Background Ischemia-reperfusion injury (IRI) is a major risk factor for chronic renal failure. Here, we characterize the different modes of programmed cell death in the tubular and microvascular compartments during the various stages of IRI-induced AKI, and their relative importance to renal fibrogenesis.

Methods We performed unilateral renal artery clamping for 30 minutes and contralateral nephrectomy in wild-type mice (C57BL/6) or caspase-3^−/− mice.

Results Compared with their wild-type counterparts, caspase-3^−/− mice in the early stage of AKI had high urine cystatin C levels, tubular injury scores, and serum creatinine levels. Electron microscopy revealed evidence of tubular epithelial cell necrosis in caspase-3^−/− mice, and immunohistochemistry showed upregulation of the necroptosis marker receptor-interacting serine/threonine-protein kinase 3 (RIPK3) in renal cortical sections. Western blot analysis further demonstrated enhanced levels of phosphorylated RIPK3 in the kidneys of caspase-3^−/− mice. In contrast, caspase-3^−/− mice had less microvascular congestion and activation in the early and extension phases of AKI. In the long term (3 weeks after IRI), caspase-3^−/− mice had reduced microvascular rarefaction and renal fibrosis, as well as decreased expression of α-smooth muscle actin and reduced collagen deposition within peritubular capillaries. Moreover, caspase-3^−/− mice exhibited signs of reduced tubular ischemia, including lower tubular expression of hypoxia-inducible factor-1α and improved tubular injury scores.

Conclusions These results establish the pivotal importance of caspase-3 in regulating microvascular endothelial cell apoptosis and renal fibrosis after IRI. These findings also demonstrate the predominant role of microvascular over tubular injury as a driver of progressive renal damage and fibrosis after IRI.

背景技术缺血再灌注损伤（IRI）是慢性肾功能衰竭的主要危险因素。在这里，我们表征了IRI诱导的AKI各个阶段中肾小管和微血管区室中程序性细胞死亡的不同模式，以及它们对肾纤维化的相对重要性。

方法我们对野生型小鼠（C57BL / 6）或caspase-3 ^-/-小鼠进行了单侧肾动脉钳夹30分钟和对侧肾切除术。

结果与野生型相比，AKI早期的caspase-3 ^-/-小鼠尿液胱抑素C水平，肾小管损伤评分和血清肌酐水平较高。电子显微镜显示了caspase-3 ^-/-小鼠中肾小管上皮细胞坏死的证据，免疫组织化学显示，在肾皮质切片中，与肾病标志物受体相互作用的丝氨酸/苏氨酸蛋白激酶3（RIPK3）上调。蛋白质印迹分析进一步证实了caspase-3 ^-/-小鼠肾脏中磷酸化RIPK3的水平提高。相反，caspase-3 ^-/-小鼠在AKI的早期和延伸期的微血管充血和激活较少。从长远来看（IRI后3周），caspase-3^-/-小鼠的微血管稀疏性和肾纤维化程度降低，α-平滑肌肌动蛋白的表达降低，肾小管周围毛细血管内的胶原蛋白沉积减少。另外，胱天蛋白酶-3 ^{- / -}小鼠表现出降低的管状缺血征象，包括低氧诱导因子1的下部管状表达α和改进的肾小管损伤分数。

结论这些结果确立了caspase-3在调节IRI后微血管内皮细胞凋亡和肾纤维化中的关键作用。这些发现还证明了微血管在肾小管损伤中的主要作用是IRI后进行性肾脏损害和纤维化的驱动因素。