Skip to main content
NCBI home page
JBRA Assisted Reproduction logoLink to JBRA Assisted Reproduction
. 2024 Jul-Sep;28(3):471–482. doi: 10.5935/1518-0557.20240033

Omega 3 fatty acids preserve testicular function by ameliorating BPF-induced dysthyroidism: role of p53/Bcl-2 signaling and proton pump activities

Adeyemi Fatai Odetayo 1,2,✉, Luqman Aribidesi Olayaki 1
PMCID: PMC11349269  PMID: 38801312

Abstract

Objective

Bisphenol F (BPF) is an endocrinedisrupting chemical, but information about its effect on thyroid hormones has not been fully explored. Omega 3 fatty acids (O3FA), on the other hand, are antioxidant and antiapoptotic agents. Therefore, this study explored the role and associated molecular mechanism of O3FA in BPF-induced hypothyroidism-mediated testicular dysfunction in male Wistar rats.

Methods

Twenty (20) male Wistar rats were randomized into four groups (n=5/group), namely: the control group; the BPF treated group (30 mg/kg of BPF); and the intervention groups (30mg/kg BPF + 100mg/kg O3FA (BPF+O3FA-L) and 30mg/kg BPF + 300mg/kg of O3FA for 28 days).

Results

Low and high doses of O3FA ameliorated BPF-induced hypothyroidism-mediated reduction in sperm quality, testosterone, luteinizing hormone, follicle-stimulating hormone, catalase, superoxide dismutase, total antioxidant capacity, and nuclear factor erythroid 2-related factor 2 and increases in estrogen, malondialdehyde, c-reactive protein, interleukin 1 beta, caspase 3. Furthermore, O3FA prevented BPF-induced Na+/K+−ATPase and Ca2+−ATPase dysfunction, estrogen receptor beta overexpression, and tumor protein P53 (p53)/ b-cell lymphoma 2 (Bcl-2) imbalance.

Conclusions

This study showed that O3FA ameliorated BPF-induced dysthyroidism-mediated testicular dysfunction by preventing proton pump dysfunction and p53/BCl-2 imbalance.

Keywords: apoptosis, bisphenol F, hypothyroidism, omega 3 fatty acid, p53/BCl-2 signaling, proton pump

INTRODUCTION

Bisphenol A (BPA) and its analogs, such as bisphenol F (BPF), are raw materials widely used in the food, pharmaceutical, chemical, and canning industries. BPA is the most widely used bisphenol, which has been implicated in various human diseases, such as metabolic syndrome (Caporossi & Papaleo, 2017). Thus, many countries have restricted BPA usage (Fatai & Aribidesi, 2022). Alternatively, BPF, considered the major replacement for BPA, has been introduced in the industries.

With the increasing usage, BPF occurrence in the environment is rising. In addition to its presence in food and medical devices, BPF has also been found in environmental media, such as water, dust, thermal paper, and water (Yuan et al., 2019). Despite the increasing usage of BPF, its gonadotoxic effect remains underexplored. Previous findings have shown that BPF impaired testicular function by disrupting the hypothalamic-pituitary-gonadal (HPG)-axis (Fatai & Aribidesi, 2022), redox balance (Ullah et al., 2019; Odetayo & Olayaki, 2022), and apoptotic markers activities (Ferreira et al., 2022; Odetayo et al., 2023a). Despite these findings, the effect of BPF on the thyroid gland has not been fully explored, nor has it been established whether BPF-induced testicular dysfunction is associated by with dysthyroidism.

Thyroid hormone (TH) has important roles in growth, oxygen consumption regulation, mitochondrial energy metabolism, and other biological processes (Meng et al., 2016). TH has also been described as one of the major hormones responsible for testicular maturation and growth (Sahoo et al., 2008), with dysthyroidism linked with testicular dysfunction (El-Kashlan et al., 2015). The presence of thyroid receptors (TRs) and thyroid hormone transporters in the Sertoli, germ, and Leydig cells further substantiates the role of TH in testicular function (Gao et al., 2014). Aside from the presence of TRs, these testicular cells also contain deiodinases, responsible for converting thyroxine (T4) into active triiodothyronine (T3) and vice versa. Hence, testicular cells are equipped with the transporters and enzymes required to maintain thyroid hormone homeostasis within the testes (Gao et al., 2014). Physiologically, TH is an important factor in redox balance, and alteration in thyroid homeostasis can lead to oxidative stress, apoptosis, and proton pump dysfunction (Chang et al., 2019). Hypothyroidism has been shown to disrupt proton pump (such as Ca2+-ATPase) activities, which in turn disrupts calcium homeostasis (Chang et al., 2019). Maintaining intracellular calcium is important for mitochondrial function since calcium imbalance is a major trigger for mitochondrial dysfunction associated with oxidative stress and apoptosis (Nazıroğlu et al., 2012).

Hence, it is plausible to predict that BPF-impaired testicular function might be associated with thyroid dyshomeostasis and proton pump dysfunction since BPF has been linked with testicular oxidative damage and apoptosis (Odetayo et al., 2023b).

