Formation of Protein Corona on Nanoparticle Affects Different Complement Activation Pathways Mediated by C1q.

As nanoparticles (NPs) are intravenously entering the bloodstream, proteins in the plasma can recognize and bind them to form a protein corona that affects how NPs are perceived by biological systems. The complement is an essential part of the innate immunity that contributes to non-specific host defense. How complement recognizes NPs has not been elucidated. Here, we developed a proteomics and biochemical approach to understand the applied risk of activated complement by NPs. Complement proteins absorbed on Hydroxyapatite Nanoparticles (HAP-NPs) and Silicon dioxide Nanoparticles (SiO2-NPs) were analyzed by proteomics with LC-MS. The effect of complement activation was studied by iC3b/Sc5b-9/C3a/C4a/C5a with ELISA. An inhibitory model was established via EDTA and EGTA to confirm the selective pathway activation of both NPs. Finally, the regulation of complement by NPs was analyzed by western blot. The results indicate that HAP-NPs start the activation of the complement through the classical pathway because of the absorption of C1q and the release of C1r/C1s. Meanwhile, the soluble regulatory molecules such as CFI, C4bp, and CFH tried to resist the complement system activation by the cleavage of C3 convertase. In contrast, SiO2-NPs can activate the alternative pathway of the complement through the absorption of CFD and CFB. It was clarified that HAP-NPs and SiO2-NPs activate complement through different mechanisms. These studies provide a scientific basis for the design and modification of nano-drug carriers for delaying their recognition and clearance by the mononuclear phagocytic system and simultaneously reducing the immunotoxicity of NPs. The understanding of protein corona is conducive to innovation in the field of "immune-safe-by-design" nanomedicines.