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Spine (Phila Pa 1976). 2019 May 15;44(10):E585-E595. doi: 10.1097/BRS.0000000000002930.

Mitochondrial Pathway Is Involved in Advanced Glycation End Products-Induced Apoptosis of Rabbit Annulus Fibrosus Cells.

Author information

1
Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
2
Department of Orthopaedic Surgery, Yan-Tai-Shan Hospital, Yantai, China.
3
Collaborative Innovation Center for Biomedical Engineering, Wuhan National Laboratory for Optoelectronics-Huazhong University of Science and Technology, Wuhan, China.
4
Dongfeng Honda Automobile Co., Wuhan, Hubei, China.
5
Department of Orthopaedic Surgery, The First People's Hospital of Jingmen, Jingmen, Hubei, China.

Abstract

STUDY DESIGN:

Experimental study.

OBJECTIVE:

The purposes of this study were to evaluate whether advanced glycation end-products (AGEs) induce annulus fibrosus (AF) cell apoptosis and further to explore the mechanism by which this process occurs.

SUMMARY OF BACKGROUND DATA:

Recent studies revealed that AGEs accumulation is considered an important factor in diabetic intervertebral disc (IVD) degeneration. However, the effect of AGEs on intervertebral disc remains unclear.

METHODS:

AF cells were treated with various concentrations of AGEs for 3 days. Cell viability and cell proliferation were measured by Cell Counting Kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays, respectively. Cell apoptosis was examined by Annexin V/PI apoptosis detection kit and Hoechst 33342. The expression of apoptosis-related proteins, including Bax, Bcl-2, cytochrome c, caspase-3, and caspase-9, was detected by western blotting. In addition, Bax and Bcl-2 mRNA expression levels were detected by real-time PCR (RT-PCR). Mitochondrial membrane potential (MMP) and intracellular reactive oxygen species (ROS) production of AF cell were examined by 5,5',6,6' -Tetrachloro-1,1',3,3'- tetraethyl-imidacarbocyanine iodide (JC-1) staining and 2',7'-Dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescent probes, respectively.

RESULTS:

Our results indicated that AGEs had inhibitory effects on AF cell proliferation and induced AF cell apoptosis. The molecular data showed that AGEs significantly up-regulated Bax expression and inhibited Bcl-2 expression. In addition, AGEs increased the release of cytochrome c into the cytosol and enhanced caspase-9 and caspase-3 activation. Moreover, treatment with AGEs resulted in a decrease in MMP and the accumulation of intracellular ROS in AF cells. The antioxidant N-acetyl-L-cysteine (NAC) significantly reversed AGE-induced MMP decrease and AF cell apoptosis.

CONCLUSION:

These results suggested that AGEs induce rabbit AF cell apoptosis and mitochondrial pathway may be involved in AGEs-mediated cell apoptosis, which may provide a theoretical basis for diabetic IVD degeneration.

LEVEL OF EVIDENCE:

N/A.

PMID:
30407277
PMCID:
PMC6504123
DOI:
10.1097/BRS.0000000000002930
[Indexed for MEDLINE]
Free PMC Article

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