Converting Nanotoxicity Data to Information Using Artificial Intelligence and Simulation.

Decades of nanotoxicology research have generated extensive and diverse data sets. However, data is not equal to information. The question is how to extract critical information buried in vast data streams. Here we show that artificial intelligence (AI) and molecular simulation play key roles in transforming nanotoxicity data into critical information, i.e., constructing the quantitative nanostructure (physicochemical properties)-toxicity relationships, and elucidating the toxicity-related molecular mechanisms. For AI and molecular simulation to realize their full impacts in this mission, several obstacles must be overcome. These include the paucity of high-quality nanomaterials (NMs) and standardized nanotoxicity data, the lack of model-friendly databases, the scarcity of specific and universal nanodescriptors, and the inability to simulate NMs at realistic spatial and temporal scales. This review provides a comprehensive and representative, but not exhaustive, summary of the current capability gaps and tools required to fill these formidable gaps. Specifically, we discuss the applications of AI and molecular simulation, which can address the large-scale data challenge for nanotoxicology research. The need for model-friendly nanotoxicity databases, powerful nanodescriptors, new modeling approaches, molecular mechanism analysis, and design of the next-generation NMs are also critically discussed. Finally, we provide a perspective on future trends and challenges.

Phosphatidylinositol 4,5-Bisphosphate Sensing Lipid Raft via Inter-Leaflet Coupling Regulated by Acyl Chain Length of Sphingomyelin.

Phosphatidylinositol 4,5-bisphosphate (PIP2) is an important molecule located at the inner leaflet of cell membrane, where it serves as anchoring sites for a cohort of membrane-associated molecules and as a broad-reaching signaling intermediate. The lipid raft is thought as the major platform recruiting proteins for signal transduction and also known to mediate PIP2 accumulation across the membrane. While the significance of this cross-membrane coupling is increasingly appreciated, it remains unclear whether and how PIP2 senses the dynamic change of the ordered lipid domains over the packed hydrophobic core of the bilayer. Herein, by means of molecular dynamic simulation, we reveal that inner PIP2 molecules can sense the outer lipid domain via inter-leaflet coupling, and the coupling manner is dictated by the acyl chain length of sphingomyelin (SM) partitioned to the lipid domain. Shorter SM promotes membrane domain registration, whereby PIP2 accumulates beneath the domain across the membrane. In contrast, the anti-registration is thermodynamically preferred if the lipid domain has longer SM due to the hydrophobic mismatch between the corresponding acyl chains in SM and PIP2. In this case, PIP2 is expelled by the domain with a higher diffusivity. These results provide molecular insights into the regulatory mechanism of correlation between the outer lipid domain and inner PIP2, both of which are critical components for cell signal transduction.

Competitive and/or cooperative interactions of graphene-family materials and benzo[a]pyrene with pulmonary surfactant: a computational and experimental study.

BACKGROUND: Airborne nanoparticles can be inhaled and deposit in human alveoli, where pulmonary surfactant (PS) molecules lining at the alveolar air-water interface act as the first barrier against inhaled nanoparticles entering the body. Although considerable efforts have been devoted to elucidate the mechanisms underlying nanoparticle-PS interactions, our understanding on this important issue is limited due to the high complexity of the atmosphere, in which nanoparticles are believed to experience transformations that remarkably change the nanoparticles' surface properties and states. By contrast with bare nanoparticles that have been extensively studied, relatively little is known about the interactions between PS and inhaled nanoparticles which already adsorb contaminants. In this combined experimental and computational effort, we investigate the joint interactions between PS and graphene-family materials (GFMs) with coexisting benzo[a]pyrene (BaP). RESULTS: Depending on the BaP concentration, molecular agglomeration, and graphene oxidation, different nanocomposite structures are formed via BaPs adsorption on GFMs. Upon deposition of GFMs carrying BaPs at the pulmonary surfactant (PS) layer, competition and cooperation of interactions between different components determines the interfacial processes including BaP solubilization, GFM translocation and PS perturbation. Importantly, BaPs adsorbed on GFMs are solubilized to increase BaP's bioavailability. By contrast with graphene adhering on the PS layer to release part of adsorbed BaPs, more BaPs are released from graphene oxide, which induces a hydrophilic pore in the PS layer and shows adverse effect on the PS biophysical function. Translocation of graphene across the PS layer is facilitated by BaP adsorption through segregating it from contact with PS, while translocation of graphene oxide is suppressed by BaP adsorption due to the increase of surface hydrophobicity. Graphene extracts PS molecules from the layer, and the resultant PS depletion declines with graphene oxidation and BaP adsorption. CONCLUSION: GFMs showed high adsorption capacity towards BaPs to form nanocomposites. Upon deposition of GFMs carrying BaPs at the alveolar air-water interface covered by a thin PS layer, the interactions of GFM-PS, GFM-BaP and BaP-PS determined the interfacial processes of BaP solubilization, GFM translocation and PS perturbation.

