Rapamycin prevents the intervertebral disc degeneration via inhibiting differentiation and senescence of annulus fibrosus cells.

Changhong Gao,Ying Zhang,Bin Ning,Chenglin Sang

doi:10.18632/aging.101364

Changhong Gao, Ying Zhang + Show 2 more

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https://doi.org/10.18632/aging.101364

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Abstract

The effects of bleomycin and rapamycin on cellular senescence and differentiation of rabbit annulus fibrosus stem cells (AFSCs) were investigated using a cell culture model. The results showed that bleomycin induced cellular senescence in AFSCs as evidenced by senescence-associated secretory phenotype. The morphology of AFSCs was changed from cobblestone-like cells to pancake-like cells. The senescence-associated β-galactosidase activity, the protein expression of P16 and P21, and inflammatory-related marker gene levels IL-1β, IL-6, and TNF-α were increased in bleomycin-treated AFSCs in a dose-dependent manner. Rapamycin treatment decreased the gene expression of MMP-3, MMP-13, IL-1β, IL-6, TNF-α, and protein levels of P16 and P21 in bleomycin-treated AFSCs. Furthermore, neither bleomycin nor rapamycin changed the ribosomal S6 protein level in AFSCs. However, the phosphorylation of the ribosomal S6 protein was increased in bleomycin-treated AFSCs and decreased in rapamycin-treated AFSCs. AFSCs differentiated into adipocytes, osteocytes, and chondrocytes when they were cultured with respective differentiation media. Rapamycin inhibited multi-differentiation potential of AFSCs in a concentration-dependent manner. Our findings demonstrated that mammalian target of rapamycin (mTOR) signaling affects cellular senescence, catabolic and inflammatory responses, and multi-differentiation potential, suggesting that potential treatment value of rapamycin for disc degenerative diseases, especially lower back pain.

Highlights

Lower back pain is a prevalent disc disorder that affects millions of Americans and costs billions of healthcare dollars every year
The proliferation of annulus fibrosus stem cells (AFSCs) was not enhanced by adding rapamycin into bleomycin-containing medium (Fig. 1L), the pancake-like cell numbers were decreased and the spindle-like cell numbers were increased (Fig. 1D, 1E, 1I, 1J) in rapamycin-treated AFSCs
Growth medium only (Control); (B, G) bleomycin 10 μg/ml (B‐10); (C, H) bleomycin 50 μg/ml (B‐50); (D, I) Bleomycin 50 μg/ml and rapamycin 25 nM (B‐50 + R‐25); (E, J) Bleomycin 50 μg/ml and rapamycin 50 nM (B‐50 + R‐50); (L) Population doubling time (PDT) of AFSCs grown in five different conditions for 6 days

Summary

Introduction

Lower back pain is a prevalent disc disorder that affects millions of Americans and costs billions of healthcare dollars every year. Current clinical treatments for lower back pain are largely palliative because the precise cellular and molecular mechanisms of the diseases are not clear. Advanced age is thought to be a primary risk factor for intervertebral disc disorders (IVDD) [1, 2]. Organismal aging results from time-dependent accumulation of molecular and cellular damage that leads to impaired tissue homeostasis and eventual physiological and functional decline [3]. In humans, aging is associated with increased incidence of disc pathology, including abnormal collagen and proteoglycan expression, degeneration and calcification [4, 5]. It has been reported that decreased extracellular matrix production, increased production of degrading enzymes, and increased expression of inflammatory cytokines contribute to the loss of structural integrity and accelerate IVDD [5]

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