Despite the available information on dysthyroidism and oxidative stress, the role of thyroid homeostasis in apoptosis has not been fully explored. Although hypothyroidism has been associated with increased apoptotic markers (Mukherjee et al., 2014), the role of tumor protein P53 (p53)/ b-cell lymphoma 2 (Bcl-2) signaling has not been explored. P53 signaling is responsible for regulating different cell reactions related to apoptosis. It is a core participant in apoptosis regulation by stimulating the caspase-dependent pathway via the modulation of multiple apoptotic markers such as BCl-2 (Muller & Vousden, 2013). Thus, P53 directly or indirectly regulates apoptosis at different levels (Moulder et al., 2018).

On the other hand, omega 3 fatty acids (O3FA) are antioxidants that can be obtained from diet. O3FA are polyunsaturated fatty acids (PUFA) with established protective roles in the cardiovascular system (Jain et al., 2015), liver (Parker et al., 2012), kidney (Hu et al., 2017), and testes (Akhigbe et al., 2021a; Odetayo & Olayaki, 2023). In addition, O3FA are antioxidant and anti-inflammatory agents considered the precursors of key active metabolites for treating several diseases (Abdel-Baky et al., 2023). This information suggests that O3FA might be a promising supplement for protecting cells from extrinsic toxic stimuli such as BPF.

Despite these pieces of information, no study has explored the possible role of TH homeostasis on BPF-induced testicular dysfunction or the role of proton pump activity on BPF-induced gonadotoxicity. Although O3FA are anti-apoptotic agents, the role of P53/BCl-2 signaling in O3FA-mediated anti-apoptotic effect has not been explored. Hence, this study was designed to establish the effect of dysthyroidism in BPF-induced testicular dysfunction and the possible ameliorative effect of O3FA. Additionally, the roles of proton pumps and P53/BCl-2 signaling as the possible mechanism of action were explored.

MATERIALS AND METHODS

Chemicals

Each capsule of O3FA used in this study contained eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) at a ratio of 3:2. The capsules were procured from Gujarat Liqui Pharmacaps Pvt. Ltd. Vadodara, Gujarat, India. All other chemicals except otherwise stated were purchased from Sigma Aldrich.

Animals

The twenty male Wistar rats (age: 10-13 weeks, weights: 160-180g) obtained from the Biochemistry Department of the University of Ilorin were randomized into four groups (n=5) after two weeks of acclimatization. The animals in Group 1 (Control) were treated with 0.5 ml of corn oil; Group 2 (BPF) received 30 mg/kg of BPF; Group 3 (BPF+O3FA-L) and Group 4 (BPF+O3A-H) received BPF + low (100 mg/kg) and high (300 mg/kg) doses of O3FA respectively. The BPF dosage used in this study was similar to the doses previously reported by Higashihara et al. (2007), Ullah et al. (2019), and Fatai & Aribidesi (2022), while the dosage of O3FA was earlier reported and used by Adeyemi & Olayaki (2017).

The designed experimental protocol was approved by the University of Ilorin Review and Ethical Committee and followed the “National Institute of Health guidelines using the guide for the care and handling of laboratory animals (NIH Publication No. 80-23; amended 1978)”. The experimental protocol complied with the National Research Council’s guidelines for the Care and Use of Laboratory Animals and the ARRIVE guidelines for reporting experimental findings.

Sample Collection

The calculated dosage of BPF for each animal was dissolved in corn oil so that each received 0.5 ml of the solution. The solution was administered orally to mimic the main route of BPF exposure. The rats were given the solution for 28 days, and the overnight fasted animals were sacrificed after 24 hours from the last treatment with intraperitoneal ketamine (40 mg/kg) and xylazine (4 mg/kg) (Afolabi et al., 2022a). The blood samples obtained via cardiac puncture were centrifuged at 3000 rpm to obtain serum for hormonal assays. The left testes were homogenized in a cold phosphate buffer solution for biochemical analysis. The right testes were harvested for immunohis-tochemistry testing, while the epididymides were removed for sperm analysis.

Sperm analysis

Each caudal epididymis was prepared in a clean petri dish, and sperm count, motility, and morphology were estimated according to previous methods (Akhigbe et al., 2021b; Afolabi et al., 2022b).

Hormonal Assay

Serum T3, T4, thyroid stimulating hormone (TSH) (Carlbiotech, USA) and luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone, and estradiol (Bio-Inteco, UK) were determined using ELISA kits according to the manufacturer’s guidelines.

Oxidative stress, inflammatory, and apoptotic markers

Testicular malondialdehyde (MDA), superoxide dismutase (SOD), and catalase (CAT) were determined as previously established (Afolabi et al., 2022c; Akhigbe et al., 2023; Olayaki et al., 2023). Total antioxidant capacity (TAC) (Fortress Diagnostic Kit, Switzerland) was estimated using a colorimetry method. Testicular interleukin-1 beta (IL-1P) (Nanjing Mornmed Medical, China), C-reactive protein (CRP), nuclear factor erythroid 2-related factor 2 (Nrf2), and caspase-3 (Elabscience, USA) were determined as described by the test kit manufacturers.

Proton pump

Testicular transmembrane protein (Na+/K+−ATPase and Ca2+−ATPase) activity was determined spectrophotometrically using the method of Torlinska & Grochowalska (2004).

