Recently, we demonstrated that lipid droplets (LDs) are rapidly upregulated by host cells very early following viral infection, and this upregulation underpins the production of antiviral cytokines (type-I and -III interferons) and eventual restriction of viral replication (ZIKV, DENV and HSV-1). However, the specific role of lipids in LD-mediated viral restriction remains unknown.
We performed lipidomics analysis to fully characterise the lipid landscape of virally induced LDs, isolated following ZIKV infection of astrocytes (8 and 24 hpi). A significant shift in the lipidome of antiviral LDs was observed which was underpinned by (1) a global upregulation of long, and very-long polyunsaturated fatty acids, including docosahexaenoic acid and arachidonic acid, known antiviral lipids; and (2) a small but significant increase in PI and PE phospholipids to the membrane. Since no system exists to experimentally investigate the role of individual antiviral lipids accumulated in LDs, we next sought to develop an artificial lipid droplet (aLD) delivery system. Leveraging our new understanding of the phospholipid exterior of native LDs following viral infection, we optimised aLDs to contain complex phospholipid membranes incorporated with a novel fluorescent phospholipid tag which enabled enhanced aLD delivery to a wide range of cell types, and antiviral activity when tested against ZIKV (increased IFN-β and a 50% decrease in ZIKV-mRNA) in vitro. These aLDs were also successfully delivered in vivo, showing biodistributions to the brain, lungs, heart and liver in mice, and a wider organ distribution in zebrafish, with aLD entry to all organ systems over 48 hours.
As a small particle delivery system, aLDs containing complex phospholipids demonstrate superior organ delivery to other small particles, such as extracellular vesicles (EVs) and lipid nanoparticles (LNPs). Future work has begun to test our novel aLDs in delivering antiviral long-chain fatty acid cargo to the brain in viral infection models.