Sphingolipids-enriched rafts domains are proposed to occur in plasma membranes and to mediate important cellular functions. Notwithstanding, the asymmetric transbilayer distribution of phospholipids that exists in the membrane confers the two leaflets different potentials to form lateral domains as next to no sphingolipids are present in the inner leaflet. How the physical properties of one leaflet can influence the properties of the other and its importance on signal transduction across the membrane are questions still unresolved. In this work, we combined AFM imaging and Force spectroscopy measurements to assess domain formation and to study the nanomechanical properties of asymmetric supported lipid bilayers (SLBs) mimicking membrane rafts. Asymmetric SLBs were formed by incorporating N-palmitoyl-sphingomyelin (16:0SM) into the outer leaflet of preformed 1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC)/Cholesterol SLBs through methyl-β-cyclodextrin–mediated lipid exchange. Lipid domains were detected after incorporation of 16:0SM though their phase state varied from gel to liquid ordered (Lo) phase if the procedure was performed at 24 or 37 °C, respectively. When comparing symmetric and asymmetric Lo domains, differences in size and morphology were observed, with asymmetric domains being smaller and more interconnected. Both types of Lo domains showed similar mechanical stability in terms of rupture forces and Young's moduli. Notably, force curves in asymmetric domains presented two rupture events that could be attributed to the sequential rupture of a liquid disordered (Ld) and a Lo phase. Interleaflet coupling in asymmetric Lo domains could also be inferred from those measurements. The experimental approach outlined here would significantly enhance the applicability of membrane models.

富含鞘脂的筏域被提议在质膜中发生并介导重要的细胞功能。尽管如此，存在于膜中的磷脂的不对称跨双分子层分布赋予两个小叶以不同的电位来形成侧向结构域，因为内部小叶中不存在鞘脂。一个小叶的物理特性如何影响另一个小叶的特性及其对跨膜信号转导的重要性仍未解决。在这项工作中，我们结合了原子力显微镜成像和力谱测量，以评估域形成并研究模仿膜筏的不对称支撑脂质双层（SLB）的纳米力学性能。。通过将N-棕榈酰基-鞘磷脂（16：0SM）掺入预先形成的1,2-二油酰基-sn-甘油-3-磷酸胆碱（DOPC）/胆固醇的SLB中，通过甲基-β-环糊精介导的脂质形成不对称的SLB。交换。如果分别在24或37°C下进行操作，则在掺入16：0SM后可检测到脂质结构域，尽管它们的相态从凝胶到液序（Lo）相均不同。比较对称和不对称Lo域时，观察到大小和形态的差异，不对称域较小且相互连接。两种类型的罗在断裂力和杨氏模量方面，磁畴表现出相似的机械稳定性。值得注意的是，非对称区域中的力曲线呈现两个破裂事件，这可以归因于液体无序（Ld）和Lo相的连续破裂。从这些测量结果也可以推断出非对称Lo域的小叶间偶联。这里概述的实验方法将大大提高膜模型的适用性。