Effects of melatonin on follicular atresia and granulosa cell apoptosis in the porcine

Abstract

The accumulation of reactive oxygen species is detrimental to the health of the ovarian follicle. The protective, antioxidant properties of melatonin, an endogenous component of porcine follicular fluid, on apoptosis of granulosa cells were evaluated in this study. Porcine granulosa cells from medium-sized (3-5 mm), healthy follicles were cultured in serum-free conditions with melatonin (0, 0.01, 0.1, 1.0, 10, and 100 ng/mL) with or without its receptor antagonist, luzindole, followed by evaluation of apoptotic markers in the treated cells. Results revealed that endogenous, intrafollicular melatonin concentration decreased as follicular atresia progressed, whereas the percentage of apoptotic granulosa cells increased. Spontaneous apoptosis of granulosa cells, triggered by serum deprivation in vitro, was remarkably blocked by melatonin (1.0 ng/mL melatonin, 32.7 ± 0.5%, vs. control, 47.0 ± 1.0%; P < 0.05). Treatment with 1.0 ng/mL of melatonin also significantly elevated MT2, SOD1, and GPX4 while lowering FASL, CHOP, and GRP78 mRNA abundance compared to the untreated control. The anti-apoptotic effect and some changes of apoptotic-relevant genes in granulosa cells invoked by melatonin supplementation were markedly blocked by luzindole, suggesting that melatonin could prevent the apoptosis of porcine granulosa cells during follicular atresia via its membrane receptors and its free-radical-scavenging activity. These findings provide new insights into the regulatory mechanism of melatonin in follicular atresia-related functions. Mol. Reprod. Dev. 83: 692-700, 2016 © 2016 Wiley Periodicals, Inc.

The role of Melatonin receptor MTNR1A in the action of Melatonin on bovine granulosa cells

“…Further, MTNR1A is reportedly responsible for the anti‐apoptotic effects of Melatonin on ejaculated human spermatozoa (Espino et al, ), as well as the downstream Melatonin‐related activity following human Chorionic gonadotropin stimulation of female reproductive processes (He, Ma, et al, ). On the other hand, MTNR1B was reported to predominantly mediate the in vitro effect of Melatonin on porcine GC proliferation, apoptosis, and steroidogenesis (He, Deng, et al, ). Melatonin also improved egg‐laying rates by hens as a result of MTNR1B activity, leading to increased serum estradiol and decreased ovarian GNIHR (Gonadotropin‐inhibitory hormone receptor) expression (Jia et al, ).…”

“…Further, MTNR1A is reportedly responsible for the anti-apoptotic effects of Melatonin on ejaculated human spermatozoa (Espino et al, 2011), as well as the downstream Melatonin-related activity following human Chorionic gonadotropin stimulation of female reproductive processes (He, Ma, et al, 2016). On the other hand, MTNR1B was reported to predominantly mediate the in vitro effect of Melatonin on porcine GC proliferation, apoptosis, and steroidogenesis (He, Deng, et al, 2016).…”

“…In animals, Melatonin was recently demonstrated to directly influence ovarian function (Dubocovich & Markowska, ; Tamura et al, ). Our previous research showed that exogenous Melatonin significantly suppressed GCs apoptosis and stimulated progesterone production (Wang, Liu, Wu, et al, ), while a more recent study demonstrated that Melatonin prevented porcine GC apoptosis during follicular atresia via its membrane receptors and its free‐radical‐scavenging activity (He, Deng, et al, ). Some of these activities may be achieved by the down‐regulation of TP53 and up‐regulation of BCL2 and LHR gene expression, as reported in ovine GCs following thermal stress (Fu et al, ), although Melatonin may also protect the integrity of GCs by reducing intracellular ROS (Tanabe et al, ).…”

Research progress on the role of melatonin and its receptors in animal reproduction: A comprehensive review

“…() has shown that MLT/MT1 regulates luteinizing hormone (LH) and subsequently luteinization both in bovine and mice ovaries. A group of researchers reported the anti‐apoptotic property of melatonin is by its strong ability to form affinity to its both receptors 1 and 2 in bovine (Wang et al., ) and porcine (He, Deng, Jiang et al., ; He, Deng, Shi et al., ). Moreover, the combined effect of FSH and melatonin has also been explored on pre‐antral follicle growth of caprine, in vitro , and the result revealed that the interaction of FSH and melatonin was vital for integrity and growth of follicles (Rocha et al., ).…”

“…In this research, authors described that melatonin binds exactly to rat ovarian granulosa cell membranes with high affinity (K D = 83 PM; B max = 3.28fmol/mg protein). Furthermore, immunoblot analysis and anti-MT1 melatonin receptors antibody were used in the research Clemens et al (2001) its strong ability to form affinity to its both receptors 1 and 2 in bovine (Wang et al, 2012) and porcine (He, Deng, Jiang et al, 2016;. Moreover, the combined effect of FSH and melatonin has also been explored on pre-antral follicle growth of caprine, in vitro, and the result revealed that the interaction of FSH and melatonin was vital for integrity and growth of follicles (Rocha et al, 2013).…”