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. 2019 Feb 22;39(2):BSR20182375.
doi: 10.1042/BSR20182375. Print 2019 Feb 28.

Responses of apoptosis and matrix metabolism of annulus fibrosus cells to different magnitudes of mechanical tension in vitro

Yanhai Jiang  1 , Lianqiang Fu  2 , Yeliang Song  2
Affiliations

Responses of apoptosis and matrix metabolism of annulus fibrosus cells to different magnitudes of mechanical tension in vitro

Yanhai Jiang et al. Biosci Rep. .

Retraction in

Abstract

Background: Annulus fibrosus (AF) is important to confine disc nucleus pulposus (NP) tissue during mechanical load experience. However, the knowledge on AF cell biology under mechanical load is much limited compared with disc NP.

Objective: The present study aimed to investigate responses of apoptosis and matrix metabolism of AF cells to different magnitudes of mechanical tension in vitro Methods: Rat AF cells were subjected to different magnitudes (5, 10, and 20% elongations at a frequency of 1.0 Hz for 6 h per day) of mechanical tension for 7 days. Control AF cells were cultured without mechanical tension. Cell apoptosis ratio, caspase-3 activity, gene/protein expression of apoptosis-related molecules (Bcl-2, Bax, caspase-3/cleaved caspase-3 and cleaved PARP), matrix macromolecules (aggrecan and collagen I) and matrix metabolism-related enzymes (TIMP-1, TIMP-3, MMP-3, and ADAMTS-4) were analyzed.

Results: Compared with 5% tension group and control group, 10 and 20% tension groups significantly increased apoptosis ratio, caspase-3 activity, up-regulated gene/protein expression of Bax, caspase-3/cleaved caspase-3, cleaved PARP, MMP-3, and ADAMTS-4, whereas down-regulated gene/protein expression of Bcl-2, aggrecan, collagen I, TIMP-1, and TIMP-3. No significant difference was found in these parameters apart from Bcl-2 expression between the control group and 5% tension group.

Conclusion: High mechanical tension promotes AF cell apoptosis and suppresses AF matrix synthesis compared with low mechanical tension. The present study indirectly indicates how mechanical overload induces disc degeneration through affecting AF biology.

Keywords: annulus fibrosus; cell apoptosis; intervertebral disc degeneration; matrix; mechanical tension.

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Conflict of interest statement

The authors declare that there are no competing interests associated with the manuscript.

Figures

Figure 1
Figure 1. Cell apoptosis analysis
AF cell apoptosis ratio under different magnitudes (5, 10, and 20% elongations) of mechanical tension was analyzed by flow cytometry. Data are presented as mean ± S.D. (n=3). *: Indicates a statistical difference (P<0.05) between two groups.
Figure 2
Figure 2. Caspase-3 activity analysis
Caspase-3 activity of AF cells under different magnitudes (5, 10, and 20% elongations) of mechanical tension was analyzed. Data are presented as mean ± S.D. (n=3). *: Indicates a statistical difference (P<0.05) between two groups.
Figure 3
Figure 3. Expression of apoptosis-related molecules
Gene/protein expression of Bcl-2, Bax, caspase-3/cleaved caspase-3, and cleaved PARP in AF cells under different magnitudes (5, 10, and 20% elongations) of mechanical tension was analyzed. (A) Real-time PCR analysis. (B) Western blot analysis. Data are presented as mean ± S.D. (n=3). *: Indicates a statistical difference (P<0.05) between two groups.
Figure 4
Figure 4. Expression of matrix macromolecules
Gene and protein expression of aggrecan and collagen I in AF cells under different magnitudes (5, 10, and 20% elongations) of mechanical tension was analyzed. (A) Real-time PCR analysis. (B) Western blot analysis. Data are presented as mean ± S.D. (n=3). *: Indicates a statistical difference (P<0.05) between two groups.
Figure 5
Figure 5. Expression of matrix metabolism-related enzymes
Gene expression of TIMP-1, TIMP-3, MMP-3, and ADAMTS-4 in AF cells under different magnitudes (5, 10, and 20% elongations) of mechanical tension was analyzed by real-time PCR. Data are presented as mean ± S.D. (n=3). *: Indicates a statistical difference (P<0.05) between two groups.
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