAT activity and GSH level in comparison to the negative control group (Table 2 and Figure 1(b) and (c)). Coadministration of either Se or NSe to BPA-treated groups significantly reversed the values of CAT and GSH toward the control levels. Treatment of male rats with NSe showed a significant increase in CAT and GSH values compared to the negative control group.

Lipid peroxidation

The present data showed a significant increase (p < 0.05) in malondialdehyde (MDA) level in testicular tissue of BPA-exposed rats. However, coadministration of Se or NSe with BPA caused a significant reduction in lipid peroxidation level when compared to the BPA-treated group. However, such reduction in the co-treated groups was still higher (p = 0.05) than the control group (Table 2 and Figure 1(d)).

Expression levels of COX-2 and ER-2 in testes

The level of mRNA of COX-2 showed sixfold increase in the BPA-treated group compared to the control group, whereas both the ionized Se and NSe groups showed down expression by 4- and 2.3-folds, respectively, compared to the BPA-treated group. In the same time, the ER-2 showed a significant increase in its expression in the BPA-treated group by 7.7-fold, whereas coadministrated ionized Se or NSe caused down expression of ER-2 by 3- and 1.8-folds compared to the control group (Table 3 and Figure 2).

Table 3.

The relative m-RNA expression level of COX-2 and ER-2 in testes.^a

Groups	Relative m-RNA expression
Groups	COX-2	ER-2
Control negative	1^bef	1^bef
Bisphenol	6^acdef	7.7^acdef
Se	1.1^bef	1^be
NSe	1^bef	0.9^bf
BAP + Se	4^abcdf	3^abcd
BAP + NSe	2.3^abcde	1.8^adb

Se: selenium; BPA: bisphenol A.

^aSignificantly different from BPA group at p < 0.05.

^bSignificantly different from NSe group at p < 0.05.

^cSignificantly different from BPA + Se at p < 0.05.

^dSignificantly different from BPA + NSe group at p < 0.05.

^eSignificantly different from the corresponding control negative group at p < 0.05.

^fSignificantly different from Se group at p < 0.05 compared with NSe.

A value of 1.0 was used for the control (calibrator).

Figure 2.

The relative mRNA expression level of COX-2 and ER-2 genes; (a) fold change of mRNA expression of COX-2 and (b) ER-2. Values are presented as mean ± SE. (n = 15 rats/group).

DNA fragmentation assay

DNA laddering is used as the basic feature for apoptosis. DNA is degraded by caspase-activated DNase at inter-nucleosomal linkers into multiple fragments of about 180-bp fragments. The results of DNA fragmentation assay showed a significant and marked increase in the DNA laddering among the bisphenol-treated group. NSe was proved to be a potent protective agent as it decreased the fragmentation percentage in bisphenol + NSe-treated group compared to other groups (Table 2 and Figure 3).

Figure 3.

(a) Electrophoretic mobility of fragmented DNA on 2% agarose gel. Lane M: 100 bp DNA ladder; lanes 1–2: control group; lane 3: NSe group; lane 4: Se group; lanes 5 and 6: BAP group; lane 7: BAP +NSe group; lane 8: BAP+ Se group. (b) Internucleosomal DNA fragmentation percentage among different groups. Se: selenium; BPA: bisphenol A.

