Elsevier

Neurotoxicology and Teratology

Volume 88, November–December 2021, 107030
Neurotoxicology and Teratology

Full Length Article
Effects of short-term exposure to genistein and overfeeding diet on the neural and retinal progenitor competence of adult zebrafish (Danio rerio)

https://doi.org/10.1016/j.ntt.2021.107030Get rights and content

Highlights

  • OFD induced obesity, general growth and fatty liver.

  • GEN induced anti-adipogenic effects on obesity.

  • OFD induced deleterious effects on retina and brain homeostasis.

  • GEN mitigate the negative impacts of OFD on retinogenesis.

  • GEN exacerbated the deleterious effects of OFD on neurogenesis.

Abstract

Neurogenesis is a process that occurs throughout the life of a vertebrate. Among the different factors that may affect the natural occurrence of neurogenesis, obesity seems to decrease the proliferation capacity of progenitor neuronal cells. Conversely, the phytoestrogen genistein is known to attenuate some obesity effects beyond its neuroprotective action.

Aiming to improve the understanding of how obesity and genistein trigger an impact on the neural and retinal progenitor competence of adult zebrafish, fish were exposed to genistein (GEN - 2 μg L−1) alone or combined with two dietary groups (control and overfeed - OFD) for up to 9 weeks. Zebrafish were fed once per day with Artemia sp. in the control and GEN (2% of BW, control diet), and three times per day in the OFD and OFD + GEN groups (12% BW, overfeeding diet). To assess obesity induction, BMI, biometric parameters, and PPAR-γ protein were quantified. Afterwards, qRT-PCR and immunohistochemistry were performed to determine the cell proliferation and the presence of stem cells through PCNA and Sox-2. Our findings proved that overfeeding adult zebrafish increased the general growth and induced the development of fatty liver. However, for OFD + GEN, this effect was assuaged through the anti-adipogenic effect of GEN. This finding suggests that phytoestrogens could be beneficial to reduce the negative effects of obesity. Moreover, OF induced negative effects on retinal and brain homeostasis, decreasing the proliferation capacity of progenitor neuronal cells. With regard to retinal progenitor competence, genistein seems to mitigate the negative impacts of obesity, whereas the effects of obesity on the brain were exacerbated by this phytoestrogen which negatively influenced the homeostasis of zebrafish neural progenitor competence. This study highlighted the fact that the effects of phytoestrogens in adult neural progenitor competence are complex and could exhibit dissimilar effects depending on the tissue.

Introduction

Neurogenesis is a complex and highly regulated process resulting in the production of new neurons (Otteson and Hitchcock, 2003; Zupanc and Zupanc, 2006; Zupanc, 2006). Particularly interesting from a comparative point of view to mammals, teleost fish exhibit remarkable adult neurogenesis and neuronal regeneration capacity (Hitchcock et al., 2004; Otteson and Hitchcock, 2003; Zupanc and Zupanc, 2006; Zupanc, 2006). In fact, in contrast to mammals, the brain of adult zebrafish displays an important number of neurogenic niches throughout (Dorsemans et al., 2017; Ghaddar et al., 2020). Such neurogenic capacities in this species are related to the maintenance of several neural stem cells (radial glial cells) during adulthood and further committed progenitors (neuroblasts) (Dorsemans et al., 2017; Ghaddar et al., 2020; Schmidt et al., 2013). However, this process might be affected, in both the adult and embryonic hypothalamus, by nutritional cues and metabolic disorders such as obesity. In recent years, several publications have reported that obesity has been strongly associated, in zebrafish and other species, with an impairment of hippocampal synaptic plasticity (Lindqvist et al., 2006; Molteni et al., 2002), alterations of cognitive abilities such as learning and memory (Stranahan et al., 2008), and ocular diseases including age-related cataract and maculopathy, glaucoma, and diabetic retinopathy (Castro-Sánchez et al., 2019; Centanin et al., 2011; Chow et al., 2009; Guo and Rahmouni, 2011; Huang et al., 2012; Otteson and Hitchcock, 2003). Moreover, several studies have provided evidence that obesity negatively affects neurogenesis, decreasing the proliferation capacity of progenitor neuronal cells (Ghaddar et al., 2020; Park et al., 2010; Tozuka et al., 2009).