Immunohistochemistry

Testicular estrogen receptor β (Erβ), P53, and BCl-2 were determined as previously described by Odetayo et al. (2023a). “Formalin-fixed and paraffin-embedded testicular tissues were sectioned at 4 µm for immunohistochemistry. Immunohistochemical procedures were performed using appropriate antibodies; anti-mouse Erβ monoclonal for Erβ expression (1:100), anti-mouse p53 monoclonal for p53 expression (1:100), and anti-mouse Bcl-2 monoclonal for Bcl-2 expression (1:200). The formalin-fixed and paraffin-embedded testicular tissues were sectioned at 4 µm for immunohistochemistry. Appropriate antibodies; anti-mouse Erβ monoclonal for Erβ expression (1:100) (Leica Biosystems, USA with CAT NO: 6069100), anti-mouse p53 monoclonal for p53 expression (1:100) (Espredia with CAT NO: 186P2105D), and anti-mouse Bcl-2 monoclonal for Bcl-2 expression (1:200) (Thermo Fisher Scientific, USA)”.

Statistical analysis

Software package GraphPad PRISM 5 (GraphPad Software, La Jolla, California, USA) was used in statistical analysis with one-way analysis of variance (ANOVA) and Tukey’s post hoc test. Data were reported as mean ± standard deviation. Values of p below 0.05 were considered statistically significant.

RESULTS

Sperm quality

Low (O3FA-L) and high doses (OFA3-H) of O3FA ameliorated BPF-induced decrease in sperm count, motility, and morphology compared with controls (Figure 1). Although there was no significant difference in sperm motility and morphology in the rats given O3FA-L and OFA3-H, the animals treated with a high dose of O3FA had better sperm counts than their counterparts treated with a low dose.

Figure 1.

Figure 1

Open in a new tab

Effect of omega 3 fatty acids on (a) sperm count (b) motility (c) morphology in BPF-exposed rats. a. p<0 .05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Hormones

BPF disrupted thyroid homeostasis, as evidenced by the significant decrease in T4 and T3 and the increase in TSH compared with controls (Figure 2). The observed alterations were prevented by the co-treatment of BPF with low and high doses of O3FA. However, animals treated with O3FA-H had better circulatory T3 levels than those treated with O3FA-L. Furthermore, the observed decrease in serum LH, FSH, and testosterone and increase in estradiol following BPF treatment compared with controls were blunted by both doses of O3FA. Although both doses of O3FA prevented the observed reproductive hormonal imbalance, the ameliorative effect was more pronounced in animals treated with high doses.

Figure 2.

Figure 2

Open in a new tab

Effect of omega 3 fatty acids on serum (a) thyroxine (b) triiodothyronine (c) thyroid stimulating hormone (TSH) (d) luteinizing hormone (LH) (e) follicle stimulating hormone (FSH) (f) testosterone (g) estradiol in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using oneway analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Oxidative stress

BPF exposure led to a significant increase in testicular MDA and a decrease in CAT, SOD, TAC, and Nrf2 compared with controls (Figure 3). Both doses of O3FA ameliorated the redox imbalance, although animals treated with O3FA-H were significantly different in TAC and Nrf2 levels compared with those treated with O3FA-L.

Figure 3.

Figure 3

Open in a new tab

Effect of omega 3 fatty acids on testicular (a) malondialdehyde (MDA) (b) catalase (CAT) (c) superoxide dismutase (SOD) (d) total antioxidant capacity (TAC) (e) nuclear factor erythroid 2-related factor 2 (nrf2) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using oneway analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Inflammatory and Apoptotic markers

O3FA prevented BPF-induced increase in testicular IL-1β, CRP, and caspase 3 compared with controls (Figures 4 and 5), although except for caspase 3, animals treated with a high dose of O3FA exhibited a better ameliorative effect in testicular parameters.

Figure 4.

Figure 4

Open in a new tab

Effect of omega 3 fatty acids on testicular (a) interleukin 1 beta (IL-1β) (b) c-reactive protein (CRP) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Figure 5.

Figure 5

Open in a new tab

Effect of omega 3 fatty acids on testicular caspase 3 in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Proton pump

BPF affected proton pump activity compared with controls (Figure 6). While both doses of O3FA prevented the observed proton pump Na+−K+ ATPase and Ca2+ ATPase dysfunction, animals treated with high doses of O3FA exhibited a better ameliorative effect than their counterparts treated with a low dose.

Figure 6.

Figure 6

Open in a new tab

Effect of omega 3 fatty acids on testicular (a) sodium-potassium ATPase (Na-K ATPase) (b) calcium ATPase (Ca-ATPase) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Er-β and P53/BCI-2 signaling

Low and high doses of O3FA blunted the observed increase in Er-β and P53 and decreased BCL-2 following BPF exposure (Figures 7, 8, and 9). However, these ameliorative effects were more pronounced in animals treated with high doses of O3FA.

Figure 7.

Figure 7

Open in a new tab

Effect of omega 3 fatty acids on testicular estrogen receptor beta (ER β) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Figure 8.

Figure 8

Open in a new tab

Effect of omega 3 fatty acids on testicular tumor protein p53 (p53) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

Figure 9.

Figure 9

Open in a new tab

Effect of omega 3 fatty acids on testicular B-cell lymphoma 2 (BCl-2) in BPF-exposed rats. a. p<0.05 versus control, b. p<0.05 versus BPF, c. p<0.05 versus BPF + O3FA-L using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for pairwise comparison. BPF: Bisphenol F, O3FA-L: omega-3 fatty acid low dose, O3FA-H: omega-3 fatty acid high dose.

