Hypoxia-induced expression of zinc finger protein A20 inhibits apoptosis and attenuates inflammation in nucleus pulposus cells

Abstract

Objective To explore the effect and mechanism of zinc finger protein A20 in the nucleus pulposus cells of intervertebral disc with hypoxic conditions. Methods Rat intervertebral discs were cultured in normoxic (21% O2) and hypoxic (1% O2) conditions treated with dimethyloxalylglycine (DMOG) and divided into normoxia group, normoxia+DMOG group, hypoxia group and hypoxia+DMOG group. Histological staining and Western blotting were used to detect the histomorphological changes of the nucleus pulposus and the differences in expression of HIF-1α and A20. A20 interfering adenovirus were used to treat human nucleus pulposus cells with hypoxic and normoxic condition respectively, and divided into hypoxia+empty adenovirus group, hypoxia+A20 interference adenovirus group, normoxia+empty adenovirus group and normoxia+A20 interference adenovirus group. Flow cytometry, JC-1 detection, EdU detection and Western blotting were used to detect the differences in apoptosis, mitochondrial membrane potential, proliferation, extracellular matrix and inflammatory cytokine expression of nucleus pulposus cells. Results Compared with the normoxia group, the morphological degeneration of the other 3 groups was milder, and the protein levels of HIF-1α and A20 were significantly increased (P < 0.05); Compared with the hypoxia+empty adenovirus group, the hypoxia+A20 interference adenovirus group showed the increased protein levels of TNF-α and P-p65 in nucleus pulposus cells (P < 0.05), increased apoptosis (P < 0.05), decreased cell proliferation (P < 0.05) and decreased protein levels of extracellular matrix proteins (P < 0.05). In the normoxia+empty adenovirus group and the normoxia+A20 interference adenovirus group, there was no significant difference of the indicators mentioned above. Conclusion Hypoxia can inhibit the apoptosis and attenuate inflammation and promote the proliferation and the anabolism of extracellular matrix in nucleus pulposus cells by inducing the expression of A20 protein.

Keywords

• zinc finger protein a20
• intervertebral disc degeneration
• organ culture in vitro
• hypoxia
• nucleus pulposus cells