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Published on: December 16, 2016
Hypoxia-induced inhibition of cell proliferation mediated by TFRC in mouse spermatogonial stem cells
Lin Qian1, Yunchao Zhang1,2, Xu Guo3
1Department of Andrology, the Affiliated Hospital of Qingdao University, Qingdao, Shandong Province 266000, China.
Context:
Hypoxia could have an impact on spermatogenesis. However, the mechanism by which hypoxia affects spermatogonial stem cells (SSC) is still unknown.
Aims:
This study aims to elucidate the impact of hypoxia on the proliferation of mouse SSC and explore the underlying mechanism.
Methods:
We established the hypoxia cultured C18-4 cell model using CoCl2. Cell counting kit-8 and 5-ethynyl-2'-deoxyuridine assays were used to assess C18-4 cells proliferation. RNA sequencing was performed to explore the differentially expressed genes. Western blotting and real-time quantitative polymerase chain reaction were used to detect the expression of hypoxia-inducible factor-1α and iron metabolism-related proteins. The small interfering RNA assay was used to knock down the expression of transferrin receptor complex (TFRC) in hypoxia cultured C18-4 cells.
Key Results:
We found that CoCl2 decreased the proliferation of C18-4 cells. Based on the RNA sequencing, we found that iron metabolism imbalance and ferroptosis might play crucial roles in the hypoxia-induced injury to C18-4 cells. Western blotting and real-time quantitative polymerase chain reaction analyses detected the upregulation of TFRC, ferritin heavy chain, ferritin light chain and ACSL4, as well as the downregulation of ferroportin and glutathione peroxidase 4 in hypoxia cultured C18-4 cells. Notably, knock down of Tfrc restored C18-4 cell proliferation, which had been inhibited by CoCl2.
Conclusions:
This study demonstrates that hypoxia inhibits C18-4 cell proliferation through the effects of TFRC on iron metabolism and ferroptosis.
Implications:
The hypoxia could induce iron metabolism imbalance and ferroptosis in SSC. Regulating the expression of TFRC could protect SSC from hypoxic injury.
Insights
Hypoxia inhibits mouse spermatogonial stem cell (SSC) proliferation by disrupting iron metabolism and inducing ferroptosis. Targeting the transferrin receptor complex (TFRC) can protect SSC from hypoxic damage.
Area of Science:
- Reproductive Biology
- Cellular Biology
- Molecular Biology
Background:
- Hypoxia (low oxygen) is known to affect spermatogenesis.
- The precise mechanisms by which hypoxia impacts spermatogonial stem cells (SSCs) remain unclear.
Purpose of the Study:
- To investigate the effects of hypoxia on mouse SSC proliferation.
- To elucidate the underlying molecular mechanisms, focusing on iron metabolism and ferroptosis.
Main Methods:
- Established a hypoxia-induced C18-4 mouse SSC model using cobalt chloride (CoCl2).
- Assessed cell proliferation using CCK-8 and EdU assays.
- Utilized RNA sequencing to identify differentially expressed genes.
- Analyzed protein expression of key factors including hypoxia-inducible factor-1α (HIF-1α) and iron metabolism regulators via Western blotting and qRT-PCR.
- Investigated the role of the transferrin receptor complex (TFRC) using small interfering RNA (siRNA) knockdown.
Main Results:
- Hypoxia significantly decreased C18-4 cell proliferation.
- RNA sequencing revealed that iron metabolism imbalance and ferroptosis are implicated in hypoxia-induced SSC injury.
- Upregulation of TFRC, ferritin, and ACSL4, alongside downregulation of ferroportin and glutathione peroxidase 4, was observed under hypoxic conditions.
- Knockdown of TFRC expression rescued the proliferation of hypoxia-inhibited C18-4 cells.
Conclusions:
- Hypoxia inhibits SSC proliferation by altering iron metabolism and inducing ferroptosis, mediated by TFRC.
- Modulating TFRC expression offers a potential strategy to protect SSCs from hypoxia-induced damage.
