Tabbaa Suzanne M, Guilak Farshid, Lemmerman Luke R, Glembotski Nicholas, D'Lima Darryl D, Wang Tong, Bugbee William D
University of California, San Francisco, San Francisco, California, USA.
Washington University, St. Louis, Missouri, USA.
Am J Sports Med. 2024 Jul;52(8):2119-2128. doi: 10.1177/03635465241252653. Epub 2024 Jun 10.
A major limitation of osteochondral allografts (OCA) is the deterioration of cartilage health associated with cell death during prolonged storage. However, little is known about the mechanisms that contribute to chondrocyte death during storage.
PURPOSE/HYPOTHESIS: This study aimed to determine whether bioactive lipid metabolites accumulate in the storage media of OCA and whether they are associated with a loss of chondrocyte viability during prolonged storage. It was hypothesized that free fatty acids (FFAs) would accumulate over time in the storage media of OCA and adversely affect cartilage health during storage.
Controlled laboratory study.
A group of 21 (n = 6-8 OCA/treatment group) fresh human hemicondylar OCA tissues and media were analyzed after 7, 28, and 68 days of prolonged cold (4°C) storage. Targeted mass spectrometry analysis was used to quantify bioactive FFAs, as well as primary (lipid hydroperoxide [ROOH]) and secondary (malondialdehyde) lipid oxidation products. Chondrocyte viability was measured using a fluorescence-based live/dead assay and confocal microscopy.
The concentration of all targeted fatty acid metabolites in storage media was significantly increased with increased cold storage time ( < .05). ROOH was significantly higher on day 28 of cold storage. No difference in secondary ROOH products in storage media was observed. Chondrocyte viability significantly declined in both the en face and the vertical cross-sectional analysis with increased cold storage time and inversely correlated with fatty acid metabolites ( < .05).
It is well established that elevated levels of certain FFAs and lipid oxidation products can alter cell function and cause cell death via lipotoxicity and other mechanisms. This work is the first to identify elevated levels of FFA metabolites and primary oxidation lipid products in the storage media from clinical OCA. The concentrations of FFA metabolites were measured at levels (>100 µM) known to induce cell death and were directly correlated with chondrocyte viability.
These findings provide important targets for understanding why cartilage health declines during cold storage, which can be used to optimize media formulations and improve graft health.
骨软骨异体移植（OCA）的一个主要局限是在长时间储存过程中，与细胞死亡相关的软骨健康状况恶化。然而，对于储存期间导致软骨细胞死亡的机制知之甚少。
目的/假设：本研究旨在确定生物活性脂质代谢产物是否在OCA的储存培养基中积累，以及它们是否与长时间储存期间软骨细胞活力的丧失有关。研究假设游离脂肪酸（FFA）会随着时间在OCA的储存培养基中积累，并在储存期间对软骨健康产生不利影响。
对照实验室研究。
一组21个（每个治疗组n = 6 - 8个OCA）新鲜人半髁状突OCA组织及培养基在4℃长时间冷藏7、28和68天后进行分析。采用靶向质谱分析法对生物活性FFA以及初级（脂质过氧化氢[ROOH]）和次级（丙二醛）脂质氧化产物进行定量。使用基于荧光的活/死检测和共聚焦显微镜测量软骨细胞活力。
随着冷藏时间延长，储存培养基中所有靶向脂肪酸代谢产物的浓度显著增加（P <.05）。冷藏第28天时ROOH显著更高。未观察到储存培养基中次级ROOH产物有差异。随着冷藏时间延长，在表面和垂直横截面分析中软骨细胞活力均显著下降，且与脂肪酸代谢产物呈负相关（P <.05）。
众所周知，某些FFA和脂质氧化产物水平升高可通过脂毒性和其他机制改变细胞功能并导致细胞死亡。本研究首次确定临床OCA储存培养基中FFA代谢产物和初级氧化脂质产物水平升高。所测量的FFA代谢产物浓度处于已知可诱导细胞死亡的水平（>100 μM），且与软骨细胞活力直接相关。
这些发现为理解冷藏期间软骨健康为何下降提供了重要靶点，可用于优化培养基配方并改善移植物健康状况。
