Mechanism underlying basic fibroblast growth factor and
insulin-like growth factor-1 effects on proliferation and apoptosis of
spermatogonial stem cells

doi:10.3969/j.issn.2095-4344.2066

Abstract

Abstract:

BACKGROUND: As a necessary regulatory factor of in vitro cell culture, and in vivo cell growth and proliferation, growth factors have attracted much attention.

OBJECTIVE: To investigate the effects of basic fibroblast growth factor (bFGF) combined with insulin-like growth factor-1 (IGF-1) on the proliferation and apoptosis of spermatogonial stem cells in mice.

METHODS: Spermatogonial stem cells were isolated and cultured from the testis of 6-8 days old male Kunming mice and were identified. Spermatogonial stem cells were inoculated into the feeder layer of embryonic fibroblasts treated with mitomycin C, and were then divided into four groups. Control group was cultured with normal DMEM medium; bFGF and IGF-1 groups were cultured with DMEM medium containing

20 μg/L bFGF and 20 μg/L IGF-1, respectively; bFGF+IGF-1 group was cultured with DMEM medium containing 20 μg/L bFGF and 20 μg/L IGF-1. The proliferation activity of spermatogonial stem cells was detected by cell counting kit-8 assay and EDU staining. The growth cycle and apoptosis of spermatogonial stem cells were detected by flow cytometry. The expression levels of PCNA, Bax and Bcl-2 proteins were detected by western blot assay.

RESULTS AND CONCLUSION: Compared with the control group, the absorbance values in the bFGF, IGF-1 and bFGF+IGF-1 groups were significantly increased. Compared with the bFGF and IGF-1 groups, the absorbance values in the bFGF+IGF-1 group were further increased (P < 0.05). EDU staining results showed the same conclusion as cell counting kit-8 assay results. The proportion of S+G₂/M phase cells in the bFGF+IGF-1 group was significantly higher than that in the other three groups (P < 0.05). The proportion in the IGF-1 and bFGF groups was significantly higher than that in control group (P < 0.05). Compared with the control group, the number of apoptotic cells in the bFGF, IGF-1 and bFGF+IGF-1 groups was decreased. Compared with the bFGF and IGF-1 groups, the number of apoptotic cells in the bFGF+IGF-1 group was further decreased. Compared with the control group, the relative expression levels of Bax protein in the bFGF, IGF-1 and bFGF+IGF-1 groups were significantly decreased (P < 0.01), and the expression levels of Bcl-2 and PCNA proteins were significantly increased (P < 0.05). Compared with the bFGF and IGF-1 groups, the relative expression level of Bax protein in the bFGF + IGF-1 group was decreased further (P < 0.01), and the relative expression levels of Bcl-2 and PCNA proteins were increased further (P < 0.05). These results indicate that bFGF and IGF-1 can promote cell proliferation and inhibit cell apoptosis by up-regulating the expression of PCNA and Bcl-2 proteins and down-regulating the expression of Bax protein. The combination of bFGF and IGF-1 can achieve favorable effects.

Key words: spermatogonial stem cells, insulin-like growth factor 1, basic fibroblast growth factor, cell proliferation, cell apoptosis

CLC Number:

Li Hong, Wu Shaohua, Qiu Mingxing, Deng Qingfu, Lei Guolin, Li Qinglong. Mechanism underlying basic fibroblast growth factor and insulin-like growth factor-1 effects on proliferation and apoptosis of spermatogonial stem cells[J]. Chinese Journal of Tissue Engineering Research, 2020, 24(19): 3017-3022.

Figures/Tables 7

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