Integrating Human and Nonhuman Primate Data to Estimate Human Tolerances for Traumatic Brain Injury.

Traumatic brain injury (TBI) contributes to a significant portion of the injuries resulting from motor vehicle crashes, falls, and sports collisions. The development of advanced countermeasures to mitigate these injuries requires a complete understanding of the tolerance of the human brain to injury. In this study, we developed a new method to establish human injury tolerance levels using an integrated database of reconstructed football impacts, subinjurious human volunteer data, and nonhuman primate data. The human tolerance levels were analyzed using tissue-level metrics determined using harmonized species-specific finite element (FE) brain models. Kinematics-based metrics involving complete characterization of angular motion (e.g., diffuse axonal multi-axial general evaluation (DAMAGE)) showed better power of predicting tissue-level deformation in a variety of impact conditions and were subsequently used to characterize injury tolerance. The proposed human brain tolerances for mild and severe TBI were estimated and presented in the form of injury risk curves based on selected tissue-level and kinematics-based injury metrics. The application of the estimated injury tolerances was finally demonstrated using real-world automotive crash data.

Influences of Hydrogen Bonding-Based Stabilization of Bolaamphiphile Layers on Molecular Diffusion within Organic Nanotubes Having Inner Carboxyl Groups.

This paper reports molecular diffusion behavior in two bolaamphiphile-based organic nanotubes having inner carboxyl groups with different inner dimeters (10 and 20 nm) and wall structures, COOH-ONT10nm and COOH-ONT20nm, using imaging fluorescence correlation spectroscopy (imaging FCS). The results were compared to those previously obtained in a similar nanotube with inner amine groups (NH2-ONT10nm). COOH-ONT10nm, as with NH2-ONT10nm, were formed from a rolled bolaamphiphile layer incorporating triglycine moieties, whereas COOH-ONT20nm consisted of four stacks of triglycine-free bolaamphiphile layers. Imaging FCS measurements were carried out for anionic sulforhodamine B (SRB), zwitterionic/cationic rhodamine B (RB), and cationic rhodamine-123 (R123) diffusing within ONTs (1-9 µm long) at different pH (3.4-8.4) and ionic strengths (1.6-500 mM). Diffusion coefficients (D) of these dyes in the ONTs were very small (0.01-0.1 µm2/s), reflecting the significant contributions of molecule-nanotube interactions to diffusion. The D of SRB was larger at higher pH and ionic strength, indicating the essential role of electrostatic repulsion that was enhanced by the deprotonation of the inner carboxyl groups. Importantly, the D of SRB was virtually independent of nanotube inner diameter and wall structure, indicating the diffusion of the hydrophilic molecule was controlled by short time scale adsorption/desorption processes onto the inner surface. In contrast, pH effects on D were less clear for relatively hydrophobic R123 and RB, suggesting the significant contributions of non-Coulombic interactions. Interestingly, the diffusion of these molecules in COOH-ONT20nm was slower than in COOH-ONT10nm. Slower diffusion in COOH-ONT20nm was attributable to relatively efficient partitioning of the hydrophobic dyes into the bolaamphiphile layers, which was reduced in COOH-ONT10nm due to the stabilization of its layer by polyglycine-II-type hydrogen bonding networks. These results show that, by tuning the bolaamphiphile structures and their intermolecular interactions, unique environments can be created within the nanospaces for enhanced molecular separations and reactions.

Diffusion Behavior of Differently Charged Molecules in Self-Assembled Organic Nanotubes Studied Using Imaging Fluorescence Correlation Spectroscopy.

