Toll-like Receptor Activation Induces Degeneration of Human Intervertebral Discs

Emerson Krock,Derek H Rosenzweig,Jean A Ouellet,Lisbet Haglund,Daniel G Bisson,J Brooke Currie

doi:10.1038/s41598-017-17472-1

Emerson Krock, Derek H Rosenzweig + Show 4 more

Open Access

https://doi.org/10.1038/s41598-017-17472-1

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Abstract

Toll-like receptors (TLR) are activated by endogenous alarmins such as fragmented extracellular matrix compounds found in the degenerating disc. TLRs regulate cytokine, neurotrophin, and protease expression in human disc cells in vitro, and thus control key factors in disc degeneration. However, whether TLR activation leads to degenerative changes in intact human discs is unclear. Nucleus pulposus (NP) cells isolated from non-degenerating discs increase IL-1β and nerve growth factor gene expression following treatment with Pam2CSK4 (TLR2/6 agonist) but not Pam3CSK4 (TLR1/2 agonist). Challenging NP cells with Pam2CSK4 or 30 kDa fibronectin fragments (FN-f, an endogenous TLR2 and TLR4 alarmin) increased secretion of proinflammatory cytokines. We then investigated the effect of TLR activation in intact, non-degenerate, ex vivo human discs. Discs were injected with PBS, Pam2CSK4 and FN-f, and cultured for 28 days. TLR activation increased proteoglycan and ECM protein release into the culture media and decreased proteoglycan content in the NP. Proteases, including MMP3, 13 and HTRA1, are secreted at higher levels following TLR activation. In addition, proinflammatory cytokine levels, including IL-6, TNFα and IFNγ, increased following TLR activation. These results indicate that TLR activation induces degeneration in human discs. Therefore, TLRs are potential disease-modifying therapeutic targets to slow disc degeneration.

Highlights

The disc is a fibrocartilaginous tissue consisting of the central, gelatinous nucleus pulposus (NP), which is surrounded by the annulus fibrosus (AF)
We found that Toll-like receptors (TLR) activation causes degenerative changes in the Nucleus pulposus (NP) tissue, increases release of specific ECM components, and increases in proteases and sterile inflammation
Use of specific agonists will give insight into which TLR2 heterodimers are more readily activated in human discs

Summary

Introduction

The disc is a fibrocartilaginous tissue consisting of the central, gelatinous nucleus pulposus (NP), which is surrounded by the annulus fibrosus (AF). TLR4 activation in human NP and AF cells increases neurotrophin and cytokine gene expression, inconsistently[22]. These previous studies demonstrate that TLR activation increases key components of disc degeneration (proteases, cytokines and neurotrophins) in vitro, which could create a proinflammatory feed-forward loop in vivo. Due to ECM fragment and alarmin turnover during physiological and pathological matrix degradation, TLRs are potentially activated during the early stages of degeneration. It remains unclear if TLR activation is sufficient to induce degenerative changes in human discs. We found that TLR activation causes degenerative changes in the NP tissue, increases release of specific ECM components, and increases in proteases and sterile inflammation

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