Temperature- and rigidity-mediated rapid transport of lipid nanovesicles in hydrogels.

Lipid nanovesicles are widely present as transport vehicles in living organisms and can serve as efficient drug delivery vectors. It is known that the size and surface charge of nanovesicles can affect their diffusion behaviors in biological hydrogels such as mucus. However, how temperature effects, including those of both ambient temperature and phase transition temperature (Tm), influence vehicle transport across various biological barriers outside and inside the cell remains unclear. Here, we utilize a series of liposomes with different Tm as typical models of nanovesicles to examine their diffusion behavior in vitro in biological hydrogels. We observe that the liposomes gain optimal diffusivity when their Tm is around the ambient temperature, which signals a drastic change in the nanovesicle rigidity, and that liposomes with Tm around body temperature (i.e., ∼37 °C) exhibit enhanced cellular uptake in mucus-secreting epithelium and show significant improvement in oral insulin delivery efficacy in diabetic rats compared with those with higher or lower Tm Molecular-dynamics (MD) simulations and superresolution microscopy reveal a temperature- and rigidity-mediated rapid transport mechanism in which the liposomes frequently deform into an ellipsoidal shape near the phase transition temperature during diffusion in biological hydrogels. These findings enhance our understanding of the effect of temperature and rigidity on extracellular and intracellular functions of nanovesicles such as endosomes, exosomes, and argosomes, and suggest that matching Tm to ambient temperature could be a feasible way to design highly efficient nanovesicle-based drug delivery vectors.

Diffusion of Nanoparticles with Activated Hopping in Crowded Polymer Solutions.

A long-distance hop of diffusive nanoparticles (NPs) in crowded environments was commonly considered unlikely, and its characteristics remain unclear. In this work, we experimentally identify the occurrence of the intermittent hops of large NPs in crowded entangled poly(ethylene oxide) (PEO) solutions, which are attributed to thermally induced activated hopping. We show that the diffusion of NPs in crowded solutions is considered as a superposition of the activated hopping and the reptation of the polymer solution. Such activated hopping becomes significant when either the PEO molecular weight is large enough or the NP size is relatively small. We reveal that the time-dependent non-Gaussianity of the NP diffusion is determined by the competition of the short-time relaxation of a polymer entanglement strand, the activated hopping, and the long-time reptation. We propose an exponential scaling law τhop/τe ∼ exp(d/dt) to characterize the hopping time scale, suggesting a linear dependence of the activated hopping energy barrier on the dimensionless NP size. The activated hopping motion can only be observed between the onset time scale of the short-time relaxation of local entanglement strands and the termination time scale of the long-time relaxation. Our findings on activated hopping provide new insights into long-distance transportation of NPs in crowded biological environments, which is essential to the delivery and targeting of nanomedicines.

Cancer-Cell-Membrane-Coated Nanoparticles with a Yolk-Shell Structure Augment Cancer Chemotherapy.

Despite rapid advancements in antitumor drug delivery, insufficient intracellular transport and subcellular drug accumulation are still issues to be addressed. Cancer cell membrane (CCM)-camouflaged nanoparticles (NPs) have shown promising potential in tumor therapy due to their immune escape and homotypic binding capacities. However, their efficacy is still limited due to inefficient tumor penetration and compromised intracellular transportation. Herein, a yolk-shell NP with a mesoporous silica nanoparticle (MSN)-supported PEGylated liposome yolk and CCM coating, CCM@LM, was developed for chemotherapy and exhibited a homologous tumor-targeting effect. The yolk-shell structure endowed CCM@LM with moderate rigidity, which might contribute to the frequent transformation into an ellipsoidal shape during infiltration, leading to facilitated penetration throughout multicellular spheroids in vitro (up to a 23.3-fold increase compared to the penetration of membrane vesicles). CCM@LM also exhibited a cellular invasion profile mimicking an enveloped virus invasion profile. CCM@LM was directly internalized by membrane fusion, and the PEGylated yolk (LM) was subsequently released into the cytosol, indicating the execution of an internalization pathway similar to that of an enveloped virus. The incoming PEGylated LM further underwent efficient trafficking throughout the cytoskeletal filament network, leading to enhanced perinuclear aggregation. Ultimately, CCM@LM, which co-encapsulated low-dose doxorubicin and the poly(ADP-ribose) polymerase inhibitor, mefuparib hydrochloride, exhibited a significantly stronger antitumor effect than the first-line chemotherapeutic drug Doxil. Our findings highlight that NPs that can undergo facilitated tumor penetration and robust intracellular trafficking have a promising future in cancer chemotherapy.

An improved surface passivation method for single-molecule studies.

We report a surface passivation method based on dichlorodimethylsilane (DDS)-Tween-20 for in vitro single-molecule studies, which, under the conditions tested here, more efficiently prevented nonspecific binding of biomolecules than the standard poly(ethylene glycol) surface. The DDS-Tween-20 surface was simple and inexpensive to prepare and did not perturb the behavior and activities of tethered biomolecules. It can also be used for single-molecule imaging in the presence of high concentrations of labeled species in solution.

Microfluidic synthesis of rigid nanovesicles for hydrophilic reagents delivery.

We present a hollow-structured rigid nanovesicle (RNV) fabricated by a multi-stage microfluidic chip in one step, to effectively entrap various hydrophilic reagents inside, without complicated synthesis, extensive use of emulsifiers and stabilizers, and laborious purification procedures. The RNV contains a hollow water core, a rigid poly (lactic-co-glycolic acid) (PLGA) shell, and an outermost lipid layer. The formation mechanism of the RNV is investigated by dissipative particle dynamics (DPD) simulations. The entrapment efficiency of hydrophilic reagents such as calcein, rhodamine B and siRNA inside the hollow water core of RNV is ≈90 %. In comparison with the combination of free Dox and siRNA, RNV that co-encapsulate siRNA and doxorubicin (Dox) reveals a significantly enhanced anti-tumor effect for a multi-drug resistant tumor model.

Monovalent, clickable, uncharged, water-soluble perylenediimide-cored dendrimers for target-specific fluorescent biolabeling.

Herein we report the synthesis of water-soluble polyglycerol-dendronized perylenediimides with a single reactive group that undergoes high-yielding click reactions. Single-molecule studies and target-specific biolabeling are reported, including the highly specific labeling of proteins on the surface of living bacterial and mammalian cells.

Rapid transport of germ-mimetic nanoparticles with dual conformational polyethylene glycol chains in biological tissues.

Polyethylene glycols (PEGs) can improve the diffusivity of nanoparticles (NPs) in biological hydrogels, while extended PEG chains severely impede cellular uptake of NPs. Inspired by invasive germs with flagellum-driven mucus-penetrating and fimbriae-mediated epithelium-adhering abilities, we developed germ-mimetic NPs (GMNPs) to overcome multiple barriers in mucosal and tumor tissues. In vitro studies and computational simulations revealed that the tip-specific extended PEG chains on GMNP functioned similarly to flagella, facilitating GMNP diffusion (up to 83.0-fold faster than their counterparts). Meanwhile, the packed PEG chains on the bodies of GMNP mediated strong adhesive interactions with cells similarly to the fimbriae, preserving cellular uptake efficiency. The in vivo results proved the superior tumor permeability and improved oral bioavailability provided by the GMNP (21.9-fold over administration of crystalline drugs). These findings offer useful guidelines for the rational design of NPs by manipulating surface polymer conformation to realize multiple functions and to enhance delivery efficacy.

A non-conjugated polyethylenimine copolymer-based unorthodox nanoprobe for bioimaging and related mechanism exploration.

Unconventional non-conjugated photoluminescent polymers have attracted increasing attention in bioimaging application, however their nonclassical photoluminescence mechanisms remain largely unclear. Herein, an amphiphilic copolymer polyethyleneimine-poly(d,l-lactide) (PEI-PDLLA) was synthesized and the obtained PEI-PDLLA copolymer exhibited intrinsic visible blue luminescence in the solid and concentrated solution states under 365 nm UV light irradiation. Using a computational assay approach, we investigated the unconventional photoluminescence mechanism of PEI-PDLLA. The results revealed that such photoluminescence should be related to the "clustered heteroatom chromophores" formed by through-space electronic interactions of N-heteroatoms in PEI. The copolymers can function as a fluorescent nanoprobe (PEI-PDLLA NPs) via a facile nanoprecipitation method and the self-assembly mechanism of PEI-PDLLA NPs was also investigated in-depth by molecular dynamics simulation. Intriguingly, the PEI-PDLLA NPs exhibited a remarkable excitation-dependent multi-wavelength emission characteristic, which was promising in acquiring a high precision imaging effect. Moreover, in contrast with conventional organic dyes with aggregation-caused quenching (ACQ), the fluorescence intensity of the PEI-PDLLA NPs was enhanced with increasing solution concentration. Furthermore, their applications in bioimaging indicated that PEI-PDLLA NPs could be utilized as a lysosome-specific and tumor-targeted nanoprobe with excellent photostability and good biocompatibility.

Chain-Length- and Saturation-Tuned Mechanics of Fluid Nanovesicles Direct Tumor Delivery.

Small unilamellar vesicles (SUVs), ubiquitous in organisms, play key and active roles in various biological processes. Although the physical properties of the constituent lipid molecules (i.e., the acyl chain length and saturation) are known to affect the mechanical properties of SUVs and consequently regulate their biological behaviors and functions, the underlying mechanism remains elusive. Here, we combined theoretical modeling and experimental investigation to probe the mechanical behaviors of SUVs with different lipid compositions. The membrane bending rigidity of SUVs increased with increasing chain length and saturation, resulting in differences in the vesicle rigidity and deformable capacity. Furthermore, we tested the tumor delivery capacity of liposomes with low, intermediate, and high rigidity as typical models for SUVs. Interestingly, liposomes with intermediate rigidity exhibited better tumor extracellular matrix diffusion and multicellular spheroid (MCS) penetration and retention than that of their stiffer or softer counterparts, contributing to improved tumor suppression. Stiff SUVs had superior cellular internalization capacity but intermediate tumor delivery efficacy. Stimulated emission depletion microscopy directly showed that the optimal formulation was able to transform to a rod-like shape in MCSs, which stimulated fast transport in tumor tissues. In contrast, stiff liposomes hardly deformed, whereas soft liposomes changed their shape irregularly, which slowed their MCS penetration. Our findings introduce special perspectives from which to map the detailed mechanical properties of SUVs with different compositions, provide clues for understanding the biological functions of SUVs, and suggest that liposome mechanics may be a design parameter for enhancing drug delivery.

Rapid transport of deformation-tuned nanoparticles across biological hydrogels and cellular barriers.

To optimally penetrate biological hydrogels such as mucus and the tumor interstitial matrix, nanoparticles (NPs) require physicochemical properties that would typically preclude cellular uptake, resulting in inefficient drug delivery. Here, we demonstrate that (poly(lactic-co-glycolic acid) (PLGA) core)-(lipid shell) NPs with moderate rigidity display enhanced diffusivity through mucus compared with some synthetic mucus penetration particles (MPPs), achieving a mucosal and tumor penetrating capability superior to that of both their soft and hard counterparts. Orally administered semi-elastic NPs efficiently overcome multiple intestinal barriers, and result in increased bioavailability of doxorubicin (Dox) (up to 8 fold) compared to Dox solution. Molecular dynamics simulations and super-resolution microscopy reveal that the semi-elastic NPs deform into ellipsoids, which enables rotation-facilitated penetration. In contrast, rigid NPs cannot deform, and overly soft NPs are impeded by interactions with the hydrogel network. Modifying particle rigidity may improve the efficacy of NP-based drugs, and can be applicable to other barriers.

c-axis preferential orientation of hydroxyapatite accounts for the high wear resistance of the teeth of black carp (Mylopharyngodon piceus).

Biological armors such as mollusk shells have long been recognized and studied for their values in inspiring novel designs of engineering materials with higher toughness and strength. However, no material is invincible and biological armors also have their rivals. In this paper, our attention is focused on the teeth of black carp (Mylopharyngodon piceus) which is a predator of shelled mollusks like snails and mussels. Nanoscratching test on the enameloid, the outermost layer of the teeth, indicates that the natural occlusal surface (OS) has much higher wear resistance compared to the other sections. Subsequent X-ray diffraction analysis reveals that the hydroxyapatite (HAp) crystallites in the vicinity of OS possess c-axis preferential orientation. The superior wear resistance of black carp teeth is attributed to the c-axis preferential orientation of HAp near the OS since the (001) surface of HAp crystal, which is perpendicular to the c-axis, exhibits much better wear resistance compared to the other surfaces as demonstrated by the molecular dynamics simulation. Our results not only shed light on the origin of the good wear resistance exhibited by the black carp teeth but are of great value to the design of engineering materials with better abrasion resistance.

Notch strengthening or weakening governed by transition of shear failure to normal mode fracture.

It is generally observed that the existence of geometrical discontinuity like notches in materials will lead to strength weakening, as a resultant of local stress concentration. By comparing the influence of notches to the strength of three typical materials, aluminum alloys with intermediate tensile ductility, metallic glasses with no tensile ductility, and brittle ceramics, we observed strengthening in aluminum alloys and metallic glasses: Tensile strength of the net section in circumferentially notched cylinders increases with the constraint quantified by the ratio of notch depth over notch root radius; in contrast, the ceramic exhibit notch weakening. The strengthening in the former two is due to resultant deformation transition: Shear failure occurs in intact samples while samples with deep notches break in normal mode fracture. No such deformation transition was observed in the ceramic, and stress concentration leads to its notch weakening. The experimental results are confirmed by theoretical analyses and numerical simulation. The results reported here suggest that the conventional criterion to use brittleness and/or ductility to differentiate notch strengthening or weakening is not physically sound. Notch strengthening or weakening relies on the existence of failure mode transition and materials exhibiting shear failure while subjected to tension will notch strengthen.

Effects of graphene oxide nanosheets on the ultrastructure and biophysical properties of the pulmonary surfactant film.

Graphene oxide (GO) is the most common derivative of graphene and has been used in a large range of biomedical applications. Despite considerable progress in understanding its cytotoxicity, its potential inhalation toxicity is still largely unknown. As the pulmonary surfactant (PS) film is the first line of host defense, interaction with the PS film determines the fate of the inhaled nanomaterials and their potential toxicity. Using a coarse-grained molecular dynamics model, we reported, for the first time, a novel mechanism of toxicity caused by the inhaled GO nanosheets. Upon deposition, the GO nanosheets induce pores in the PS film and thus have adverse effects on the ultrastructure and biophysical properties of the PS film. Notably, the pores induced by GO nanosheets result in increasing the compressibility of the PS film, which is an important indication of surfactant inhibition. In vitro experiments have also been conducted to study the interactions between GO and animal-derived natural PS films, qualitatively confirming the simulation results.

Microfluidic Synthesis of Hybrid Nanoparticles with Controlled Lipid Layers: Understanding Flexibility-Regulated Cell-Nanoparticle Interaction.

The functionalized lipid shell of hybrid nanoparticles plays an important role for improving their biocompatibility and in vivo stability. Yet few efforts have been made to critically examine the shell structure of nanoparticles and its effect on cell-particle interaction. Here we develop a microfluidic chip allowing for the synthesis of structurally well-defined lipid-polymer nanoparticles of the same sizes, but covered with either lipid-monolayer-shell (MPs, monolayer nanoparticles) or lipid-bilayer-shell (BPs, bilayer nanoparticles). Atomic force microscope and atomistic simulations reveal that MPs have a lower flexibility than BPs, resulting in a more efficient cellular uptake and thus anticancer effect than BPs do. This flexibility-regulated cell-particle interaction may have important implications for designing drug nanocarriers.

Tunable rigidity of (polymeric core)-(lipid shell) nanoparticles for regulated cellular uptake.

Core-shell nanoparticles (NPs) with lipid shells and varying water content and rigidity but with the same chemical composition, size, and surface properties are assembled using a microfluidic platform. Rigidity can dramatically alter the cellular uptake efficiency, with more-rigid NPs able to pass more easily through cell membranes. The mechanism accounting for this rigidity-dependent cellular uptake is revealed through atomistic-level simulations.

A microfluidic tubing method and its application for controlled synthesis of polymeric nanoparticles.

This report describes a straightforward but robust tubing method for connecting polydimethylsiloxane (PDMS) microfluidic devices to external equipment. The interconnection is irreversible and can sustain a pressure of up to 4.5 MPa that is characterized experimentally and theoretically. To demonstrate applications of this high-pressure tubing technique, we fabricate a semicircular microfluidic channel to implement a high-throughput, size-controlled synthesis of poly(lactic-co-glycolic acid) (PLGA) nanoparticles ranging from 55 to 135 nm in diameter. This microfluidic device allows for a total flow rate of 410 mL h(-1), resulting in enhanced convective mixing which can be utilized to precipitate small size nanoparticles with a good dispersion. We expect that this tubing technique would be widely used in microfluidic chips for nanoparticle synthesis, cell manipulation, and potentially nanofluidic applications.

Physicochemical properties of nanoparticles regulate translocation across pulmonary surfactant monolayer and formation of lipoprotein corona.

Interaction with the pulmonary surfactant film, being the first line of host defense, represents the initial bio-nano interaction in the lungs. Such interaction determines the fate of the inhaled nanoparticles and their potential therapeutic or toxicological effect. Despite considerable progress in optimizing physicochemical properties of nanoparticles for improved delivery and targeting, the mechanisms by which inhaled nanoparticles interact with the pulmonary surfactant film are still largely unknown. Here, using combined in vitro and in silico methods, we show how hydrophobicity and surface charge of nanoparticles differentially regulate the translocation and interaction with the pulmonary surfactant film. While hydrophilic nanoparticles generally translocate quickly across the pulmonary surfactant film, a significant portion of hydrophobic nanoparticles are trapped by the surfactant film and encapsulated in lipid protrusions upon film compression. Our results support a novel model of pulmonary surfactant lipoprotein corona associated with inhaled nanoparticles of different physicochemical properties. Our data suggest that the study of pulmonary nanotoxicology and nanoparticle-based pulmonary drug delivery should consider this lipoprotein corona.