Rapamycin mitigates inflammation-mediated disc matrix homeostatic imbalance by inhibiting mTORC1 and inducing autophagy through Akt activation.

Background

Low back pain is a global health problem that originated mainly from intervertebral disc degeneration (IDD). Autophagy, negatively regulated by the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway, prevents metabolic and degenerative diseases by removing and recycling damaged cellular components. Despite growing evidence that autophagy occurs in the intervertebral disc, the regulation of disc cellular autophagy is still poorly understood.

Methods

Annulus fibrosus (rAF) cell cultures derived from healthy female rabbit discs were used to test the effect of autophagy inhibition or activation on disc cell fate and matrix homeostasis. Specifically, different chemical inhibitors including rapamycin, 3-methyladenine, MK-2206, and PP242 were used to modulate activities of different proteins in the PI3K/Akt/mTOR signaling pathway to assess IL-1β-induced cellular senescence, apoptosis, and matrix homeostasis in rAF cells grown under nutrient-poor culture condition.

Results

Rapamycin, an inhibitor of mTOR complex 1 (mTORC1), reduced the phosphorylation of mTOR and its effector p70/S6K in rAF cell cultures. Rapamycin also induced autophagic flux as measured by increased expression of key autophagy markers, including LC3 puncta number, LC3-II expression, and cytoplasmic HMGB1 intensity and decreased p62/SQSTM1 expression. As expected, IL-1β stimulation promoted rAF cellular senescence, apoptosis, and matrix homeostatic imbalance with enhanced aggrecanolysis and MMP-3 and MMP-13 expression. Rapamycin treatment effectively mitigated IL-1β-mediated inflammatory stress changes, but these alleviating effects of rapamycin were abrogated by chemical inhibition of Akt and mTOR complex 2 (mTORC2).

Conclusions

These findings suggest that rapamycin blunts adverse effects of inflammation on disc cells by inhibiting mTORC1 to induce autophagy through the PI3K/Akt/mTOR pathway that is dependent on Akt and mTORC2 activities. Hence, our findings identify autophagy, rapamycin, and PI3K/Akt/mTOR signaling as potential therapeutic targets for IDD treatment.

Informations de copyright

© 2024 The Authors. JOR Spine published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.

Déclaration de conflit d'intérêts

Takashi Yurube, William J. Buchser, Zhongying Zhang, Prashanta Silwal, Michael T. Lotze, and Nam V. Vo declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. James D. Kang has received research grants from Stryker and Synthes. Gwendolyn A. Sowa has received a manuscript royalty from UpToDate and a speaking honorarium from Cytonics.