Inkjet printing of bioadhesives

Over the past century, synthetic adhesives have largely displaced their natural counterparts in medical applications. However, rising concerns over the environmental and toxicological effects of the solvents, monomers, and additives used in synthetic adhesives have recently led the scientific community to seek natural substitutes. Marine mussel adhesive protein is a formaldehyde-free natural adhesive that demonstrates excellent adhesion to several classes of materials, including glasses, metals, metal oxides, and polymers. In this study, we have demonstrated computer aided design (CAD) patterning of various biological adhesives using piezoelectric inkjet technology. A MEMS-based piezoelectric actuator was used to control the flow of the mussel adhesive protein solution through the ink jet nozzles. Fourier transform infrared spectroscopy (FTIR), microscopy, and adhesion studies were performed to examine the chemical, structural, and functional properties of these patterns, respectively. FTIR revealed the piezoelectric inkjet technology technique to be nondestructive. Atomic force microscopy was used to determine the extent of chelation caused by Fe(III). The adhesive strength in these materials was correlated with the extent of chelation by Fe(III). Piezoelectric inkjet printing of naturally-derived biological adhesives may overcome several problems associated with conventional tissue bonding materials. This technique may significantly improve wound repair in next generation eye repair, fracture fixation, wound closure, and drug delivery devices.     

Controlled Release in Transdermal Pressure Sensitive Adhesives using Organosilicate Nanocomposites

Polydimethyl siloxane (PDMS) based pressure sensitive adhesives (PSA) incorporating organo-clays at different loadings were fabricated via solution casting. Partially exfoliated nanocomposites were obtained for the hydroxyl terminated PDMS in ethyl acetate solvent as determined by X-ray diffraction and atomic force microscopy. Drug release studies showed that the initial burst release was substantially reduced and the drug release could be controlled by the addition of organo-clay. Shear strength and shear adhesion failure temperature (SAFT) measurements indicated substantial improvement in adhesive properties of the PSA nanocomposite adhesives. Shear strength showed more than 200% improvement at the lower clay loadings and the SAFT increased by about 21% due to the reinforcement provided by the nano-dispersed clay platelets. It was found that by optimizing the level of the organosilicate additive to the polymer matrix, superior control over drug release kinetics and simultaneous improvements in adhesive properties could be attained for a transdermal PSA formulation.     

Adhesive bond strength and compliance for denture soft lining materials.

Peel bond strength and tensile bond strength between three polyvinylsiloxane denture soft liners and a heat-cured acrylic resin denture base were measured using two adhesive systems. The soft lining materials differed only in regard of their filler content and compliance. The values of bond strength and mode of failure were explained in terms of the inherent strength of the bond and varying compliance and tear strength of the soft material. For tensile testing, when bond failure occurred through an adhesive debonding mechanism, materials of low compliance (stiffer materials) produced the greatest tensile bond strength. Conversely, when the same materials were subjected to peel testing a different trend emerged; the material with lowest compliance produced the lowest peel bond strength. When de-bonding occurs by tearing or snapping of the soft material, the measured value of bond strength was controlled by the tear strength of the soft material. The results could be explained by a consideration of stress concentrations at the soft-hard material interface during 180 degrees peel testing. Adhesives based on ethyl acetate solvents produced stronger bond strengths than equivalent toluene based adhesives, particularly for materials of low compliance. Bond failure for toluene based adhesives was predominantly adhesive, whereas that for ethyl acetate based adhesives was predominantly cohesive. Overall, the least resistance to peeling was exhibited by a material of low compliance (i.e. relatively stiff) bonded with a toluene based adhesive. When an ethyl acetate based adhesive was used, all materials exhibited a resistance to peeling with a predominantly cohesive mode of failure.     

