Microfluidic generation of lipid nanoparticles to facilitate DNA entry into human mast cells

Abstract

Abstract Mast cells are highly granulated immune cells that can be targeted for transfection using lipid nanoparticles (LNP). Lipid nanoparticles (LNPs) were composed of cholesterol and 1-Stearoyl-2-oleoyl-sn-glycero-3-phosphocholine, the ionizable lipid 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane, and the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene-glycol)-2000] using microfluidic mixing and used to encapsulate a plasmid encoding enhanced green fluorescent protein (EGFP) under the control of a human cytomegalovirus promoter (pEGFP-C1). Encapsulation efficiency was determined using PicoGreen and LNP size was measured using dynamic light scattering. Human mast cells-1 (HMC-1) were transfected with loaded or empty LNPs in the presence or absence of apolipoprotein E (ApoE) for 24, 48 or 72 hours in medium. Cells were analyzed by fluorescence microscopy and flow cytometry. LNPs were approximately 100 nm in diameter (ideal) and efficiently encapsulated plasmids pEGFP-C1 (75.38%). ApoE4 appeared to slightly augment (>10%) LNP uptake after 24 hr. Our study demonstrates that LNPs generated by microfluidic mixing efficiently encapsulate a plasmid payload and can be used to successfully transfect human mast cells. Carrier molecules such as ApoE can be used to augment LNP uptake by mast cells. Further functionalization and optimization of these LNPs can provide powerful new tools for targeting other immune cells.