Vara E, Arias-Díaz J, Villa N, Hernández J, García C, Ortiz P, Balibrea J L
Department of Biochemistry (Facultad de Medicina), Universidad Complutense, Madrid, Spain.
Cryobiology. 1995 Oct;32(5):422-7. doi: 10.1006/cryo.1995.1041.
Oxygen free radicals appear to be the prime cell-toxic products during cold preservation. Glutathione (GSH) seems to play a critical role in cell protection against oxidant stress. The experimental decrease of intracellular GSH in vivo may be prevented by the administration of S-adenosylmethionine (SAMe), which seems also to play an important role in preserving the structure of cell membranes. We designed our study to investigate whether the addition of SAMe to EuroCollins solution (EC) could provide a similar degree of protection as the more complex University of Wisconsin (UW) solution during cold preservation. In addition, we have investigated a possible protective action of SAMe during hepatocyte cryopreservation. Wistar rat hepatocytes (10(6) cells/ml) were stored in either EC (+/- 12 mumol/l SAMe) or UW. In parallel, hepatocytes (10(6) cells/ml) were cryopreserved in M199 culture medium (+/- SAMe) using dimethyl sulfoxide as cryoprotectant. LDH release, viability, and hepatocyte GSH and malondialdehyde (MDA) content were sequentially determined during cold preservation. There were no differences on viability or GSH and MDA content between EC+SAMe and UW stored cells, although LDH release was slightly higher in the first group. The addition of SAMe also attenuated the decrease in both viability (37 +/- 0.8 vs 53.0 +/- 7.4%, mean +/- SEM, N = 5, P < 0.05) and GSH content (13.4 +/- 15.1 vs 45.1 +/- 16.8%, mean +/- SEM, N = 5, P < 0.01), observed after thawing. Our results suggest that SAMe could be a useful additive for both cold storage and cryopreservation solutions of hepatocytes.
氧自由基似乎是冷保存过程中主要的细胞毒性产物。谷胱甘肽（GSH）在细胞对抗氧化应激的保护中似乎起着关键作用。体内细胞内GSH的实验性降低可通过给予S-腺苷甲硫氨酸（SAMe）来预防，SAMe在维持细胞膜结构方面似乎也起着重要作用。我们设计本研究以调查在冷保存期间向EuroCollins溶液（EC）中添加SAMe是否能提供与更复杂的威斯康星大学（UW）溶液相似程度的保护。此外，我们还研究了SAMe在肝细胞冷冻保存期间可能的保护作用。将Wistar大鼠肝细胞（10⁶个细胞/毫升）保存在EC（±12微摩尔/升SAMe）或UW中。同时，使用二甲基亚砜作为冷冻保护剂，将肝细胞（10⁶个细胞/毫升）在M199培养基（±SAMe）中进行冷冻保存。在冷保存期间依次测定乳酸脱氢酶（LDH）释放、活力以及肝细胞GSH和丙二醛（MDA）含量。EC + SAMe组和UW保存的细胞在活力、GSH和MDA含量方面没有差异，尽管第一组的LDH释放略高。添加SAMe也减轻了解冻后观察到的活力降低（37±0.8对53.0±7.4%，平均值±标准误，N = 5，P < 0.05）和GSH含量降低（13.4±15.1对45.1±16.8%，平均值±标准误，N = 5，P < 0.01）。我们的结果表明，SAMe对于肝细胞的冷藏和冷冻保存溶液可能是一种有用的添加剂。
