Klauke H, Minor T, Vollmar B, Isselhard W, Menger M D
Institute for Experimental Medicine, University of Cologne, Cologne, 50931, Germany.
Cryobiology. 1998 Mar;36(2):108-14. doi: 10.1006/cryo.1997.2068.
Gaseous insufflation of oxygen via the venous vascular system is thought to be an useful tool for preventing anoxic tissue injury during extended time periods of ischemic preservation and for allowing for an improved recovery of organ function after transplantation. The present study aimed at the application of a noninvasive technique for monitoring effectiveness and homogeneity of gaseous areation by using an epiillumination microscopic technique for assessment of tissue nicotinamide adenine dinucleotide (NADH) fluorescence. Rat livers were flushed with and stored in University of Wisconsin solution at 4 degrees C for 48 h (n = 20). In half of the experiments (n = 10) gaseous oxygen was applied subsequent to organ harvest. Using ultraviolet-excitation high-resolution microscopy and computer-assisted image analysis liver surfaces were scanned for NADH intensity and spatial heterogeneity at 1, 24, and 48 h preservation time. Livers simply stored without aeration served as controls (n = 10). NADH intensity data were compared with corresponding data of tissue adenosine triphosphate (ATP) concentrations determined enzymatically. NADH fluorescence already differed at 1 h preservation between the two groups with significantly lower values in the aerobically stored livers. NADH fluorescence further decreased between 1 and 24 h preservation and remained low until 48 h, whereas in the anaerobically stored livers NADH fluorescence was found to be constantly high over the entire observation period. Aerobic storage resulted in rather homogeneous tissue oxygenation with an intrahepatic variation of NADH fluorescence <20%. In parallel, oxygen persufflation appropriately restored tissue ATP content within 1 to 24 h of preservation, while the simply stored livers exhibited pronounced depletion of ATP. We demonstrate for the first time that by means of retrograde gaseous oxygenation, ischemic livers can be readily and effectively oxygenated. Our study further indicates that the noninvasive microscopic analysis of tissue NADH fluorescence may be an useful tool for estimating efficiency of strategies in organ preservation.
通过静脉血管系统进行氧气吹入被认为是一种有用的工具，可用于在长时间缺血保存期间预防缺氧性组织损伤，并使移植后器官功能得到更好的恢复。本研究旨在应用一种非侵入性技术，通过落射光显微镜技术评估组织烟酰胺腺嘌呤二核苷酸（NADH）荧光，来监测气体通气的有效性和均匀性。将大鼠肝脏用威斯康星大学溶液冲洗并在4℃下保存48小时（n = 20）。在一半的实验（n = 10）中，在器官收获后施加气态氧。使用紫外线激发高分辨率显微镜和计算机辅助图像分析，在保存1、24和48小时时扫描肝脏表面的NADH强度和空间异质性。仅储存而未通气的肝脏用作对照（n = 10）。将NADH强度数据与通过酶法测定的组织三磷酸腺苷（ATP）浓度的相应数据进行比较。两组在保存1小时时NADH强度就已出现差异，有氧储存的肝脏中NADH值明显更低。在1至24小时的保存期间，NADH荧光进一步降低并一直保持在低水平直至48小时，而在无氧储存的肝脏中，在整个观察期内NADH荧光一直很高。有氧储存导致组织氧合较为均匀，肝内NADH荧光变化<20％。同时，在保存1至24小时内，持续吹入氧气可适当恢复组织ATP含量，而仅储存的肝脏则表现出明显的ATP消耗。我们首次证明，通过逆行气态氧合，缺血肝脏能够轻松且有效地实现氧合。我们的研究还表明，对组织NADH荧光进行非侵入性显微镜分析可能是评估器官保存策略效率的有用工具。
