BMSCs reduce rat granulosa cell apoptosis induced by cisplatin and perimenopause

Abstract

Background: The objective of this study was to evaluate the effect of bone marrow mesenchymal stem cells (BMSCs) on the apoptosis of granulosa cells (GCs) in rats.

Results: Cisplatin increased GC apoptosis from 0.59% to 13.04% in the control and cisplatin treatment groups, respectively, which was significantly reduced upon co-culture with BMSCs to 4.84%. Cisplatin treatment increased p21 and bax and decreased c-myc mRNA expression, which was reversed upon co-culture with BMSCs. As compared to young rats, increased apoptosis was observed in the perimenopausal rats (P < 0.001). After 3 months, the apoptosis rate in the BMSC group was significantly lower than that of the control group (P = 0.007).

Conclusions: BMSC therapy may protect against GC apoptosis induced by cisplatin and perimenopause. Further studies are necessary to evaluate therapeutic efficacy of BMSCs.

Figure 1

BMSC morphology and surface marker expression. BMSC morphology and differentiation was assessed by microscopy of BMSCs (A) in the first passage, (B) third passage, (C) after Von Kossa staining for visualization of osteoblasts after induction, (D) and after Oil red staining for adipocytes after induction (200×). (E) CD90, CD45, CD34, and Cd29 surface market expression was determined using flow cytometry.

Figure 2

GC morphology and FSHR expression. (A) Primary GC morphology was assessed by microscopy. (B) Intracellular expression of FSHR by GCs was observed by immunocytochemistry (400×).

Figure 3

Effects of BMSCs on GC apoptosis in vitro. The apoptosis rate of GCs in the control (A), cisplatin (B) and BMSC co-culture (C) groups.

Figure 4

Effects of BMSCs on apoptosis-related genes in GCs. Relative gene expression levels were determined using real-time PCR among the control (lanes 1), cisplatin (lanes 2), and BMSC co-culture (lanes 3) groups. (A) Representative images for each gene analyzed were shown. (B) Relative gene expressions levels by using real-time PCR. ^* indicates a significant difference from the control group; P < 0.05, ^† indicates a significant difference from the cisplatin group; P < 0.05. N = 6 for each group.

Figure 5

Effects of BMSCs in an in vivo model of perimenopause. GC apoptosis was assessed by TUNEL staining. (A) Representative images of ovarian tissues obtained from the young, BMSC, control, and estrogen groups one (top panels) and three (bottom panels) months after the intervention are shown (200×). (B) GC apoptosis rates among the control, BMSC, estrogen, and young groups after 1 and 3 months intervention. Values are presented as mean ± SD. ^* indicates a significant difference from control; P < 0.05 (Two-way ANOVA: treatment effect, P < 0.0001; time effect, P = 0.0008). N = 5 for each group.