Crossing Biological Barriers by Engineered Nanoparticles.

Engineered nanoparticles (ENPs) may cause toxicity if they cross various biological barriers and are accumulated in vital organs. Which factors affect barrier crossing efficiency of ENPs are crucial to understand. Here, we present strong data showing that various nanoparticles crossed biological barriers to enter vital animal organs and cause toxicity. We also point out that physicochemical properties of ENPs, modifications of ENPs in biofluid, and physiological and pathological conditions of the body all affect barrier crossing efficiency. We also summarized our limited understanding of the related mechanisms. On the basis of this summary, major research gaps and direction of further efforts are then discussed.

Microplastics Reduce Lipid Digestion in Simulated Human Gastrointestinal System.

Microplastics (MPs) are unavoidably ingested by humans, and their gastrointestinal processes and impact on lipid digestion are unknown. In the present work, all five MP types used, including polystyrene (PS), polyethylene terephthalate, polyethylene, polyvinyl chloride, and poly(lactic-co-glycolic acid) (80 mg/L in small intestine), significantly reduced lipid digestion in the in vitro gastrointestinal system. PS MPs exhibited the highest inhibition (12.7%) among the five MPs. Lipid digestion decreased with increasing PS concentration, but independent of PS size (50 nm, 1 µm, 10 µm). PS MPs after photoaging by simulated sunlight also significantly decreased lipid digestion. Confocal imaging shows that PS MPs could interact with both lipid droplets and lipases. Two mechanisms underlying the PS-induced digestion inhibition were revealed using both experimental and molecular dynamics simulation approaches: (1) PS MPs decreased the bioavailability of lipid droplets via forming large lipid-MPs heteroaggregates due to the high MP hydrophobicity; and (2) PS MPs adsorbed lipase, and reduced its activity by changing the secondary structure and disturbing the essential open conformation. The first mechanism (MP-lipid interaction) played a more important role in lipid digestion reduction based on interaction energy calculation. These findings reveal potential risk of MPs to human digestion health and nutrient assimilation.

Mechanics of the Formation, Interaction, and Evolution of Membrane Tubular Structures.

Membrane nanotubes, also known as membrane tethers, play important functional roles in many cellular processes, such as trafficking and signaling. Although considerable progresses have been made in understanding the physics regulating the mechanical behaviors of individual membrane nanotubes, relatively little is known about the formation of multiple membrane nanotubes due to the rapid occurring process involving strong cooperative effects and complex configurational transitions. By exerting a pair of external extraction upon two separate membrane regions, here, we combine molecular dynamics simulations and theoretical analysis to investigate how the membrane nanotube formation and pulling behaviors are regulated by the separation between the pulling forces and how the membrane protrusions interact with each other. As the force separation increases, different membrane configurations are observed, including an individual tubular protrusion, a relatively less deformed protrusion with two nanotubes on its top forming a V shape, a Y-shaped configuration through nanotube coalescence via a zipper-like mechanism, and two weakly interacting tubular protrusions. The energy profile as a function of the separation is determined. Moreover, the directional flow of lipid molecules accompanying the membrane shape transition is analyzed. Our results provide new, to our knowledge, insights at a molecular level into the interaction between membrane protrusions and help in understanding the formation and evolution of intra- and intercellular membrane tubular networks involved in numerous cell activities.

Directional and Rotational Motions of Nanoparticles on Plasma Membranes as Local Probes of Surface Tension Propagation.