Electromechanical properties of dried tendon and isoelectrically focused collagen hydrogels

Assembling artificial collagenous tissues with structural, functional, and mechanical properties which mimic natural tissues is of vital importance for many tissue engineering applications. While the electro-mechanical properties of collagen are thought to play a role in, for example, bone formation and remodeling, this functional property has not been adequately addressed in engineered tissues. Here the electro-mechanical properties of rat tail tendon are compared with those of dried isoelectrically focused collagen hydrogels using piezoresponse force microscopy under ambient conditions. In both the natural tissue and the engineered hydrogel D-periodic type I collagen fibrils are observed, which exhibit shear piezoelectricity. While both tissues also exhibit fibrils with parallel orientations, Fourier transform analysis has revealed that the degree of parallel alignment of the fibrils in the tendon is three times that of the dried hydrogel. The results obtained demonstrate that isoelectrically focused collagen has similar structural and electro-mechanical properties to that of tendon, which is relevant for tissue engineering applications.     

Biomimetic remineralization as a progressive dehydration mechanism of collagen matrices – Implications in the aging of resin–dentin bonds

Biomineralization is a dehydration process in which water from the intrafibrillar compartments of collagen fibrils are progressively replaced by apatites. As water is an important element that induces a lack of durability of resin–dentin bonds, this study has examined the use of a biomimetic remineralization strategy as a progressive dehydration mechanism to preserve joint integrity and maintain adhesive strength after ageing. Human dentin surfaces were bonded with dentin adhesives, restored with resin composites and sectioned into sticks containing the adhesive joint. Experimental specimens were aged in a biomimetic analog-containing remineralizing medium and control specimens in simulated body fluid for up to 12 months. Specimens retrieved after the designated periods were examined by transmission electron microscopy for the presence of water-rich regions using a silver tracer and for collagen degradation within the adhesive joints. Tensile testing was performed to determine the potential loss of bond integrity after ageing. Control specimens exhibited severe collagen degradation within the adhesive joint after ageing. Remineralized specimens exhibited progressive dehydration, as manifested by silver tracer reduction and partial remineralization of water-filled microchannels within the adhesive joint, as well as intrafibrillar remineralization of collagen fibrils that were demineralized initially as part of the bonding procedure. Biomimetic remineralization as a progressive dehydration mechanism of water-rich, resin-sparse collagen matrices enables these adhesive joints to resist degradation over a 12-month ageing period, as verified by the conservation of their tensile bond strength. The ability of the proof of concept biomimetic remineralization strategy to prevent bond degradation warrants further development of clinically relevant delivery systems.     

Failure characteristics of multiple-component fibrin-based adhesives.

A series of analyses were performed on fibrin-based adhesives to describe their failure characteristics. Two test methods were used: uniaxial, monotonic tensile testing of the bulk material, and blister testing using fresh porcine-source skin graft as the adherend. Two fibrin concentrations, high (HFC), and low (LFC), were used to investigate the effects of the gel matrix density upon mechanical properties. In tensile tests, fibrin gels strain hardened, as functions of percent strain and of strain rate. An increase in modulus of elasticity (E) was seen with increasing strain and strain rate at both tested fibrin concentrations. Mode I failure mechanisms were predominant. Both adhesives appeared to fracture from the outer edge to the interior of the specimen at slower strain rate tests. This trend reversed as strain rate increased, becoming a classic "cup and cone" ductile fracture. Syneresis occurred at both concentrations at lower strain rates, but was more pronounced for the LFC. Ultimate tensile strength and E were greater for the HFC than for the LFC at all strain rates, decreasing with increasing strain rate. In the blister test, the failure locus changed from cohesive to adhesive as the strain rate was increased for the HFC. Failure of fibrin gels likely occurs by percolation of the pressurized saline, displacing the entrapped liquid phase of the gel in regions of relatively low moduli and strength, leading to fracture of the matrix. For LFC, the overall fracture locus remained predominantly cohesive regardless of strain rate. Burst strength and failure energy were higher for HFC than for LFC. It would appear that fibrin acts more as a viscous liquid than a rubberlike/elastic material at lower concentrations because adhesive failures had a higher burst strength and fracture energy (Gc) than did cohesive failures.     

