1 Human stem cells

Over the last ten years, extensive research has been carried out on stem cell therapy for improving ovarian function. Human amniotic mesenchymal stem cells (hAMSCs) have been shown to be able to improve ovarian function in multiple studies. For instance, Liu et al. have provided evidence showing an improved ovarian function of POF (premature ovarian failure) mice through hAMSC transplantation. Such a beneficial effect is related to an improved intraovarian microenvironment because of the promotion of follicular development and an increase in granulosa cell proliferation and secretion in the mouse ovary.¹⁰ Additionally, the secretion level of EGF and HGF from hAMSCs was higher than other growth factors which can improve the proliferation rate and more effectively inhibit the apoptosis rate of granulosa cells.¹¹ Human umbilical cord-derived mesenchymal stem cells (hUCMSCs) have been shown to improve ovarian function through secreting several paracrine factors, such as HGF, vascular endothelial growth factor (VEGF), and insulin-like growth factor 1 (IGF-1).¹² Similarly, human amniotic fluid mesenchymal stem cells (hAFMSCs) are reported to be helpful in the treatment of ovarian physiologic aging (OPA) by means of resisting DNA damage.¹³ Human bone marrow-derived stem cells (BMDSCs) produce higher numbers of preovulatory follicles, metaphase II oocytes, and 2-cell embryos in mice with chemotherapy-induced ovarian damage. Meanwhile, stem cells promote ovarian vascularization and cell proliferation and reduced apoptosis during the treatment, indicating that promoting ovarian angiogenesis is crucial for improving follicular development.¹⁴ The effects of bone marrow-derived stem cells on ovarian function in patients with DOR has been confirmed by a prospective observational pilot study. The ovarian function improves in 81.3% of DOR patients with an increase in antral follicle count of three or more follicles and/or two consecutive increases in AMH levels as success criteria after autologous stem cell ovarian transplantation (ASCOT). Meanwhile, the improvement is accompanied by a higher concentration of fibroblast growth factor-2 and thrombospondin. In addition, ASCOT increases the number of stimulable antral follicles and oocytes; however, the embryo euploidy rate was low (16.1%) during controlled ovarian stimulation.¹⁵

Exosomes derived from different stem cells also seem to be related to ovarian functions. Human adipose stem cell-derived exosome (hADSC-exo) transplantation significantly exerts better therapeutic effects on mouse ovarian function. Furthermore, hADSC-exos also significantly promoted the proliferation rate and inhibited the apoptosis of granulosa cells via the SMAD-dependent signaling pathway.¹⁶ Similarly, animal studies using a rat model show that bone marrow mesenchymal stem cell-derived exosomes prevent ovarian follicular atresia via the delivery of miR-144–5p in chemotherapy-induced ovarian failure.¹⁷ The exosomes derived from human umbilical cord mesenchymal stem cells (HucMSC-exos) are reported to stimulate primordial follicles by activating the phosphatidylinositol 3-kinase (PI3K)/mTOR signaling pathway in oocytes. Furthermore, HucMSC-exos stimulates primordial follicles by delivering functional microRNAs, such as miR-146a-5p or miR-21–5p. In addition, HucMSC-exos are capable of increasing oocyte production and enhance oocyte quality in elderly female mice.¹⁸ The effects of stem cells and their exosomes on inhibiting ovarian damage and alleviating age-related decline in fertility indicate that stem cells exert paracrine effects.

Researchers began to explore the better strategy to improve ovarian function with stem cells given that the stem cell therapy is effective. A study comparing two methods (intravenous injection or IV vs. in situ ovarian microinjection or MI) reveals that hUCMSCs IV is more effective to restore ovarian function with a lower rate of ovarian granulosa cell apoptosis accompanied by higher levels of SOD (superoxide dismutase) and Bcl2 and higher efficiency in regular cycle recovery (25–37.5% with IV treatment vs. 12.5–25% with MI treatment).¹⁹ Nevertheless, the in-depth analysis of the stem cell and identification of key molecules need further exploration to provide a new paradigm basis for cell-free treatment strategies.

