Messier R H, Domkowski P W, Aly H M, Jones J L, Hilbert S L, Crescenzo D G, Abd-Elfattah A S, Wallace R B, Bass B L, Hopkins R A
Department of Surgery, Georgetown University Medical Center, Washington, DC 20007, USA.
Cryobiology. 1995 Jun;32(3):199-208. doi: 10.1006/cryo.1995.1019.
Preparation protocols for human cardiac valves are intended to minimize cytotoxicity because it has been thought that viable leaflet interstitial cells may enhance homograft durability. Preimplantation factors influencing the status of these cells at the time of transplantation include ischemia, disinfection, and cryopreservation freezing programs. In these experiments, adenine nucleotide quantitation was undertaken to assess metabolic consequences of preparation; preharvest ischemia served as an independent variable to examine the relationship between time of procurement (postmortem) and high-energy phosphate status of the cryopreserved leaflets at thaw. Nucleotides were measured using high-performance liquid chromatography performed on extracts of semilunar cusps from 25 cryopreserved human valves with documented ischemic times. Results indicate total adenine nucleotides (TAN; [ATP] + [ADP] + [AMP], in nmol TAN/mg leaflet protein) are higher (P < 0.05) after < 2 h of harvest ischemia (1.16 +/- 0.36) than with ischemic times of 3-6 h (undetected), 7-12 h (0.18 +/- 0.07), and 13-20 h (0.06 +/- 0.06). Depletion of ATP was similar, with many leaflets devoid of detectable levels. Net utilization of leaflet energy stores demonstrates time dependency when assayed after completed processing. However, relatively elevated catabolites, even with brief ischemia, and infrequently identified ATP, ADP, and AMP, suggest a consumption so accelerated that the following cryopreservation it is virtually independent of procurement-associated ischemia. We conclude resumption of a functional cell population obligates significant de novo phosphoanhydride boned reformation or a repopulation of dead/dying interstitial cells from a subset surviving the apparently severe rigors of valve preparation.
人类心脏瓣膜的制备方案旨在将细胞毒性降至最低，因为人们认为活瓣叶间质细胞可能会提高同种异体移植物的耐久性。移植时影响这些细胞状态的植入前因素包括缺血、消毒和冷冻保存程序。在这些实验中，进行了腺嘌呤核苷酸定量分析以评估制备的代谢后果；收获前缺血作为一个自变量，以检查获取时间（死后）与解冻时冷冻保存瓣叶的高能磷酸盐状态之间的关系。使用高效液相色谱法对25个有记录缺血时间的冷冻保存人类瓣膜的半月瓣尖提取物进行核苷酸测量。结果表明，收获后缺血时间<2小时（1.16±0.36）时，总腺嘌呤核苷酸（TAN；[ATP]+[ADP]+[AMP]，单位为nmol TAN/mg瓣叶蛋白）高于缺血时间为3 - 6小时（未检测到）、7 - 12小时（0.18±0.07）和13 - 20小时（0.06±0.06）时（P<0.05）。ATP的消耗情况相似，许多瓣叶检测不到可检测水平。在完成处理后进行测定时，瓣叶能量储存的净利用率显示出时间依赖性。然而，即使是短暂缺血，分解代谢产物相对升高，且很少能鉴定出ATP、ADP和AMP，这表明消耗加速，以至于随后的冷冻保存实际上与获取相关的缺血无关。我们得出结论，要恢复功能性细胞群体，需要大量重新形成磷酸酐键或从在瓣膜制备的明显严峻条件下存活下来的亚群中重新填充死亡/濒死的间质细胞。