Interaction force measurements for the design of tissue adhesives

Synthesis of tissue adhesives had been carried out in various laboratories in the past decades but the development is currently stalled. One of the key reasons, it is believed, is that researchers have not fully understood and resolved the role of the functional groups that are responsible for good adhesion to biological tissues. Further progress in synthesis is significantly hindered without this fundamental understanding. With this aim in mind, atomic force microscopy (AFM) has been exploited in this work to study the interactions between functional groups that are common to biological tissues. In this work, the AFM tip and substrates were functionalized and used to measure the non-specific interaction among these common functional groups. The ultimate aim of the study is to calculate the interaction force between a single pair of functional groups. A novel calculation method based on the AFM data and probe geometry is presented. The results provide insights into the strength of the bond between different functional groups and the could serve as a guide in selecting the appropriate functional groups in tissue adhesive synthesis. This method could be further applied to studies involving interfaces of biomedical devices where intermolecular interactions are of concern.     

A challenge to the conventional wisdom that simultaneous etching and resin infiltration always occurs in self-etch adhesives

This study provided morphological evidence that discrepancies between the depth of demineralisation and the depth of resin infiltration can occur in some mild self-etch adhesives. Sound dentine specimens derived from extracted human third molars were bonded with 5 one-step and 5 two-step self-etch adhesives. One millimeter thick slabs containing the resin-dentine interfaces were immersed in 50 wt% aqueous ammoniacal silver nitrate and processed for TEM examination. A zone of partially etched but uninfiltrated dentine was identified beneath the hybrid layers in the milder versions of both one-step and two-step self-etch adhesives. This zone was characterised by the occurrence of silver deposits along the interfibrillar spaces of mineralised collagen fibrils. The silver infiltrated interfibrillar spaces were clearly identified from the one-step self-etch adhesives Xeno III, iBond, Brush&Bond and the experimental adhesive, and were thinner and only occasionally observed in the two-step self-etch adhesives Clearfil SE Bond and Clearfil Protect Bond. The more aggressive one-step and two-step adhesives that exhibit more abrupt transitions from completely demineralised to mineralised dentin were devoid of these silver-infiltrated interfibrillar spaces beneath the hybrid layers. Incomplete resin infiltration observed in some self-etch adhesives may be caused by the reduced etching potential of the acidic monomers toward the base of hybrid layers, or the presence of acidic but non-polymerisable hydrolytic adhesive components, creating potential sites for the degradation of the bonded created by these self-etch adhesives.     

Degradation patterns of different adhesives and bonding procedures

Various types of resin adhesives and procedures are available in the clinical field, so comprehensive understanding of degradation is required for each material and bonding procedure. The objective of this study was to investigate the bond durability for different adhesives and bonding procedures. Resin-dentin bonded beams were prepared with the use of two adhesives (One-Up Bond F/self-etching primer system and One Bond/total-etch adhesive) and two experimental groups for the bonding procedure (wet and dry bonding of the total-etch adhesive). Those samples were soaked in water for 24 h(control), 6 and 12 months. After the water immersion, the bond strengths were measured by the microtensile bond test, and subsequently fractography was performed with the use of SEM. Statistically significant reduction of the bond strength (p < 0.05) was apparent after 12 months of water exposure in the range 22-48% of the control. The bonding resin was eluted from the hybrid layer of the self-etching and the total-etch adhesives for the wet bonding. Micromorphological alterations were found due to the hydrolysis of collagen fibrils with the total-etch adhesive for the dry bonding mode. These pathologic alterations were in accord with the bond strength.     

High-Performance Supramolecular Adhesives

Supramolecular adhesives have gained broad attention from both scientific and industrial aspects by virtue of their dynamic and reversible bonding properties. Although considerable achievements have been made in the exploration of noncovalent interactions for supramolecular adhesives, developing high-performance supramolecular adhesives has remained a great challenge. Recently, by combining well-developed noncovalent interactions with rational design strategies, strong and functional supramolecular adhesives have been constructed. From this perspective, the latest developments of high-performance supramolecular adhesives and their design notions will be discussed. At first, supramolecular adhesives with high bonding strengths are focused. By improving mechanical properties and enhancing interfacial interactions, the adhesion strength of supramolecular adhesive has been increased substantially. Secondly, supramolecular adhesives with various functionalities, such as underwater adhesion and conductivity, are introduced. By removing the hydration layer and incorporating conductive components, underwater and conductive supramolecular adhesives have been fabricated. At last, current challenges and future opportunities in this rapidly developing field will be discussed. It is highly anticipated that this perspective will provide guidance for the construction of supramolecular adhesives with desirable bonding strengths and tailor-made functionalities.     

