β2-Microglobulin Amyloid Fibril-Induced Membrane Disruption Is Enhanced by Endosomal Lipids and Acidic pH

Although the molecular mechanisms underlying the pathology of amyloidoses are not well understood, the interaction between amyloid proteins and cell membranes is thought to play a role in several amyloid diseases. Amyloid fibrils of β2-microglobulin (β2m), associated with dialysis-related amyloidosis (DRA), have been shown to cause disruption of anionic lipid bilayers in vitro. However, the effect of lipid composition and the chemical environment in which β2m-lipid interactions occur have not been investigated previously. Here we examine membrane damage resulting from the interaction of β2m monomers and fibrils with lipid bilayers. Using dye release, tryptophan fluorescence quenching and fluorescence confocal microscopy assays we investigate the effect of anionic lipid composition and pH on the susceptibility of liposomes to fibril-induced membrane damage. We show that β2m fibril-induced membrane disruption is modulated by anionic lipid composition and is enhanced by acidic pH. Most strikingly, the greatest degree of membrane disruption is observed for liposomes containing bis(monoacylglycero)phosphate (BMP) at acidic pH, conditions likely to reflect those encountered in the endocytic pathway. The results suggest that the interaction between β2m fibrils and membranes of endosomal origin may play a role in the molecular mechanism of β2m amyloid-associated osteoarticular tissue destruction in DRA.

尽管淀粉样变性病（amyloidoses）的病理分子机制尚未完全阐明，但淀粉样蛋白与细胞膜的相互作用被认为在多种淀粉样疾病的发病过程中发挥作用。与透析相关淀粉样变性（dialysis-related amyloidosis, DRA）相关的β2微球蛋白（β2-microglobulin, β2m）淀粉样原纤维，已被证实可在体外（in vitro）破坏阴离子脂质双层。然而，此前尚未有研究探讨脂质组成以及β2m与脂质相互作用所处的化学环境对该过程的影响。本研究针对β2m单体与原纤维分别与脂质双层相互作用所引发的膜损伤展开探究。我们采用染料释放实验、色氨酸荧光猝灭实验及荧光共聚焦显微镜（fluorescence confocal microscopy）成像实验，考察了阴离子脂质组成与pH值对脂质体易受原纤维诱导膜损伤程度的影响。结果显示，β2m原纤维诱导的膜破坏过程受阴离子脂质组成调控，且酸性pH可显著增强该效应。尤为引人注目的是，在酸性pH条件下，含有双（单酰甘油）磷酸酯（bis(monoacylglycero)phosphate, BMP）的脂质体膜破坏程度最高，该环境条件与内吞途径（endocytic pathway）中实际遭遇的生理环境高度相似。本研究结果提示，β2m原纤维与内体来源的细胞膜的相互作用，可能在透析相关淀粉样变性中β2m淀粉样相关骨关节组织破坏的分子机制中发挥重要作用。