The diffusion behavior of fluorescent molecules within bolaamphiphile-based organic nanotubes (ONTs) was systematically investigated using imaging fluorescence correlation spectroscopy (imaging FCS). Anionic sulforhodamine B, zwitterionic/cationic rhodamine B, or cationic rhodamine 123 was loaded into ONTs having cylindrical hollow structures (ca. 10 nm in inner diameter) with amine and glucose groups on the inner and outer surfaces, respectively. Wide-field fluorescence video microscopy was used to acquire imaging FCS data for dye-doped ONTs in aqueous solutions of different ionic strengths (1-500 mM) at different pH (3.4-8.4). The diffusion behavior of these dyes was discussed on the basis of their apparent diffusion coefficients ( D) that were determined by autocorrelating the time transient of fluorescence intensity at each pixel on an ONT. Molecular diffusion in the ONTs was significantly slowed by the molecule-nanotube interactions, as shown by the very small D (10-1 to 10-2 µm2/s). The pH dependence of D revealed that dye diffusion was basically controlled by electrostatic interactions associated with the protonation of the amine groups on the ONT inner surface. The pH-dependent change in D was observed over a wide pH range, possibly because of electrostatically induced variations in the p Ka of the densely packed ammonium ions on the ONT inner surface. On the other hand, the influence of ionic strength on D was relatively unclear, suggesting the involvement of non-Coulombic interactions with the ONTs in molecular diffusion. Importantly, individual ONTs of different lengths (1-5 µm) afforded similar diffusion coefficients for each type of dye at each solution condition, implying that the properties of the ONTs were uniform in terms of solute loading and release. These results highlight the characteristics of the molecular diffusion behavior within the ONTs and will help in the design of ONTs better suited for use as drug vehicles and contaminant adsorbents.

Preparation and Formation Process of Zn(II)-Coordinated Nanovesicles.

Mixing a glycylglycine lipid and zinc acetate has been reported to form novel supramolecular Zn(II)-coordinated nanovesicles in ethanol. In this study, we investigate in detail the formation of nanovesicles by using three lipids at different temperatures and discuss their formation process. The original lipids show extremely low solubilities and appear as plate structures in ethanol. Within a small window of lipid solubility, the formation of lipid-Zn(II) complexes occurs mainly on the solid surfaces of plate structures. Controlling of the lipid solubility by temperature affects the kinetics of complex formation and the subsequent transformation of the complexes into nanovesicles and nanotubes. An improved method of two-step control of temperature is developed for preparing all the three kinds of nanovesicles. We provide new insights into the formation process of nanovesicles based on several control experiments. A tetrahedral lipid-cobalt(II) complex similarly produces nanovesicles, whereas an octahedral complex gives sheet structures. Mixing of zinc acetate with a ß-alanyl-ß-alanine lipid can only give sheet structures, which lack a polyglycine II hydrogen-bond network and induce no morphological changes. We conclude that the formation of the lipid-Zn(II) complexes on solid plate structures, tetrahedral geometry, and polyglycine II hydrogen-bond network in the complexes shall work cooperatively for the formation of Zn(II)-coordinated nanovesicles.

Spectroscopic imaging studies of nanoscale polarity and mass transport phenomena in self-assembled organic nanotubes.

Synthetic organic nanotubes self-assembled from bolaamphiphile surfactants are now being explored for use as drug delivery vehicles. In this work, several factors important to their implementation in drug delivery are explored. All experiments are performed with the nanotubes immersed in ethanol. First, Nile Red (NR) and a hydroxylated Nile Red derivative (NR-OH) are loaded into the nanotubes and spectroscopic fluorescence imaging methods are used to determine the apparent dielectric constant of their local environment. Both are found in relatively nonpolar environments, with the NR-OH molecules preferring regions of relatively higher dielectric constant compared to NR. Unique two-color imaging fluorescence correlation spectroscopy (imaging FCS) measurements are then used along with the spectroscopic imaging results to deduce the dielectric properties of the environments sensed by mobile and immobile populations of probe molecules. The results reveal that mobile NR molecules pass through less polar regions, likely within the nanotube walls, while immobile NR molecules are found in more polar regions, possibly near the nanotube surfaces. In contrast, mobile and immobile NR-OH molecules are found to locate in environments of similar polarity. The imaging FCS results also provide quantitative data on the apparent diffusion coefficient for each dye. The mean diffusion coefficient for the NR dye was approximately two-fold larger than that of NR-OH. Slower diffusion by the latter could result from its additional hydrogen bonding interactions with polar triglycine, amine, and glucose moieties near the nanotube surfaces. The knowledge gained in these studies will allow for the development of nanotubes that are better engineered for applications in the controlled transport and release of uncharged, dipolar drug molecules.

