Abstract
Dialysis-related amyloidosis is a major complication in long-term hemodialysis patients. In dialysis-related amyloidosis, β2-microglobulin (β2-m) amyloid fibrils deposit in the osteoarticular tissue, leading to carpal tunnel syndrome and destructive arthropathy with cystic bone lesions, but the mechanism by which these amyloid fibrils destruct bone and joint tissue is not fully understood. In this study, we assessed the cytotoxic effect of β2-m amyloid fibrils on the cultured rabbit synovial fibroblasts. Under light microscopy, the cells treated with amyloid fibrils exhibited both necrotic and apoptotic changes, while the cells treated with β2-m monomers and vehicle buffer exhibited no morphological changes. As compared to β2-m monomers and vehicle buffer, β2-m amyloid fibrils significantly reduced cellular viability as measured by the lactate dehydrogenase release assay and the 3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay and significantly increased the percentage of apoptotic cells as measured by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling method. β2-m amyloid fibrils added to the medium adhered to cell surfaces, but did not disrupt artificial plasma membranes as measured by the liposome dye release assay. Interestingly, when the cells were incubated with amyloid fibrils for several hours, many endosomes/lysosomes filled with amyloid fibrils were observed under confocal laser microscopy and electron microscopy, Moreover, some endosomal/lysosomal membranes were disrupted by intravesicular fibrils, leading to the leakage of the fibrils into the cytosol and adjacent to mitochondria. Inhibition of actin-dependent endocytosis by cytochalasin D attenuated the toxicity of amyloid fibrils. These results suggest that endocytosed β2-m amyloid fibrils induce necrosis and apoptosis by disrupting endosomal/lysosomal membranes, and this novel mechanism on the cytotoxicity of amyloid fibrils is described.
Highlights
Dialysis-related amyloidosis (DRA), is a systemic and nonhereditary amyloidosis that is a major and serious complication in long-term hemodialysis patients [1,2,3]
There were many intracytoplasmic endosomes/lysosomes filled with amyloid fibrils (Fig 7B) and some endosomal/lysosomal membranes were disrupted by intravesicular fibrils (Fig 7C)
We assessed the cytotoxic effect of β2-m amyloid fibrils on the cultured rabbit synovial fibroblasts (HIG-82 cells)
Summary
Dialysis-related amyloidosis (DRA), is a systemic and nonhereditary amyloidosis that is a major and serious complication in long-term hemodialysis patients [1,2,3]. A proinflammatory state induced by dialyzer membranes and contaminated dialysate may contribute to the pathogenesis of DRA [14] This type of amyloidosis is characterized by marked infiltration of activated macrophages around the amyloid deposits [15,16,17,18]. These macrophages are considered to cause chronic destructive inflammation in the osteoarticular tissue [15, 16], and/ or to play a role in the formation or degradation of β2-m amyloid fibrils [17, 18]. The molecular basis of the formation of β2-m amyloid fibrils has been explored intensively [19,20,21], but the mechanism by which the deposition of these amyloid fibrils causes the destruction of bone and joint tissue is not fully understood
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