Effects of cholesterol on the bending modulus of membranes and the electroporation induced pore formation of vesicles

Abstract

The effects of cholesterol (chol) in the membranes of giant unilamellar vesicles (GUVs) on the size distribution of vesicles and the irreversible electroporation (IRE)-induced pore formation in GUVs were investigated. Dioleoylphosphatidylcholine (DOPC)/chol-GUVs and dioleoylphosphatidylglycerol (DOPG)/DOPC/chol-GUVs were prepared using the natural swelling method. The average sizes of GUVs were obtained from the size distribution histograms of vesicles, which were fitted by the lognormal distribution. The size of GUVs increased with the increase of chol in neutral and charged membranes. Using the theorical approach, the values of bending modulus (Kben) were obtained 19.1  0.1, 23.1 ± 0.1, 28.6 ± 0.01 and 31.0 ± 0.1 kBT (here kB is the Boltzmann constant and T is the absolute temperature) for DOPC/chol-GUVs ratios of 100/0, 85/15, 71/29 and 60/40, respectively. The values of Kben were also found to increase with chol for charged membrane. IRE signal (pulsating direct current) induced lateral electric tension in the membranes of GUVs. The time dependent fraction of intact GUVs among all the examined GUVs was fitted to a single exponential decay function from where the rate constant (kp) of pore formation was obtained. The values of kp were obtained (1.0 ± 0.1)×10-2 s-1 at 7.0 mN/m, (2.9 ± 0.3)×10-2 s-1 at 8.0 mN/m, (11.0 ± 0.3)×10-2 s-1 at 9.0 mN/m for ratio of DOPG/DOPC/chol-GUVs as 46/39/15. Similar increase of kp was also obtained for other chol content. The estimated line tension was observed to increase from 12.9 to 14.6 pN with the corresponding increase of chol from 15 to 40 mole%. The increased energy barrier in the prepore state, due to the increase of chol, was the main factor for decreasing the kp. Hence, the increase of Kben due to chol controlled the kp of IRE-induced pore formation in GUVs.