Mechanical heterogeneity is ubiquitous in plasma membranes and of essential importance to cellular functioning. As a feedback of mechanical stimuli, local surface tension can be readily changed and immediately propagated through the membrane, influencing structures and dynamics of both inclusions and membrane-associated proteins. Using the nonequilibrium coarse-grained membrane simulation, here we investigate the inter-related processes of tension propagation, lipid diffusion, and transport of nanoparticles (NPs) adhering on the membrane of constant tension gradient, mimicking that of migrating cells or cells under prolonged stimulation. Our results demonstrate that the lipid bilayer membrane can by itself propagate surface tension in defined rates and pathways to reach a dynamic equilibrium state where surface tension is linearly distributed along the gradient maintained by the directional flow-like motion of lipids. Such lipid flow exerts shearing forces to transport adhesive NPs toward the region of a larger surface tension. Under certain conditions, the shearing force can generate nonzero torques driving the rotational motion of NPs, with the direction of the NP rotation determined by the NP-membrane interaction state as functions of both NP property and local membrane surface tension. Such features endow NPs with promising applications ranging from biosensing to targeted drug delivery.

Stabilizing Effect of Inherent Knots on Proteins Revealed by Molecular Dynamics Simulations.

A growing number of proteins have been identified as knotted in their native structures, with such entangled topological features being expected to play stabilizing roles maintaining both the global fold and the nature of proteins. However, the molecular mechanism underlying the stabilizing effect is ambiguous. Here, we combine unbiased and mechanical atomistic molecular dynamics simulations to investigate how a protein is stabilized by an inherent knot by directly comparing chemical, thermal, and mechanical denaturing properties of two proteins having the same sequence and secondary structures but differing in the presence or absence of an inherent knot. One protein is YbeA from Escherichia coli, containing a deep trefoil knot within the sequence, and the other is the modified protein with the knot of YbeA being removed. Under certain chemical denaturing conditions, the unknotted protein fully unfolds whereas the knotted protein does not, suggesting a higher intrinsic stability for the protein having a knot. Both proteins unfold under enhanced thermal fluctuations but at different rates and with distinct pathways. Opening the hydrophobic core via separation between two α-helices is identified as a crucial step initiating the protein unfolding, which, however, is restrained for the knotted protein by topological and geometrical frustrations. Energy barriers for denaturing the protein are reduced by removing the knot, as evidenced by mechanical unfolding simulations. Finally, yet importantly, no obvious change in size or location of the knot was observed during denaturing processes, indicating that YbeA may remain knotted for a relatively long time during and after denaturation.

Role of Lipid Coating in the Transport of Nanodroplets across the Pulmonary Surfactant Layer Revealed by Molecular Dynamics Simulations.

Hydrophilic drugs can be delivered into lungs via nebulization for both local and systemic therapies. Once inhaled, ultrafine nanodroplets preferentially deposit in the alveolar region, where they first interact with the pulmonary surfactant (PS) layer, with nature of the interaction determining both efficiency of the pulmonary drug delivery and extent of the PS perturbation. Here, we demonstrate by molecular dynamics simulations the transport of nanodroplets across the PS layer being improved by lipid coating. In the absence of lipids, bare nanodroplets deposit at the PS layer to release drugs that can be directly translocated across the PS layer. The translocation is quicker under higher surface tensions but at the cost of opening pores that disrupt the ultrastructure of the PS layer. When the PS layer is compressed to lower surface tensions, the nanodroplet prompts collapse of the PS layer to induce severe PS perturbation. By coating the nanodroplet with lipids, the disturbance of the nanodroplet on the PS layer can be reduced. Moreover, the lipid-coated nanodroplet can be readily wrapped by the PS layer to form vesicular structures, which are expected to fuse with the cell membrane to release drugs into secondary organs. Properties of drug bioavailability, controlled drug release, and enzymatic tolerance in real systems could be improved by lipid coating on nanodroplets. Our results provide useful guidelines for the molecular design of nanodroplets as carriers for the pulmonary drug delivery.

Membrane nanotube pearling restricted by confined polymers.