Effect of Photoinduced Size Changes on Protein Refolding and Transport Abilities of Soft Nanotubes.

Self-assembly of azobenzene-modified amphiphiles (Glyn Azo, n=1-3) in water at room temperature in the presence of a protein produced nanotubes with the protein encapsulated in the channels. The Gly2 Azo nanotubes (7 nm internal diameter [i.d.]) promoted refolding of some encapsulated proteins, whereas the Gly3 Azo nanotubes (13 nm i.d.) promoted protein aggregation. Although the 20 nm i.d. channels of the Gly1 Azo nanotubes were too large to influence the encapsulated proteins, narrowing of the i.d. to 1 nm by trans-to-cis photoisomerization of the azobenzene units of the Gly1 Azo monomers packed in the solid bilayer membranes led to a squeezing out of the proteins into the bulk solution and simultaneously enhanced their refolding ratios. In contrast, photoinduced transformation of the Gly2 Azo nanotubes to short nanorings (<40 nm) with a large i.d. (28 nm) provided no further refolding assistance. We thus demonstrate that pertubation by the solid bilayer membrane wall of the nanotubes is important to accelerate refolding of the denatured proteins during their transport in the narrow nanotube channels.

Lipid Nanotube Tailored Fabrication of Uniquely Shaped Polydopamine Nanofibers as Photothermal Converters.

Helically coiled and linear polydopamine (PDA) nanofibers were selectively fabricated with two different types of lipid nanotubes (LNTs) that acted as templates. The obtained coiled PDA-LNT hybrid showed morphological advantages such as higher light absorbance and photothermal conversion effect compared to a linear counterpart. Laser irradiation of the coiled PDA-LNT hybrid induced a morphological change and subsequent release of the encapsulated guest molecule. In cellular experiments, the coiled PDA-LNT efficiently eliminated HeLa cells because of its strong affinity with the tumor cells. This work illustrates the first approach to construct characteristic morphologies of PDA nanofibers using LNTs as simple templates, and the coiled PDA-LNT hybrid exhibits attractive photothermal features derived from its unique coiled shape.

Supramolecular Self-Assembly into Biofunctional Soft Nanotubes: From Bilayers to Monolayers.

The inner and outer surfaces of bilayer-based lipid nanotubes can be hardly modified selectively by a favorite functional group. Monolayer-based nanotubes display a definitive difference in their inner and outer functionalities if bipolar wedge-shaped amphiphiles, so-called bolaamphiphiles, as a constituent of the monolayer membrane pack in a parallel fashion with a head-to-tail interface. To exclusively form unsymmetrical monolayer lipid membranes, we focus herein on the rational molecular design of bolaamphiphiles and a variety of self-assembly processes into tubular architectures. We first describe the importance of polymorph and polytype control and then discuss diverse methodologies utilizing a polymer template, multiple hydrogen bonds, binary and ternary coassembly, and two-step self-assembly. Novel biologically important functions of the obtained soft nanotubes, brought about only by completely unsymmetrical inner and outer surfaces, are discussed in terms of protein refolding, drug nanocarriers, lectin detection, a chiral inducer for achiral polymers, the tailored fabrication of polydopamine, and spontaneous nematic alignment.

Imaging fluorescence correlation spectroscopy studies of dye diffusion in self-assembled organic nanotubes.

