One strike loading organ culture model to investigate the post-traumatic disc degenerative condition

Zhou, Zhiyu; Cui, Shangbin; Du, Jie; Richards, R. Geoff; Alini, Mauro; Grad, Sibylle; Li, Zhen

doi:10.1016/j.jot.2020.08.003

Cited by 22 publications

(52 citation statements)

References 50 publications

(32 reference statements)

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“…A possible explanation for the poor metabolic activity in the AF region may be due to constant compression of the disc, where the pressure created in the NP is balanced by tensile stresses in the lamella of the AF. A recent study, providing support for this hypothesis, found a significant reduction in viable cells (lactate dehydrogenase staining) in the inner and outer AF after severe static compressive loading 95 . However, the effects of dynamic loading and fluid flow on solute transport in avascular tissues and constructs has not been fully elucidated.…”

Section: Discussionmentioning

confidence: 99%

“…A recent study, providing support for this hypothesis, found a significant reduction in viable cells (lactate dehydrogenase staining) in the inner and outer AF after severe static compressive loading. 95 However, the effects of dynamic loading and fluid flow on solute transport in avascular tissues and constructs has not been fully elucidated. A key early in vivo study looking at the effects of physiological loads and fluid transport on tracer concentrations found that fluid "pumping" during movement had an insignificant effect on small solutes such as oxygen and glucose.…”

Section: Discussionmentioning

confidence: 99%

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Investigating the physiological relevance of ex vivo disc organ culture nutrient microenvironments using in silico modeling and experimental validation

McDonnell

Buckley

2021

JOR Spine

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Background: Ex vivo disc organ culture systems have become a valuable tool for the development and pre-clinical testing of potential intervertebral disc (IVD) regeneration strategies. Bovine caudal discs have been widely selected due to their large availability and comparability to human IVDs in terms of size and biochemical composition. However, despite their extensive use, it remains to be elucidated whether their nutrient microenvironment is comparable to human degeneration.Aims: This work aims to create the first experimentally validated in silico model which can be used to predict and characterize the metabolite concentrations within ex vivo culture systems.Materials & Methods: Finite element models of cultured discs governed by previously established coupled reaction-diffusion equations were created using COMSOL Multiphysics. Experimental validation was performed by measuring oxygen, glucose and pH levels within discs cultured for 7 days, in a static compression bioreactor. Results:The in silico model was successfully validated through good agreement between the predicted and experimentally measured concentrations. For an ex vivo organ cultured in high glucose medium (4.5 g/L or 25 mM) and normoxia, a larger bovine caudal disc (Cd1-2 to Cd3-4) had a central concentration of 2.6 %O 2 , 8 mM of glucose and a pH value of 6.7, while the smallest caudal discs investigated (Cd6-7 and Cd7-8), had a central concentration of 6.5 %O 2 , 12 mM of glucose and a pH value of 6.9.Discussion: This work advances the knowledge of ex vivo disc culture microenvironments and highlights a critical need for optimization and standardization of culturing conditions. Conclusion:Ultimately, for assessment of cell-based therapies and successful clinical translation based on nutritional demands, it is imperative that the critical metabolite values within organ cultures (minimum glucose, oxygen and pH values) are physiologically relevant and comparable to the stages of human degeneration.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Investigating the physiological relevance of ex vivo disc organ culture nutrient microenvironments using in silico modeling and experimental validation

McDonnell

Buckley

2021

JOR Spine

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Section: Introductionmentioning

confidence: 99%

“…Several studies have shown that high-impact loading is a significant factor contributing to DDD [ 18 – 24 ]. Based on this finding, we successfully established a disc degeneration organ culture model via one-strike loading [ 24 ]. This model showed signs of early degeneration, including annulus fibrosus (AF) fissures, ECM degradation, glycosaminoglycan (GAG) release, and upregulated catabolic marker gene expression.…”

Section: Introductionmentioning

confidence: 99%

Programmed NP Cell Death Induced by Mitochondrial ROS in a One‐Strike Loading Disc Degeneration Organ Culture Model