Increasing evidence showed that membrane nanotubes readily undergo pearling in response to external stimuli, while long tubular membrane structures have been observed connecting cells and functioning as channels for intercellular transport, raising a fundamental question of how the stability of membrane nanotubes is maintained in the cellular environment. Here, combining dissipative particle dynamics simulations, free energy calculations, and a force analysis, we propose and demonstrate that nanotube pearling can be restricted by confined polymers, which can be DNA and protein chains transported through the nanotubes, or actin filaments participating in tube formation and elongation. Thermodynamically, nanotube pearling releases the membrane surface energy, but costs bending energies of both the membrane and the confined polymers. Following the mechanism, the pearling of nanotubes confining longer and stiffer polymers is more difficult as it costs larger polymer bending energies. In dynamics, nanotube pearling occurs by repelling polymers from the region of nanotube shrinking to that of swelling. Shorter polymers can be readily repelled owing to the unbalanced force exerted by the shrinking tube region, whereas longer polymers tend to be trapped at the shrinking region to retard the nanotube pearling. Besides the low surface tension maintained by lipid reservoirs kept in living cells, our results supplement the explanation for the stability of membrane nanotubes, and open up a new avenue to manipulate the shape deformation of tubular membrane structures for study of many biological processes.

Size-dependent formation of membrane nanotubes: continuum modeling and molecular dynamics simulations.

Membrane nanotubes play important functional roles in numerous cell activities such as cellular transport and communication. By exerting an external pulling force over a finite region in a membrane patch, here we investigate the size dependence of the membrane nanotube formation under the continuum and atomistic modeling frameworks. It is shown that the membrane undergoes a discontinuous shape transition as the size of the pulling region and the membrane tension increase. A formula characterizing the nonlinear relationship between the maximum static pulling force and pulling size is identified. During the membrane extraction, lipids in the upper and lower leaflets exhibit different behaviors of structural rearrangements. Moreover, our computational simulations indicate that the steady state pulling force increases linearly with the pulling velocity as well as the size of the pulling region.

Interaction pathways between soft lipid nanodiscs and plasma membranes: A molecular modeling study.

Lipid nanodisc, a model membrane platform originally synthesized for study of membrane proteins, has recently been used as the carrier to deliver amphiphilic drugs into target tumor cells. However, the central question of how cells interact with such emerging nanomaterials remains unclear and deserves our research for both improving the delivery efficiency and reducing the side effect. In this work, a binary lipid nanodisc is designed as the minimum model to investigate its interactions with plasma membranes by using the dissipative particle dynamics method. Three typical interaction pathways, including the membrane attachment with lipid domain exchange of nanodiscs, the partial membrane wrapping with nanodisc vesiculation, and the receptor-mediated endocytosis, are discovered. For the first pathway, the boundary normal lipids acting as ligands diffuse along the nanodisc rim to gather at the membrane interface, repelling the central bola lipids to reach a stable membrane attachment. If bola lipids are positioned at the periphery and act as ligands, they diffuse to form a large aggregate being wrapped by the membrane, leaving the normal lipids exposed on the membrane exterior by assembling into a vesicle. Finally, by setting both central normal lipids and boundary bola lipids as ligands, the receptor-mediated endocytosis occurs via both deformation and self-rotation of the nanodiscs. All above pathways for soft lipid nanodiscs are quite different from those for rigid nanoparticles, which may provide useful guidelines for design of soft lipid nanodiscs in widespread biomedical applications.

Identifying Cu(ii)-amyloid peptide binding intermediates in the early stages of aggregation by resonance Raman spectroscopy: a simulation study.

The aggregation of amyloid beta (Aß) peptides plays a crucial role in the pathology and etiology of Alzheimer's disease. Experimental evidence shows that copper ion is an aggregation-prone species with the ability to coordinately bind to Aß and further induce the formation of neurotoxic Aß oligomers. However, the detailed structures of Cu(ii)-Aß complexes have not been illustrated, and the kinetics and dynamics of the Cu(ii) binding are not well understood. Two Cu(ii)-Aß complexes have been proposed to exist under physiological conditions, and another two might exist at higher pH values. By using ab initio simulations for the spontaneous resonance Raman and time domain stimulated resonance Raman spectroscopy signals, we obtained the characteristic Raman vibronic features of each complex. These signals contain rich structural information with high temporal resolution, enabling the characterization of transient states during the fast Cu-Aß binding and interconversion processes.

Transport of nanoparticles across pulmonary surfactant monolayer: a molecular dynamics study.

