Reactive Oxygen Species Regulate Endoplasmic Reticulum Stress and ER-Mitochondrial Ca²⁺ Crosstalk to Promote Programmed Necrosis of Rat Nucleus Pulposus Cells under Compression

Figure 1

ERS plays a critical role in compression-induced programmed necrosis of NP cells. (a) Representative fluorescent images of GRP78 and CHOP staining in normal and degenerated discs detected by immunohistochemistry. (b) Representative western blot graph of the levels of p-PERK, GRP78, and CHOP protein expression in NP cells exposed to mechanical load for 0, 12, 24, and 36 h. (c) The mRNA levels of PERK, GRP78, and CHOP detected by RT-qPCR in NP cells under compression conditions. NS means no significant difference. The values are expressed as the from three biological replicates (, , and vs. control, ANOVA/LSD). (d) Ultrastructural observations of rat NP cell morphology by TEM (ER: white arrowheads; mitochondria: black arrowheads; mitochondria interacting with ER: red arrowheads). (e, f) Representative dot plots and quantitative analysis of PI uptake in NP cells. Cells were pretreated with 200 μM 4-PBA for 1 h and then subjected to compression for 0, 12, 24, and 36 h. The values are expressed as the from three biological replicates ( and vs. control, Student’s -test). (g) Representative western blot graphs of the levels of HMGB1 content in the media. Cells were pretreated with 200 μM 4-PBA for 1 h and then subjected to compression for 36 h. (h) Histogram for statistical analysis of the LDH leakage in compression-treated NP cells. NP cells were treated as in (g). The values are expressed as the from three biological replicates ( vs. control, ^## vs. compression alone, ANOVA/LSD).

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