Ripk3-mediated cellular apoptosis is a major contributor to the pathogenesis of myocardial ischemia reperfusion (IR) injury. However, the mechanisms by which Ripk3 influences microvascular homeostasis and endothelial apoptosis are not completely understood. In this study, loss of Ripk3 inhibited endothelial apoptosis, alleviated luminal swelling, maintained microvasculature patency, reduced the expression of adhesion molecules and limited the myocardial inflammatory response. In vitro, Ripk3 deficiency protected endothelial cells from apoptosis and migratory arrest induced by HR injury. Mechanistically, Ripk3 had the ability to migrate onto the endoplasmic reticulum (ER), leading to ER damage, as evidenced by increased IP3R and XO expression. The higher IP3R content was associated with cellular calcium overload, and increased XO expression was involved in cellular oxidative injury. Furthermore, IP3R-mediated calcium overload and XO-dependent oxidative damage were able to initiate cellular apoptosis. More importantly, IP3R and XO also caused F-actin degradation into G-actin via post-transcriptional modification of cofilin, impairing the formation of the filopodia and limiting the migratory response of endothelial cells. Altogether, our data confirmed that Ripk3 was involved in microvascular IR injury via regulation of IP3R-mediated calcium overload, XO-dependent oxidative damage and filopodia-related cellular migration, ultimately leading to endothelial apoptosis and migratory inhibition. These findings provide a potential target for treating cardiac microcirculatory IR injury.

Ripk3 介导的细胞凋亡是心肌缺血再灌注 (IR) 损伤发病机制的主要贡献者。然而，Ripk3 影响微血管稳态和内皮细胞凋亡的机制尚不完全清楚。在这项研究中，Ripk3的缺失抑制了内皮细胞凋亡，减轻了管腔肿胀，维持了微血管通畅，减少了粘附分子的表达并限制了心肌炎症反应。在体外，Ripk3 缺陷可保护内皮细胞免受 HR 损伤引起的凋亡和迁移停滞。从机制上讲，Ripk3 具有迁移到内质网 (ER) 上的能力，导致 ER 损伤，IP3R 和 XO 表达增加证明了这一点。较高的IP3R含量与细胞钙超载有关，XO表达增加与细胞氧化损伤有关。此外，IP3R 介导的钙超载和 XO 依赖性氧化损伤能够引发细胞凋亡。更重要的是，IP3R和XO还通过丝切蛋白的转录后修饰导致F-肌动蛋白降解为G-肌动蛋白，损害丝状伪足的形成并限制内皮细胞的迁移反应。总之，我们的数据证实，Ripk3 通过调节 IP3R 介导的钙超载、XO 依赖性氧化损伤和丝状伪足相关的细胞迁移参与微血管 IR 损伤，最终导致内皮细胞凋亡和迁移抑制。这些发现为治疗心脏微循环IR损伤提供了潜在的靶点。