Taking into account this emergent issue, soy and phytoestrogens have received increasing attention due to the health benefits associated with their consumption, leading to them being considered potential therapeutic compounds for several metabolic disorders (Cederroth and Nef, 2009; Tuli et al., 2019), including obesity (Behloul and Wu, 2012; Rietjens et al., 2017; Shen et al., 2019; Velasquez and Bhathena, 2007; Xin et al., 2019). Soybeans contain isoflavones, mainly daidzein and genistein, a class of phytoestrogens structurally similar to estradiol and that mimic its effects (Cederroth and Nef, 2009; Vitale et al., 2013). As a phytoestrogen, genistein can bind to estrogen receptors and activate estrogen receptor-dependent pathways, which exert pleiotropic effects on many target organs such as the nervous system (Diotel et al., 2013). Indeed, several reports have provided evidence that soybean has a neuroprotective effect on both the brain (Park et al., 2016; Soltani et al., 2015; Uddin and Kabir, 2019) and retina (Elgayar et al., 2015; Hayashi et al., 1997; Ibrahim et al., 2010) beyond its health benefits on the prevention of obesity (Rietjens et al., 2017; Shen et al., 2019; Velasquez and Bhathena, 2007; Xin et al., 2019).

Within this conceptual framework, zebrafish (Danio rerio) emerged as an interesting and recognized model for studying overweight/obesity (Faillaci et al., 2018; Ghaddar et al., 2020; Landgraf et al., 2017; Montalbano et al., 2019; Oka et al., 2010; Tainaka et al., 2011; Zang et al., 2018) and related metabolic disorders like adult neurogenesis (Dorsemans et al., 2017; Ghaddar et al., 2020; Schmidt et al., 2013) or retinogenesis alterations (Castro-Sánchez et al., 2019; Centanin et al., 2011; Chow et al., 2009; Guo and Rahmouni, 2011; Huang et al., 2012; Otteson and Hitchcock, 2003). Aiming to improve the understanding of how obesity and genistein trigger an impact on adult zebrafish neural and retinal progenitor competence, fish were exposed to genistein (GEN - 2 μg L−1) alone or combined with two dietary groups (control and overfeed - OFD) up to 9 weeks. Zebrafish were fed once per day with Artemia sp. in the control and GEN (2% of BW, control diet), and three times per day in the OFD and OFD + GEN groups (12% BW, overfeeding diet). In this sense, gene expression analysis was performed by qRT-PCR and stereological analysis was based on immunohistochemistry. The proliferating cell nuclear antigen (PCNA) and sox transcription factor-2 (Sox-2) were evaluated in the zebrafish brain and retina to assess the impact of obesity and/or genistein on both cell proliferation and the presence of neuronal progenitor cells, respectively. On the other hand, in an attempt to explore the effects of obesity on zebrafish, biometric parameters were assessed, and the liver was used as an indicator of lipid accumulation rate, with peroxisome proliferator-activated receptor gamma (PPAR-γ), a regulator of lipid metabolism, also being analyzed through immunohistochemistry.

Section snippets

Chemicals

Genistein (GE) (≥98% purity) was purchased from Sigma (St. Louis, MO, USA) and diluted in ethanol. Working solutions were prepared in MilliQ water and all, including the control water, had an insignificant final ethanol volume (0.000002%) in the experimental system.

Zebrafish maintenance

Adult zebrafish (Danio rerio) AB strain were maintained under a controlled temperature (28 ± 1 °C) and photoperiod 14:10 h (light: dark cycle) in dechlorinated and aerated water in a recirculation system with both mechanical and

Biometric parameters

Zebrafish body weights (BW) and total lengths (TL) were assessed (Fig. 2), and body mass index (BMI) was calculated (Table 1). The one-way factorial ANOVA of BMI revealed significant differences between groups (F(3,11) = 26.681 p < 0.001). Firstly, overfeeding diet (12% BW) induced obesity in zebrafish compared to the control diet (2% BW) (Fig. 2 and Table 1). In fact, zebrafish exposed to OFD and OFD + GEN showed a significant increase in body mass index (BMI, Table 1) (p < 0.05). At week 0,

Discussion

Over the last few years, zebrafish (Danio rerio) emerged as an interesting and recognized model for studying overweight/obesity (Faillaci et al., 2018; Ghaddar et al., 2020; Landgraf et al., 2017; Montalbano et al., 2019; Oka et al., 2010; Tainaka et al., 2011; Zang et al., 2018). Additionally, while Artemia nauplii is considered an aquaculture live food with a high nutritional content, previous studies reported that zebrafish overfed with Artemia, as high-fat diet, induced obesity (Landgraf et

Conclusion

In summary, to the best of our knowledge, this is the first study to evaluate the interaction between overfeeding diet, genistein and t effects on zebrafish neural and retinal progenitor competence. The findings suggest that phytoestrogens can represent a resilience strategy, aiming to improve the use of these natural bioactive compounds as phytopharmaceuticals to counteract the negative effects of obesity. From a biological perspective, our results show that zebrafish overfed with an

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work is supported by National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UIDB/04033/2020.

