Bone marrow mesenchymal stem cells derived from juvenile macaques reversed ovarian ageing in elderly macaques

Abstract

Background: Female sex hormone secretion and reproductive ability decrease with ageing. Bone marrow mesenchymal stem cells (BMMSCs) have been postulated to play a key role in treating ovarian ageing.

Methods: We used macaque ovarian ageing models to observe the structural and functional changes after juvenile BMMSC treatment. Moreover, RNA-seq was used to analyse the ovarian transcriptional expression profile and key pathways through which BMMSCs reverse ovarian ageing.

Results: In the elderly macaque models, the ovaries were atrophied, the regulation ability of sex hormones was reduced, the ovarian structure was destroyed, and only local atretic follicles were observed, in contrast with young rhesus monkeys. Intravenous infusion of BMMSCs in elderly macaques increased ovarian volume, strengthened the regulation ability of sex hormones, reduced the degree of pulmonary fibrosis, inhibited apoptosis, increased density of blood vessels, and promoted follicular regeneration. In addition, the ovarian expression characteristics of ageing-related genes of the elderly treatment group reverted to that of the young control group, 1258 genes that were differentially expressed, among which 415 genes upregulated with age were downregulated, 843 genes downregulated with age were upregulated after BMMSC treatment, and the top 20 differentially expressed genes (DEGs) in the protein-protein interaction (PPI) network were significantly enriched in oocyte meiosis and progesterone-mediated oocyte maturation pathways.

Conclusion: The BMMSCs derived from juvenile macaques can reverse ovarian ageing in elderly macaques.

Keywords: BMMSCs; Macaque; Ovarian ageing.

Fig. 1

Evaluation of elderly macaques as research models of ovarian ageing. a Facial features of young macaques. b Facial features of elderly macaques. c Back features of young macaques. d Back features of elderly macaques. e Ovarian morphology of young macaques. f Ovarian morphology of elderly macaques. g HE staining of a young macaque ovary (100×). h HE staining of a young macaque ovary (400×). i HE staining of an elderly macaque ovary (100×). j HE staining of an elderly macaque ovary (400×). k Statistical analysis of ovarian organ index of young and elderly macaques. l Statistical analysis of sex hormone secretion levels in young and elderly macaques

Fig. 2

Morphology of BMMSCs. a Primary generation of BMMSCs (100×); b P4 BMMSCs (100×); c E-GFP-labelled BMMSCs (100×)

Fig. 3

Changes in ovarian structure and function after BMMSC treatment. a PECT-CT was used to observe the changes in ovarian SUVmax and volume and the CT value; white areas represent the ovarian volume, and red areas represent metabolically active regions. b Statistical analysis of CT value, volume, and SUVmax. c Changes in hFSH, hLH, Testo, Prog, PRL, E2, and AMH levels at 3, 6, and 8 months after BMMSC treatment

Fig. 4

Ovarian histopathological observation after BMMSC treatment. a The morphology of ovaries. b HE staining was performed to observe ovarian structure (100 μm). c Masson staining was performed to observe the degree of fibrosis (50 μm). d TUNEL assay was performed to analyse apoptosis (100 μm). e Immunohistochemical staining was performed to observe the density of blood vessels (50 μm). f Immunofluorescence staining was performed to track BMMSCs in the ovary (50 μm). g Statistical analyses of the degree of fibrosis, the percentage of apoptotic cells, the density of blood vessels, and the ovarian organ index (^*p < 0.05, ^**p < 0.01, and ^***p < 0.001)

Fig. 5

Transcriptome sequencing of ovarian tissues. a Cluster plot showing the 1258 genes differentially expression. b 3D-PCA analysis of ageing-related genes, light green represents the young control group, purple represents the elderly model group, and dark blue represents the elderly treatment group. c ClueGO analysis functional enrichment of 1258 genes. d Trajectory analysis the ovarian expression characteristics of 415 upregulated genes with age and BMMSC treatment, and the GO enrichment. e Trajectory analysis of the ovarian expression characteristics of 843 downregulated genes with age and BMMSC treatment, and the GO enrichment. f Cytohub screened the top20 PPI. g PPI top 20 gene function and pathway