Tissue distribution, biochemical properties, and transmission of mouse type A AApoAII amyloid fibrils

In mouse strains with the amyloidogenic apolipoprotein A-II (ApoA-II) gene (Apoa2c), the type C ApoA-II protein (APOAIIC) associates to form amyloid fibrils AApoAII(C) that lead to development of early onset and systemic amyloidosis with characteristic heavy amyloid deposits in the liver and spleen. We found age-associated heavy deposition of amyloid fibrils [AApoAII(A)] composed of type A ApoA-II protein (APOAIIA) in BDF1 and C57BL/6 mice reared at one of our institutes. AApoAII(A) fibrils were deposited in the intestine, lungs, tongue, and stomach but not in the liver or spleen. AApoAII(A) fibrils were isolated, and morphological, biochemical, and structural characteristics distinct from those seen in AApoAII(C) and mouse AA amyloid fibrils were found. Transmission electron and atomic force microscopy showed that the majority of isolated AApoAII(A) amyloid fibrils featured fine, protofibril-like shapes. AApoAII(A) fibrils have a much weaker affinity for thioflavine T than for AApoAII(C), whereas APOAIIA protein contains less of the beta-pleated sheet structure than does APOAIIC. The injection of AApoAII(A) fibrils induced amyloid deposition in C57BL/6 and DBA2 mice (Apoa2a) as well as in R1.P1-Apoa2c mice (Apoa2c), but AApoAII(A) induced more severe amyloidosis in Apoa2a strains than in the Apoa2c strain. It was found that AApoAII(A) fibrils isolated from mice with mildly amyloidogenic APOAIIA protein have distinct characteristics. Induction of amyloidosis by heterologous amyloid fibrils clearly showed interactions between amyloid protein monomers and fibrils having different primary structures.     

In vitro effect of nanoleakage expression on resin-dentin bond strengths analyzed by microtensile bond test, SEM/EDX and TEM

This study evaluated the effect of multiple consecutive adhesive resin coatings of adhesive bonded to human dentin on nanoleakage and resin-dentin bond strength. Resin bonded dentin specimens were prepared using a total-etch adhesive (One-Step Plus) applied as multiple consecutive coating, or using two self-etch adhesive systems (iBond or Fluoro Bond). For the total-etch adhesive, resin application and air evaporation were performed 1, 2, 3, or 4 times. The self-etch adhesives were applied according to manufacturers' instructions. Resin-dentin bonded beams were prepared and immersed in water (control) or ammoniacal silver nitrate. After storage, microtensile bond strengths were measured. The fractured surfaces were examined by scanning and transmission electron microscopy (SEM and TEM), and energy-dispersive X-ray spectrometry (EDX). No significant differences in bond strength were found between water and silver nitrate storage groups. Several types of silver depositions (spotted, reticular, or water trees) were found in adhesive joints. The bond strengths of the single coated specimens of the total-etch adhesive were significantly lower than those receiving 2-4 coatings. Single coats produced more nanoleakage than multiple coats. However, no correlation was found between the bond strengths and nanoleakage between the different adhesives (total-etch adhesive with different conditions or self-etch adhesives).     