References (72)

  • C. Lawrence et al.

    The effects of feeding frequency on growth and reproduction in zebrafish (Danio rerio)

    Aquaculture

    (2012)
  • Z. Liu et al.

    A review of phytoestrogens: their occurrence and fate in the environment

    Water Res.

    (2010)
  • K.J. Livak et al.

    Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method

    Methods

    (2001)
  • A. Luzio et al.

    Zebrafish sex differentiation and gonad development after exposure to 17 α-ethinylestradiol, fadrozole and their binary mixture: A stereological study

    Aquat Toxicol

    (2015)
  • P. Makantasi et al.

    Estradiol treatment decreases cell proliferation in the neurogenic zones of adult female zebrafish (Danio rerio) brain

    Neurosci.

    (2014)
  • R. Molteni et al.

    A high-fat, refined sugar diet reduces hippocampal brain-derived neurotrophic factor, neuronal plasticity, and learning

    Neurosci.

    (2002)
  • B.K. Ormerod et al.

    Estradiol enhances neurogenesis in the dentate gyri of adult male meadow voles by increasing the survival of young granule neurons

    Neurosci.

    (2004)
  • D.C. Otteson et al.

    Stem cells in the teleost retina: persistent neurogenesis and injury-induced regeneration

    Vis. Res.

    (2003)
  • H.J. Park et al.

    Genistein inhibits differentiation of primary human adipocytes

    J. Nutr. Biochem.

    (2009)
  • H.R. Park et al.

    A high-fat diet impairs neurogenesis: involvement of lipid peroxidation and brain-derived neurotrophic factor

    Neurosci. Lett.

    (2010)
  • H.-H. Shen et al.

    Genistein ameliorated obesity accompanied with adipose tissue browning and attenuation of hepatic lipogenesis in ovariectomized rats with high-fat diet

    J. Nutr. Biochem.

    (2019)
  • Z. Soltani et al.

    Is genistein neuroprotective in traumatic brain injury?

    Physiol. Behav.

    (2015)
  • A. Tingaud-Sequeira et al.

    Zebrafish obesogenic test: a tool for screening molecules that target adiposity

    J. Lipid Res.

    (2011)
  • L. Wang et al.

    Natural product agonists of peroxisome proliferator-activated receptor gamma (PPARγ): a review

    Biochem. Pharmacol.

    (2014)
  • X. Xin et al.

    Protective effect of genistein on nonalcoholic fatty liver disease (NAFLD)

    Biomed. Pharmacother.

    (2019)
  • P. Aleström et al.

    Zebrafish: housing and husbandry recommendations

    Lab. Anim.

    (2020)
  • S. Balbuena-Pecino et al.

    Genistein induces adipogenic and autophagic effects in rainbow trout (Oncorhynchus mykiss) adipose tissue: in vitro and in vivo models

    Int. J. Mol. Sci.

    (2020)
  • N. Behloul et al.

    Genistein a promising therapeutic agent for obesity and diabetes treatment

    Eur. J. Pharmacol.

    (2012)
  • S. Castro-Sánchez et al.

    Functional analysis of new human Bardet-Biedl syndrome loci specific variants in the zebrafish model

    Sci. Rep.

    (2019)
  • Q. Chen et al.

    Direct interaction of receptor tyrosine kinases, EphA4 and PDGFRβ, plays an important role in the proliferation of neural stem cells

    J. Neuro-Oncol.

    (2017)
  • E.S.H. Chow et al.

    Cadmium affects retinogenesis during zebrafish embryonic development

    Toxicol. Appl. Pharmacol.

    (2009)
  • Z.C. Dang

    Dose-dependent effects of soy phytoestrogen genistein on adipocytes: mechanisms of action

    Obes. Rev.

    (2009)
  • A.-C. Dorsemans et al.

    Diabetes, adult neurogenesis and brain remodeling: new insights from rodent and zebrafish models

    Neurogenesis

    (2017)
  • S.A.M. Elgayar et al.

    Genistein treatment confers protection against gliopathy and vasculopathy of the diabetic retina in rats

    Ultrastruct. Pathol.

    (2015)
  • F. Faillaci et al.

    Obese zebrafish: a small fish for a major human health condition

    Animal Model Exp. Med.

    (2018)
  • A. Fickler et al.

    Screening dietary biochanin A, daidzein, equol and genistein for their potential to increase DHA biosynthesis in rainbow trout (Oncorhynchus mykiss)

    PLoS One

    (2019)
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