Discussion

The xenoestrogenic compound BPA has injurious impacts on the reproductive system of both males and females.²⁹ The precise mechanisms by which BPA triggers reproductive influences remain inadequately understood. Nonetheless, it has been reported to produce reactive oxygen species (ROS) that alter genes expression and impair gametogenesis in males.³⁰ The current study was designed to evaluate the effects of BPA exposure on testicular tissue. New insights were gained into the potential roles of both Se and its form against BPA as well as underlying whether NSe or Se was more effective. Our findings indicated that BPA can affect the oxidative stress state in the testis. The testes are considered highly sensitive to oxidative stress due to the presence of abundant polyunsaturated fatty acids. To counteract the effects of ROS, the testes contain a high concentration of antioxidant defense systems that prevent cellular damage.³¹ BPA caused toxic effects to the testicular tissue of the exposed rats by decreasing the activity of CAT and GSH level and elevating the MDA level. BPA was previously reported to induce oxidative damage in several tissues.³² Sperm plasma membrane is extremely susceptible to lipid peroxidation due to the presence of high concentration of polyunsaturated fatty acids (PUFAs); those PUFAs give to the membrane a high level of fluidity and elasticity necessary for sperm motility and their fusion with oocytes. Lipid peroxidation can lead to loss of membrane fluidity and integrity and thus to reducing of sperm–oocyte fusion ability.³³ Excessive ROS triggers the apoptosis pathways and even further induces cell death. Thus, oxidative stress may be the main reason for multiple toxicities.³⁴ The decrease in the activity of catalase enzyme may be attributed to the exhaustion of the enzyme in trials to eliminate the hydrogen peroxide (H₂O₂) generated after BPA treatment. Also, this may be due to enzyme inactivation caused by increased level of ROS production in mitochondria and microsomes.²⁹ The current result revealed a significant depletion in GSH levels after BPA treatment. It may be revealed that GSH, in this study, is exhausted during the conversion of H₂O₂ into hydrogen oxide. The peroxide is readily converted to the hydroxyl radical, which may be involved in the observed decrease in GSH levels as GSH itself is also a general hydroxyl-radical scavenger and its depletion increases the sensitivity of cells to various injuries. This observation is supported by various studies demonstrating that GSH is reduced in the tissues by oxidative stress.³⁵ Our results came in harmony with those reported by Kabuto et al.³⁶ who found that intraperitoneal administration of 50 mg/kg body weight of BPA decreased the activity of GSH and catalase in mouse tissue. These findings were also correlated with the previous studies.^37
–39

The significant increase in lipid peroxidation level in BPA-exposed rats reflected the state of oxidative stress in testicular tissue. This action may be the mechanism by which BPA induced testicular toxicity in rat and mice.⁴⁰ Oxidative damage might be induced by free-radical generation via metabolic redox cycling between quinine and hydroquinone forms of BPA.⁴¹ Previous studies revealed that BPA produced ROS, as superoxide anions, hydroxyl radicals, and H₂O₂, and that it induced oxidative stress.⁴² MDA, the product of aldehydic lipid peroxidation generated by the action of ROS on membrane lipid, is one of the reactive and mutagenic aldehydic lipid peroxidation products in seminal plasma and can be considered as a diagnostic tool in male infertility.⁴³ The present study showed a significant increase in lipid peroxidation in BPA-administered rats that agreed with previous studies.^44,45 Other investigations showed that BPA injection resulted in elevated lipid peroxidation levels in the brain, liver, kidney, and testes of treated rats.⁴¹ The current study showed a significant increase in serum PAP in BPA-treated groups that were in agreement with Jorge et al.⁴⁶ Alteration in PAP expression observed in BPA-exposed groups could be mediated by either a direct effect of BPA on the columnar epithelial cells or through an indirect consequence of primary events occurring in the stroma.⁴⁷ Se and NSe co-treatment to the BPA-intoxicated rats on the following biochemical markers: (a) the PAP level and (b) the level of CAT activity provided adequate protection through inhibition of ROS overproduction in testis.⁴⁸ Numerous studies on Se and its nano form have shown its powerful protective effects against oxidative stress, DNA damage, and apoptosis in response to BPA⁴⁹ cisplatin and ionizing radiation⁵⁰ exposure using in vivo models. Se played important roles in the enhancement of the antioxidant defense system and scavenge free radicals.⁴⁸ Oxidative stress might prevent the RNA polymerase transcribing the DNA and thus inhibit the synthesis of mRNA. A failure in mRNA formation can bring about inhibition of protein synthesis which may cause tissue damage.³¹ In the current study, qRT-PCR showed significant increase in the levels of testicular COX-2 and ER-2 among the BPA-treated group. BPA might bind to estrogen receptors and modulate gene transcription, which corresponds to the classic genomic mechanism of steroid action. The upregulation of the ER-2 receptor in the BPA-exposed group could promote apoptosis because selective activation of estrogen receptor promotes apoptosis of germ cells. BPA is a potential estrogen receptor agonist⁵¹ that modifies hormonal signal as a result of binding to the estrogen receptors on male germ cells. Accordingly, BPA provoked devastating effects as oligospermia,⁵² altered sperm protein,⁵³ abnormal development in spermatozoa, which may be attributed to DNA damage, oxidative stress, and disruption of spermatozoa development during spermatogenesis.⁵⁴ BPA induced oxidative stress and disrupted the estrogenic functions in rats.⁵⁵ Cyclooxygenase catalyzes the rate-limiting step in the production of eicosanoids; however, COX2 is believed to produce mainly thromboxanes and prostaglandins (PGs). In addition to their involvement in the control of testicular activity under pathological disorders and aging, COX2 might play a critical role in fertility. A study on mice lacking COX2 expression revealed severe fertility deficits represented by subfertility, with impaired oocyte release, oocyte nuclear maturation, and fertilization suggesting a link between COX2 activity and these essential reproductive processes in female.⁵⁶ However, in male, it might play a role in the regulation of testicular function mainly in cancer, aging, and fertility disorders.⁵⁷ It is expressed in Leydig cells and macrophages in testes of men suffering from idiopathic infertility.⁵⁸ Our results revealed marked upregulation in COX-2 gene in the BPA-intoxicated group. In this context, COX2 has been found to be upregulated of testicular cancer and fibrosis.⁵⁹ COX2 has also been associated with the age-related decline in testosterone biosynthesis.⁵⁷ Overexpression of COX-2 might be contributed to the oxidative stress mediated damage by ROS production. A persistent overexpression of COX-2 initiates degenerative disorders.⁶⁰ Our results proved that Se and its nano form greatly modulated the mRNA levels of both genes under study. Yet the NSe was capable of bringing the COX-2 gene back to the control level.

Several studies have indicated that mechanism of BPA-induced toxicity may be via the induction of oxidative stress and the depletion of antioxidant enzymes.⁶¹ Previous investigations suggested that BPA can induce oxidative stress, which could in turn be one of the mechanisms underlying DNA damage in germ cells. DNA fragmentation into nucleosomal fragments is the most distinctive biochemical mark of apoptosis. In the current study, exposure of male rats to BPA-induced marked DNA damage in testicular tissue in comparison to that of negative control one. These findings were in agreement with other studies which showed that DNA damage in MCF7 cells was after BPA exposure.⁶² Also, our results were correlated with findings obtained in a comet assay showing that BPA induced DNA migration in the inner organs of rats and mice.⁶³ It has been proposed that BPA may either affect DNA synthesis in germinal cells or interfere with the normal replacement of nuclear histones in germ cells for transition.⁶⁴ The DNA laddering in the groups administered either Se or NSe, besides BPA showed a significant decrease to a great extent. However, the NSe co-treatment conquered the Se in protection against BPA.

Conclusion

In conclusion, the protective effects of both Se and its nano form were assessed for the first time against the reproductive toxicity induced by BPA in male rats. Our findings obviously demonstrated that they attenuated the development of testicular toxicity through inhibition of the oxidative stress, which is believed to be the main mechanism by which BPA produces reproductive toxicity. They also upregulated the COX-2 and ER-2 genes and reduced the DNA laddering percentage compared to BPA-treated rats. Moreover, it is worth mentioning that NSe potentially surpassed the elemental Se in providing protection against the deleterious effect of BPA.

Footnotes

Acknowledgments

Sincere gratitude is to the members of the Department of Nanotechnology, Faculty of Science, Beni-Suef University, for their kind supervision and their helpfulness during conducting this study.

Authors’ contributions

AAK designed the the study; WM, BAH, and WM evaluated the biochemical parameters; And MI performed the molecular genetics parameters. All authors read and approved the final manuscript.

Declaration of Conflicting Interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The author(s) received no financial support for the research, authorship, and/or publication of this article.

ORCID iD

WA Moselhy

MA Ibrahim

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Protective effects of selenium and nano-selenium on bisphenol-induced reproductive toxicity in male rats

Abstract

Keywords

Introduction

Materials and methods

Animals

Chemicals

Experimental treatment and sample collection

Estimation of PAP activity

Determination of testicular antioxidant enzymes

Assessment of lipid peroxidation

mRNA expression analysis

DNA fragmentation assay

Statistical analysis

Results

PAP activity

Activity of CAT and GSH level in rat’s testicular tissue

Lipid peroxidation

Expression levels of COX-2 and ER-2 in testes

DNA fragmentation assay

Discussion

Conclusion

Footnotes

Acknowledgments

Authors’ contributions

Declaration of Conflicting Interests

Funding

ORCID iD

References