The rate and mechanism of diffusion by anionic sulforhodamine B (SRB) dye molecules within organic nanotubes self-assembled from bolaamphiphile surfactants were investigated by imaging fluorescence correlation spectroscopy (imaging-FCS). The inner and outer surfaces of the nanotubes are terminated with amine and glucose groups, respectively; the former allow for pH-dependent manipulation of nanotube surface charge while the latter enhance their biocompatibility. Wide-field fluorescence video microscopy was used to locate and image dye-doped nanotubes dispersed on a glass surface. Imaging-FCS was then used to spatially resolve the SRB transport dynamics. Mobilization of the dye molecules was achieved by immersion of the nanotubes in buffer solution. Experiments were performed in pH 6.4, 7.4 and 8.4 buffers, at ionic strengths ranging from 1.73 mM to 520 mM. The results show that coulombic interactions between cationic ammonium ions on the inner nanotube surface and the anionic SRB molecules play a critical role in governing mass transport of the dye. The apparent dye diffusion coefficient, D, was found to generally increase with increasing ionic strength and with increasing pH. The D values obtained were found to be invariant along the nanotube length. Mass transport of the SRB molecules within the nanotubes is concluded to occur by a desorption-mediated Fickian diffusion mechanism in which dye motion is slowed by its coulombic interactions with the inner surfaces of the nanotubes. The results of these studies afford information essential to the use of organic nanotubes in controlled drug release applications.

Photoinduced morphological transformations of soft nanotubes.

In water, synthetic amphiphiles composed of a photoresponsive azobenzene moiety and an oligoglycine hydrogen-bonding moiety selectively self-assembled into nanotubes with solid bilayer membranes. The nanotubes underwent morphological transformations induced by photoisomerization of the azobenzene moiety within the membranes, and the nature of the transformation depended on the number of glycine residues in the oligoglycine moiety (i.e., on the strength of intermolecular hydrogen bonding). Upon UV-light irradiation of nanotubes prepared from amphiphiles with the diglycine residue, trans-to-cis isomerization induced a transformation from nanotubes (inner diameter (i.d.) 7 nm), several hundreds of nanometers to several tens of micrometers in length, to imperfect nanorings (i.d. 21-38 nm). The cis-to-trans isomerization induced by continuous visible-light irradiation resulted in the stacking of the imperfect nanorings to form nanotubes with an i.d. of 25 nm and an average length of 310 nm, which were never formed by a self-assembly process. Time-lapse fluorescence microscopy enabled us to visualize the transformation of nanotubes with an i.d. of 20 nm (self-assembled from amphiphiles with the monoglycine residue) to cylindrical nanofibers with an i.d. of 1 nm; shrinkage of the hollow cylinders started at the two open ends with simultaneous elongation in the direction of the long axis.

Spontaneous nematic alignment of a lipid nanotube in aqueous solutions.

The dispersibility and liquid crystal formation of a self-assembled lipid nanotube (LNT) was investigated in a variety of aqueous solutions. As the lipid component, we chose a bipolar lipid with glucose and tetraglycine headgroups, which self-assembled into an LNT with a small outer diameter of 16 to 17 nm and a high axial ratio of more than 310. The LNT gave a stable colloidal dispersion in its dilute solutions and showed spontaneous liquid crystal (LC) alignment at relatively low concentrations and in a pH region including neutral pH. The LNT samples with shorter length distributions were prepared by sonication, and the relationship between the LNT axial ratio and the minimum LC formation concentration was examined. The robustness of the LNT made the liquid crystal stable in mixed solvents of water/ethanol, water/acetone, and water/tetrahydrofuran (1:1 by volume) and at a temperature of up to 90 °C in water. The observed colloidal behavior of the LNT was compared to those of similar 1D nanostructures such as a phospholipid tubule.

Control of self-assembled morphology and molecular packing of asymmetric glycolipids by association/dissociation with poly(thiopheneboronic acid).

