To study the intervention effect of highly active mesenchymal stem cells (HA-MSCs) on the structure and function of the ovary in aging macaques.

Macaque HA-MSCs and bone marrow mesenchymal stem cells (BMSCs) were isolated and expanded from young macaque bone marrow using TeSR-E8 medium and DMEM/F12 medium containing 10% fetal bovine serum, respectively. Cell surface markers were detected by flow cytometry, and adipogenic, osteogenic, and chondrogenic differentiation capacities were identified. These two kinds of cells were then infused intravenously into ovarian senescent macaques at a dose of 1×10⁷ cells/kg, once a day for 3 consecutive days. The peripheral blood anti-Mullerian hormone (AMH) levels were measured before and 1, 2, 3 and 5 months after treatment, and the macaques were euthanized at the 5th month after treatment. The histopathological structure of the ovaries was observed by HE staining, and the degree of ovarian fibrosis alteration was evaluated by Masson staining. Immunohistochemical staining was employed to detect the expression levels of ageing-associated markers p16 and p21 in the ovaries.

HA-MSCs are short and fusiform with a high nuclear/cytoplasmic ratio and uniform morphology. BMSCs are large in size, showing typical fibroblast morphology, with few a small number of miscellaneous cells. The adipogenic, osteogenic, and chondrogenic differentiation capacities of HA-MSCs are all superior to those of BMSCs. Compared with the aged model group, the HA-MSCs treatment group showed an increase in AMH levels [ (1.85 ± 0.21) vs (0.29 ± 0.15) ng/mL] within two months post-treatment, with ovarian tissues exhibiting follicles of various stages and a decreased degree of fibrosis (all P < 0.001) . Compared to the aged model group, the BMSCs treatment group demonstrated an increase in AMH levels [ (1.26 ± 0.28) vs (0.36 ± 0.16) ng/mL] one month post-treatment. When compared with the HA-MSCs treatment group, the BMSCs treatment group showed a decrease in AMH levels [ (1.85 ± 0.21) vs (0.60 ± 0.20) ng/mL] two months after treatment. The ovarian tissues of this group revealed a small number of immature and primordial follicles, along with a reduced dreduction in fibrosis (all P < 0.001) . The elderly model group had the lowest AMH levels, with occasional atretic follicles and extensive fibrosis in the ovarian tissues. The expression levels of p16 and p21 proteins in the ovaries were highest in the elderly model group, lowest in the HA-MSCs treatment group, and intermediate in the BMSCs treatment group.

HA-MSCs can increase the reserve capacity of the aging ovary, and significantly mitigate ovarian fibrosis and the expression levels of ovarian aging-related proteins, and restart the follicle formation function. HA-MSCs transplantation therapy can be a better intervention for the treatment of ovarian aging than BMSCs.