Verhoven Bret M, Karim Aos S, Bath Natalie M, Sarabia Fahl Carol J, Wilson Nancy A, Redfield Robert R, Fahl William E
Division of Organ Transplant, Department of Surgery, University of Wisconsin-Madison, Madison, WI.
Wisconsin Institutes for Medical Research, Department of Oncology, University of Wisconsin-Madison, Madison, WI.
Transplant Direct. 2020 Jul 15;6(8):e578. doi: 10.1097/TXD.0000000000001032. eCollection 2020 Aug.
Ischemia-reperfusion injury, including injury from warm- and cold-ischemia (CI) organ storage, remains a significant problem for all solid organ transplants. Suppressing CI damage would reduce delayed graft function and increase the donor organ pool size. PrC-210 has demonstrated superior prevention of damage in several preclinical studies as an immediate-acting free-radical scavenger. Here, we describe its profound efficacy in suppressing CI injury in a rat kidney model.
Kidneys in 300 gm Sprague-Dawley rats were perfused in situ with UW solution with or without added PrC-210 and then stored at 4°C in the same solution for 0 to 48 hours. When procured, kidney-activated caspase-3 level (a marker of cell death) was measured, and direct histological analysis of kidneys was performed to assess PrC-210 protective efficacy. In vitro analyses of PrC-210-conferred protection to isolated rat kidneys or naked DNA were also performed.
A single 15 seconds in situ perfusion of kidneys with 20 mmol/L PrC-210 in UW solution resulted in significant reductions in (1) 30-hour CI-induced kidney-activated caspase level ( < 0.0001); activated caspase was reduced to levels not significantly different than control activated caspase levels seen in unperturbed kidneys, (2) 30-hour CI-induced renal Tubular Injury Scores ( = 0.0004) where brush border and tubular necrosis were markedly reduced, (3) PrC-210 conferred 100% protection against ·OH damage to naked DNA and isolated kidney mitochondria while current UW solution antioxidants were without protective effect.
A single PrC-210-UW solution perfusion of rat kidneys upon removal from the rat profoundly reduced caspase and renal tubular injury in kidneys exposed to 30 hours of CI organ storage. These findings support further development of the PrC-210 molecule to suppress or to prevent ischemia-reperfusion injury in organ transplant and other ischemia-reperfusion injury settings.
缺血再灌注损伤，包括热缺血和冷缺血（CI）器官保存引起的损伤，仍然是所有实体器官移植面临的重大问题。抑制CI损伤将减少移植器官延迟功能，并增加供体器官库规模。在多项临床前研究中，PrC-210作为一种速效自由基清除剂，已证明对损伤具有卓越的预防作用。在此，我们描述了其在大鼠肾脏模型中抑制CI损伤的显著疗效。
对体重300克的Sprague-Dawley大鼠的肾脏进行原位灌注，灌注液为添加或未添加PrC-210的UW溶液，然后在4℃下于相同溶液中保存0至48小时。获取肾脏时，测量肾脏激活的半胱天冬酶-3水平（细胞死亡标志物），并对肾脏进行直接组织学分析，以评估PrC-210的保护效果。还对PrC-210对分离的大鼠肾脏或裸DNA的保护作用进行了体外分析。
用含20 mmol/L PrC-210的UW溶液对肾脏进行单次15秒原位灌注，导致以下情况显著减少：（1）30小时CI诱导的肾脏激活半胱天冬酶水平（<0.0001）；激活的半胱天冬酶水平降低至与未受干扰的肾脏中对照激活半胱天冬酶水平无显著差异的水平；（2）30小时CI诱导的肾小管损伤评分（=0.0004），其中刷状缘和肾小管坏死明显减少；（3）PrC-210对裸DNA和分离的肾脏线粒体的·OH损伤提供100%保护，而目前的UW溶液抗氧化剂则无保护作用。
大鼠肾脏从大鼠体内取出后，用PrC-210-UW溶液进行单次灌注，可显著降低暴露于30小时CI器官保存的肾脏中的半胱天冬酶和肾小管损伤。这些发现支持进一步开发PrC-210分子，以抑制或预防器官移植及其他缺血再灌注损伤情况下的缺血再灌注损伤。