The molecular packing and self-assembled morphologies of asymmetric bolaamphiphiles, N-(2-aminoethyl)-N'-(ß-d-glucopyranosyl)alkanediamide [1(n), n = 12, 14, 16, 17, 18, and 20], were precisely controlled by association/dissociation with poly(thiopheneboronic acid) (PTB). Differential scanning calorimetry, X-ray diffraction, and infrared spectroscopy revealed that the starting film of 1(n) associated with 1 equiv of the boronic acid moiety of PTB, (Film-1(n)PTB), had antiparallel molecular packing of 1(n) moiety within the monolayer membranes. However, the molecular packing of the starting film that contained 0.5 equiv of the boronic acid moiety of PTB (Film-2eq1(n)PTB) was parallel. The dispersion of Film-1(n)PTB in water gave only nanotapes, whereas that of Film-2eq1(n)PTB in water selectively formed nanotubes, through a dissociation reaction of PTB based on the hydrolysis of the boronate esters in the complexes. The nanotapes and nanotubes memorized the antiparallel and parallel molecular packing of the starting films, respectively. Changes in the length of the oligomethylene spacer of 1(n) never affected the molecular packing or self-assembled morphologies. However, the inner diameters of the nanotubes increased irregularly in the range of 67.9-79.6 nm as the length of the oligomethylene spacer of 1(n) increased from n = 12 to n = 18.

Reinforcement of a sugar-based bolaamphiphile/functionalized graphene oxide composite gel: rheological and electrochemical properties.

A sugar-based bolaamphiphile/graphene oxide composite hydrogel has been prepared using simple mixing. Unlike the corresponding sugar-based native gel, the composite gel exhibits a fibrillar structure with a 10-20 nm fiber diameter. The composite gel forms an interdigitated bilayer structure incorporating intermolecular hydrogen-bonding interactions. The composite gel formation did not change the beneficial electrical properties of graphene offering the potential for integration of this new material into electronic systems. Interestingly, the mechanical and electrochemical properties of the composite gel are both dramatically enhanced when compared to the native gel, thereby reflecting that the functionalized graphene oxide layers are efficiently intercalated within the composite gel structure.

Buffers to suppress sodium dodecyl sulfate adsorption to polyethylene oxide for protein separation on capillary polymer electrophoresis.

Although polyethylene oxide (PEO) offers several advantages as a sieving polymer in SDS capillary polymer electrophoresis (SDS-CPE), solution properties of PEO cause deterioration in the electrophoresis because PEO in solution aggregates itself, degrades into smaller pieces, and forms polymer-micelle complexes with SDS. We examined protein separation on SDS-CPE with PEO as a sieving matrix in four individual buffer solutions: Tris-CHES, Tris-Gly, Tris-Tricine, and Tris-HCl buffers. The solution properties of PEO as a sieving matrix in those buffers were examined by dynamic light scattering (DLS) and by surface tension. Preferential SDS adsorption onto PEO disturbed protein-SDS complexation and impaired the protein separation efficiency. Substantial adsorption of SDS to PEO was particularly observed in Tris-Gly buffer. The Tris-CHES buffer prevented SDS from adsorbing onto the PEO. Only Tris-CHES buffer achieved separation of six proteins. This study demonstrated efficient protein separation on SDS-CPE with PEO.

Glycolipid nanotube templates for the production of hydrophilic/hydrophobic and left/right-handed helical polydiacetylene nanotubes.

Encapsulation and preorganization of diacetylene monomers in glycolipid nanotubes allows for the production of polydiacetylene nanotubes with hydrophilic/hydrophobic surfaces and left/right-handed helicities.

Confinement effect of organic nanotubes toward green fluorescent protein (GFP) depending on the inner diameter size.

Transportation, release behavior, and stability of a green fluorescent protein (GFP, 3x4 nm) in self-assembled organic nanotubes with three different inner diameters (10, 20, and 80 nm) have been studied in terms of novel nanocontainers. Selective immobilization of a fluorescent acceptor dye on the inner surface enabled us to not only visualize the transportation of GFP in the nanochannels but to also detect release of the encapsulated GFP to the bulk solution in real time, based on fluorescence resonance energy transfer (FRET). Obtained diffusion constants and release rates of GFP markedly decreased as the inner diameter of the nanotubes was decreased. An endo-sensing procedure also clarified the dependence of the thermal and chemical stabilities of the GFP on the inner diameters. The GFP encapsulated in the 10 nm nanochannel showed strong resistance to heat and to a denaturant. On the other hand, the 20 nm nanochannel accelerated the denaturation of the encapsulated GFP compared with the rate of denaturation of the free GFP in bulk and the encapsulated GFP in the 80 nm nanochannels. The confinement effect based on rational fitting of the inner diameter to the size of GFP allowed us to store it stably and without denaturation under high temperatures and high denaturant concentrations.