DISCUSSION

The present study explored the effects of BPF on thyroid and testicular function. It also determined the role of Na+−K+ ATPase and Ca2+ ATPase activities and P53/BCl-2 signaling in testicular function. A possible ameliorative effect of O3FA on BPF-induced hypothyroidism was also established. It further hypothesized that BPF-induced testicular dysfunction was due to dysthyroidism-mediated Na+−K+ and Ca2+ ATPase dysfunction and P53/BCl-2 imbalance and that O3FA might ameliorate BPF-induced dysthyroidism. Our findings confirmed that BPF-induced hypothyroidism was associated with impaired testicular function. They also established that O3FA ameliorated BPF-induced hypothyroidism, oxidative stress, inflammatory response, and apoptosis in testicular tissues, as evidenced by the significant decreases in testicular CAT, SOD, and TAC, Nrf2, and the increases in MDA, IL-1β, CRP, and caspase 3. These events were accompanied by Na+−K+ and Ca2+ ATPase dysfunction and P53/BCl-2 imbalance in testicular tissues. In sum, our data suggest that O3FA restored testicular function by preventing BPF-induced hypothyroidism via Na+−K+ and Ca2+ ATPase and P53/BCl-2 mediated oxidative stress and apoptosis.

As expected, the results from this study showed that hypothyroidism (decreased T4 and T3 levels) was associated with testicular dysfunction, as evidenced by a significant decline in sperm quality and reproductive hormones. This confirms previous findings (Mazzilli et al., 2023; El-Kashlan et al., 2015; La Vignera et al., 2017) demonstrating testicular dysfunction and male infertility in hypothyroidism. Hyperprolactinemia might be the link between the observed primary hypothyroidism and hypogonadotropic hypogonadism (Brown et al., 2023) since BPF has been shown to increase circulatory prolactin (Odetayo & Olayaki, 2023). Hypothyroidism is associated with an increase in circulatory TRH, which in turn increases circulatory prolactin (Tashjian Jr et al., 1971). This hypothyroidism-mediated hyperprolactinemia might directly impair the HPG-axis by inhibiting GnRH (Brown et al., 2019) and LH (Gregory et al., 2004) secretion, leading to a decline in circulatory testosterone. Furthermore, the findings from this study that BPF-induced hypothyroidism was associated with testicular oxidative damage, inflammation, and apoptosis aligns with the study of Kochman et al. (2021) and Bowman-Colin et al. (2016), which reported similar findings in dysthyroidism.

Although compelling evidence established that hypothyroidism induces testicular dysfunction, information about the associated mechanisms is still lacking. Therefore, the finding that BPF-induced hypothyroidism is associated with testicular transmembrane protein dysfunction is noteworthy.

BPF-induced testicular injury may be a consequence of dysthyroidism-induced testicular Na+/K+−ATPase and Ca2+−ATPase dysfunction (Chang et al., 2019). Impairment of these transmembrane proteins depresses electrochemical gradient generation and maintenance across the testicular cell membrane and key organelles such as the mitochondria (Therien et al., 1997). Additionally, Ca2+−ATPase maintains calcium homeostasis, which plays a key role in male fertility. The prostate gland (the main source of calcium for human semen), epididymis, and seminal vesicles require optimal calcium levels (Valsa et al., 2016). Impaired steroidogenesis, sperm motility, chemotaxis, capacitation, and acrosome reaction have been reported in hypocalcemia (Beigi Harchegani et al., 2019; Naz et al., 2022). Also, men with hypomotility show lower calcium levels in semen than those with typical motility (Naz et al., 2022), illustrating the importance of calcium homeostasis in male reproduction. Ca2+−ATPase is a key factor for regulating calcium homeostasis, and its dysfunction has been reported in calcium overload (Chang et al., 2019). The testicular Na+/K+−ATPase and Ca2+−ATPase dysfunction in this study might be a consequence of the observed BPF-induced oxidative stress or increase in serum estrogen and the subsequent ER3 overexpression. Thyroid hormones play a major role in the maintenance of reactive oxygen species (ROS) generation, and dysfunction has been reported to cause the overproduction of ROS, leading to oxidative stress (Chang et al., 2019), which might impair proton pump function (Zaidi, 2010). Proton pump inhibition has been linked with increased circulatory estrogen (Ding et al., 2023). Thus, the observed increase in estrogen and estrogen receptors observed in this study might result from the observed proton pump dysfunction. Interestingly enough, both oxidative stress and ER3 are triggers of apoptosis (Odetayo et al., 2023a), which culminates in the stimulation of p53/BCl-2-mediated apoptosis.

p53 is a key protein that regulates apoptosis via direct induction of Bax transcription, which can, in turn, overwhelm the antiapoptotic effects of BCl-2 (McCurrach et al., 1997; Giorgi et al., 2015). The activation of Bax leads to the release of mitochondrial cytochrome c, leading to the activation of caspase 3-mediated apoptosis (Schuler et al., 2000). In addition, p53 can directly inhibit Bcl-2 activities by mimicking the proapoptotic ‘BH3-only’ class of Bcl-2 to bring about mitochondria permeabilization and apoptosis (Hemann & Lowe, 2006). Again, p53-induced PUMA protein can distort cytosolic p53-Bcl-2 complexes, eventually leading to the direct induction of p53-mediated apoptosis in mitochondria (Chipuk et al., 2005). Furthermore, p53 can directly disrupt proton pump activity in the endoplasmic reticulum, leading to calcium overload and increased transfer to the mitochondria, leading to apoptosis induction (Giorgi et al., 2015). Hence, the increase in p53 expression observed in this study might also explain the proton pump dysfunction following BPF-induced hypothyroidism. Our findings that BPF-induced hypothyroidism disrupted p53/BCl-2 signaling agreed with the study of Singh et al. (2003), in which increased apoptosis following a decrease in thyroid hormones was described.

