Wang Xiuxian, Zhang Peicheng, Xie Jing, Zuo Xiangrong
Department of Ophthalmology, Xingtai People's Hospital, Xingtai, 054001, Hebei, China.
Department of Ophthalmology, Hebei Provincial Key Laboratory of Ophthalmology, Hebei Provincial Clinical Medical Research Center for Ocular Diseases, Hebei Eye Hospital, No.399, Quanbei East Road, Xingtai, 054001, Hebei, China.
Int Ophthalmol. 2025 Jan 24;45(1):39. doi: 10.1007/s10792-025-03410-y.
Retinopathy of prematurity (ROP) is a major cause of childhood blindness worldwide, highlighted by retinal neovascularization. Ubiquitin is present throughout the retina. The deubiquitinating enzyme ubiquitin-specific protease 39 (USP39) has been reported to be involved in angiogenesis. Here, this study aimed to investigate the effects of USP39 on ROP and its associated mechanism.
Hypoxia-induced human retinal microvascular endothelial cells (hRMECs) were adopted for functional analyses. Detection of mRNA and protein was conducted using quantitative real-time PCR and western blotting. Cell migration, invasion and angiogenesis were evaluated using transwell and tube formation assays. Protein interaction was determined by immunoprecipitation assay. Oxygen-induced retinopathy (OIR) mouse models were used for in vivo analysis.
USP39 level was higher in hypoxia-induced hRMECs, functionally, USP39 silencing reversed hypoxia-induced migration, invasion and angiogenesis in hRMECs. In further mechanism analysis, we found that USP39 stabilized SIRT2 protein expression in hRMECs by inducing SIRT2 deubiquitination. Moreover, SIRT2 up-regulation abated hypoxia-evoked migration, invasion and angiogenesis in hRMECs. Besides that, the inhibitory effects of USP39 silencing on hypoxia-induced metastatic and angiogenic behaviors were abolished after SIRT2 overexpression. In addition, USP39 silencing blocked the activation of phosphoinositide 3-kinase (PI3K)/protein kinase B pathway (AKT) by regulating SIRT2. In vivo assay showed that levels of USP39, SIRT2, matrix metalloproteinase (MMP)-2 (MMP-2), MMP-9 and Vascular endothelial growth factor A (VEGFA) were increased in the retinas of OIR mice, while intravitreal injection of USP39 short hairpin RNA (shRNA) could reduce their expression.
USP39 stabilized SIRT2 expression by deubiquitination and promoted hypoxia-induced metastatic and angiogenic behaviors of RMECs in vitro, as well as retinal angiogenesis in vivo.
早产儿视网膜病变（ROP）是全球儿童失明的主要原因，其特征为视网膜新生血管形成。泛素存在于整个视网膜中。据报道，去泛素化酶泛素特异性蛋白酶39（USP39）参与血管生成。在此，本研究旨在探讨USP39对ROP的影响及其相关机制。
采用缺氧诱导的人视网膜微血管内皮细胞（hRMECs）进行功能分析。使用定量实时PCR和蛋白质印迹法检测mRNA和蛋白质。使用Transwell和管形成试验评估细胞迁移、侵袭和血管生成。通过免疫沉淀试验确定蛋白质相互作用。使用氧诱导视网膜病变（OIR）小鼠模型进行体内分析。
缺氧诱导的hRMECs中USP39水平较高，在功能上，USP39沉默可逆转缺氧诱导的hRMECs迁移、侵袭和血管生成。在进一步的机制分析中，我们发现USP39通过诱导SIRT2去泛素化来稳定hRMECs中SIRT2蛋白的表达。此外，SIRT2上调减弱了缺氧诱导的hRMECs迁移、侵袭和血管生成。除此之外，SIRT2过表达后，USP39沉默对缺氧诱导的转移和血管生成行为的抑制作用被消除。此外，USP39沉默通过调节SIRT2阻断磷酸肌醇3激酶（PI3K）/蛋白激酶B通路（AKT）的激活。体内试验表明，OIR小鼠视网膜中USP39、SIRT2、基质金属蛋白酶（MMP）-2、MMP-9和血管内皮生长因子A（VEGFA）的水平升高，而玻璃体内注射USP39短发夹RNA（shRNA）可降低它们的表达。
USP39通过去泛素化稳定SIRT2表达，促进缺氧诱导的RMECs体外转移和血管生成行为以及体内视网膜血管生成。
