17β-Estradiol Inhibites Tumor Necrosis Factor-α Induced Apoptosis of Human Nucleus Pulposus Cells via the PI3K/Akt Pathway

Background: Tumor necrosis factor-alpha (TNF-alpha) has been widely known to induce degeneration of nucleus pulposus cells (NPCs). 17 beta-estradiol (17 beta-E2) has been broadly proven for its function of suppressing cell apoptosis. The aim of this study is to explore whether 17 beta-E2 protects apoptosis of human NPCs induced by TNF-alpha via the PI3K/AKT pathway. Material/Methods: NPCs were divided into four groups: control, TNF-alpha (100 ng/mL), TNF-alpha (100 ng/mL) with pretreated 17 beta-E2 (10 um/L), TNF-alpha (100 ng/mL) with pretreated 17 beta-E2 (10 um/L) and MK2206 (10 um/L, inhibitor of the PI3K/AKT pathway). Flow cytometry was used to measure the apoptotic incidence. Inverted phase-contrast microscopy was used to accomplish the morphological observation for apoptosis of treated cells. Additionally, Cell Counting Kit 8 (CCK-8) assay was used to detected cell proliferation. Western blot and quantitative real-time PCR (qRT-PCR) were applied to explore the expression of pro-caspase-3, caspase-3/p17, cleaved PARP, PARP, Akt, and phospho-Akt (p-Akt). Results: First, inverted phase-contrast microscopy, CCK-8, and flow cytometry showed that TNF-a induced marked apoptosis, which was abolished by 17 beta-E2. Furthermore, Western blot and qRT-PCR showed that 17 beta-E2 protects TNF-alpha which can induced apoptosis by upregulating p-Akt, whereas Akt was essentially constant. Our data revealed that p-Akt expression peaked at 24 hours in a time-dependent manner (0-48 hours) after treating with TNF-alpha; and the p-Akt expression generally increased in a time-dependent manner (0-48 hours) after treating with TNF-alpha and 17 beta-E2. Conclusions: 17 beta-E2 is shown to protect NPCs against TNF-alpha induced apoptosis by upregulating p-Akt in the PI3K/AKT pathway. 17 beta-E2 generally increases expression of p-Akt.