Another important finding is the protective role of O3FA against BPF-induced hypothyroidism. This study revealed that O3FA prevented BPF-induced hypothyroidism-mediated testicular injury by restoring hormonal and redox balance and suppressing inflammatory and apoptotic markers, thus improving sperm parameters and testosterone synthesis. Although this is the first study to demonstrate the ameliorative effect of O3FA on BPF-induced hypothyroidism, our findings agreed with those of previous studies that described the antioxidant (Meital et al., 2019), anti-inflammatory (Calder, 2017), antiapoptotic (Wendel & Heller, 2009), and thyroid protective (Abd Allah et al., 2014; Benvenga et al., 2022) effects of O3FA. In addition, O3FA prevents calcium overload during ischemic insult (Kromhout et al., 2012), which further supports our claim that O3FA prevents BPF-induced dysthyroidism by preventing proton pump dysfunction.

CONCLUSION

O3FA prevented BPF-induced primary (peripheral) hypothyroidism and secondary (hypogonadotropic) testicular dysfunction. O3FA also reversed the deleterious effects of BPF on testicular oxidative stress, inflammation, and apoptosis markers.

Footnotes

Ethical Approval

The animals were purchased from the University of Ilorin and carefully handled as required by the National Institute of Health (NIH). The ARRIVE guidelines for reporting experimental findings were strictly followed. The experimental research protocol was designed according to the National Research Council’s guidelines for the Care and Use of Laboratory Animals, and ethical approval was obtained from the University of Ilorin Ethical Review Committee.

CONFLICT OF INTERESTS

The authors have no conflicts of interest to declare.

