Fluorescent Labeling and Imaging of IL-22 mRNA-Loaded Lipid Nanoparticles

Background

In ulcerative colitis, interleukin-22 (IL-22) plays a crucial role by promoting mucosal healing and regulating the inflammatory response. Lipid nanoparticles (LNPs) offer a targeted delivery platform for IL-22 in this context, effectively harnessing the cytokine's therapeutic potential to address mucosal healing and inflammation precisely at the site of injury [1]. Despite the increasing interest in utilizing LNPs for targeted delivery of therapeutic agents in ulcerative colitis, our understanding of their in vivo behavior is limited, hindering the clinical translation of LNP-based therapies. The tracking of LNPs in vivo can provide crucial insights into their biodistribution, migration abilities, and mechanism of action [2]. Therefore, the development of efficient and sensitive techniques for labeling LNPs is highly desired.

To date, several methods have been developed to unravel the in vivo dynamics of LNPs. Notably, the use of fluorescent dyes to label LNPs stands out as an effective approach for confirming successful therapeutic delivery. This highly sensitive and selective technique enables real-time monitoring and visualization of nanoparticle behavior and distribution in biological systems. However, a potential limitation of fluorescent labeling is the risk of dye leakage from nanoparticles in vivo, resulting in diminished brightness over time and the development of a background signal that may hinder accurate nanoparticle localization [3].

In a prior study, we engineered LNPs loaded with IL-22 mRNA for treating ulcerative colitis, evaluating the biodistribution of DiR-labeled IL-22/LNP [4]. In this protocol, we will describe the detailed process of labeling and imaging IL-22/LNP in the gastrointestinal (GI) tract using a fluorescent dye via the IVIS spectrum. In our previously published study [4], LNPs loaded with mRNA as described in this protocol displayed a distinct signal in the targeted organ (colon) and exhibited therapeutic efficacy.