Salzer Elias, Mouser Vivian H M, Bulsink Jurgen A, Tryfonidou Marianna A, Ito Keita
Orthopaedic Biomechanics, Department of Biomedical Engineering Eindhoven University of Technology Eindhoven The Netherlands.
Department of Clinical Sciences, Faculty of Veterinary Medicine Utrecht University Utrecht The Netherlands.
JOR Spine. 2023 Jan 3;6(1):e1240. doi: 10.1002/jsp2.1240. eCollection 2023 Mar.
Nucleus pulposus (NP) cell density is orchestrated by an interplay between nutrient supply and metabolite accumulation. Physiological loading is essential for tissue homeostasis. However, dynamic loading is also believed to increase metabolic activity and could thereby interfere with cell density regulation and regenerative strategies. The aim of this study was to determine whether dynamic loading could reduce the NP cell density by interacting with its energy metabolism.
Bovine NP explants were cultured in a novel NP bioreactor with and without dynamic loading in milieus mimicking the pathophysiological or physiological NP environment. The extracellular content was evaluated biochemically and by Alcian Blue staining. Metabolic activity was determined by measuring glucose and lactate in tissue and medium supernatants. A lactate-dehydrogenase staining was performed to determine the viable cell density (VCD) in the peripheral and core regions of the NP.
The histological appearance and tissue composition of NP explants did not change in any of the groups. Glucose levels in the tissue reached critical values for cell survival (≤0.5 mM) in all groups. Lactate released into the medium was increased in the dynamically loaded compared to the unloaded groups. While the VCD was unchanged on Day 2 in all regions, it was significantly reduced in the dynamically loaded groups on Day 7 ( ≤ 0.01) in the NP core, which led to a gradient formation of VCD in the group with degenerated NP milieu and dynamic loading ( ≤ 0.05).
It was demonstrated that dynamic loading in a nutrient deprived environment similar to that during IVD degeneration can increase cell metabolism to the extent that it was associated with changes in cell viability leading to a new equilibrium in the NP core. This should be considered for cell injections and therapies that lead to cell proliferation for treatment of IVD degeneration.
髓核（NP）细胞密度由营养供应和代谢物积累之间的相互作用所调控。生理负荷对组织稳态至关重要。然而，动态负荷也被认为会增加代谢活性，从而可能干扰细胞密度调节和再生策略。本研究的目的是确定动态负荷是否会通过与其能量代谢相互作用而降低NP细胞密度。
将牛NP外植体在一种新型NP生物反应器中培养，在模拟病理生理或生理NP环境的条件下，分别进行动态负荷和无动态负荷培养。通过生化分析和阿尔辛蓝染色评估细胞外成分。通过测量组织和培养基上清液中的葡萄糖和乳酸来确定代谢活性。进行乳酸脱氢酶染色以确定NP外周和核心区域的活细胞密度（VCD）。
各实验组中NP外植体的组织学外观和组织组成均未发生变化。所有组中组织内葡萄糖水平均达到细胞存活的临界值（≤0.5 mM）。与未加载组相比，动态加载组释放到培养基中的乳酸增加。虽然所有区域在第2天VCD均未改变，但在第7天，动态加载组NP核心区域的VCD显著降低（≤0.01），这导致在退变NP环境和动态加载组中形成VCD梯度（≤0.05）。
结果表明，在类似于椎间盘退变期间的营养缺乏环境中进行动态负荷可增加细胞代谢，其程度与细胞活力变化相关，导致NP核心区域达到新的平衡。在旨在通过细胞增殖治疗椎间盘退变的细胞注射和治疗中应考虑这一点。