Pulmonary nanodrug delivery is an emerging concept, especially for targeted lung cancer therapy. Once inhaled, the nanoparticles (NPs) acting as drug carriers need to efficiently cross the pulmonary surfactant monolayer (PSM) of lung alveoli, which act as the first barrier for external particles entering the lung. Herein, by performing molecular dynamics simulations, we study how inhaled NPs interact with the PSM, particularly focusing on the transport of NPs with different properties across the PSM. While hydrophilic NPs translocate directly across the PSM, transport of hydrophobic NPs is achieved as the PSM wraps them. Intriguingly, when hydrophilic NPs are decorated with lipid molecules (LCNPs), they are wrapped by the PSM efficiently with mild PSM perturbation. Moreover, the structure formed is like a vesicle, which will likely fuse with cell membranes to accomplish the transport of hydrophilic NPs into secondary organs. This behavior makes the LCNP a prospective candidate for pulmonary nanodrug delivery. Herein, the effects of the physical properties of LCNPs on their transport are investigated. Increasing the LCNP size promotes its wrapping by reducing the PSM bending energy. The binding energy that drives transport can be strengthened by increasing the lipid coating density and the lipid tail length, both of which also reduce the risk of PSM rupture during transport. These results should help researchers understand how to better use surface decorations to achieve efficient pulmonary entry, which may provide useful guidance for the design of nano-based platforms for inhaled drug delivery.

Pulling force and surface tension drive membrane fusion.

Despite catalyzed by fusion proteins of quite different molecular architectures, intracellular, viral, and cell-to-cell fusions are found to have the essential common features and the nearly same nature of transition states. The similarity inspires us to find a more general catalysis mechanism for membrane fusion that minimally depends on the specific structures of fusion proteins. In this work, we built a minimal model for membrane fusion, and by using dissipative particle dynamics simulations, we propose a mechanism that the pulling force generated by fusion proteins initiates the fusion process and the membrane tension regulates the subsequent fusion stages. The model shows different features compared to previous computer simulation studies: the pulling force catalyzes membrane fusion through lipid head overcrowding in the contacting region, leading to an increase in the head-head repulsion and/or the unfavorable head-tail contacts from opposing membranes, both of which destabilize the contacting leaflets and thus promote membrane fusion or vesicle rupture. Our simulations produce a variety of shapes and intermediates, closely resembling cases seen experimentally. Our work strongly supports the view that the tight pulling mechanism is a conserved feature of fusion protein-mediated fusion and that the membrane tension plays an essential role in fusion.

Exploring the shape deformation of biomembrane tubes with theoretical analysis and computer simulation.

The shape deformation of membrane nanotubes is studied by a combination of theoretical analysis and molecular simulation. First we perform free energy analysis to demonstrate the effects of various factors on two ideal states for the pearling transition, and then we carry out dissipative particle dynamics simulations, through which various types of membrane tube deformation are found, including membrane pearling, buckling, and bulging. Different models for inducing tube deformation, including the osmotic pressure, area difference and spontaneous curvature models, are considered to investigate tubular instabilities. Combined with free energy analysis, our simulations show that the origin of the deformation of membrane tubes in different models can be classified into two categories: effective spontaneous curvature and membrane tension. We further demonstrate that for different models, a positive membrane tension is required for the pearling transition. Finally we show that different models can be coupled to effectively deform the membrane tube.

Inter-tube adhesion mediates a new pearling mechanism.

A common mechanism for intracellular transport is the controlled shape transformation, also known as pearling, of membrane tubes. Exploring how tube pearling takes place is thus of quite importance to not only understand the bio-functions of tubes, but also promote their potential biomedical applications. While the pearling mechanism of one single tube is well understood, both the pathway and the mechanism of pearling of multiple tubes still remain unclear. Herein, by means of computer simulations we show that the tube pearling can be mediated by the inter-tube adhesion. By increasing the inter-tube adhesion strength, each tube undergoes a discontinuous transition from no pearling to thorough pearling. The discontinuous pearling transition is ascribed to the competitive variation between tube surface tension and the extent of inter-tube adhesion. Besides, the final pearling instability is also affected by tube diameter and inter-tube orientation. Thinner tubes undergo inter-tube lipid diffusion before completion of pearling. The early lipid diffusion reduces the extent of inter-tube adhesion and thus restrains the subsequent pearling. Therefore, only partial or no pearling can take place for two thinner tubes. For two perpendicular tubes, the pearling is also observed, but with different pathways and higher efficiency. The finite size effect is discussed by comparing the pearling of tubes with different lengths. It is expected that this work will not only provide new insights into the mechanism of membrane tube pearling, but also shed light on the potential applications in biomaterials science and nanomedicine.