2 Dehydroepiandrosterone（DHEA）

DHEA, a C19 androgenic steroid, has a pro-inflammatory immune function against cortisol. Researchers have reported a series of potential mechanisms by which DHEA modulates ovarian function, including improving follicular steroidogenesis, increasing IGF-1 (insulin-like growth factor (IGF)-1) acting as a prehormone for follicular testosterone, reducing aneuploidy, and increasing AMH levels and antral follicle count.^20,21 Also, Kuan-Hao Tsui et al. found that DHEA can restore starvation-induced reactive oxygen species (ROS) production and mitochondrial membrane potential imbalance via upregulation of cytochrome c and downregulation of BAX in mitochondria.²²

DHEA has been used in the treatment of patients with DOR during IVF in recent years.²³ A retrospective cohort study showed that DHEA supplementation increased oocyte and embryo yields, as well as cumulative pregnancy rates in patients with DOR.²⁴ However, a meta-analysis evaluated the effect of DHEA therapy in patients with DOR. The results showed that clinical pregnancy rates were increased significantly in patients who were pretreated with DHEA. In contrast, no differences were found in the number of oocytes retrieved, the cancellation rate of IVF cycles, or the miscarriage rate. Furthermore, the difference in the clinical pregnancy rate disappeared when the data were restricted to RCTs.²⁵ Taken together, DHEA might have a beneficial effect in patients with DOR, but the method of DHEA treatment and its in-depth mechanism still need further research to confirm these effects.

3 Growth hormone (GH)

GH has been used as an adjuvant in the treatment of infertility for more than 25 years.²⁶ It was initially used for patients with GH deficiency, but recent studies found that GH and GH/IGF axes might improve ovarian function.²⁷ GH and IGF modulate key signal pathways, such as the MAPK/ERK, Jak/STAT, and PI3K/Akt signaling pathway along with the subsequent effects on cell division and steroidogenesis related to ovarian function.²⁸ Medium- and high-dose recombinant human GH (rhGH) significantly increase the level of antral follicles but not AMH in aged mice. In addition, several parameters, including retrieved oocytes, MII-stage oocyte rate, ATP levels, mitochondrial membrane potential, and frequencies of homogeneous mitochondrial distribution, increase after rhGH treatment.²⁹ Clinical studies also present a higher number of grade 1 embryos (70.7% vs. 50.6%) and levels of estradiol and progesterone on the day of HCG administration in patients receiving GH. The rates of cumulative clinical pregnancy were higher in patients who received GH, yet the clinical pregnancy rates were not significantly different.^30,31 However, a meta-analysis analyzing GH application for patients with POR during their IVF treatment showed that there was no evidence for an increased live birth rate for a woman who received GH.²⁶ In this regard, the role of GH treatment for patients with DOR awaits further investigation and clarification.

4 Melatonin

Melatonin is an indoleamine produced by all cells.³² It has potent antioxidant activity and can directly scavenge free radicals without relying on receptors, acting as ROS.³³ Previous studies discovered that melatonin promote oocyte maturation, fertilization and embryonic development by protecting oocytes and other follicle cells from oxidative damage. In this regard, melatonin eliminates free radicals and reduces oxidative stress in ovarian follicles, thereby protecting oocytes and granulosa cells.^34,35 In addition, PAR (poly adp-ribose) expression and AIF nuclear translocation are significantly higher in cumulus GCs (granulosa cells) of DOR patients, indicating PARP1-dependent cell death may be associated with DOR. However, melatonin treatment effectively inhibits polyADP-ribosylation (PARylation) and blocks translocation of AIF into the nucleus, thereby reducing the risk of apoptosis in GCs.³⁶ A double-blinded, placebo-controlled clinical trial has been conducted to verify the effectiveness of melatonin. The results showed that patients who received melatonin had higher serum estradiol levels on the triggering day and increased numbers of good-quality oocytes and embryos. However, there were no differences in clinical pregnancy rate or other ART outcomes.³⁷ Therefore, the effectiveness of melatonin in the treatment of DOR still needs more clinical trials to confirm.