Dynamic light-scattering measurement of sieving polymer solutions for protein separation on SDS CE.

We evaluated the mesh size and homogeneity of polymer network by dynamic light scattering and discussed the relationship between the physical properties of polymer network and the protein separation behavior by capillary polymer electrophoresis. We compared three kinds of sieving polymers in solutions with a wide range of molecular weights and concentrations: polyacrylamide and polyethylene oxide as flexible polymers, and hydroxyethyl cellulose as a semiflexible polymer. We found that the mobility of protein was dominated primarily by the mesh size xi, irrespective of the type of sieving polymers, and the peak spacing between protein peaks increased drastically in the range of xi<10 nm, where the mobility also decreased. And the peak widths were dependent on the molecular species of sieving polymers and their homogeneity of polymer network. We proposed that a polymer network with a homogenous mesh size of less than 10 nm is the best sieving medium for separation of the proteins in the molecular weight range 14,300-97,200 Da from the view point of the resolution in protein separation.

Relationship between frontal car-to-car test result and vehicle crash compatibility evaluation in mobile progressive deformable barrier test.

Objective: In 2020, the world's first crash compatibility rating test will be introduced in the European mobile progressive deformable barrier (MPDB) test. In this research, the quantitative change in partner protection performance of large vehicles in car-to-car (C2C) impacts was studied if these large vehicles were designed in future based on MPDB tests addressing crash compatibility ratings. Methods: Representative vehicles of the European fleet were selected and a Computer Aided Engineering (CAE) parameter study was conducted. In particular, by changing an indicator of structural interaction performance (SD; i.e., the degree of uniformity of barrier deformation)/mass/stiffness of large vehicles systematically in a step-by-step approach, the compatibility evaluation results of large vehicles in MPDB and the occupant injury score of small vehicles in C2C impacts were compared. The CAE result was evaluated compared to that of C2C physical impact tests. Results: The CAE parameter study showed that in the C2C impact condition, the effects on occupant injury in a small vehicle due to changes in the large vehicle were as follows: (1) SD change: The effect was minor except for small overlap condition. (2) Mass and stiffness change: The effect was relatively major. On the other hand, compatibility evaluation in the MPDB showed a tendency to overestimate the effect of SD change in comparison with the above-mentioned C2C impact condition. In addition, physical impact tests showed that, based on SD evaluation, the large vehicle with a relatively inferior compatibility rating compared to those with superior compatibility ratings showed a contradicting trend of better compatibility performance in the C2C test. Conclusions: The currently proposed compatibility evaluation method of the MPDB test showed some tendency to overestimate the effect of SD change and resulted in quantitatively inconsistent outcomes regarding occupant injury in the partner car in C2C impact conditions.

Controllable biomolecule release from self-assembled organic nanotubes with asymmetric surfaces: pH and temperature dependence.

The release behavior of fluorescent dyes, oligo DNAs and spherical proteins from self-assembled organic nanotubes having 7-9 nm inner diameters has been studied in terms of novel nanocontainers with high-axial ratios. Both much smaller inner diameters and asymmetric inner and outer surfaces are characteristic of the nanotubes. The acid-dissociation constant (pKa) of the amino groups located at the inner surface and the thermal phase transition temperature (Tg-l) of the nanotube were evaluated based on the pH titration and variable-temperature circular dichroism (CD) spectroscopic experiments, respectively. Each guest was slowly released from both open ends of the nanotube under weak alkaline conditions (pH 8.5), as a result of the decrease in electrostatic attraction between the inner surface and the guests. Elevated temperatures above the obtained Tg-l converted the monolayer membrane of the nanotube from a solid state to a fluid one, promoting the remarkably fast release of the guests. The unique release properties of the nanotube as a nanocontainer with two terminal open ends were compared with those of liposomes that posses a closed hollow space covered with fluid bilayer membranes.