REFERENCES

  1. Abd Allah ES, Gomaa AM, Sayed MM. The effect of omega-3 on cognition in hypothyroid adult male rats. Acta Physiol Hung. 2014;101:362–376. doi: 10.1556/APhysiol.101.2014.3.11. [DOI] [PubMed] [Google Scholar]
  2. Abdel-Baky ES, Radwan SA, Ibrahim MF, Abdel-Rahman ON. Influence of omega-3 fatty acids, soya isoflavones and their combination for abrogating carbon tetrachloride hazards in male rats. Braz J Biol. 2023;84:e266024–e266024. doi: 10.1590/1519-6984.266024. [DOI] [PubMed] [Google Scholar]
  3. Adeyemi WJ, Olayaki LA. Diclofenac-induced hepatotoxicity: Low dose of omega-3 fatty acids have more protective effects. Toxicol Rep. 2017;5:90–95. doi: 10.1016/j.toxrep.2017.12.002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Afolabi OA, Hamed MA, Anyogu DC, Adeyemi DH, Odetayo AF, Akhigbe RE. Atorvastatin-mediated downregulation of VCAM-1 and XO/UA/caspase 3 signaling averts oxidative damage and apoptosis induced by ovarian ischaemia/reperfusion injury. Redox Rep. 2022a;27:212–220. doi: 10.1080/13510002.2022.2129192. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Afolabi OA, Anyogu DC, Hamed MA, Odetayo AF, Adeyemi DH, Akhigbe RE. Glutamine prevents upregulation of NF-kB signaling and caspase 3 activation in ischaemia/reperfusion-induced testicular damage: An animal model. Biomed Pharmacother. 2022b;150:113056–113056. doi: 10.1016/j.biopha.2022.113056. [DOI] [PubMed] [Google Scholar]
  6. Afolabi AO, Akhigbe TM, Odetayo AF, Anyogu DC, Hamed MA, Akhigbe RE. Restoration of Hepatic and Intestinal Integrity by Phyllanthus amarus Is Dependent on Bax/Caspase 3 Modulation in Intestinal Ischemia-/Reperfusion-Induced Injury. Molecules. 2022c;27:5073–5073. doi: 10.3390/molecules27165073. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Akhigbe RE, Hamed MA, Odetayo AF, Akhigbe TM, Ajayi AF, Ajibogun FAH. Omega-3 fatty acid rescues ischaemia/perfusion-induced testicular and sperm damage via modulation of lactate transport and xanthine oxidase/uric acid signaling. Biomed Pharmacother. 2021a;142:111975–111975. doi: 10.1016/j.biopha.2021.111975. [DOI] [PubMed] [Google Scholar]
  8. Akhigbe RE, Hamed MA, Odetayo AF. HAART and anti-Koch’s impair sexual competence, sperm quality and offspring quality when used singly and in combination in male Wistar rats. Andrologia. 2021b;53:e13951–e13951. doi: 10.1111/and.13951. [DOI] [PubMed] [Google Scholar]
  9. Akhigbe RE, Hamed MA, Odetayo AF, Akhigbe TM, Oyedokun PA. Zinc improves sexual and erectile function in HAART-treated rats via the upregulation of erectogenic enzymes and maintenance of redox balance. Aging Male. 2023;26:2205517–2205517. doi: 10.1080/13685538.2023.2205517. [DOI] [PubMed] [Google Scholar]
  10. Beigi Harchegani A, Irandoost A, Mirnamniha M, Rahmani H, Tahmasbpour E, Shahriary A. Possible Mechanisms for The Effects of Calcium Deficiency on Male Infertility. Int J Fertil Steril. 2019;12:267–272. doi: 10.22074/ijfs.2019.5420. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Benvenga S, Famà F, Perdichizzi LG, Antonelli A, Brenta G, Vermiglio F, Moleti M. Fish and the Thyroid: A Janus Bifrons Relationship Caused by Pollutants and the Omega-3 Polyunsaturated Fatty Acids. Front Endocrinol (Lausanne) 2022;13:891233–891233. doi: 10.3389/fendo.2022.891233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Bowman-Colin C, Salazar LA, Martins JO. The Role of Endocrine System in the Inflammatory Process. Mediators Inflamm. 2016;2016:6081752–6081752. doi: 10.1155/2016/6081752. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Brown EDL, Obeng-Gyasi B, Hall JE, Shekhar S. The Thyroid Hormone Axis and Female Reproduction. Int J Mol Sci. 2023;24:9815–9815. doi: 10.3390/ijms24129815. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Brown RSE, Khant Aung Z, Phillipps HR, Barad Z, Lein HJ, Boehm U, Szawka RE, Grattan DR. Acute Suppression of LH Secretion by Prolactin in Female Mice Is Mediated by Kisspeptin Neurons in the Arcuate Nucleus. Endocrinology. 2019;160:1323–1332. doi: 10.1210/en.2019-00038. [DOI] [PubMed] [Google Scholar]
  15. Calder PC. Omega-3 fatty acids and inflammatory processes: from molecules to man. Biochem Soc Trans. 2017;45:1105–1115. doi: 10.1042/BST20160474. [DOI] [PubMed] [Google Scholar]
  16. Caporossi L, Papaleo B. Bisphenol A and Metabolic Diseases: Challenges for Occupational Medicine. Int J Environ Res Public Health. 2017;14:959–959. doi: 10.3390/ijerph14090959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Chang XR, Yao YL, Wang D, Ma HT, Gou PH, Li CY, Wang JL. Influence of hypothyroidism on testicular mitochondrial oxidative stress by activating the p38 mitogen-activated protein kinase and c-Jun NH2-terminal kinase signaling pathways in rats. Hum Exp Toxicol. 2019;38:95–105. doi: 10.1177/0960327118781927. [DOI] [PubMed] [Google Scholar]
  18. Chipuk JE, Bouchier-Hayes L, Kuwana T, Newmeyer DD, Green DR. PUMA couples the nuclear and cytoplasmic proapoptotic function of p53. Science. 2005;309:1732–1735. doi: 10.1126/science.1114297. [DOI] [PubMed] [Google Scholar]