Gelatin–alginate novel tissue adhesives and their formulation–strength effects

Interest in tissue adhesives as alternatives for conventional wound-closing applications such as sutures and staples has increased in the last few decades due to numerous possible advantages, including less discomfort and lower cost. Novel tissue adhesives based on gelatin, with alginate as a polymeric additive and crosslinked by carbodiimide, were recently developed by our research group. The effects of the formulation parameters on the adhesives’ function were investigated in the current study. We examined the effects of gelatin and alginate concentrations and their viscosities on the ability of the bioadhesives to bind soft tissues. The effect of the crosslinking agent’s concentration was studied as well. A qualitative model describing these effects in terms of adherence mechanisms was developed. Our results show that the adherence properties of our new bioadhesives are achieved by a combination of two main mechanisms: mechanical interlocking and chemical adsorption. The former mechanism is probably more dominant. The polymer’s molecular weight and concentration affect the mechanical interlocking through mobility and penetration ability, entanglement of the three-dimensional structure and crosslinking density. The crosslinking agent’s concentration as well as the polymer’s concentration affect the crosslinking density and contribute to higher strength, achieved through both the mechanical interlocking and the chemical adsorption mechanisms. Understanding the effects of the adhesives’ components and their viscosities on the bonding strength enabled us to elucidate the bonding strength mechanisms. This can lead to proper selection of the adhesive formulation and may enable tailoring the bioadhesives to the desired applications.     

Effect of solvent content on resin hybridization in wet dentin bonding

With wet bonding techniques, the channels between the demineralized dentin collagen fibrils are filled with debris, solvent, and water. Commercial adhesives include solvents such as ethanol or acetone to facilitate resin-infiltration into this wet substrate. Under in vivo conditions, the solvent may be diluted because of repeated exposure of the material to the atmosphere, or concentrated because of separation of the bonding liquids into layers within the bottle. The purpose of this study was to investigate the effect of different concentrations of ethanol (10-50%) on infiltration of the adhesive resin and collagen fibril encapsulation in the adhesive/dentin interface using light microscopy, micro-Raman spectroscopy, and scanning electron microscopy. The results indicated that under wet bonding conditions the hybridization process was highly sensitive to the initial solvent concentration in the adhesive system. The staining and scanning electron microscopy results showed that the quality of the interfacial hybrid layer was poor at the lower (10%) or higher (50%) ethanol content. Micro-Raman analysis indicated that there was a distinct difference in the degree of adhesive penetration among adhesives containing different concentrations of ethanol. Adhesives containing 10 or 50% ethanol did not realize effective penetration; the penetration of the adhesive monomers increased dramatically when the initial ethanol content was 30%. The amount of solvents are essential for achieving effective bonding to dentin.     

Failure of Aβ(1-40) amyloid fibrils under tensile loading

Amyloid fibrils and plaques are detected in the brain tissue of patients affected by Alzheimer's disease, but have also been found as part of normal physiological processes such as bacterial adhesion. Due to their highly organized structures, amyloid proteins have also been used for the development of nanomaterials, for a variety of applications including biomaterials for tissue engineering, nanolectronics, or optical devices. Past research on amyloid fibrils resulted in advances in identifying their mechanical properties, revealing a remarkable stiffness. However, the failure mechanism under tensile loading has not been elucidated yet, despite its importance for the understanding of key mechanical properties of amyloid fibrils and plaques as well as the growth and aggregation of amyloids into long fibers and plaques. Here we report a molecular level analysis of failure of amyloids under uniaxial tensile loading. Our molecular modeling results demonstrate that amyloid fibrils are extremely stiff with a Young's modulus in the range of 18-30 GPa, in good agreement with previous experimental and computational findings. The most important contribution of our study is our finding that amyloid fibrils fail at relatively small strains of 2.5%-4%, and at stress levels in the range of 1.02 to 0.64 GPa, in good agreement with experimental findings. Notably, we find that the strength properties of amyloid fibrils are extremely length dependent, and that longer amyloid fibrils show drastically smaller failure strains and failure stresses. As a result, longer fibrils in excess of hundreds of nanometers to micrometers have a greatly enhanced propensity towards spontaneous fragmentation and failure. We use a combination of simulation results and simple theoretical models to define critical fibril lengths where distinct failure mechanisms dominate.     

