Triglyceride Lenses at the Air-Water Interface as a Model System for Studying the Initial Stage in the Biogenesis of Lipid Droplets

Lipid droplets (LD) are intracellular structures consisting of an apolar lipid core, composed mainly of triglycerides (TG) and steryl esters, coated by a lipid-protein mixed monolayer. The mechanisms underlying LD biogenesis at the endoplasmic reticulum membrane are a matter of many current investigations. Although models explaining the budding-off of protuberances of phase-segregated TG inside bilayers have been proposed recently, the assumption of such initial blisters needs further empirical support. Here, we study mixtures of egg phosphatidylcholine (EPC) and TG at the air-water interface in order to describe some physical properties and topographic stability of TG bulk structures in contact with interfaces. Brewster angle microscopy images revealed the appearance of microscopic collapsed structures (CS) with highly reproducible lateral size (similar to 1 mu m lateral radius) not varying with lateral packing changes and being highly stable at surface pressures (p) beyond collapse. By surface spectral fluorescence microscopy, we were able to characterize the solvatochromism of Nile Red both in monolayers and inside CS. This allowed to conclude that CS corresponded to a phase of liquid TG and to characterize them as lenses forming a three-phase (oil-water-air) system. Thereby, the thicknesses of the lenses could be determined, observing that they were dramatically flattened when EPC was present (6-12 nm compared to 30-50 nm for lenses on EPC/TG and TG films, respectively). Considering the shape of lenses, the interfacial tensions, and the Neumann's triangle, this experimental approach allows one to estimate the oil-water interfacial tension acting at each individual microscopic lens and at varying compression states of the surrounding monolayer. Thus, lenses formed on air-water Langmuir films can serve to assess variables of relevance to the initial step of LD biogenesis, such as the degree of dispersion of excluded-TG phase and shape, spatial distribution, and oil-water interfacial tension of lenses.