A preliminary study of the innate immune memory of Kupffer cells induced by PEGylated nanoemulsions

The accelerated blood clearance (ABC) phenomenon describes a dilemma of polyethylene glycol (PEG) applied in drug delivery system (DDS) caused by its immunogenicity, that results in the enhanced blood clearance rate and increased hepatic and splenic accumulation after secondary injection of PEGylated nanocarriers. However, the ABC index, as the judgement of ABC phenomenon, only describes the accelerated blood clearance rate, but ignores the enhanced hepatic and splenic accumulation. Therefore, we proposed the hepatic accumulation (HA) index and the splenic accumulation (SA) index as supplements for assessing the ABC phenomenon, to emphasize the contribution of liver and spleen, especially the liver, possessing the most population of tissue resident macrophages. By altering the first injection site from the tail vein to the liver portal vein, there was no impact on anti-PEG IgM production, and the secondary hepatic accumulation of PEGylated nanoemulsions (PE) was observed to be proportionate to the first PE stimulation strength on the liver. We also determined that Kupffer cells (KCs) were the main contributor to this enhancement. On this basis, we revealed a definite phenomenon that PE could induce innate immune memory in KCs, by enhancing the phagocytosis of KCs toward PE during the secondary stimulation. The PE-stimulated KCs could carry this memory to the naïve rats through adoptive transfer, resulting in increased hepatic accumulation in the recipient rats without antibody production. Studies examining the phagocytosis of KCs in vivo, ex vivo and in vitro revealed that the memory of KCs against PE triggered by first-stimulated PE could be maintained independently of other cells or components until 21 days after the first stimulation, and possessing specificity to PEG, which was invalid to long-circulating GE (GM1 modified nanoemulsions). The discovery of immune memory in KCs induced by PE highlights the importance of focusing on the relationship between the innate immune system and PEGylated nanocarriers during the development of DDS to improve medication safety in the clinic.