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Surface curvature both emerges from, and influences the behavior of, living objects at length scales ranging from cell membranes to single cells to tissues and organs. The relevance of surface curvature in biology has been supported by numerous recent experimental and theoretical investigations in recent years. In this review, we first give a brief introduction to the key ideas of surface curvature in the context of biological systems and discuss the challenges that arise when measuring surface curvature. Giving an overview of the emergence of curvature in biological systems, its significance at different length scales becomes apparent. On the other hand, summarizing current findings also shows that both single cells and entire cell sheets, tissues or organisms respond to curvature by modulating their shape and their migration behavior. Finally, we address the interplay between the distribution of morphogens or micro-organisms and the emergence of curvature across length scales with examples demonstrating these key mechanistic principles of morphogenesis. Overall, this review highlights that curved interfaces are not merely a passive by-product of the chemical, biological and mechanical processes but that curvature acts also as a signal that co-determines these processes. This article is protected by copyright. All rights reserved.

INTRODUCTION: The etiology of lower back pain (LBP) is multifactorial. The cartilage endplate (CEP) is the spinal tissue receiving the least attention in scientific literature. Yet it plays a crucial role in keeping the intervertebral disc (IVD) healthy since it is the main gateway to nutrients and waste in and out of the nucleus pulposus in the avascular IVD1 . It is also suggested that imperfections and weaknesses in the CEP can be a better anticipator of pain than IVD degeneration 2 since chemical changes to the CEP are directly related to the intervertebral disc degeneration3 . Immunohisto-chemical research showed that the decline of CEP permeability due to calcification causes blood vessels and nerve fibers to infiltrate the IVD from the CEP and subchondral bone in an attempt to repair the damage and this infiltration leads to LBP3 . In this study we aim to investigate the mass transport through the CEP in and out of the IVD starting from CT images. METHODS: Given the scarcity of human CEP, a fresh bovine tail was obtained from the local slaughterhouse and 6 CEPs were harvested from it. 8 mm punches were taken from the CEPs. The samples were incubated with 20% Hexabrix, a contrast agent to visualize sulfated glycosaminoglycans. The samples were imaged on a Nanotom® M (GE nanoCT©) using in-house optimized settings for osteochondral tissues. The CT images were exported to Salome and manually curated to generate a high-quality volume mesh of the CEP. OpenFOAM® was used to perform Computational Fluid Dynamics (CFD) Simulations to quantify the mass transport through the CEP. The CEP samples were fitted into the 8 mm silicon tubes of a custom-made mesofluidic setup and fixed in place using clamps. physiological fluid (PBS) and standard growth medium were passed through the specimens separately. The specimens were left to fill then the time to pass 10 ml of solution was recorded for each specimen in the forward and reverse flows to calculate the volumetric flow rate and the permeability using Darcy’s law. RESULTS: The nCT imaging settings used resulted in a good quality image of the scanned samples. The 20% Hexabrix was unable to fully stain the CEP and a higher concentration is currently being tested. Both the CFD simulations and the in vitro diffusion tests are ongoing. Figure 1: nCT-scan of CEP with vertebrate bone attached (sagittal view) DISCUSSION & CONCLUSIONS: Combining image-based in silico modelling with in vitro experiments, as proposed in this study, will allow to quantify the fluid dynamics across the CEP, which is the gateway controlling mass transport in and out of the IVD. ACKNOWLEDGEMENTS: This project received funds from the European Commission (H2020-MSCA-ITN-ETN-2020-955735). REFERENCES: 1 Roberts S, et al. Spine. 1989;14(2):166-174. 2 Lakstins K, et al. J Orthop Res. 2021;39(9):1898-1907. 3 Yao Y, et al. Stem Cell Rep. 2016;7(2):249-262.

Low back pain is the leading cause of morbidity worldwide. Its aetiology is complex and multifactorial with degeneration of the intervertebral disc (IVD) being a key cause. Causality between infection of the IVD and its degenerative process gained great interest. In a systematic review, Granville Smith et al. 1 (2021) identified 36 articles from 34 research studies investigating bacteria in human IVDs. In 27 studies bacteria were identified. To date, all the studies detecting bacteria found Gram-positive bacteria with Cutibacterium acnes being the most abundant. However, there are too few quantitative studies investigating bacterial infection to show whether bacteria are present in vivo or represent perioperative contamination. This study aims to investigate bacterial presence within the disc and their potential influence. METHODS: Immunohistochemical staining for Gram-positive bacteria was performed on 100 human disc specimens to identify whether bacteria are present. Only samples with an intact annulus fibrosus (AF) were included to minimize the risk of contamination from adjacent tissue. Staining with a second anti Gram-positive bacteria antibody is ongoing. Nucleus pulposus (NP) cells in monolayer were stimulated with lipopolysaccharides (LPS) and peptidoglycans (PPG) (0.1-50 µg/ml) for 24, 48 and 72 hours. Factors associated with disc degeneration were analyzed by qPCR, ELISA and Luminex. Experiments with NP cells resuspended in 1.2 w/v alginate treated with 5 µg/ml and 50 µg/ml LPS respectively PPG are ongoing. RESULTS: Gram-positive bacteria were internalized by human disc cells and influenced the cell and nuclei morphology (Fig.1, a). Luminex data supported by qPCR and ELISA results shows an increase in cytokines such as IL-1β, IL-6, IL-8, TNF-α, IFN-γ, IL-4, IL-17A, matrix degrading enzymes MMP1, MMP9 and neurotrophic and angiogenic factors NGF and VEGF in treated samples compared to the untreated control, with greatest effects seen following 50 µg/ml PPG stimulation. DISCUSSION & CONCLUSIONS: This study demonstrated the presence of bacteria within human IVD tissue samples of patients undergoing spinal surgery. Whether the detected bacteria are representative for the disc microbiome or for an infection is unclear and will be further investigated. Exposure of human NP cells to the bacterial components LPS and PPG showed enhancement of several cytokines, chemokines, matrix degrading enzymes and other factors associated with disc degeneration. Demonstrating the capacity of both LPS and PPG to induce catabolism within disc cells.