  19. Ding F, Liu Y, Chen J, Li Y, Guo X, Wu J, Liu Q. Proton pump inhibitors affect sperm parameters by regulating aquaporins. Heliyon. 2023;9:e17911–e17911. doi: 10.1016/j.heliyon.2023.e17911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. El-Kashlan AM, Nooh MM, Hassan WA, Rizk SM. Therapeutic Potential of Date Palm Pollen for Testicular Dysfunction Induced by Thyroid Disorders in Male Rats. PLoS One. 2015;10:e0139493–e0139493. doi: 10.1371/journal.pone.0139493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Fatai OA, Aribidesi OL. Effect of bisphenol F on sexual performance and quality of offspring in Male Wistar rats. Ecotoxicol Environ Saf. 2022;244:114079–114079. doi: 10.1016/j.ecoenv.2022.114079. [DOI] [PubMed] [Google Scholar]
  22. Ferreira R, Amaral C, Correia-da-Silva G, Almada M, Borges M, Cunha SC, Fernandes JO, Teixeira N. Bisphenols A, F, S and AF trigger apoptosis and/or endoplasmic reticulum stress in human endometrial stromal cells. Toxicology. 2022;478:153282–153282. doi: 10.1016/j.tox.2022.153282. [DOI] [PubMed] [Google Scholar]
  23. Gao Y, Lee WM, Cheng CY. Thyroid hormone function in the rat testis. Front Endocrinol (Lausanne) 2014;5:188–188. doi: 10.3389/fendo.2014.00188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Giorgi C, Bonora M, Sorrentino G, Missiroli S, Poletti F, Suski JM, Galindo Ramirez F, Rizzuto R, Di Virgilio F, Zito E, Pandolfi PP, Wieckowski MR, Mammano F, Del Sal G, Pinton P. p53 at the endoplasmic reticulum regulates apoptosis in a Ca2+-dependent manner. Proc Natl Acad Sci U S A. 2015;112:1779–1784. doi: 10.1073/pnas.1410723112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Gregory SJ, Townsend J, McNeilly AS, Tortonese DJ. Effects of prolactin on the luteinizing hormone response to gonadotropin-releasing hormone in primary pituitary cell cultures during the ovine annual reproductive cycle. Biol Reprod. 2004;70:1299–1305. doi: 10.1095/biolreprod.103.022806. [DOI] [PubMed] [Google Scholar]
  26. Hemann MT, Lowe SW. The p53-Bcl-2 connection. Cell Death Differ. 2006;13:1256–1259. doi: 10.1038/sj.cdd.4401962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Higashihara N, Shiraishi K, Miyata K, Oshima Y, Minobe Y, Yamasaki K. Subacute oral toxicity study of bisphenol F based on the draft protocol for the “Enhanced OECD Test Guideline no. 407”. Arch Toxicol. 2007;81:825–832. doi: 10.1007/s00204-007-0223-4. [DOI] [PubMed] [Google Scholar]
  28. Hu J, Liu Z, Zhang H. Omega-3 fatty acid supplementation as an adjunctive therapy in the treatment of chronic kidney disease: a meta-analysis. Clinics (Sao Paulo) 2017;72:58–64. doi: 10.6061/clinics/2017(01)10. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jain AP, Aggarwal KK, Zhang PY. Omega-3 fatty acids and cardiovascular disease. Eur Rev Med Pharmacol Sci. 2015;19:441–445. [PubMed] [Google Scholar]
  30. Kochman J, Jakubczyk K, Bargiel P, Janda-Milczarek K. The Influence of Oxidative Stress on Thyroid Diseases. Antioxidants (Basel) 2021;10:1442–1442. doi: 10.3390/antiox10091442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kromhout D, Yasuda S, Geleijnse JM, Shimokawa H. Fish oil and omega-3 fatty acids in cardiovascular disease: do they really work? Eur Heart J. 2012;33:436–443. doi: 10.1093/eurheartj/ehr362. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. La Vignera S, Vita R, Condorelli RA, Mongioì LM, Presti S, Benvenga S, Calogero AE. Impact of thyroid disease on testicular function. Endocrine. 2017;58:397–407. doi: 10.1007/s12020-017-1303-8. [DOI] [PubMed] [Google Scholar]
  33. Mazzilli R, Medenica S, Di Tommaso AM, Fabozzi G, Zamponi V, Cimadomo D, Rienzi L, Ubaldi FM, Watanabe M, Faggiano A, La Vignera S, Defeudis G. The role of thyroid function in female and male infertility: a narrative review. J Endocrinol Invest. 2023;46:15–26. doi: 10.1007/s40618-022-01883-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. McCurrach ME, Connor TM, Knudson CM, Korsmeyer SJ, Lowe SW. bax-deficiency promotes drug resistance and oncogenic transformation by attenuating p53-dependent apoptosis. Proc Natl Acad Sci U S A. 1997;94:2345–2349. doi: 10.1073/pnas.94.6.2345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Meital LT, Windsor MT, Perissiou M, Schulze K, Magee R, Kuballa A, Golledge J, Bailey TG, Askew CD, Russell FD. Omega-3 fatty acids decrease oxidative stress and inflammation in macrophages from patients with small abdominal aortic aneurysm. Sci Rep. 2019;9:12978–12978. doi: 10.1038/s41598-019-49362-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Meng L, Rijntjes E, Swarts H, Bunschoten A, van der Stelt I, Keijer J, Teerds K. Dietary-Induced Chronic Hypothyroidism Negatively Affects Rat Follicular Development and Ovulation Rate and Is Associated with Oxidative Stress. Biol Reprod. 2016;94:90–90. doi: 10.1095/biolreprod.115.136515. [DOI] [PubMed] [Google Scholar]
  37. Moulder DE, Hatoum D, Tay E, Lin Y, McGowan EM. The Roles of p53 in Mitochondrial Dynamics and Cancer Metabolism: The Pendulum between Survival and Death in Breast Cancer? Cancers (Basel) 2018;10:189–189. doi: 10.3390/cancers10060189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Mukherjee S, Samanta L, Roy A, Bhanja S, Chainy GB. Supplementation of T3 recovers hypothyroid rat liver cells from oxidatively damaged inner mitochondrial membrane leading to apoptosis. Biomed Res Int. 2014;2014:590897–590897. doi: 10.1155/2014/590897. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Muller PA, Vousden KH. p53 mutations in cancer. Nat Cell Biol. 2013;15:2–8. doi: 10.1038/ncb2641. [DOI] [PubMed] [Google Scholar]
