Speaker
Description
Introduction
Lipid nanoparticles (LNPs) represent a promising and versatile platform for the non-viral delivery of genetic material, including microRNA (miRNA), which plays a crucial regulatory role in gene expression. While LNPs have already demonstrated clinical success as mRNA carriers in vaccines, most notably in COVID-19 immunization strategies, their application in miRNA delivery opens new opportunities in regenerative medicine, such as wound healing, based on cellular reprogramming. Unlike mRNA, which typically aims to express therapeutic proteins transiently, miRNA can modulate entire gene networks, making LNP-based delivery systems particularly attractive for fine-tuned, endogenous control of cellular processes. The primary aim of this study is to develop effective LNP-based carriers for miRNA delivery and to investigate how various post-processing techniques influence their physicochemical properties. This work focuses on assessing the relationship between processing conditions and key characteristics such as particle size, polydispersity, surface charge, and ultimately, transfection efficiency. Additionally, the performance of the optimized LNPs is compared to a commercial transfection reagent—Lipofectamine™ 3000—to evaluate their potential as a viable alternative in non-viral gene delivery systems.
Methods
miRNA-loaded LNPs (miRNA-LNPs) and blank LNPs (without miRNA) were synthesized using a microfluidic approach. The samples were subjected to four post-processing techniques: sonication, filtration, dialysis, and thermal treatment. Key physicochemical parameters, including average particle size, polydispersity index (PDI), and zeta potential, were evaluated using dynamic light scattering (DLS) and electrophoretic mobility measurements. The optimized LNP formulations were assessed for colloidal stability and homogeneity. Transfection efficiency was tested in L929 fibroblast cells using Cy3-labeled miRNA, with comparative analysis against transfection mediated by Lipofectamine™ 3000.
Results
Post-processing significantly affected LNP properties: sonication and filtration improved size uniformity and colloidal stability, while dialysis effectively reduced PDI without altering surface charge. In contrast, thermal treatment compromised particle stability. The optimized miRNA-LNPs successfully delivered Cy3-miRNA into L929 cells, demonstrating effective intracellular uptake. When compared to Lipofectamine™ 3000, the developed LNPs exhibited comparable—or in some cases superior—transfection efficiency, while showing lower cytotoxicity and better biocompatibility.
Discussion
These findings emphasize the importance of post-processing strategies in tuning the physicochemical properties of LNPs to enhance their performance as gene delivery vehicles. Microfluidic synthesis enabled the fabrication of uniform and scalable LNP formulations suitable for in vitro applications. The comparable performance of the developed LNPs to the commercial Lipofectamine™ 3000 highlights their potential as safer, customizable alternatives for non-viral miRNA delivery in biomedical research. This study provides a systematic framework for designing and optimizing LNPs tailored for nucleic acid delivery, reinforcing the value of microfluidic technology in nanomedicine development.
References
Kulkarni, J. A. et al. Nature Nanotechnology, 2021.
Pattipeiluhu, R. et al. Advanced Drug Delivery Reviews, 2023
Acknowledgments
This work was supported by the National Science Center (NCN) (2020/38/E/ST5/00456).
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