Differential viscoelastic response of gel and fluid phase lipid vesicles probed by atomic force microscopy

Biological membranes, primarily composed of lipid bilayers, exhibit complex viscoelastic properties that are critical for cellular functions such as migration, endocytosis, and mechanotransduction. Although lipid composition is recognized as a key regulator of membrane mechanics, its role in shaping the apparent viscoelastic properties of membrane vesicles, particularly the distinct contributions of gel-phase vs fluid-phase lipids, remains inadequately characterized at the whole-vesicle level. To address this issue, we used atomic force microscopy (AFM) combined with a power-law rheological model to investigate the creep deformation processes of two typical types of lipid giant unilamellar vesicles (GUVs): gel-phase dipalmitoylphosphatidylcholine (DPPC) and fluid-phase 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). Quantitative measurements of rheological characteristics reveal distinct viscoelastic responses in gel-phase and fluid-phase GUVs during AFM indentation. Interestingly, fluid-phase DOPC GUVs exhibit nearly elastic mechanical behavior, characterized by a very low fluidity value of ∼0.06. In contrast, gel-phase (solid) DPPC GUVs demonstrate significantly more pronounced creep, accompanied by a higher fluidity value of around 0.12. Moreover, the analyzed storage modulus, loss modulus, and loss factor further confirm that DPPC vesicles exhibit significantly greater energy dissipation than DOPC vesicles. These distinct viscoelastic properties can be understood through the well-established difference in water permeability between the two lipid phases: the high permeability of fluid-phase DOPC membranes enables rapid hydraulic equilibration, leading to a near-elastic response, whereas the extremely low permeability of gel-phase DPPC membranes kinetically traps water, resulting in poroelastic relaxation and more pronounced creep. Additionally, gel-phase DPPC GUVs exhibited a steep initial stiffening regime followed by softening, which we interpret as pre-buckling stiffening of a pressurized elastic shell under indentation, a structural mechanical response distinct from that of fluid-phase DOPC vesicles. All extracted parameters represent apparent mechanical quantities that reflect the integrated structural response of the whole vesicle (membrane together with encapsulated fluid), rather than intrinsic material properties of the lipid bilayer alone. These findings advance the fundamental understanding of membrane vesicle mechanics under mechanical stress, offering insights into the viscoelastic regulation of vesicle-based biological processes.