  40. Naz T, Chakraborty S, Saha S. Role of fatty acids and calcium in male reproduction. Reprod Dev Med. 2022;6:57–64. doi: 10.1097/RD9.0000000000000003. [DOI] [Google Scholar]
  41. Nazıroğlu M, Dikici DM, Dursun S. Role of oxidative stress and Ca2+ signaling on molecular pathways of neuropathic pain in diabetes: focus on TRP channels. Neurochem Res. 2012;37:2065–2075. doi: 10.1007/s11064-012-0850-x. [DOI] [PubMed] [Google Scholar]
  42. Odetayo AF, Adeyemi WJ, Olayaki LA. In vivo exposure to bisphenol F induces oxidative testicular toxicity: role of Erβ and p53/Bcl-2 signaling pathway. Front Reprod Health. 2023a;5:1204728–1204728. doi: 10.3389/frph.2023.1204728. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Odetayo AF, Adeyemi WJ, Olayaki LA. Omega-3 fatty acid ameliorates bisphenol F-induced testicular toxicity by modulating Nrf2/NFkB pathway and apoptotic signaling. Front Endocrinol (Lausanne) 2023b;14:1256154–1256154. doi: 10.3389/fendo.2023.1256154. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Odetayo A, Olayaki LA. Bisphenol F Induced Reproductive Toxicity by Disrupting Steroidogenic Enzymes Activities and Upregulating Xanthine Oxidase/Uric Acid Signaling. Fertil Steril. 2022;118:e75–e75. doi: 10.1016/j.fertnstert.2022.08.230. [DOI] [Google Scholar]
  45. Odetayo AF, Olayaki LA. Omega 3 fatty acid improves sexual and erectile function in BPF-treated rats by upregulating NO/cGMP signaling and steroidogenic enzymes activities. Sci Rep. 2023;13:18060–18060. doi: 10.1038/s41598-023-45344-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Olayaki LA, Okesina KB, Jesubowale JD, Ajibare AJ, Odetayo AF. Orange Peel Extract and Physical Exercise Synergistically Ameliorate Type 2 Diabetes Mellitus-Induced Dysmetabolism by Upregulating GLUT4 Concentration in Male Wistar Rats. J Med Food. 2023;26:470–479. doi: 10.1089/jmf.2023.0061. [DOI] [PubMed] [Google Scholar]
  47. Parker HM, Johnson NA, Burdon CA, Cohn JS, O’Connor HT, George J. Omega-3 supplementation and non-alcoholic fatty liver disease: a systematic review and meta-analysis. J Hepatol. 2012;56:944–951. doi: 10.1016/j.jhep.2011.08.018. [DOI] [PubMed] [Google Scholar]
  48. Sahoo DK, Roy A, Bhanja S, Chainy GB. Hypothyroidism impairs antioxidant defence system and testicular physiology during development and maturation. Gen Comp Endocrinol. 2008;156:63–70. doi: 10.1016/j.ygcen.2007.11.007. [DOI] [PubMed] [Google Scholar]
  49. Schuler M, Bossy-Wetzel E, Goldstein JC, Fitzgerald P, Green DR. p53 induces apoptosis by caspase activation through mitochondrial cytochrome c release. J Biol Chem. 2000;275:7337–7342. doi: 10.1074/jbc.275.10.7337. [DOI] [PubMed] [Google Scholar]
  50. Singh R, Upadhyay G, Kumar S, Kapoor A, Kumar A, Tiwari M, Godbole MM. Hypothyroidism alters the expression of Bcl-2 family genes to induce enhanced apoptosis in the developing cerebellum. J Endocrinol. 2003;176:39–46. doi: 10.1677/joe.0.1760039. [DOI] [PubMed] [Google Scholar]
  51. Tashjian AH Jr, Barowsky NJ, Jensen DK. Thyrotropin releasing hormone: direct evidence for stimulation of prolactin production by pituitary cells in culture. Biochem Biophys Res Commun. 1971;43:516–523. doi: 10.1016/0006-291X(71)90644-9. [DOI] [PubMed] [Google Scholar]
  52. Therien AG, Goldshleger R, Karlish SJ, Blostein R. Tissue-specific distribution and modulatory role of the gamma subunit of the Na,K-ATPase. J Biol Chem. 1997;272:32628–32634. doi: 10.1074/jbc.272.51.32628. [DOI] [PubMed] [Google Scholar]
  53. Torlińska T, Grochowalska A. Age-related changes of NA(+), K(+) - ATPase, Ca(+2) - ATPase and Mg(+2) - ATPase activities in rat brain synaptosomes. J Physiol Pharmacol. 2004;55:457–465. [PubMed] [Google Scholar]
  54. Ullah A, Pirzada M, Afsar T, Razak S, Almajwal A, Jahan S. Effect of bisphenol F, an analog of bisphenol A, on the reproductive functions of male rats. Environ Health Prev Med. 2019;24:41–41. doi: 10.1186/s12199-019-0797-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Valsa J, Skandhan KP, Sumangala B, Amith S, Skandhan Avni KP. Effect of different timings of the day on semen and calcium and magnesium in it. Urologia. 2016;83:207–210. doi: 10.5301/uro.5000029. [DOI] [PubMed] [Google Scholar]
  56. Wendel M, Heller AR. Anticancer actions of omega-3 fatty acids--current state and future perspectives. Anticancer Agents Med Chem. 2009;9:457–470. doi: 10.2174/1871520610909040457. [DOI] [PubMed] [Google Scholar]
  57. Yuan L, Qian L, Qian Y, Liu J, Yang K, Huang Y, Wang C, Li Y, Mu X. Bisphenol F-Induced Neurotoxicity toward Zebrafish Embryos. Environ Sci Technol. 2019;53:48–48. doi: 10.1021/acs.est.9b04097. [DOI] [PubMed] [Google Scholar]
  58. Zaidi A. Plasma membrane Ca-ATPases: Targets of oxidative stress in brain aging and neurodegeneration. World J Biol Chem. 2010;1:271–280. doi: 10.4331/wjbc.v1.i9.271. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from JBRA Assisted Reproduction are provided here courtesy of Brazilian Society of Assisted Reproduction

RESOURCES