Impact of Protein Coronas on Lipid Nanoparticle Uptake and Endocytic Pathways in Cells

Lipid nanoparticles (LNPs), widely used in disease diagnosis and drug delivery, face the challenge of being surrounded by biological macromolecules such as proteins upon entering the human body. These molecules compete for binding sites on the nanoparticle surfaces, forming a protein corona. The impact of different types of protein coronas on LNP delivery remains unclear. In this study, we employed a newly developed, highly sensitive LNP labeling platform and analyzed the endocytosis of HeLa cells under different nutritional conditions using proteomics to address this critical issue. Our research found that under conditions of complete medium and amino acid starvation, most DNA-FITC vesicles in HeLa cells were located in the perinuclear region 4 h after transfection. In contrast, under serum starvation conditions, only a small portion of DNA-FITC vesicles were in the perinuclear region. On the other hand, through proteomics, we discovered that cells that were enriched in amino acids and complete medium contained more proteins, whereas those under serum starvation had relatively fewer enriched proteins. Through KEGG pathway enrichment analysis, we identified the phagosome and endocytosis pathways as particularly important. Lastly, differential analysis of proteins in these pathways revealed that proteins such as F-actin, Coronin, vATPase, TUBA, TUBB, MHCII, and TSP may have significant impacts on cellular endocytosis. Our research findings indicate that it is necessary to regulate cellular endocytosis based on different protein coronas to achieve optimal cytoplasmic release.