Effects of Di-Isononyl Phthalate (DiNP) on Follicular Atresia in Zebrafish Ovary.

Filipe G Andrade Godoi,Hamid R Habibi,Renata Guimarães Moreira,Oliana Carnevali,Fabiana L Lo Nostro,Isabel Forner-Piquer,Basilio Randazzo

doi:10.3389/fendo.2021.677853

Abstract

Di-isononyl phthalate (DiNP) is a plasticizer reported to elicit hormone-like activity and disrupt metabolism and reproduction in fish and other vertebrates. In general, phthalates have been used at high concentrations beyond reported environmental levels to assess their adverse effects on fish gonadal physiology. The present study exposed adult female zebrafish to a wide range of DiNP concentrations [0.42 µg L−1 (10−9 M), 4.2 µg L−1 (10−8 M), and 42 µg L−1 (10−7 M)] for 21 days. We evaluated gene expression profiles related to apoptosis, autophagy, and oxidative stress; DNA fragmentation (TUNEL assay: terminal deoxynucleotidyl transferase dUTP nick end labeling) and caspase activity (CAS3) were also examined. Exposure to 0.42 and 4.2 µg L−1 upregulated the genes coding for tnfa and baxa, sod1, prkaa1, respectively. CAS3 immunohistochemistry revealed a higher number of positive vitellogenic oocytes in ovaries exposed to 0.42 µg L−1. Subsequently, we examined the relationship between CAS3 signaling and DNA fragmentation. Accordingly, DNA fragmentation was observed in vitellogenic follicles of fish exposed to 0.42 and 4.2 μg L−1. Our results demonstrate that follicular atresia can occur after exposure to environmental levels of DiNP for 21 days, which may adversely affect the reproductive performance of female zebrafish in a non-monotonic manner.

Highlights

6 million tons of phthalate esters are produced worldwide [1] and used primarily to improve the elasticity, strength, and durability of plastic items
Our results demonstrate that follicular atresia can occur after exposure to environmental levels of di-isononyl phthalate (DiNP) for 21 days, which may adversely affect the reproductive performance of female zebrafish in a nonmonotonic manner
We examined the effects of DiNP on the transcriptomic profile to unravel candidate markers for apoptosis, autophagy, and oxidative stress in the D. rerio ovaries

Summary

Introduction

6 million tons of phthalate esters are produced worldwide [1] and used primarily to improve the elasticity, strength, and durability of plastic items. Many phthalate esters, i.e., dimethyl phthalate (DMP), diethyl phthalate (DEP), di-n-butyl phthalate (DnBP), butyl benzyl phthalate (BBP), di-n-octyl phthalate (DnOP), and di (2-ethylhexyl) phthalate (DEHP), can be detected in aquatic ecosystems [4,5,6]. Considering the major effects of DEHP on aquatic life, the di-isononyl phthalate (DiNP) was introduced in the plastic industry as a DEHP substitute [12] due to its similar applications [13], and described as less harmful to reproduction when compared to other phthalates [14, 15]. DiNP can be detected in wastewater, soil [7, 16], and surface waters with concentrations ranging from 0.23 to 85 μg L−1 (see Materials and Methods)

Methods

Results

Discussion

Conclusion