Abstract：
In this study we investigate the role of Zipper-interacting protein kinase (ZIPK) in high glucose-induced vascular injury, focusing on its interaction with STAT5A and its effects on p53 and inducible nitric oxide synthase (NOS2) expression. Human umbilical vein endothelial cells (HUVECs) are cultured under normal (5 mM) and high (25 mM) glucose conditions. Protein and gene expression levels are assessed by western blot analysis and qPCR respectively, while ROS levels are measured via flow cytometry. ZIPK expression is manipulated using overexpression plasmids, siRNAs, and shRNAs. The effects of the ZIPK inhibitor TC-DAPK6 are evaluated in a diabetic rat model. Our results show that high glucose significantly upregulates ZIPK, STAT5A, p53, and NOS2 expressions in HUVECs, thus increasing oxidative stress. Silencing of STAT5A reduces p53 and NOS2 expressions and reactive oxygen species (ROS) accumulation. ZIPK is essential for high glucose-induced p53 expression and ROS accumulation, while silencing of ZIPK reverses these effects. Overexpression of ZIPK combined with STAT5A silencing attenuates glucose-induced alterations in p53 and NOS2 expression, thereby preventing cell damage. Coimmunoprecipitation reveals a direct interaction between ZIPK and STAT5A in the nucleus under high-glucose condition. In diabetic rats, TC-DAPK6 treatment significantly decreases ZIPK, p53, and NOS2 expressions. Our findings suggest that ZIPK plays a critical role in high glucose-induced vascular injury via STAT5A-mediated pathways, proposing that ZIPK is a potential therapeutic target for diabetic vascular complications.
摘要：
在这项研究中，我们研究了拉链相互作用蛋白激酶（ZIPK）在高糖诱导的血管损伤中的作用。重点研究其与STAT5A的相互作用及其对p53和诱导型一氧化氮合酶（NOS2）表达的影响。在正常(5mM)和高(25mM)葡萄糖条件下培养人脐静脉内皮细胞(HUVEC)。蛋白质和基因表达水平分别通过蛋白质印迹分析和qPCR评估，而ROS水平通过流式细胞术测量。使用过表达质粒操纵ZIPK表达，siRNA,还有shRNA.在糖尿病大鼠模型中评价ZIPK抑制剂TC-DAPK6的作用。我们的结果表明，高葡萄糖显著上调ZIPK，STAT5A,p53和NOS2在HUVECs中的表达，从而增加氧化应激。STAT5A的沉默减少了p53和NOS2的表达以及活性氧（ROS）的积累。ZIPK对于高糖诱导的p53表达和ROS积累至关重要，而ZIPK的沉默逆转了这些影响。ZIPK过表达联合STAT5A沉默可减弱葡萄糖诱导的p53和NOS2表达改变,从而防止细胞损伤。免疫共沉淀揭示了在高糖条件下细胞核中ZIPK和STAT5A之间的直接相互作用。在糖尿病大鼠中，TC-DAPK6治疗显著降低ZIPK,p53和NOS2表达。我们的研究结果表明，ZIPK通过STAT5A介导的途径在高糖诱导的血管损伤中起关键作用。提出ZIPK是糖尿病血管并发症的潜在治疗靶点。