A novel age-related venous amyloidosis derived from EGF-containing fibulin-like extracellular matrix protein 1

Most intractable tissue-degenerative disorders share a common pathogenic condition, so-called proteinopathy. Amyloid-related disorders are the most common proteinopathies and are characterized by amyloid fibril deposits in the brain or other organs. Aging is generally associated with the development of these amyloid-related disorders, but we still do not fully understand how functional proteins become pathogenic amyloid deposits during the human aging process. We identified a novel amyloidogenic protein, named epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1), in massive venous amyloid deposits in specimens that we obtained from an autopsied patient who died of gastrointestinal bleeding. Our postmortem analyses of additional patients indicate that EFEMP1 amyloid deposits frequently developed in systemic venous walls of elderly people. EFEMP1 was highly expressed in veins, and aging enhanced venous EFEMP1 expression. In addition, biochemical analyses indicated that these venous amyloid deposits consisted of C-terminal regions of EFEMP1. In vitro studies showed that C-terminal regions formed amyloid fibrils, which inhibited venous tube formation and cell viability. EFEMP1 thus caused a novel age-related venous amyloid-related disorder frequently found in the elderly population. Understanding EFEMP1 amyloid formation provides new insights into amyloid-related disorders occurring during the aging process. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Immunoelectron microscopic identification of human AA-type amyloid: exploration of various monoclonal AA-antibodies, methods of fixation, embedding and of other parameters for the protein-A gold method

Using the postembedding protein-A gold technique ten monoclonal antibodies directed against amyloid-A protein (AA) were examined by immunoelectron microscopy to identify amyloid-A (AA) amyloid fibrils in plastic-embedded renal tissue of five patients and two controls. Two monoclonal antibodies (mc1, mc20) specifically labeled these amyloid deposits; two additional ones (mc4, mc13) bound with an intermediate rabbit anti-mouse IgG antiserum. These monoclonal anti-AA antibodies clearly separate amyloid fibrils from morphologically similar fibrils in the vicinity. Employing varying embedding media, fixation techniques, as well as etching and staining protocols, we adapted this method for the immunoelectron microscopic identification of AA-type amyloid fibrils and for the antigenic diagnosis of AA-type amyloid on routinely processed ultrathin sections.     

Immunogold labeling of cerebrovascular and neuritic plaque amyloid fibrils in Alzheimer's disease with an anti-beta protein monoclonal antibody

A monoclonal antibody raised to a synthetic peptide consisting of residues 8 to 17 of the amyloid beta protein of Alzheimer's disease was employed for immunogold electron microscopic studies on amyloid fibrils of cerebrovascular walls and neuritic plaques in this disease. Electron microscopy revealed a specific gold labeling of the amyloid fibrils in these structures. This provides ultrastructural evidence that beta protein is intimately associated with the amyloid fibril. With previous chemical evidence, this observation supports the concentration that it is an intrinsic component of the fibril.     

Effect of the hydrophilic nanofiller loading on the mechanical properties and the microtensile bond strength of an ethanol-based one-bottle dentin adhesive

This study evaluated the hypothesis that if hydrophilic nanofillers were dispersed evenly within the adhesive layer under moist conditions, adding them to a one-bottle dentin adhesive might improve the mechanical properties of the adhesive layer, and accordingly increase the bond strength. The flexural strength (FS), the degree of conversion (DC), and the microtensile bond strength (MTBS) to the dentin of four experimental ethanol-based one-bottle dentin adhesives containing 0, 0.5, 1.0, and 3.0 wt % of 12-nm hydrophilic fumed silica were evaluated, and the distribution of the nanofillers were compared using transmission electron microscopy (TEM). Although the nanofiller content did not affect the DC, the FS tended to increase with increasing nanofiller content. The MTBS appeared to increase when up to 1.0 wt % of the nanofillers were added, but they were statistically not significant. However, when 3.0 wt % of the nanofillers were added, the MTBS decreased significantly comparing to the adhesive containing 0.5 wt % nanofillers (p < 0.05). The TEM image suggested that if the nanofillers within the adhesive were 3.0 wt % and applied to a wet dentin surface, they aggregated easily into large clusters and would decrease the MTBS.