Parallel networks to predict TIMP and protease cell activity of Nucleus Pulposus cells exposed and not exposed to pro-inflammatory cytokines

Abstract

Background: Intervertebral disc (IVD) degeneration is characterized by a disruption of the balance between anabolic and catabolic cellular processes. Within the Nucleus Pulposus (NP), this involves increased levels of the pro-inflammatory cytokines Interleukin 1beta (IL1B) and Tumor Necrosis Factor (TNF) and an upregulation of the protease families MMP and ADAMTS. Primary inhibitors of those proteases are the tissue inhibitors of matrix metalloproteinases (TIMP). This work aims at contributing to a better understanding of the dynamics among proteases, TIMP and proinflammatory cytokines within the complex, multifactorial environment of the NP. Methods: The Parallel Network (PN)-Methodology was used to estimate relative mRNA expressions of TIMP1-3, MMP3 and ADAMTS4 for five simulated human activities; walking, sitting, jogging, hiking with 20 kg extra weight, and exposure to high vibration. Simulations were executed for nutrient conditions in non- and early-degenerated IVD approximations. To estimate the impact of cytokines, the PN-Methodology inferred relative protein levels for IL1B and TNF, re-integrated as secondary stimuli into the network. Results: TIMP1 and TIMP2 expression were found to be overall lower than TIMP3 exp. In absence of pro-inflammatory cytokines, MMP3 and/or ADAMTS4 expression were strongly downregulated in all conditions but vibration and hiking with extra weight. Pro-inflammatory cytokine exposure resulted in an impaired inhibition of MMP3, rather than of ADAMTS4, progressively rising with increasing nutrient deprivation. TNF mRNA was less expressed than IL1B. However, at the protein level, TNF was mainly responsible for the catabolic shift in the simulated pro-inflammatory environment. Overall, results agreed with previous experimental findings. Conclusions: The PN-Methodology successfully allowed the exploration of the relative dynamics of TIMP and protease regulations in different mechanical, nutritional, and inflammatory environments, in the NP. It shall stand for a comprehensive tool to integrate in vitro model results in IVD research and approximate NP cell activities in complex multifactorial environments.