Lipid Peroxidation Drives Liquid-Liquid Phase Separation and Disrupts Raft Protein Partitioning in Biological Membranes.

The peroxidation of membrane lipids by free radicals contributes to aging, numerous diseases, and ferroptosis, an iron-dependent form of cell death. Peroxidation changes the structure and physicochemical properties of lipids, leading to bilayer thinning, altered fluidity, and increased permeability of membranes in model systems. Whether and how lipid peroxidation impacts the lateral organization of proteins and lipids in biological membranes, however, remains poorly understood. Here, we employ cell-derived giant plasma membrane vesicles (GPMVs) as a model to investigate the impact of lipid peroxidation on ordered membrane domains, often termed membrane rafts. We show that lipid peroxidation induced by the Fenton reaction dramatically enhances the phase separation propensity of GPMVs into coexisting liquid-ordered (Lo) and liquid-disordered (Ld) domains and increases the relative abundance of the disordered phase. Peroxidation also leads to preferential accumulation of peroxidized lipids and 4-hydroxynonenal (4-HNE) adducts in the disordered phase, decreased lipid packing in both Lo and Ld domains, and translocation of multiple classes of raft proteins out of ordered domains. These findings indicate that the peroxidation of plasma membrane lipids disturbs many aspects of membrane rafts, including their stability, abundance, packing, and protein and lipid composition. We propose that these disruptions contribute to the pathological consequences of lipid peroxidation during aging and disease and thus serve as potential targets for therapeutic intervention.

Lipid peroxidation drives liquid-liquid phase separation and disrupts raft protein partitioning in biological membranes.

The peroxidation of membrane lipids by free radicals contributes to aging, numerous diseases, and ferroptosis, an iron-dependent form of cell death. Peroxidation changes the structure, conformation and physicochemical properties of lipids, leading to major membrane alterations including bilayer thinning, altered fluidity, and increased permeability. Whether and how lipid peroxidation impacts the lateral organization of proteins and lipids in biological membranes, however, remains poorly understood. Here, we employ cell-derived giant plasma membrane vesicles (GPMVs) as a model to investigate the impact of lipid peroxidation on ordered membrane domains, often termed membrane rafts. We show that lipid peroxidation induced by the Fenton reaction dramatically enhances phase separation propensity of GPMVs into co-existing liquid ordered (raft) and liquid disordered (non-raft) domains and increases the relative abundance of the disordered, non-raft phase. Peroxidation also leads to preferential accumulation of peroxidized lipids and 4-hydroxynonenal (4-HNE) adducts in the disordered phase, decreased lipid packing in both raft and non-raft domains, and translocation of multiple classes of proteins out of rafts. These findings indicate that peroxidation of plasma membrane lipids disturbs many aspects of membrane rafts, including their stability, abundance, packing, and protein and lipid composition. We propose that these disruptions contribute to the pathological consequences of lipid peroxidation during aging and disease, and thus serve as potential targets for therapeutic intervention.

Enhancement of cell migration and wound healing by nano-herb ointment formulated with biosurfactant, silver nanoparticles and Tridax procumbens.

Introduction: Ointments are generally used as a therapeutic agent for topical medication or transdermal drug delivery, such as wound healing and skin lesions. Methods: In this study, Tridax procumbens plant extract (0.7 g/mL) was used to prepare herbal-infused oil as the oil phase and gelatin-stabilized silver nanoparticle (G-AgNPs) (0.3 g/mL) as the aqueous phase. To blend the oil and aqueous phases, rhamnolipid biosurfactant with a critical micelle concentration of 55 mg/L from strain Pseudomonas aeruginosa PP4 has been used for herb ointment preparation. The average size of the synthesized G-AgNPs was observed between 10-30 nm and confirmed as spherical-shaped particles by TEM analysis. Subsequently, GC-MS and FTIR characterization are used to confirm herb ointment's chemical and functional characteristics. Results: Based on the antibacterial studies, the highest microbial growth inhibition was observed for herb ointment, about 19.5 mm for the pathogen Staphylococcus aureus at the concentration of 100 µg/mL, whereas 15.5 mm was obtained for Escherichia coli, respectively. In addition, the minimum inhibitory concentration (MIC) assay showed negligible bacterial growth at 100 µg/mL for S. aureus and E. coli, respectively. Moreover, the cell viability assay for herb ointment exhibited low cytotoxic activity at higher concentrations (100 µg/mL) in Vero cell lines. In this study, wound scratch assay showed a significant cell migration rate (90 ± 2%) in 3 days of incubation than the control (62 ± 2%). Discussion: As a result, the biosurfactant-based nano-topical herb ointment revealed a low cytotoxic and higher cell migration capacity. Altogether, these findings highlighted the utility of this herb ointment in therapeutic applications such as wound healing.

High-Content Imaging Platform to Discover Chemical Modulators of Plasma Membrane Rafts.

Plasma membrane organization profoundly impacts cellular functionality. A well-known mechanism underlying this organization is through nanoscopic clustering of distinct lipids and proteins in membrane rafts. Despite their physiological importance, rafts remain a difficult-to-study aspect of membrane organization, in part because of the paucity of chemical tools to experimentally modulate their properties. Methods to selectively target rafts for therapeutic purposes are also currently lacking. To tackle these problems, we developed a high-throughput screen and an accompanying image analysis pipeline to identify small molecules that enhance or inhibit raft formation. Cell-derived giant plasma membrane vesicles were used as the experimental platform. A proof-of-principle screen using a bioactive lipid library demonstrates that this method is robust and capable of validating established raft modulators including C6- and C8-ceramide, miltefosine, and epigallocatechin gallate as well as identifying new ones. The platform we describe here represents a powerful tool to discover new chemical approaches to manipulate rafts and their components.