Inertial Imaging with Nanomechanical Systems

Mass sensing with nanoelectromechanical systems has advanced significantly during the last decade. With nanoelectromechanical systems sensors it is now possible to carry out ultrasensitive detection of gaseous analytes, to achieve atomic-scale mass resolution and to perform mass spectrometry on single proteins. Here, we demonstrate that the spatial distribution of mass within an individual analyte can be imaged—in real time and at the molecular scale—when it adsorbs onto a nanomechanical resonator. Each single-molecule adsorption event induces discrete, time-correlated perturbations to all modal frequencies of the device. We show that by continuously monitoring a multiplicity of vibrational modes, the spatial moments of mass distribution can be deduced for individual analytes, one-by-one, as they adsorb. We validate this method for inertial imaging, using both experimental measurements of multimode frequency shifts and numerical simulations, to analyse the inertial mass, position of adsorption and the size and shape of individual analytes. Unlike conventional imaging, the minimum analyte size detectable through nanomechanical inertial imaging is not limited by wavelength-dependent diffraction phenomena. Instead, frequency fluctuation processes determine the ultimate attainable resolution. Advanced nanoelectromechanical devices appear capable of resolving molecular-scale analytes

Gelatin methacryloyl hydrogels for the localized delivery of cefazolin

The tuneability of hydrogels renders them promising candidates for local drug delivery to prevent and treat local surgical site infection (SSI) while avoiding the systemic side-effects of intravenous antibiotic injections. Here, we present a newly developed gelatin methacryloyl (GelMA)-based hydrogel drug delivery system (GelMA-DDS) to locally deliver the broad-spectrum antibiotic cefazolin for SSI prophylaxis and treatment. Antibiotic doses from 3 µg to 90 µg were loaded in photocrosslinked GelMA hydrogel discs with 5 to 15% w/v polymer concentration and drug encapsulation efficiencies, mechanical properties, crosslinking and release kinetics, as well as bacterial growth inhibition were assessed. Our results demonstrate that all GelMA groups supported excellent drug encapsulation efficiencies of up to 99%. Mechanical properties of the GelMA-DDS were highly tuneable and unaffected by the loading of small to medium doses of cefazolin. The diffusive and the proteolytic in vitro drug delivery of all investigated cefazolin doses was characterized by a burst release, and the delivered cefazolin amount was directly proportional to the encapsulated dose. Accelerated enzymatic degradation of the GelMA-DDS followed zero-order kinetics and was dependent on both the cefazolin dose and GelMA concentration (3–13 h). Finally, we demonstrate that cefazolin delivered from GelMA induced a dose-dependent antibacterial efficacy against S. aureus, in both a broth and a diffusive assay. The cefazolin-loaded GelMA-DDS presented here provides a highly tuneable and easy-to-use local delivery system for the prophylaxis and treatment of SSI.</p

3D printed multiphasic scaffolds for osteochondral repair: Challenges and opportunities

Osteochondral (OC) defects are debilitating joint injuries characterized by the loss of full thickness articular cartilage along with the underlying calcified cartilage through to the subchondral bone. While current surgical treatments can provide some relief from pain, none can fully repair all the components of the OC unit and restore its native function. Engineering OC tissue is challenging due to the presence of the three distinct tissue regions. Recent advances in additive manufacturing provide unprecedented control over the internal microstructure of bioscaffolds, the patterning of growth factors and the encapsulation of potentially regenerative cells. These developments are ushering in a new paradigm of ‘multiphasic’ scaffold designs in which the optimal micro-environment for each tissue region is individually crafted. Although the adoption of these techniques provides new opportunities in OC research, it also introduces challenges, such as creating tissue interfaces, integrating multiple fabrication techniques and co-culturing different cells within the same construct. This review captures the considerations and capabilities in developing 3D printed OC scaf-folds, including materials, fabrication techniques, mechanical function, biological components and design

Characterization of polycaprolactone nanohydroxyapatite composites with tunable degradability suitable for indirect printing

Degradable bone implants are designed to foster the complete regeneration of natural tissue after large-scale loss trauma. Polycaprolactone (PCL) and hydroxyapatite (HA) composites are promising scaffold materials with superior mechanical and osteoinductive properties compared to the single materials. However, producing three-dimensional (3D) structures with high HA content as well as tuneable degradability remains a challenge. To address this issue and create homogeneously distributed PCL-nanoHA (nHA) scaffolds with tuneable degradation rates through both PCL molecular weight and nHA concentration, we conducted a detailed characterisation and comparison of a range of PCL-nHA composites across three molecular weight PCLs (14, 45, and 80 kDa) and with nHA content up to 30% w/w. In general, the addition of nHA results in an increase of viscosity for the PCL-nHA composites but has little effect on their compressive modulus. Importantly, we observe that the addition of nHA increases the rate of degradation compared to PCL alone. We show that the 45 and 80 kDa PCL-nHA groups can be fabricated via indirect 3D printing and have homogenously distributed nHA even after fabrication. Finally, the cytocompatibility of the composite materials is evaluated for the 45 and 80 kDa groups, with the results showing no significant change in cell number compared to the control. In conclusion, our analyses unveil several features that are crucial for processing the composite material into a tissue engineered implant

