Hu Qianchao, Chen Hao, Lan Jia'nan, Chen Yiwen, Liu Zhongzhong, Xiong Yan, Zhou Wei, Zhong Zibiao, Ye Qifa
Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Hubei Key Laboratory of Medical Technology on Transplantation, Engineering Research Center of Natural Polymer-based Medical Materials in Hubei Province, Wuhan, China.
The 3rd Xiangya Hospital of Central South University, Research Center of National Health Ministry on Transplantation Medicine Engineering and Technology, Changsha, China.
Transplantation. 2025 Jun 1;109(6):e273-e286. doi: 10.1097/TP.0000000000005314. Epub 2024 Dec 24.
Hypothermic machine perfusion (HMP) is becoming the main preservation method for donation after circulatory death (DCD) kidneys. It can provide continuous flow and form shear stress (SS) upon endothelial cells (ECs), thereby regulating EC injury. Krüppel-like factor 10 (KLF10) has been shown to lessen vascular damage. However, how SS and KLF10 impact HMP-regulated injury is unclear.
In this study, we investigated the influences of KLF10 on HMP in animal models and human renal biopsy and explored how SS affected KLF10 expression in a parallel-plate flow chamber system. Chromatin Immunoprecipitation sequencing and luciferase assay were performed to seek the target genes of KLF10. The influences of KLF10 on HMP-regulated injury were investigated by transfecting si-KLF10 adeno-associated virus serotype 9 into rat kidneys. The molecular expression was examined using immunofluorescence staining, Western blotting, and quantitative polymerase chain reaction.
Our results show KLF10 expression was augmented in human, rabbit, and rat DCD kidneys after HMP. HMP improved ECs and tubule injury and attenuated inflammation; however, the knockdown of KLF10 reversed this effect. SS regulated KLF10 expression in ECs by affecting F-actin, and KLF10 could maintain ECs homeostasis. Chromatin Immunoprecipitation sequencing and luciferase assay revealed that baculoviral inhibitor of apoptosis protein repeat-containing 2 (BIRC2) is a target gene of KLF10. Furthermore, BIRC2 linked to nuclear factor kappa B (NF-κB)-inducing kinase, induced NF-κB)-inducing kinase ubiquitination, and resulted in inhibiting the noncanonical NF-κB pathway.
SS can mediate KLF10 expression, whereas HMP can protect against warm ischemic injury by reducing inflammation via KLF10/BIRC2/noncanonical NF-κB pathway. Therefore, KLF10 might be a novel target for improving DCD kidney quality.
低温机器灌注（HMP）正成为循环性死亡（DCD）供肾的主要保存方法。它能提供持续血流并在内皮细胞（ECs）上形成剪切应力（SS），从而调节EC损伤。已有研究表明，Krüppel样因子10（KLF10）可减轻血管损伤。然而，SS和KLF10如何影响HMP调节的损伤尚不清楚。
在本研究中，我们在动物模型和人类肾活检中研究了KLF10对HMP的影响，并在平行板流动腔系统中探索了SS如何影响KLF10的表达。进行染色质免疫沉淀测序和荧光素酶测定以寻找KLF10的靶基因。通过将si-KLF10腺相关病毒血清型9转染到大鼠肾脏中，研究KLF10对HMP调节损伤的影响。使用免疫荧光染色、蛋白质免疫印迹和定量聚合酶链反应检测分子表达。
我们的结果显示，HMP后人类、兔和大鼠DCD肾脏中KLF10表达增加。HMP改善了ECs和肾小管损伤并减轻了炎症；然而，敲低KLF10可逆转这种效应。SS通过影响F-肌动蛋白调节ECs中KLF10的表达，而KLF10可维持ECs的稳态。染色质免疫沉淀测序和荧光素酶测定显示，含杆状病毒凋亡蛋白重复序列2（BIRC2）是KLF10的靶基因。此外，BIRC2与核因子κB（NF-κB）诱导激酶相关，诱导NF-κB诱导激酶泛素化，并导致非经典NF-κB途径受到抑制。
SS可介导KLF10表达，而HMP可通过KLF10/BIRC2/非经典NF-κB途径减轻炎症，从而预防热缺血损伤。因此，KLF10可能是改善DCD肾脏质量的新靶点。
