C89 Induces Autophagy of Female Germline Stem Cells via Inhibition of the PI3K-Akt Pathway In Vitro

Abstract

Postnatal female germline stem cells (FGSCs) are a type of germline stem cell with self-renewal ability and the capacity of differentiation toward oocyte. The proliferation, differentiation, and apoptosis of FGSCs have been researched in recent years, but autophagy in FGSCs has not been explored. This study investigated the effects of the small-molecule compound 89 (C89) on FGSCs and the underlying molecular mechanism in vitro. Cytometry, Cell Counting Kit-8 (CCK8), and 5-ethynyl-2’-deoxyuridine (EdU) assay showed that the number, viability, and proliferation of FGSCs were significantly reduced in C89-treated groups (0.5, 1, and 2 µM) compared with controls. C89 had no impact on FGSC apoptosis or differentiation. However, C89 treatment induced the expression of light chain 3 beta II (LC3BII) and reduced the expression of sequestosome-1 (SQSTM1) in FGSCs, indicating that C89 induced FGSC autophagy. To investigate the mechanism of C89-induced FGSC autophagy, RNA-seq technology was used to compare the transcriptome differences between C89-treated FGSCs and controls. Bioinformatics analysis of the sequencing data indicated a potential involvement of the phosphatidylinositol 3 kinase and kinase Akt (PI3K-Akt) pathway in the effects of C89′s induction of autophagy in FGSCs. Western blot confirmed that levels of p-PI3K and p-Akt were significantly reduced in the C89- or LY294002 (PI3K inhibitor)-treated groups compared with controls. Moreover, we found cooperative functions of C89 and LY294002 in inducing FGSC autophagy through suppressing the PI3K-Akt pathway. Taken together, this research demonstrates that C89 can reduce the number, viability, and proliferation of FGSCs by inducing autophagy. Furthermore, C89 induced FGSC autophagy by inhibiting the activity of PI3K and Akt. The PI3K-Akt pathway may be a target to regulate FGSC proliferation and death.

C28-induced autophagy of female germline stem cells in vitro and its potential mechanisms

“…Many studies have provided convincing evidence that small molecular compounds affect the physiological functions of mammalian cells, such as proliferation, differentiation, autophagy, and apoptosis [13,14,29,45]. In this study, we found that C28 induced significant autophagy, and H3K27ac and ER stress might play roles in C28-induced autophagy of FGSCs in vitro.…”

“…Compound 28 (C28) synthesized from 6-formylbenzoxaborol acts as a Tb LeuRS inhibitor and can be used as a potential novel antitrypanosomal agent. Two benzoxaboroles, C89 and ZCL-082, induce FGSC autophagy in vitro, and the molecular mechanism has been preliminary elucidated [13,29]. However, the epigenetic mechanism of benzoxaborole-induced autophagy of FGSCs and the effects of C28 on FGSCs are largely unknown.…”

Fucoxanthinol from the Diatom Nitzschia Laevis Ameliorates Neuroinflammatory Responses in Lipopolysaccharide-Stimulated BV-2 Microglia

“…Akt, a serine/threonine kinase, is activated by PI3K. It has been reported that PI3K/Akt pathway is engaged in various cellular processes such as apoptosis, inflammatory responses, differentiation, and tumor angiogenesis [46][47][48]. As the upstream gene of NF-κB, Akt could modulate IκB degradation via phosphorylation of IKK in activated BV-2 microglia [49].…”

Ubiquitin-Specific-Processing Protease 7 Regulates Female Germline Stem Cell Self-Renewal Through DNA Methylation

“…Several studies have revealed the mechanism underlying FGSC self-renewal, differentiation, and autophagy [ 19 , 46 , 51 , 52 ]. Additionally, FGSCs were demonstrated to have the potential to differentiate into oocytes, which provided a new strategy to research oogenesis [ 7 , 41 , 52 ].…”

“…Comparing FGSCs and spermatogonial stem cells (SSCs), Li et al uncovered the transcript structures, genetic variants, and interaction between microRNAs and circRNAs [ 20 ]. Li et al suggested that chemical compounds play important roles in modulating FGSCs via the PI3K/Akt pathway [ 19 ]. Additionally, Ma et al revealed that Etv5 , Foxo1 , and Akt genes positively regulate FGSC self-renewal [ 27 ].…”