背景:在糖尿病性视网膜病变(DR)中,缺氧诱导因子(HIF-1α)通过上调糖酵解诱导氧化应激。这个过程导致神经变性,特别是感光细胞损伤,这进一步导致视网膜微血管恶化。Further,Wnt抑制因子1(WIF1)的调节,分泌的Wnt信号拮抗剂,在神经退行性眼病中尚未完全表征。我们旨在探讨在DR背景下WIF1对光感受器功能的影响。
方法:将12周龄的C57BL/KsJ-db/db小鼠玻璃体内注射WIF1过表达慢病毒。4周后,光学相干断层扫描(OCT),透射电子显微镜(TEM),H&E染色,和视网膜电图(ERG)用于评估视网膜组织和功能。使用单细胞RNA测序探索了WIF1在感光细胞中的潜在作用机制。在高葡萄糖条件下,661W细胞作为体外DR模子。使用定量实时PCR评估WIF1介导的信号通路成分,免疫染色,和西方印迹。
结果:在db/db小鼠中观察到典型的糖尿病表现。值得注意的是,WIF1在mRNA和蛋白水平上的表达降低。在db/db小鼠中WIF1过表达后,这些病理表现和视觉功能得到改善。TEM显示WIF1恢复了受损的线粒体,高尔基体,和感光体外段。此外,ERG显示a波电位振幅的恢复。单细胞RNA测序和体外实验表明,WIF1过表达通过抑制经典的Wnt信号通路来阻止糖酵解酶的表达和乳酸的产生,HIF-1α,和Glut1,从而降低视网膜和细胞活性氧水平并维持661W细胞活力。
结论:WIF1对Wnt/β-catenin-HIF-1α-Glut1糖酵解通路有抑制作用,从而减轻氧化应激水平并减轻视网膜感光细胞的病理结构特征。这种机制有助于保持DR中感光细胞的功能,并表明WIF1有望成为DR和其他神经退行性眼部疾病的潜在治疗候选药物。
In diabetic retinopathy (DR), hypoxia-inducible factor (HIF-1α) induces oxidative stress by upregulating glycolysis. This process leads to neurodegeneration, particularly photoreceptor cell damage, which further contributes to retinal microvascular deterioration. Further, the regulation of Wnt-inhibitory factor 1 (WIF1), a secreted Wnt signaling antagonist, has not been fully characterized in neurodegenerative eye diseases. We aimed to explore the impact of WIF1 on photoreceptor function within the context of DR.
Twelve-week-old C57BL/KsJ-db/db mice were intravitreally injected with WIF1 overexpression lentivirus. After 4 weeks, optical coherence tomography (OCT), transmission electron microscopy (TEM), H&E staining, and electroretinography (ERG) were used to assess the retinal tissue and function. The potential mechanism of action of WIF1 in photoreceptor cells was explored using single-cell RNA sequencing. Under high-glucose conditions, 661 W cells were used as an in vitro DR model. WIF1-mediated signaling pathway components were assessed using quantitative real-time PCR, immunostaining, and western blotting.
Typical diabetic manifestations were observed in db/db mice. Notably, the expression of WIF1 was decreased at the mRNA and protein levels. These pathological manifestations and visual function improved after WIF1 overexpression in db/db mice. TEM demonstrated that WIF1 restored damaged mitochondria, the Golgi apparatus, and photoreceptor outer segments. Moreover, ERG indicated the recovery of a-wave potential amplitude. Single-cell RNA sequencing and in vitro experiments suggested that WIF1 overexpression prevented the expression of glycolytic enzymes and lactate production by inhibiting the canonical Wnt signaling pathway, HIF-1α, and Glut1, thereby reducing retinal and cellular reactive oxygen species levels and maintaining 661 W cell viability.
WIF1 exerts an inhibitory effect on the Wnt/β-catenin-HIF-1α-Glut1 glycolytic pathway, thereby alleviating oxidative stress levels and mitigating pathological structural characteristics in retinal photoreceptor cells. This mechanism helps preserve the function of photoreceptor cells in DR and indicates that WIF1 holds promise as a potential therapeutic candidate for DR and other neurodegenerative ocular disorders.