Numerical Investigation of the Permeation Characteristics of Reactive Oxygen and Nitrogen Species into Biological Membrane under Electric Field Using Classical Molecular Dynamics

Recently, plasma medicine has been rapidly developing as a new research field due to the establishment of discharge technology for the stable formation of cold atmospheric pressure plasma (CAP) [1]. In fact, useful effects for medical applications have been confirmed [2]. These phenomena are strongly related to radicals and ions generated from CAP. The plasma species penetrate the cell membrane and react with intracellular substances. As a result, a variety of cellular functions are induced. However, the detailed mechanisms of biological reactions are not well understood because of the complicated physicochemical processes in living cells. Thus, the aim of our study is to obtain the basic properties of membrane permeability of reactive oxygen and nitrogen species (RONS) in an all-atom model. In this report, the transport properties of RONS in the cell membrane, especially under electric field, are investigated using classical molecular dynamics (MD).<br/>In this analysis, 1-palmitoyl-2-oleoyl-sn-glyoero-3-phosphocholine (POPC) was chosen as the plasma-irradiated target. POPC is one of the major phospholipids in human cell membranes. The present membrane model consisted of 128 POPCs surrounded by 3812 H₂O molecules. The investigated RONS are O, OH, HO₂, and H₂O₂. To stabilize the molecular system, the total energy was minimized by using the steepest descent and conjugate gradient methods. The temperature and pressure were controlled by Langevin dynamics and Berendsen methods, respectively. 8 nanosecond equilibrium calculations were performed in the NPT ensemble at a temperature of 303 K and a pressure of 1 atm. After the above equilibration calculations, umbrella sampling (US) simulations were performed for a time period of 5 ns to obtain a histogram of the presence of RONS in the membrane. A sampling window was created every 1 Å along the perpendicular direction to the bilayer. The time increment of simulation and cutoff distance for the Coulomb and VDW forces were set to 2 fs and 1 nm, respectively. The applied electric field was set at 0.1-0.5 V/nm. The spatiotemporal trajectories of RONS were calculated using the MD software AMBER18.<br/>First, the position-dependent diffusion coefficients of OH, HO₂, and H₂O₂ were found to be low in the inner part of the membrane and high in the water layer. There was no significant difference in these diffusion coefficients. On the other hand, O showed higher diffusion coefficients in the inner part of the membrane than other RONS as described above. This result indicates non-polar molecules diffuse more easily inside the membrane than polar molecules.<br/>When an electric field was applied to the cell membrane, the free energy of the central part of the membrane decreased. At 0.3 V/nm, the membrane permeability coefficient of H₂O₂ was four times higher than that without the electric field. The electric field kept the energy bias of RONS to the membrane lowered. This effect assisted the permeation of RONS. When the electric field was increased to 0.4 V/nm, the permeation through the cell membrane was significantly enhanced.<br/>The formation of water channels at an electric field strength of 0.5 V/nm indicated that hydrophilic RONS were also significantly transported through the lipid bilayer of POPCs. In addition, differences in membrane permeability were observed depending on the RONS retention region. In particular, most of the ONOOH could easily permeate into the membrane within 1 ns after channel formation.<br/><br/>References<br/>[1] M. G Kong, G. Kroesen, G. Morfill, T. Nosenko, T. Shimizu, J. van Dijk, J. L. Zimmermann, New J. Phys. 11 (2009), 115012.<br/>[2] S. Toyokuni, Y. Ikehara, F. Kikkawa, and M. Hori, Plasma Medical Science, Academic Press (2018).