Anethole reduces oxidative stress and improves in vitro survival and activation of primordial follicles

Abstract

Primordial follicles, the main source of oocytes in the ovary, are essential for the maintenance of fertility throughout the reproductive lifespan. To the best of our knowledge, there are no reports describing the effect of anethole on this important ovarian follicle population. The aim of the study was to investigate the effect of different anethole concentrations on the in vitro culture of caprine preantral follicles enclosed in ovarian tissue. Randomized ovarian fragments were fixed immediately (non-cultured treatment) or distributed into five treatments: α-MEM+ (cultured control), α-MEM+ supplemented with ascorbic acid at 50 μg/mL (AA), and anethole at 30 (AN30), 300 (AN300), or 2000 µg/mL (AN2000), for 1 or 7 days. After 7 days of culture, a significantly higher percentage of morphologically normal follicles was observed when anethole at 2000 μg/mL was used. For both culture times, a greater percentage of growing follicles was observed with the AN30 treatment compared to AA and AN2000 treatments. Anethole at 30 and 2000 µg/mL concentrations at days 1 and 7 of culture resulted in significantly larger follicular diameter than in the cultured control treatment. Anethole at 30 µg/mL concentration at day 7 showed significantly greater oocyte diameter than the other treatments, except when compared to the AN2000 treatment. At day 7 of culture, levels of reactive oxygen species (ROS) were significantly lower in the AN30 treatment than the other treatments. In conclusion, supplementation of culture medium with anethole improves survival and early follicle development at different concentrations in the caprine species.

First pregnancy after in vitro culture of early antral follicles in goats: Positive effects of anethole on follicle development and steroidogenesis

“…Thus, we suggest that anethole probably enhanced the redox balance by increasing the total antioxidant capacity at the end of the culture period. Previous studies by our group (Sá et al, 2017, 2018) showed the protective effect of anethole against oxidative damage during the in vitro culture of caprine preantral follicles.…”

“…However, the daily follicle growth rate was greater with the presence of anethole. It has been described previously that anethole stimulates the proliferation of different cell types, such as granulosa cells from caprine preantral follicles cultured in vitro (Sá et al, 2017, 2018) and mouse hepatocytes (Marshall & Caldwell, 1996). Also, it is known that antioxidants are directly related to a cascade of events leading to cell growth and differentiation (Birben, Sahiner, Sackesen, Erzurum, & Kalayci, 2012).…”

“…Recently, the role of anethole (trans‐1‐methoxy‐4‐(1‐propenyl)benzene) as an antioxidant has been highlighted (Sá et al, 2017, 2018). Anethole is the main constituent of the essential oil extracted from several plants (Marinov & Valcheva‐Kuzmanova, 2015) and has been shown to scavenge ROS in vivo (Freire, Morais, Catunda‐Junior, & Pinheiro, 2005; Leal‐Cardoso & Fonteles, 1999) and in vitro (Chainy, Manna, Chaturvedi, & Aggarwal, 2000; Sá et al, 2017).…”

Powdered coconut water (ACP 406®) as an alternative base culture medium for in vitro culture of goat preantral follicles enclosed in ovarian tissue

“…The levels of ROS were determined in the spent culture media at Day 2 and Day 6 by a spectrofluorimetric method (Loetchutinat et al, 2005;Sá et al, 2018). Briefly, culture media for all treatments were incubated with 10 µL of 2',7'-dihydrodichlorofluorescein diacetate (DCHF-DA; 1mM).…”

Estratégias para a melhoria da eficiência do cultivo folicular in vitro: Importância da suplementação do meio e estudo das alterações epigenéticas

“…Diante disso, a utilização de substâncias com capacidade antioxidante no meio de CIV pode melhorar a sobrevivência e desenvolvimento celular, por reduzir os danos causados pelo EO. Dentre essas substâncias pode-se destacar, o antioxidante anetol e o meio condicionado (MC) obtido de células-tronco mesenquimais (CTMs), os quais reduziram níveis de EROs ao serem adicionados no meio de CIV de FOPAs e FOAs (caprinos; Sá et al, 2017;Sá et al, 2018;Sá et al, 2020) e FOPAs (ovinos; Bezerra et al, 2019). Portanto, o objetivo dessa revisão é abordar as principais características e avanços do CIV folicular, expondo as suas limitações (ex.…”