Membrane Vesicles Nucleate Mineralo-organic Nanoparticles and Induce Carbonate Apatite Precipitation in Human Body Fluids

Recent studies indicate that membrane vesicles (MVs) secreted by various cells are associated with human diseases, including arthritis, atherosclerosis, cancer, and chronic kidney disease. The possibility that MVs may induce the formation of mineralo-organic nanoparticles (NPs) and ectopic calcification has not been investigated so far. Here, we isolated MVs ranging in size between 20 and 400 nm from human serum and FBS using ultracentrifugation and sucrose gradient centrifugation. The MV preparations consisted of phospholipid-bound vesicles containing the serum proteins albumin, fetuin-A, and apolipoprotein A1; the mineralization-associated enzyme alkaline phosphatase; and the exosome proteins TNFR1 and CD63. Notably, we observed that MVs induced mineral precipitation following inoculation and incubation in cell culture medium. The mineral precipitates consisted of round, mineralo-organic NPs containing carbonate hydroxyapatite, similar to previous descriptions of the so-called nanobacteria. Annexin V-immunogold staining revealed that the calcium-binding lipid phosphatidylserine (PS) was exposed on the external surface of serum MVs. Treatment of MVs with an anti-PS antibody significantly decreased their mineral seeding activity, suggesting that PS may provide nucleating sites for calcium phosphate deposition on the vesicles. These results indicate that MVs may represent nucleating agents that induce the formation of mineral NPs in body fluids. Given that mineralo-organic NPs represent precursors of calcification in vivo , our results suggest that MVs may initiate ectopic calcification in the human body. Background: Membrane vesicles (MVs) released from various cells are associated with human diseases. Results: MVs isolated from human serum induce the formation of mineralo-organic nanoparticles in culture. Conclusion: MVs represent a nucleating factor that promotes the formation of mineralo-organic nanoparticles and the precipitation of mineral deposits in body fluids. Significance: The ectopic precipitation of carbonate apatite deposits in body fluids and tissues may be initiated in part by MVs.