5 Traditional Chinese medicines

So far, many studies have worked on the influence of pivotal ingredients in Chinese herbal medicine on ovarian function, especially its important role in the treatment of DOR. Polysaccharides of Fructus corni (PFC) were reported to be antiaging molecules, and ovarian function was protected after PFC treatment, which was assessed by sex hormone levels and follicular development. Furthermore, treatment with PFC upregulated Bcl-2 and downregulated Bax and cleaved-caspase-3, suggesting that PFC inhibited apoptosis of granulosa cells in the ovaries of aging mice.³⁸ Yifuning is a traditional Chinese medicine recipe that can significantly prevent ovarian failure in aging mice, which has been shown to maintain estrous cycling, reproductive organ weights, and serum sex hormone levels. Yifuning has been shown to reduce age-induced p19, p53, p21, and Rb activity, and DNA damage by inhibiting the expression of 8-OHDG and p53 in the ovaries.³⁹ In addition, the Kuntai capsule improves damaged ovarian function via antioxidant and antiapoptosis effects.⁴⁰ A meta-analysis study comparing the effectiveness of cotreatment with Kuntai capsule and climen for patients with POF shows that cotreatment is more effective than climen alone.⁴¹ Although traditional Chinese medicine might have a favorable effect on ovarian function, further exploring its specific mechanism is still necessary to improve its application for clinical treatment.

6 Other therapeutic options

Although age-related ovarian failure cannot be reversed, many researchers are still committed to discovering effective methods to improve or preserve ovarian function. In recent years, various molecules have been found to improve ovarian function or delay ovarian aging, but the specific mechanisms are still unclear.

Rapamycin has been reported to be detrimental to follicular development and ovulation with long-term treatment. However, shortening the administration time of rapamycin to 2 weeks causes a rebound in fertility and prolongs the reproductive lifespan in aging females, which appears after short-term ovarian dysfunction. The improvement of ovarian function is accompanied by increased oocyte numbers, higher quality of oocytes, higher levels of Gdf 9 and Bmp15 and higher mitochondrial activity.⁴²

A prospective observational cohort study has shown that intraovarian platelet-rich plasma (PRP) infusion significantly improves the hormonal profile and ovarian reserve status，as well as improves pregnancy outcomes in intracytoplasmic sperm injection (ICSI) cycles in POR participants. A similar effect on improving ovarian function can be observed in perimenopausal and premature ovarian insufficiency participants.⁴³ Similarly, a study including 38 women (31–45 years old) with low ovarian reserves and who had at least two unsuccessful attempts to receive their oocytes through IVF shows laparoscopically assisted treatment administration of 0.7 ​ml 1 ​× ​10⁶ PRP into the ovary is capable to improving ovarian function based on the improvement in the levels of FSH, luteinizing hormone (LH), estradiol, and AMH, as well as the pregnancy rate.⁴⁴

Curcumin (CRC) is a constituent of the traditional medicine known as turmeric, and capsaicin (CPS) is the active ingredient of chili peppers.^45,46 CRC and CPS also have a beneficial effect on ovarian function in a cyclophosphamide-induced POF model. A significant reduction in serum levels of FSH and LH and an increase in AMH can be observed after treatment with CRC and CPS. Moreover, malonaldehyde levels are significantly reduced, glutathione levels and superoxide dismutase activity are significantly increased, and histopathological damages, such as atresia in ovarian follicles, vascular congestion, and hemorrhage around the corpus luteum, are all attenuated by the treatment with CRC and CPS.⁴⁷

The immunopotentiator chitosan oligosaccharide (COS) is the only basic amino oligosaccharide among natural sugars.⁴⁸ Multiple studies have discovered its unique role in restoring ovarian function. With increased doses of COS, the total number of follicles and every stage of follicles show progressive tendencies. Neutral red experiments revealed that the phagocytosis ability of peritoneal macrophages becomes stronger. Additionally, the levels of dynamic germ stem cell markers have a positive correlation with the levels of immune factors.⁴⁹ In the study performed by Huang et al. COS significantly increases the organ index of the ovary and immune organs, the levels of estradiol and AMH and protein expression levels of IL-2 and TNF-α in the ovary. Additionally, COS reduces the rate of follicular atresia, hence significantly promotes the proliferation of ovarian germ stem cells (OGSCs), and protects ovarian function.⁵⁰

Funding

This work was supported by the National Science Fund for Distinguished Young Scholars (No.81925013 to R.L.)

Declaration of competing interest

We declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted.