As one of the important components of follicles, ovarian granulosa cells play an important role in signal transduction and nutritional support for oocytes. The decrease of the number of GCs and their function damage may lead to the decline of oocyte quality, and eventually the decline of ovarian function, which could cause female infertility.
The oocytes show different sensitivity to oxidative stress at different stages of follicle development . It is known that when the dominant follicle enters the first meiosis, a certain degree of OS is present to maintain a high metabolic level, while the subsequent process requires low OS to avoid cell damage . Compared with somatic cells, GCs are always at a high metabolic level during the entire process of follicular development, while the oxidation and antioxidant systems in the follicular microenvironment maintain a dynamic balance to meet the needs of oocytes. When this system is disrupted by stress stimuli, OS damage and reduced local repair capability will damage the various biomolecules in the follicular microenvironment. A large number of follicle atresia causes an irreversible decline in ovarian function and eventually leads to premature ovarian failure. Some studies have also found that the high level of ROS in follicular fluid is associated with poor follicular quality, dysontogenesis, and failure of in vitro fertilization-embryo transfer (IVF-ET). Moreover, the increase of ROS in follicular fluid was also found in common female reproductive diseases such as PCOS and EMT, indicating that ROS may be correlated with the occurrence and development of ovarian disorders. Therefore, it is important to protect ovarian granulosa cells from OS damage for female reproductive health.
L-carnitine is an amino acid analogue naturally occurring in the human body. It can promote lipid metabolism and protect plasma and mitochondrial membrane from damage by lipid peroxidation. LC has been widely used as an antioxidant and studies have found that the addition of L-carnitine in sperm culture improves sperm motility . Addition of LC in the oocyte culture can improve oocyte quality, which may be related to the reduction of GCs apoptosis and improvement of mitochondrial function . It is believed that the supplement of LC is beneficial to alleviate the delayed embryonic development, high DNA fragmentation and abnormal blastocyst development after long-term culture mediated by the increased ROS. Therefore, treatment with LC may have the potential to improve the quality of follicles and the pregnancy outcome of IVF-ET.
Oxidative stress injury of granulosa cells can lead to imbalanced follicular microenvironment, reduced function of granulosa cells, and ovarian aging . In this study, we found that LC can effectively alleviate the generation of excess oxygen free radicals induced by H2O2 and maintain the redox balance. We also found the level of MDA, the end-product of lipid peroxidation, was significantly reduced in GCs after LC treatment, indicating that LC alleviated the cell damage caused by OS and mitochondrial oxidative respiratory chain. The SOD activity and GSH content were measured and the results indicated that LC pretreatment improved the antioxidant activity and reduced the consumption of antioxidant, which is helpful to maintain the dynamic balance of redox in the follicular microenvironment.
Damaging of macromolecules such as proteins and DNA by OS will eventually lead to apoptosis of GCs . We observed that LC pretreatment can effectively prevent the formation of apoptotic bodies, mitochondrial swelling and vacuolation, nuclear membrane blurring and other phenomena induced by H2O2. The protective effect of LC was confirmed by fluorescence detection of ΔΨml and cell apoptosis study. Moreover, it is well-known that there is a significant correlation between the FSHR expression on ovarian GCs and ovarian activity . Some studies have proposed that OS can change FSH activity by regulating the expression of P450 aromatase . Our results confirmed that the expression level of FSHR in the OS group was significantly decreased compared with the control group. We found that the expression of FSHR on GCs in the presence of H2O2 was significantly increased after being pretreated with LC, further demonstrating the protective effect of LC.
In conclusion, LC was shown to have high protective effect from the injury of ovarian GCs induced by H2O2. Therefore, LC treatment has the potential to reduce OS injury of GCs, decrease GCs apoptosis, and provide better nutritional support for oocytes to improve the quality of follicles.