Effect of PI3K/AKT/HIF-1α signaling pathway on oxidative stress and apoptosis in nucleus pulposus cells.

Abstract

The aim of the study was to explore the expression of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/AKT)/hypoxia-induced factor-1 alpha (HIF-1α) signaling pathway and the apoptosis of nucleus pulposus cells (NPCs) under different oxygen concentrations to clarify the biological characteristics of NPCs and the molecular mechanism of intervertebral disc degeneration (IVDD). Normal and degenerated human NPCs were collected. Leibovitz's medium with 100 μmol/L CoCl2 was used to establish a hypoxic culture environment, and 100 μmol/L H2O2 was used to establish an oxidative stress culture environment. Third-generation NPCs were divided into 6 groups: normal NPCs + hypoxia, normal NPCs + normoxia, normal NPCs + oxidative stress, degenerated NPCs + hypoxia, degenerated NPCs + normoxia, and degenerated NPCs + oxidative stress. Normal NPCs + hypoxia was used as the experimental control group. Cell viability and proliferation were detected by using the Cell Counting Kit-8 (CCK-8) method. Cell apoptosis rate was assessed by flow cytometry, and expression levels of PI3K, AKT, and HIF-1α were determined by Real-Time-Polymerase Chain Reaction (RT-PCR) and Western blotting. The cell proliferation rate of both normal and degenerated NPCs decreased with increasing oxygen concentration. Conversely, the apoptosis rate increased as the oxygen concentration increased (p<0.05). Compared with the control group, whether the cells degenerated had a very significant impact on the apoptosis rate (p<0.001), and oxygen concentration also had a highly significant impact on both the cell proliferation rate and apoptosis rate (both p<0.001). The interaction between cell degeneration and oxygen concentration significantly affected both cell proliferation and apoptosis rates (p<0.05). Considering the expression levels of PI3K, AKT, and HIF-1α, normal NPCs had the highest levels under low oxygen concentrations, followed by oxidative stress and normoxia. In degenerated NPCs, the expression levels also decrease as the oxygen concentration increases. The PI3K/Akt/HIF-1α signaling pathway plays a significant role in inhibiting oxidative stress, antagonizing NPC apoptosis, and consequently delaying IVDD.