School of Chemical, Biological and Environmental Engineering, Oregon State University, Corvallis, OR, USA.
Department of Clinical Sciences, Carlson College of Veterinary Medicine, Oregon State University, Corvallis, OR, USA.
Cryobiology. 2023 Sep;112:104558. doi: 10.1016/j.cryobiol.2023.104558. Epub 2023 Jul 13.
The ability to cryopreserve bone marrow within the vertebral body (VB) would offer significant clinical and research benefits. However, cryopreservation of large structures, such as VBs, is challenging due to mass transport limitations that prevent the effective delivery of cryoprotectants into the tissue. To overcome this challenge, we examined the potential of vacuum infiltration, along with carbonation, to increase the penetration of cryoprotectants. In particular, we hypothesized that initial exposure to high-pressure carbon dioxide gas would introduce bubbles into the tissue and that subsequent vacuum cycling would cause expansion and contraction of the bubbles, thus enhancing the transport of cryoprotectant into the tissue. Experiments were carried out using colored dye and agarose gel as a model revealing that carbonation and vacuum cycling result in a 14% increase in dye penetration compared to the atmospheric controls. Experiments were also carried out by exposing VBs isolated from human vertebrae to 40% (v/v) DMSO solution. CT imaging showed the presence of gas bubbles within the tissue pores for carbonated VBs as well as control VBs. Vacuum cycling reduced the bubble volume by more than 50%, most likely resulting in replacement of this volume with DMSO solution. However, we were unable to detect a statistically significant increase in DMSO concentration within the VBs using CT imaging. This research suggests that there may be a modest benefit to carbonation and vacuum cycling for introduction of cryoprotectants into larger structures, like VBs.
将骨髓冷冻保存在椎体（VB）内将提供重大的临床和研究效益。然而，由于质量传输限制，使冷冻保护剂无法有效进入组织，因此对大型结构（如 VB）进行冷冻保存具有挑战性。为了克服这一挑战，我们研究了真空渗透与碳化相结合，以增加冷冻保护剂渗透的潜力。特别是，我们假设最初暴露于高压二氧化碳气体会在组织中引入气泡，随后的真空循环会导致气泡的膨胀和收缩，从而增强冷冻保护剂进入组织的传输。实验使用有色染料和琼脂糖凝胶作为模型进行，结果表明，与大气对照相比，碳化和真空循环使染料渗透增加了 14%。还对从人椎骨中分离出的 VB 进行了实验，将其暴露于 40%（v/v）DMSO 溶液中。CT 成像显示，碳化 VB 和对照 VB 的组织孔隙内存在气泡。真空循环使气泡体积减少了 50%以上，很可能是用 DMSO 溶液取代了这部分体积。然而，我们无法通过 CT 成像检测到 VB 内 DMSO 浓度的统计学显著增加。这项研究表明，对于将冷冻保护剂引入 VB 等较大结构，碳化和真空循环可能会有一定的益处。
