rhEPO Upregulates the PPARγ Pathway in Long-term Cultured Primary Nerve Cells via PI3K/Akt to Delay Cell Senescence

Previous studies have confirmed that both recombinant human erythropoietin (rhEPO) and peroxisome proliferator-activated receptors gamma (PPAR gamma) activator pioglitazone can protect senescent nerve cells, and their mechanisms involve enhancing cell antioxidant capacity and reducing cell apoptosis. However, whether the PPAR gamma pathway is involved in the rhEPO anti-aging process in neuronal cells is still unclear. In this study, to explore the relationship between rhEPO and the PPAR gamma pathway at the cellular level, primary nerve cells cultured for 22 days were used to simulate the natural aging process of nerve cells. Starting on the 11th day of culture, rhEPO, LY294002, and GW9662 were added for treatment. Immunochemical methods and SA-beta-gal staining were used to observe the changes in cellular antioxidant capacity and the fraction of senescent cells. The results showed that PPAR gamma blockade retarded the effect of rhEPO on the cellular antioxidant capacity and altered the fraction of senescent cells. It was confirmed that PPAR gamma was involved in rhEPO's anti-aging process in neuronal cells. Real-time fluorescent quantitative RT-PCR, Western blotting, and immunofluorescence staining were used to observe the changes in PPAR gamma pathway-related factors in nerve cells after rhEPO treatment. The results showed that rhEPO significantly upregulated the expression of PPAR gamma coactivator-1 alpha (PGC-1 alpha), PPAR gamma, and nuclear PPAR gamma in cells but did not affect the level of phosphorylated PPARy protein, confirming that rhEPO has the ability to upregulate the PPARy pathway. PI3K/Akt and PPAR gamma pathway blockade experiments were used to explore the relationships among rhEPO, PI3K/Akt, and PPAR gamma. The results showed that after PPAR gamma blockade, rhEPO had no significant effect on the PI3K/Akt pathway-related factor p-Akt, while after PI3K/Akt blockade, rhEPO's effects on PPAR gamma-related factors (PGC-1 alpha, PPAR gamma, and nuclear PPAR gamma) were significantly decreased. It is suggested that rhEPO delays the PI3K/Akt pathway in the process of neuronal senescence, which is located upstream of PPARy regulation. In conclusion, this study confirmed that rhEPO can upregulate the expression of PGC-1 alpha and PPAR gamma in cells and the level of PPAR gamma protein in the nucleus to enhance the antioxidant capacity of cells and delay the senescence of nerve cells through the PI3K/Akt pathway. These findings will provide ideas for finding new targets for neuroprotection research and will also provide a theoretical basis and experimental evidence for rhEPO anti-aging research in neural cells.

Automated summary

This study confirmed that rhEPO can enhance the antioxidant capacity of cells and delay the senescence of nerve cells through the PI3K/Akt pathway by upregulating the expression of PGC-1 alpha and PPAR gamma.