Vapor Phase Polymerization of EDOT from Submicrometer Scale Oxidant Patterned by Dip-Pen Nanolithography

Some of the most exciting recent advances in conducting polymer synthesis have centered around the method of vapor phase polymerization (VPP) of thin films. However, it is not known whether the VPP process can proceed using significantly reduced volumes of oxidant and therefore be implemented as part of nanolithography approach. Here, we present a strategy for submicrometer scale patterning of the conducting polymer poly­(3,4-ethylenedioxythiophene) (PEDOT) via in situ VPP. Attolitre (10^–18 L) volumes of oxidant “ink” are controllably deposited using dip-pen nanolithography (DPN). DPN patterning of the oxidant ink is facilitated by the incorporation of an amphiphilic block copolymer thickener, an additive that also assists with stabilization of the oxidant. When exposed to EDOT monomer in a VPP chamber, each deposited feature localizes the synthesis of conducting PEDOT structures of several micrometers down to 250 nm in width. PEDOT patterns are characterized by atomic force microscopy (AFM), conductive AFM, two probe electrical measurement, and micro-Raman spectroscopy, evidencing in situ vapor phase synthesis of conducting polymer at a scale (picogram) which is much smaller than that previously reported. Although the process of VPP on this scale was achieved, we highlight some of the challenges that need to be overcome to make this approach feasible in an applied setting

Matured myofibers in bioprinted constructs with in vivo vascularization and innervation

For decades, the study of tissue-engineered skeletal muscle has been driven by a clinical need to treat neuromuscular diseases and volumetric muscle loss. The in vitro fabrication of muscle offers the opportunity to test drug-and cell-based therapies, to study disease processes, and to perhaps, one day, serve as a muscle graft for reconstructive surgery. This study developed a biofabrication technique to engineer muscle for research and clinical applications. A bioprinting protocol was established to deliver primary mouse myoblasts in a gelatin methacryloyl (GelMA) bioink, which was implanted in an in vivo chamber in a nude rat model. For the first time, this work demonstrated the phenomenon of myoblast migration through the bioprinted GelMA scaffold with cells spontaneously forming fibers on the surface of the material. This enabled advanced maturation and facilitated the connection between incoming vessels and nerve axons in vivo without the hindrance of a scaffold material. Immunohistochemistry revealed the hallmarks of tissue maturity with sarcomeric striations and peripherally placed nuclei in the organized bundles of muscle fibers. Such engineered muscle autografts could, with further structural development, eventually be used for surgical reconstructive purposes while the methodology presented here specifically has wide applications for in vitro and in vivo neuromuscular function and disease modelling

In-situ handheld 3D Bioprinting for cartilage regeneration

Articular cartilage injuries experienced at an early age can lead to the development of osteoarthritis later in life. In situ 3D printing is an exciting and innovative bio-fabrication technology that enables the surgeon to deliver tissue- engineering techniques at the time and location of need. We have created a hand- held 3D printing device (Biopen) that allows the simultaneous co-axial extrusion of bioscaffold and cultured cells directly into the cartilage defect in vivo in a single session surgery. This pilot study assesses the ability of the Biopen to repair a full thickness chondral defect and the early outcomes in cartilage regeneration, and compares these results to other treatments in a large animal model. A standardised critical-sized full thickness chondral defect was created in the weight-bearing surface of the lateral and medial condyles of both femurs of 6 sheep. Each defect was treated with one of the following treatments: \ud \ud <i>(i) hand- held in situ 3D printed bioscaffold using the Biopen (HH group);</i> \ud \ud <i>(ii) pre- constructed bench-based printed bioscaffolds (BB group);</i> \ud \ud <i>(iii) micro-fractures (MF group), or;</i>\ud \ud <i>(iv) untreated (Control, C group). </i>\ud \ud At 8 weeks after surgery, macroscopic, microscopic and biomechanical tests were performed. Surgical 3D bio-printing was performed in all animals without any intra- or post- operative complication. The HH Biopen allowed early cartilage regeneration. Results of this study show that real-time, in vivo bioprinting with cells and scaffold is a feasible means of delivering a regenerative medicine strategy in a large animal model to regenerate articular cartilage. This article is protected by copyright. All rights reserved