Synergistic Entry of Individual Nanoparticles into Mammalian Cells Driven by Free Energy Decline and Regulated by Their Sizes

Abstract

Cell entry is one of the common prerequisites for nanomaterial applications. Despite extensive studies on a homogeneous group of nanoparticles (NPs), fewer studies have been performed when two or more types of NPs were coadministrated. We previously described a synergistic cell entry process for two heterogeneous groups of NPs, where NPs functionalized with TAT (transactivator of transcription) peptide (T-NPs) stimulate the cellular uptake of coadministered unfunctionalized NPs (bystander NPs, B-NPs). Here, we show that the synergistic cell entry of NPs is driven by free energy decline and depends on B-NP sizes. Simulations showed that when separately placed initially, two NPs first move toward each other instead of initiating cell entry individually. Only T-NP invokes an inward bending of membrane mimicking endocytosis, which attracts the nearby NPs into the same “vesicle”. A two-phase free energy decline of the entire system occurred as two NPs get closer until contact, which is likely the thermodynamic driver for synergistic NP coentry. Experimentally, we found that T-NPs increase the apparent affinity of B-NPs to plasma membrane, suggesting that T-NPs help B-NPs “trapped” in the endocytic vesicles. Next, we varied the sizes of B-NPs and found that bystander activity peaks around 50 nm. Simulations also showed that the size of B-NPs influences the free energy decline, and thus the tendency and dynamics of NP coentry. These efforts provide a system to further understand the synergistic cell entry among individual NPs or multiple NP types on a biophysical basis and shed light on the future design of nanostructures for intracellular delivery.

Membrane-Specific Binding of 4 nm Lipid Nanoparticles Mediated by an Entropy-Driven Interaction Mechanism

“…After membrane binding, the NPs may further demonstrate complicated interactions with the cell membrane to achieve cellular entry for the realization of biomedical functions. NPs might adopt different pathways for cellular entry, such as transmembrane diffusion and endocytosis, ,,− which is significantly influenced by NP size, as well as other characteristic properties including particle shape and surface chemistry. ,− For example, it has been reported that zwitterionic sub 10 nm NPs primarily enter cells through passive diffusion; however, for larger NPs (e.g., ∼40–50 nm in size), their internalization mainly depends on endocytic pathways. ,, Nevertheless, the successful membrane binding of NPs is always the prerequisite for the following cellular internalization and biomedical functions.…”

“…Note that, in the membrane-binding process, the NP core does not interact with the membrane directly (Figure S1). Thus, rigid spheres with distinct surficial hydrophobic degree were used as simplified models of the cLNP core for computational efficiency. ,,,,, Then, similarly to the experimental works, ,− a cLNP was obtained by the self-assembly of this spherical NP core and 200 DOTAP molecules in an aqueous solution (Figure S16). Due to the hydrophobic surface of the core, DOTAP molecules could spontaneously adsorb on the NP core surface with their tails (Figure S16).…”

“…45,50−56 For example, it has been reported that zwitterionic sub 10 nm NPs primarily enter cells through passive diffusion; 45 however, for larger NPs (e.g., ∼40−50 nm in size), their internalization mainly depends on endocytic pathways. 50,51,54 Nevertheless, the successful membrane binding of NPs is always the prerequisite for the following cellular internalization and biomedical functions. Entropy-Driven Membrane Binding of cLNPs.…”

Co-administration of Transportan Peptide Enhances the Cellular Entry of Liposomes in the Bystander Manner Both In Vitro and In Vivo

“…Using a panel of pharmacological interventions, we showed that TP-assisted liposome uptake is most sensitive to the inhibitors of the macropinocytosis pathway, whereas others have much milder effects. We have performed several studies on the mechanism of bystander uptake using computation simulation. ,, These studies showed that first, macropinocytosis plays a predominate role for mediating bystander uptake and second, there is a free energy benefit for two or more NPs (e.g., inducer and bystander NPs) to enter the same endocytic vesicle, instead of internalizing separately. This mechanism may apply to CPPs as the inducer, and soft organic NPs such as liposomes, but it remains to be further validated.…”

Computational Study on the Regulatory Mechanism of Cell Membrane Wrapping on Liposomes by Embedded Bowl-like Nanostructures for Drug Delivery

“…Simulations presented in this work were performed based on the dissipative particle dynamics (DPD), which is a CG simulation technique being widely used for the study of NP cell membrane interactions. − In DPD, the dynamics of elementary units are controlled by Newton’s equation of motion, similar to the molecular dynamics method. The interaction between beads i and j is composed of three components, namely, a conservative force F ij C , a dissipative force F ij D , and a random force F ij R .…”