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17β-雌二醇通过PI3K/Akt/mTOR通路抑制白介素1β诱导的大鼠椎间盘髓核细胞的凋亡

郭洪涛, 郭尔斐, 徐建杰, 张斌, 高雁冰, 武建忠^*

石家庄市第一医院脊柱外科，石家庄 050011

摘要

髓核细胞(nucleus pulposus cells, NPCs)的异常凋亡是导致椎间盘退变(intervertebral disc degeneration, IVDD)的主要原因。本研究组前期研究显示，17β-雌二醇(17β-estradiol, E2)能够通过PI3K/Akt信号通路抑制白介素1β (interleukin-1β, IL-1β)诱导的大鼠椎间盘NPCs凋亡。本研究旨在探讨PI3K/Akt途径的下游蛋白是否参与E2对NPCs凋亡的抑制作用。用胰蛋白酶消化法分离原代大鼠NPCs，采用E2和PI3K/Akt信号通路下游蛋白的不同抑制剂预处理后用IL-1β处理，用Annexin V/PI染色法检测凋亡率，用CCK-8法检测细胞活力，用细胞黏附试验检测NPCs与II型胶原的黏附能力，用Western blot检测哺乳动物雷帕霉素靶蛋白(mammalian target of Rapamycin, mTOR)、糖原合成酶激酶-3β (glycogen synthase kinase-3β, GSK-3β)和核因子κB (nuclear factor kappaB, NF-κB)磷酸化水平。结果显示，E2显著抑制IL-1β诱导的NPCs凋亡，逆转由IL-1β引起的细胞活力和黏附能力的降低，抑制IL-1β对mTOR磷酸化水平的下调作用，而雷帕霉素可以阻断E2的这些保护作用。以上结果提示，E2可能通过PI3K/Akt/mTOR信号通路抑制IL-1β诱导的NPCs凋亡。

关键词： 17β-雌二醇; 髓核细胞; mTOR; 白介素1β; 凋亡

分类号：R34

17β-estradiol inhibits interleukin-1β-induced rat nucleus pulposus cell apoptosis through the PI3K/Akt/mTOR signal pathway

GUO Hong-Tao, GUO Er-Fei, XU Jian-Jie, ZHANG Bin, GAO Yan-Bing, WU Jian-Zhong^*

Department of Spinal Surgery, Shijiazhuang first hospital, Shijiazhuang 050011, China

Abstract

The apoptosis of nucleus pulposus cells (NPCs) is the main cellular process of intervertebral disc degeneration (IVDD). Our previous studies showed that 17β-estradiol (E2) protects rat NPCs from interleukin-1β (IL-1β)-induced apoptosis via the PI3K/Akt signaling pathway. This study was aimed to investigate whether downstream proteins of PI3K/Akt pathway were involved in inhibition of E2 on NPCs’ apoptosis. Primary culture of rat NPCs was isolated by trypsin digestion. Being pretreated with E2 and different inhibitors of downstream proteins of PI3K/Akt pathway, the NPCs were treated with IL-1β. Cellular apoptosis was detected by Annexin V/PI staining. Cell viability was detected by CCK-8. Cell adhesion was evaluated by cell-collagen binding assay. Phosphorylation levels of mammalian target of Rapamycin (mTOR), glycogen synthase kinase-3β (GSK-3β) and nuclear factor κB (NF-κB) were detected by Western blot. The results showed that E2 significantly inhibited the IL-1β-induced apoptosis of NPCs, reversed the decrease of cell viability and adhesion induced by IL-1β, and inhibited the down-regulation of mTOR phosphorylation level induced by IL-1β. Rapamycin could block these protective effects of E2. These results suggest that E2 may inhibit IL-1β-induced NPCs’ apoptosis through the PI3K/Akt/mTOR signaling pathway.

Key words: 17β-estradiol; nucleus pulposus cells; mTOR; interleukin-1β; apoptosis

收稿日期：2020-03-03　　录用日期：2020-08-17

通讯作者：武建忠　　E-mail: 2085561950@qq.com

DOI: 10.13294/j.aps.2021.0002

引用本文：

郭洪涛, 郭尔斐, 徐建杰, 张斌, 高雁冰, 武建忠. 17β-雌二醇通过PI3K/Akt/mTOR通路抑制白介素1β诱导的大鼠椎间盘髓核细胞的凋亡[J]. 生理学报 2021; 73 (1): 62-68.

GUO Hong-Tao, GUO Er-Fei, XU Jian-Jie, ZHANG Bin, GAO Yan-Bing, WU Jian-Zhong. 17β-estradiol inhibits interleukin-1β-induced rat nucleus pulposus cell apoptosis through the PI3K/Akt/mTOR signal pathway. Acta Physiol Sin 2021; 73 (1): 62-68 (in Chinese with English abstract).