How tubular aggregates interact with biomembranes: wrapping, fusion and pearling.

How soft tubular aggregates interact with biomembranes is crucial for understanding the formation of membrane tubes connecting two eukaryotic cells, which are initially created from one cell and then connect with the other. On the other hand, recent experiments have shown that tubular polymersomes display different cellular internalization kinetics in their biomedical applications compared with spherical ones with an underlying mechanism that is not fully understood. Inspired by above observations, in this work we investigate how tubular aggregates interact with biomembranes with the aid of computer simulation techniques. We identify three different pathways for membrane interaction with parallel tubes: membrane wrapping, tube-membrane fusion and tube pearling. For the first pathway, soft tubes can be wrapped from the top side by membranes through membrane monolayer protrusion, which cooperatively leads to a heterogeneous wrapping dynamics along with tube deformation. The second pathway found is that soft tubes fuse with the membrane under certain conditions. Both wrapping and fusion have distinct influence on the third pathway, tube pearling. While a weak membrane adhesion promotes tube pearling, the strong adhesion that leads to higher extent of membrane wrapping conversely restrains tube pearling. Under highly positive membrane tension, partial tube-membrane fusion provides another way to mediate tube pearling. The findings shed light on the formation of a bridge membrane tube and the rational design of tube-based therapeutic agents with improved efficiency for targeted cellular delivery.

Lipid extraction mediates aggregation of carbon nanospheres in pulmonary surfactant monolayers.

Increasing evidence indicates that carbon nanoparticles (CNPs), which mainly originate from incomplete combustion of fossil fuels, have an adverse impact on the respiratory system. Recent in vivo experiments have shown that the pulmonary toxicity of CNPs is attributed to their aggregation in pulmonary surfactant monolayers (PSMs) while the underlying mechanism of aggregation remains unclear. Here, by performing coarse grained molecular dynamics simulations, we demonstrate for the first time that the aggregation of carbon nanospheres (CNSs) in PSMs is in fact size-dependent and mediated by lipid extractions. Upon CNS deposition, neighbouring lipid molecules are extracted from PSMs to cover CNSs from the top side. The extracted lipids induce clustering of CNSs to maximize the CNS-lipid interaction, by forming inverse micelles to wrap the aggregated CNSs cooperatively. The formed CNS clusters perturb the molecule structure of the PSM and thus affect its biofunction on respiration. Our simulations show that during the expiration process, CNSs form clusters that perturb the mechanical properties of the PSM in a manner depending on the CNS size. With deep inspiration, a high concentration of large CNSs may induce PSM rupture and thus have a potential impact on its biophysical properties.

How transmembrane peptides insert and orientate in biomembranes: a combined experimental and simulation study.

After the synthesis of transmembrane peptides/proteins (TMPs), their insertion into a lipid bilayer is a fundamental biophysical process. Moreover, correct orientations of TMPs in membranes determine the normal functions they play in relevant cellular activities. In this study, we have established a method to determine the orientation of TMPs in membranes. This method is based on the use of TAMRA, a fluorescent molecule with high extinction coefficient and fluorescence quantum yield, to act as a fluorescent probe and tryptophan as a quencher. Fluorescence quenching indicates that the model peptide displays membrane orientation with the N terminus outside and the C terminus inside dominantly. To elucidate the underlying mechanism, we performed molecular dynamics simulations. Our simulations suggest that both membrane insertion and the orientation of TMPs are determined by complex competition and cooperation between hydrophobic and electrostatic interactions. After initial membrane anchorage via electrostatic interactions of the charged residues with the lipid headgroups, further insertion is hindered by unfavorable interactions between the polar residues and lipid tails, which result in an energy barrier. Nevertheless, such a finite energy barrier is reduced by hydrophobic interactions between the non-polar residues and lipid tails. Moreover, a transient terminal flipping was captured to facilitate the membrane insertion. Once the inserted terminus reaches the opposite lipid headgroups, the hydrophobic interactions cooperate with the electrostatic interactions to complete the membrane insertion process.