Rat Nucleus Pulposus Cells

ERS Is Involved in Compression-Induced Programmed Necrosis of NP Cells

Intervertebral disc degeneration (IVDD) is a leading cause of low back pain (LBP), causing significant economic losses. The endoplasmic reticulum (ER) and mitochondria are crucial regulators of cell death, and their involvement in IVDD has been reported. However, the specific roles of ER stress (ERS) and ER-mitochondria interaction in compression-induced programmed necrosis remain unclear. Lin's team investigated the mechanisms by which ERS and ER-mitochondria interaction contribute to programmed necrosis in nucleus pulposus (NP) cells under compression.

To confirm ERS in IVDD, they compared ERS markers in normal and degenerative discs. Degenerated discs showed higher GRP78 and CHOP expression (Fig. 1a). They then tested if compression induced ERS in vitro. NP cells under compression had increased CHOP, GRP78, and p-PERK protein levels over time (Fig. 1b). Similarly, compression raised CHOP, GRP78, and PERK mRNA levels (Fig. 1c). Electron microscopy revealed ER swelling in compressed NP cells, with unclear ER-mitochondria borders at 24 and 36 hours, indicating interaction (Fig. 1d).

Fig. 1. ERS plays a critical role in compression-induced programmed necrosis of NP cells (Lin H, Peng Y Z, et al., 2021).

Effect of Ori on Apoptosis and ECM Degeneration in TBHP-stimulated NPCs

Restoration of SIRT1/AMPK was confirmed to prevent IVDD and damage via maintaining ER and extracellular homeostasis. orientin (Ori) has been shown to upregulate SIRT1. However, the effect of Ori in nucleus pulposus cells (NPCs) is not determined. Hence, in this study, Zhang's team aim to explore the function of Ori in IVDD pathological model.

The chemical structure of Ori is shown in Fig. 2A. Rat NPCs were treated with different concentrations (0, 10, 20, 40, and 100 μM) of Ori for 12, 24, and 48 hours. A CCK-8 assay showed no significant cytotoxic effects of Ori at any concentration or time point (Fig. 2B). Further experiments revealed that 40 μM Ori was optimal for NPCs (Fig. 2C). Ori effectively reduced apoptosis in TBHP-treated NPCs, as shown by TUNEL staining (Fig. 3A, B). RT-PCR and western blot results indicated that Ori reversed TBHP-induced changes in apoptosis-related and ECM degeneration-related biomarkers (Fig. 3C-L). Immunofluorescence showed that Ori enhanced Bcl-2 and reduced MMP13 fluorescence in TBHP-treated NPCs (Fig. 3M, N). These results suggest that Ori inhibits apoptosis and ECM degeneration in TBHP-treated NPCs. To explore the mechanisms, we focused on OS and OS-mediated ER stress.

Fig. 2. The Cytotoxic Effects of Ori on NPCs (Zhang Z, Wu T, et al., 2022).

Fig. 3. The Effect of Ori on Apoptosis and ECM degeneration in TBHP-Stimulated NPCs (Zhang Z, Wu T, et al., 2022).