Banking of osteochondral allografts, Part II. Preservation of chondrocyte viability during long-term storage.
Banking of osteochondral allografts, Part II. Preservation of chondrocyte viability during long-term storage.
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DOI：
10.1023/a:1023687419152
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发表时间：
2002-01-01
影响因子：
1.5
通讯作者：
Pellet, Sandor
中科院分区：
文献类型：
--
作者：
Csonge, Lajos;Bravo, Daniel;Pellet, Sandor
骨软骨同种异体移植物移植后临床结果成功的最重要因素之一是软骨的活力。冷冻保存的软骨的存活率相当差，已发表的细胞存活率为20-30%。本研究的目的是开发一种新的储存方法，提高软骨细胞的活力。尸体供体的距骨被用作模型组织，以比较冷冻保存后的人骨软骨同种异体移植物软骨活力与长期冷藏后的存活率。全层软骨穿刺活检标本已被冷冻保存，距骨被分为两组，并在+4 ℃下储存在TCM中60天，有或没有定期更换培养基。每三天对每个移植物的软骨进行活组织检查并通过MTT还原测定法测定存活率。在4degreeC储存期间，在每个储存的移植物中观察到表观软骨活力的大波动的重复模式，有或没有培养基替代。这些波动并没有出现在对照标本的新鲜或冷冻保存的人的皮肤进行了分析，在平行的软骨活检，所以活力波动似乎是一个内在的属性的软骨在这些条件下。在+4 ℃下保存60天的软骨显示出比冷冻保存的新鲜软骨（21.6 ± 1.8%）显著更高的存活率（35.2 ± 3.3%）。即使在假定有利的条件下对冷冻保存和解冻的软骨进行3天的解冻后孵育（17.7 ± 1.6%），也是如此。这些活力测定结果（反映细胞内代谢活性）通过荧光染料混合物SYTO-16和碘化丙啶得到证实。这些数据表明，长期储存的冷冻软骨似乎保持比冷冻保存的软骨更高的生存力，长达60天，也许超过60天的存储。培养基更换组和未更换组之间无活力指数差异。尽管结果异常，但在一个个体病例中，在+4 ℃下储存60天后，在距骨软骨中可检测到超过65%的活细胞。
One of the most important factors concerning the successful clinical outcome after transplantation of osteochondral allografts is the viability of the cartilage. The viability of cryopreserved cartilage is quite poor, 20-30% cell survival has been published. The purpose of this study was to develop a new storage method which improves the chondrocyte viability. The talus of cadaveric donors was used as a model tissue to compare human osteochondral allograft cartilage viability following cryopreservation with that remaining after prolonged refrigerated storage. Full-thickness cartilage punch biopsies had been cryopreserved, and tali were divided into two matched groups and stored in TCM for 60 days at +4degreeC, either with or without regular medium replacement. The cartilage of each graft was biopsied and assayed for viability on every third day by the MTT reduction assay. During 4degreeC storage, a recurring pattern of large fluctuations in apparent cartilage viability was observed in every stored graft, with or without medium replacement. These fluctuations did not appear in control specimens of either fresh or cryopreserved human skin that were assayed in parallel with the cartilage biopsies, so the viability fluctuation seems an intrinsic property of the cartilage in these conditions. Cartilage stored for 60 days at +4degreeC showed significantly higher viability (35.2+-3.3%) than fresh cartilage that had been cryopreserved (21.6+-1.8%). This was true even when cryopreserved and thawed cartilage was subjected to a 3 day post thaw incubation under presumably favorable conditions (17.7+-1.6%). These viability assay results, (reflective of intracellular metabolic activity), were corroborated by the fluorescent dye mixture SYTO-16 and propidium iodide. The data indicate that long-term stored refrigerated cartilage appears to retain a viability higher than that of cryopreserved cartilage for up to and perhaps beyond 60 days of storage. There was no viability index difference between the medium replaced and non-replaced groups. Although an exceptional result, in one individual case, more than 65% viable cells could be detected in the talar cartilage after 60 days storage at +4degreeC.