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颗粒状猪关节软骨在经过冷冻保护剂暴露、玻璃化和组织复温后,其软骨细胞活力在组织储存中得到维持。

Chondrocyte Viability of Particulated Porcine Articular Cartilage Is Maintained in Tissue Storage After Cryoprotectant Exposure, Vitrification, and Tissue Warming.

机构信息

Department of Surgery, University of Alberta, Edmonton, AB, Canada.

Sports Medicine Centre, Department of Orthopedic Surgery, First Affiliated Hospital of Shantou University Medical College, Shantou, Guangdong, China.

出版信息

Cartilage. 2024 Jun;15(2):139-146. doi: 10.1177/19476035221118656. Epub 2023 May 6.

DOI:10.1177/19476035221118656
PMID:37148124
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11368895/
Abstract

OBJECTIVE

Vitrification of articular cartilage (AC) is a promising technique which may enable long-term tissue banking of AC allografts. We previously developed a 2-step, dual-temperature, multi-cryoprotectant agent (CPA) loading protocol to cryopreserve particulated AC (1 mm cubes). Furthermore, we also determined that the inclusion of ascorbic acid (AA) effectively mitigates CPA toxicity in cryopreserved AC. Prior to clinical translation, chondrocytes must remain viable after tissue re-warming and before transplantation. However, the effects of short-term hypothermic storage of particulated AC after vitrification and re-warming are not documented. This study evaluated the chondrocyte viability of post-vitrified particulated AC during a 7-day tissue storage period at 4 °C. We hypothesized that porcine particulated AC could be stored for up to 7 days after successful vitrification without significant loss of cell viability, and these results would be enhanced when cartilage is incubated in storage medium supplemented with clinical grade AA.

DESIGN

Three experimental groups were examined at 5 time points: a fresh control (only incubated in medium), a vitrified - AA group, and a vitrified + AA group ( = 7).

RESULTS

There was a mild decline in cell viability but both treatment groups maintained a viability of greater than 80% viable cells which is acceptable for clinical translation.

CONCLUSION

We determined that particulated AC can be stored for up to 7 days after successful vitrification without a clinically significant decline in chondrocyte viability. This information can be used to guide tissue banks regarding the implementation of AC vitrification to increase cartilage allograft availability.

摘要

目的

关节软骨(AC)的玻璃化是一种很有前途的技术,它可以使 AC 同种异体移植物的长期组织库得以实现。我们之前开发了一种两步法、双温度、多冷冻保护剂(CPA)加载方案来冷冻保存颗粒状 AC(1mm 立方体)。此外,我们还确定添加抗坏血酸(AA)可以有效减轻冷冻保存 AC 中的 CPA 毒性。在临床转化之前,软骨细胞在组织复温后和移植前必须保持活力。然而,玻璃化后颗粒状 AC 短期低温储存后的效果在再复温后并未得到记录。本研究评估了玻璃化后颗粒状 AC 在 4°C 下 7 天组织储存期间的软骨细胞活力。我们假设,成功玻璃化后,猪颗粒状 AC 可以储存长达 7 天,而不会导致细胞活力显著丧失,并且当软骨在添加临床级 AA 的储存培养基中孵育时,这些结果将得到增强。

设计

在 5 个时间点检查了三个实验组:新鲜对照组(仅在培养基中孵育)、玻璃化-AA 组和玻璃化+AA 组(n=7)。

结果

细胞活力略有下降,但两组处理组均保持超过 80%活细胞的活力,这对于临床转化是可以接受的。

结论

我们确定,成功玻璃化后,颗粒状 AC 可以储存长达 7 天,而软骨细胞活力不会出现临床上显著下降。这些信息可以用于指导组织库实施 AC 玻璃化,以增加软骨同种异体移植物的供应。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/fef014e675d3/10.1177_19476035221118656-fig3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/6115cac17a4f/10.1177_19476035221118656-fig1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/a2bc1bd08fbe/10.1177_19476035221118656-fig2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/fef014e675d3/10.1177_19476035221118656-fig3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/6115cac17a4f/10.1177_19476035221118656-fig1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/a2bc1bd08fbe/10.1177_19476035221118656-fig2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7e09/11368895/fef014e675d3/10.1177_19476035221118656-fig3.jpg

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本文引用的文献

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Biopreserv Biobank. 2022 Aug;20(4):401-408. doi: 10.1089/bio.2021.0083. Epub 2021 Oct 12.
2
Fresh Osteochondral and Chondral Allograft Preservation and Storage Media: A Systematic Review of the Literature.新鲜骨软骨和软骨同种异体移植保存与储存介质:文献系统评价
Am J Sports Med. 2022 May;50(6):1702-1716. doi: 10.1177/03635465211016832. Epub 2021 Jul 26.
3
Vitrification of particulated articular cartilage via calculated protocols.
通过计算方案对颗粒状关节软骨进行玻璃化处理。
NPJ Regen Med. 2021 Mar 19;6(1):15. doi: 10.1038/s41536-021-00123-5.
4
The effect of cryoprotectant vehicle solution on cartilage cell viability following vitrification.玻璃化冷冻后冷冻保护剂载体溶液对软骨细胞活力的影响。
Cell Tissue Bank. 2022 Mar;23(1):31-41. doi: 10.1007/s10561-021-09906-y. Epub 2021 Feb 25.
5
Vitrification of Intact Porcine Femoral Condyle Allografts Using an Optimized Approach.优化方法玻璃化完整猪股骨髁异体移植物。
Cartilage. 2021 Dec;13(2_suppl):1688S-1699S. doi: 10.1177/1947603520967077. Epub 2020 Oct 26.
6
Antioxidant additives reduce reactive oxygen species production in articular cartilage during exposure to cryoprotective agents.抗氧化添加剂可减少关节软骨在暴露于冷冻保护剂时产生的活性氧物质。
Cryobiology. 2020 Oct;96:114-121. doi: 10.1016/j.cryobiol.2020.07.008. Epub 2020 Aug 8.
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