Department of Ophthalmology, The Affiliated Eye Hospital, Nanjing Medical University, #138 Han-ZhongRoad, Nanjing, 210000, China.
The Fourth School of Clinical Medicine, Nanjing Medical University, Nanjing, 210000, China.
J Transl Med. 2023 Sep 22;21(1):651. doi: 10.1186/s12967-023-04503-x.
Pathological neovascularization plays a pivotal role in the onset and progression of tumors and neovascular eye diseases. Despite notable advancements in the development of anti-angiogenic medications that target vascular endothelial growth factor (VEGF) and its receptors (VEGFRs), the occurrence of adverse reactions and drug resistance has somewhat impeded the widespread application of these drugs. Therefore, additional investigations are warranted to explore alternative therapeutic targets. In recent years, owing to the swift advancement of high-throughput sequencing technology, pan-cancer analysis and single-cell sequencing analysis have emerged as pivotal methodologies and focal areas within the domain of omics research, which is of great significance for us to find potential targets related to the regulation of pathological neovascularization.
Pan-cancer analysis and scRNA-seq data analysis were employed to forecast the association between Actin filament-associated protein 1 like 1 (AFAP1L1) and the development of tumors and endothelial cells. Tumor xenograft model and ocular pathological neovascularization model were constructed as well as Isolectin B4 (IsoB4) staining and immunofluorescence staining were used to assess the effects of AFAP1L1 on the progression of neoplasms and neovascular eye diseases in vivo. Transwell assay, wound scratch assay, tube forming assay, three-dimensional germination assay, and rhodamine-phalloidin staining were used to evaluate the impact of AFAP1L1 on human umbilical vein endothelial cells (HUVECs) function in vitro; Dual luciferase reporting, qRT-PCR and western blot were used to investigate the upstream and downstream mechanisms of pathological neovascularization mediated by AFAP1L1.
Our investigation revealed that AFAP1L1 plays a crucial role in promoting the development of various tumors and demonstrates a strong correlation with endothelial cells. Targeted suppression of AFAP1L1 specifically in endothelial cells in vivo proves effective in inhibiting tumor formation and ocular pathological neovascularization. Mechanistically, AFAP1L1 functions as a hypoxia-related regulatory protein that can be activated by HIF-1α. In vitro experiments demonstrated that reducing AFAP1L1 levels can reverse hypoxia-induced excessive angiogenic capacity in HUVECs. The principal mechanism of angiogenesis inhibition entails the regulation of tip cell behavior through the YAP-DLL4-NOTCH axis.
In conclusion, AFAP1L1, a newly identified hypoxia-related regulatory protein, can be activated by HIF-1α. Inhibiting AFAP1L1 results in the inhibition of angiogenesis by suppressing the germination of endothelial tip cells through the YAP-DLL4-NOTCH axis. This presents a promising therapeutic target to halt the progression of tumors and neovascular eye disease.
病理性血管新生在肿瘤和新生血管性眼病的发生和发展中起着关键作用。尽管针对血管内皮生长因子(VEGF)及其受体(VEGFRs)的抗血管生成药物取得了显著进展，但不良反应和耐药性的发生在一定程度上阻碍了这些药物的广泛应用。因此，有必要进行更多的研究来探索替代治疗靶点。近年来，由于高通量测序技术的快速发展，泛癌症分析和单细胞测序分析已成为组学研究领域的重要方法学和焦点领域，这对于我们寻找与病理性血管新生调节相关的潜在靶点具有重要意义。
采用泛癌症分析和 scRNA-seq 数据分析预测肌动蛋白丝相关蛋白 1 样 1(AFAP1L1)与肿瘤和内皮细胞发生发展的关系。构建肿瘤异种移植模型和眼部病理性血管新生模型，采用 Isolectin B4(IsoB4)染色和免疫荧光染色评估 AFAP1L1 对体内肿瘤和新生血管性眼病进展的影响。Transwell 实验、划痕实验、管形成实验、三维发芽实验和罗丹明-鬼笔环肽染色评估 AFAP1L1 对人脐静脉内皮细胞(HUVECs)功能的影响；双荧光素酶报告、qRT-PCR 和 Western blot 检测 AFAP1L1 介导病理性血管新生的上下游机制。
我们的研究表明，AFAP1L1 在促进各种肿瘤的发生发展中起关键作用，并与内皮细胞有很强的相关性。体内靶向抑制内皮细胞中的 AFAP1L1 可有效抑制肿瘤形成和眼部病理性血管新生。机制上，AFAP1L1 作为一种缺氧相关的调节蛋白，可被 HIF-1α激活。体外实验表明，降低 AFAP1L1 水平可逆转 HUVECs 中缺氧诱导的过度血管生成能力。抑制血管生成的主要机制是通过 YAP-DLL4-NOTCH 轴调节尖端细胞行为。
综上所述，新发现的缺氧相关调节蛋白 AFAP1L1 可被 HIF-1α激活。抑制 AFAP1L1 通过 YAP-DLL4-NOTCH 轴抑制内皮细胞尖端细胞的发芽来抑制血管生成。这为阻止肿瘤和新生血管性眼病的进展提供了一个有前途的治疗靶点。