Liu

Gao

et al. 2021

Oxidative Medicine and Cellular Longevity

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Increasing evidence has indicated that mitochondrial reactive oxygen species (ROS) play critical roles in mechanical stress-induced lumbar degenerative disc disease (DDD). However, the detailed underlying pathological mechanism needs further investigation. In this study, we utilized a one-strike loading disc degeneration organ culture model to explore the responses of intervertebral discs (IVDs) to mechanical stress. IVDs were subjected to a strain of 40% of the disc height for one second and then cultured under physiological loading. Mitoquinone mesylate (MitoQ) or other inhibitors were injected into the IVDs. IVDs subjected to only physiological loading culture were used as controls. Mitochondrial membrane potential was significantly depressed immediately after mechanical stress ( P < 0.01 ). The percentage of ROS-positive cells significantly increased in the first 12 hours after mechanical stress and then declined to a low level by 48 hours. Pretreatment with MitoQ or rotenone significantly decreased the proportion of ROS-positive cells ( P < 0.01 ). Nucleus pulposus (NP) cell viability was sharply reduced at 12 hours after mechanical stress and reached a stable status by 48 hours. While the levels of necroptosis- and apoptosis-related markers were significantly increased at 12 hours after mechanical stress, no significant changes were observed at day 7. Pretreatment with MitoQ increased NP cell viability and alleviated the marker changes by 12 hours after mechanical stress. Elevated mitochondrial ROS levels were also related to extracellular matrix (ECM) degeneration signs, including catabolic marker upregulation, anabolic marker downregulation, increased glycosaminoglycan (GAG) loss, IVD dynamic compressive stiffness reduction, and morphological degradation changes at the early time points after mechanical stress. Pretreatment with MitoQ alleviated some of these degenerative changes by 12 hours after mechanical stress. These changes were eliminated by day 7. Taken together, our findings demonstrate that mitochondrial ROS act as important regulators of programmed NP cell death and ECM degeneration in IVDs at early time points after mechanical stress.

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“…Previously, we have established an IVDD model using one strike loading to mimic the post‐traumatic pathological changes in whole organ cultured IVD. A single hyperphysiological mechanical compression applied to healthy bovine IVDs caused significant drop of cell viability, altered the mRNA expression in the IVD, and induced ECM degradation 19 . The present study aimed to identify the DEGs induced by one strike loading and further analyze their functions and pathways associated with the progression of IVDD by utilizing bioinformatics methods.…”

Section: Introductionmentioning

confidence: 99%

Transcriptional profiling of intervertebral disc in a post‐traumatic early degeneration organ culture model

et al. 2021

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Introduction The goal of this study is to characterize transcriptome changes and gene regulation networks in an organ culture system that mimics early post‐traumatic intervertebral disc (IVD) degeneration. Methods To mimic a traumatic insult, bovine caudal IVDs underwent one strike loading. The control group was cultured under physiological loading. At 24 hours after one strike or physiological loading, RNA was extracted from nucleus pulposus (NP) and annulus fibrosus (AF) tissue. High throughput next generation RNA sequencing was performed to identify differentially expressed genes (DEGs) between the one strike loading group and the control group. Gene Ontology (GO) functional and Kyoto Encyclopedia of Genes and Genomes analyses were performed to analyze DEGs and pathways. Protein‐protein interaction (PPI) network was analyzed with cytoscape software. DEGs were verified using qRT‐PCR. Degenerated human IVD tissue was collected for immunofluorescence staining to verify the expression of DEGs in human disc tissue. Results One strike loading resulted in significant gene expression changes compared with physiological loading. In total 253 DEGs were found in NP tissue and 208 DEGs in AF tissue. Many of the highly dysregulated genes have known functions in disc degeneration and extracellular matrix (ECM) homeostasis. ACTB, ACTG, PFN1, MYL12B in NP tissue and FGF1, SPP1 in AF tissue were verified by qRT‐PCR and immunofluorescence imaging. The identified DEGs were involved in focal adhesion, ECM‐receptor interaction, PI3K‐AKT, and cytokine‐cytokine receptor interaction pathways. Three clusters of PPI networks were identified. GO enrichment revealed that these DEGs were mainly involved in inflammatory response, the ECM and growth factor signaling and protein folding biological process. Conclusion Our study revealed different DEGs, pathways, biological process and PPI networks involved in post‐traumatic IVD degeneration. These findings will advance the understanding of the pathogenesis of IVD degeneration, and help to identify novel biomarkers for the disease diagnosis.

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One strike loading organ culture model to investigate the post-traumatic disc degenerative condition

Cited by 22 publications

References 50 publications

Investigating the physiological relevance of ex vivo disc organ culture nutrient microenvironments using in silico modeling and experimental validation

Investigating the physiological relevance of ex vivo disc organ culture nutrient microenvironments using in silico modeling and experimental validation

Programmed NP Cell Death Induced by Mitochondrial ROS in a One‐Strike Loading Disc Degeneration Organ Culture Model

Transcriptional profiling of intervertebral disc in a post‐traumatic early degeneration organ culture model

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