Department of Cell Biology and Molecular Genetics, Biological Sciences Graduate Program, University of Maryland, College Park, MD 20742, USA.
Arterioscler Thromb Vasc Biol. 2012 May;32(5):1178-85. doi: 10.1161/ATVBAHA.111.244186. Epub 2012 Feb 9.
Engagement of intercellular adhesion molecule 1 (ICAM-1) on endothelial cells by ICAM-1-targeted carriers induces cell adhesion molecule-mediated endocytosis, providing intraendothelial delivery of therapeutics. This pathway differs from classical endocytic mechanisms and invokes aspects of endothelial signaling during inflammation. ICAM-1 interacts with Na(+)/H(+) exchanger NHE1 during endocytosis, but it is unclear how this regulates plasmalemma and cytoskeletal changes. We studied such aspects in this work.
We used fluorescence and electron microscopy, inhibitors and knockout tools, cell culture, and mouse models. ICAM-1 engagement by anti-ICAM carriers induced sphingomyelin-enriched engulfment structures. Acid sphingomyelinase (ASM), an acidic enzyme that hydrolyzes sphingomyelin into ceramide (involved in plasmalemma deformability and cytoskeletal reorganization), redistributed to ICAM-1-engagement sites at ceramide-enriched areas. This induced actin stress fibers and carrier endocytosis. Inhibiting ASM impaired ceramide enrichment, engulfment structures, cytoskeletal reorganization, and carrier uptake, which was rescued by supplying this enzyme activity exogenously. Interfering with NHE1 rendered similar outcomes, suggesting that Na(+)/H(+) exchange might provide an acidic microenvironment for ASM at the plasmalemma.
These findings are consistent with the ability of endothelial cells to internalize relatively large ICAM- 1--targeted drug carriers and expand our knowledge on the regulation of the sphingomyelin/ceramide pathway by the vascular endothelium.
细胞间黏附分子 1（ICAM-1）与内皮细胞上的 ICAM-1 靶向载体结合，诱导细胞黏附分子介导的内吞作用，从而实现治疗药物向内皮细胞内的递送。该途径不同于经典的内吞机制，并在炎症过程中引发内皮信号转导的某些方面。ICAM-1 在胞吞作用过程中与 Na(+)/H(+)交换器 NHE1 相互作用，但尚不清楚这如何调节质膜和细胞骨架的变化。我们在这项工作中研究了这些方面。
我们使用荧光和电子显微镜、抑制剂和基因敲除工具、细胞培养和小鼠模型。抗 ICAM 载体与 ICAM-1 的结合诱导了富含鞘磷脂的吞噬结构。酸性鞘磷脂酶（ASM）是一种酸性酶，可将鞘磷脂水解为神经酰胺（参与质膜变形和细胞骨架重组），在富含神经酰胺的区域重新分布到与 ICAM-1 结合的部位。这诱导了肌动蛋白应力纤维和载体内吞作用。抑制 ASM 会损害神经酰胺的富集、吞噬结构、细胞骨架重组和载体摄取，而通过外源提供这种酶活性可以挽救这些结果。干扰 NHE1 会产生类似的结果，这表明 Na(+)/H(+) 交换可能为质膜上的 ASM 提供酸性微环境。
这些发现与内皮细胞内化相对较大的 ICAM-1 靶向药物载体的能力一致，并扩展了我们对血管内皮细胞调节鞘磷脂/神经酰胺途径的认识。
