Evidence-Based Network Modelling to Simulate Nucleus Pulposus Multicellular Activity in Different Nutritional and Pro-Inflammatory Environments

Abstract

Initiation of intervertebral disc degeneration is thought to be biologically driven. This reflects a process, where biochemical and mechanical stimuli affect cell activity (CA) that compromise the tissue strength over time. Experimental research enhanced our understanding about the effect of such stimuli on different CA, such as protein synthesis or mRNA expression. However, it is still unclear how cells respond to their native environment that consists of a “cocktail” of different stimuli that might locally vary. This work presents an interdisciplinary approach of experimental and in silico research to approximate Nucleus Pulposus CA within multifactorial biochemical environments. Thereby, the biochemical key stimuli glucose, pH, and the proinflammatory cytokines TNF-α and IL1β were considered that were experimentally shown to critically affect CA. To this end, a Nucleus Pulposus multicellular system was modelled. It integrated experimental findings from in vitro studies of human or bovine Nucleus Pulposus cells, to relate the individual effects of targeted stimuli to alterations in CA. Unknown stimulus-CA relationships were obtained through own experimental 3D cultures of bovine Nucleus Pulposus cells in alginate beads. Translation of experimental findings into suitable parameters for network modelling approaches was achieved thanks to a new numerical approach to estimate the individual sensitivity of a CA to each stimulus type. Hence, the effect of each stimulus type on a specific CA was assessed and integrated to approximate a multifactorial stimulus environment. Tackled CA were the mRNA expressions of Aggrecan, Collagen types I & II, MMP3, and ADAMTS4. CA was assessed for four different proinflammatory cell states; non-inflamed and inflamed for IL1β, TNF-α or both IL1β&TNF-α. Inflamed cell clusters were eventually predicted in a multicellular 3D agent-based model. Experimental results showed that glucose had no significant impact on proinflammatory cytokine or ADAMTS4 mRNA expression, whereas TNF-α caused a significant catabolic shift in most explored CA. In silico results showed that the presented methodology to estimate the sensitivity of a CA to a stimulus type importantly improved qualitative model predictions. However, more stimuli and/or further experimental knowledge need to be integrated, especially regarding predictions about the possible progression of inflammatory environments under adverse nutritional conditions. Tackling the multicellular level is a new and promising approach to estimate manifold responses of intervertebral disc cells. Such a top-down high-level network modelling approach allows to obtain information about relevant stimulus environments for a specific CA and could be shown to be suitable to tackle complex biological systems, including different proinflammatory cell states. The development of this methodology required a close interaction with experimental research. Thereby, specific experimental needs were derived from systematic in silico approaches and obtained results were directly used to enhance model predictions, which reflects a novelty in this research field. Eventually, the presented methodology provides modelling solutions suitable for multiscale approaches to contribute to a better understanding about dynamics over multiple spatial scales. Future work should focus on an amplification of the stimulus environment by integrating more key relevant stimuli, such as mechanical loading parameters, in order to better approximate native physiological environments.

Immuno-Modulatory Effects of Intervertebral Disc Cells

“…In IVD research, however, ABMs have been used only recently, though initial studies have demonstrated their value in predicting IVD cell responses in a pro-inflammatory environment. Baumgartner et al (2021c) coupled an ABM with mathematical network models (see Section “ Network Modeling ”) to investigate the relative mRNA expression of proteins and proteases in NP cells of different pro-inflammatory cell states, i.e., immunopositive for TNF-α, IL-1β, both, or none ( Baumgartner et al, 2021a ; Baumgartner et al, 2021b ). The ABM was used to visualize cell states within the NP through predicting how immunopositive cells could be arranged within a 3D environment.…”

“…The reductionist view, one gene to one disease, is not applicable to a highly multifactorial condition such as IDD, thus cell signaling pathway analysis is of high importance in order to understand the system as a whole. A mechanistic understanding of the condition could pave the way for a mechanism-based biomarker selection for the effective and personalized treatment of IDD ( Baumgartner et al, 2021b ).…”

“…Recently, a top-down network modeling approach was presented to approximate cell responses of NP cells, where the cell is considered as a “black-box” ( Baumgartner et al, 2021a ; Baumgartner et al, 2021b ). Approximations of cell responses were obtained by directly linking key relevant micro-environmental stimuli with cell responses of interest.…”

“…In the centre of the IVD, glucose levels, oxygen and pH can hence drop to 0.5 mM, 1% oxygen, and 6.5, respectively, although even lower values have also been reported ( Urban et al, 2004 ). It is not clear whether glucose deprivation can activate the expression of pro-catabolic factors in NPC ( Baumgartner et al, 2021b ). However, multiple studies have demonstrated that glucose deprivation impairs NP cell proliferation and survival as well as proteoglycan synthesis/degradation and collagen synthesis ( Bibby and Urban, 2004 ; Johnson et al, 2008 ; Chang et al, 2017 ; Saggese et al, 2020 ), and recent evidence highlights that non-coding RNAs (e.g., circ_0075062) may be involved in these processes ( Chang et al, 2021 ).…”

“…Therefore, low glucose and high lactic acid levels, but not hypoxia, contribute to the catabolic shift observed during IVD degeneration. In general, there is a clear need to increase the number of experimental studies where different microenvironmental factors are combined, including both nutritional and pro-inflammatory cues ( Baumgartner et al, 2021b ).…”