FSH prevents porcine granulosa cells from hypoxia‐induced apoptosis via activating mitophagy through the HIF‐1α‐PINK1‐Parkin pathway

Abstract

In developing follicles, the granulosa cells (GCs) live in a hypoxic environment due to the devoid of blood supply. Upon hypoxic conditions, several types of mammalian cells have been reported to undergo apoptosis. Follicle‐stimulating hormone (FSH) is known as the primary survival factor for antral follicles by preventing GCs apoptosis. Mitophagy is a type of organelle‐specific autophagy that removes damaged or stressed mitochondria to maintain cellular health. This study provides the first evidence suggesting that FSH‐mediated mitophagy protected porcine GCs from hypoxia‐induced apoptosis. Our data showed that the GCs apoptosis caused by mitochondrial pathway upon hypoxia stress was markedly attenuated after FSH treatment, which was correlated with enhanced activation of mitophagy. Interestingly, FSH also stimulated mitochondrial biogenesis as suggested by increased expression of mitochondrial transcription factor A and nuclear respiratory factor 1 during hypoxia exposure. Notably, the protein level of hypoxia inducible factor‐1α (HIF‐1α) was significantly increased in hypoxic GCs following FSH treatment, accompanied by elevated mitophagic activity and dampened apoptotic signaling. Blocking HIF‐1α inhibited mitophagy and restored hypoxia‐induced apoptosis despite FSH treatment. Importantly, FSH promoted the expression of serine/threonine kinase PTEN induced putative kinase 1 (PINK1) and the E3 ligase Parkin during hypoxia stress through a HIF‐1α dependent manner. This induced the mitophagic clearance of damaged mitochondria, hence inhibiting apoptosis by reducing cytochrome c releasing. The inhibition of HIF‐1α and/or PINK1 using inhibitor or RNAi further confirmed the role of the FSH‐HIF‐1α‐PINK1‐Parkin‐mitophagy axis in suppressing GC apoptosis under hypoxic conditions. These findings highlight a novel function of FSH in preserving GCs viability against hypoxic damage by activating HIF‐1α‐PINK1‐Parkin‐mediated mitophagy.

Melatonin Alleviates Hypoxia-Induced Apoptosis of Granulosa Cells by Reducing ROS and Activating MTNR1B–PKA–Caspase8/9 Pathway

“…Canonical apoptosis pathways mainly include the mitochondrial apoptosis pathway, death receptor apoptosis pathway, and endoplasmic reticulum apoptosis pathway. Our previous data showed that hypoxia acted through the mitochondrial pathway to induce apoptosis in GCs [30,31]. Consistently, the present study revealed that the mitochondrial pathway, as evidenced by activation of the caspase-9-caspase-3 axis, is required for hypoxiainduced apoptosis in GCs.…”

“…The avascular environment within ovarian follicles limits oxygen supply for GCs, and the hypoxia status of the GCs are gradually intensified as the follicle develops, during which a large amount of growing follicles undergo degeneration via atresia. In fact, our previous studies have demonstrated that under severe hypoxia stimulation, increased apoptosis occurred in ovarian GCs [30,31], indicating that hypoxia may be an important factor in initiating follicular atresia. Interestingly, some of the follicles retain the potential to develop toward ovulation.…”

“…Using an established in vitro hypoxic model [30,31], we verified the hypoxia state by detecting the protein level of hypoxia-inducible factor 1α (HIF-1α), which showed a marked upregulation in GCs cultured with 1% O 2 (Figure 1A). Concomitantly, the cell viability was significantly decreased after hypoxia incubation, along with an increased level of cleaved caspase-3 (Figure 1B,C).…”

Role of Mitofusins and Mitophagy in Life or Death Decisions

“…Importantly, the inverse correlation between apoptosis and mitophagy was also observed under physiological conditions involving hypoxia. During follicle development, the follicle stimulating hormone is responsible for ensuring survival of porcine granulosa cells, despite their hypoxic environment (Li C. et al, 2020). In fact, the follicle stimulating hormone induced mitophagy, thereby protecting from hypoxia-induced apoptosis.…”

“…The majority of the findings showing a direct regulation of these two processes support the idea that mitophagy occurs as a pro-survival mechanism. Prevention of mitophagy, by Parkin and/or PINK1 inhibition, or by cyclosporine A or 3-MA treatment, induced CytC release and caspase activity, stimulating apoptosis ( Tian et al, 2019 ; Kang et al, 2020 ; Li C. et al, 2020 ; Lin et al, 2020 ). Consistently, mitophagy induction via Urolithin A or FCCP treatment prevented apoptosis ( Tian et al, 2019 ; Lin et al, 2020 ).…”