B21-High Throughput SAXS
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Diamond Proposal Number(s):
[29767]
Open Access
Abstract: Sterilisation of implantable biomaterials such as hydrogels remains a key step towards their clinical translation. Standard sterilisation methods can significantly alter hydrogels' physicochemical and biological performance. Previously, we developed composite hydrogels based on ε-polylysine (ε-PL) and hyaluronic acid (HA). The developed hydrogels demonstrated promising antibacterial activity and in vitro cell viability and their variable properties depending on the chosen ε-PL to HA ratio. In this study, we fabricated a series of chemically cross-linked ԑ-PL/HA hydrogels with expanded ԑ-PL to HA mass ratios.
Using small-angle X-ray scattering (SAXS), we unravelled the topological differences between physically and chemically crosslinked hydrogels. We then selected the chemically crosslinked hydrogel ԑ-PL/HA series of 60:40 wt%, 70:30 wt%, and 80:20 wt% ratios, with similar network topologies, to evaluate the impact of steam sterilisation on their physicochemical and viscoelastic properties. The antibacterial activity of the sterilized hydrogels was also evaluated against Gram-negative and Gram-positive bacteria. Our results showed that steam sterilisation minimally affects structure and physicochemical properties of ԑ-PL/HA hydrogels. Furthermore, the developed hydrogel ԑ-PL/HA series of 60:40 wt%, 70:30 wt%, and 80:20 wt% ratios showed pronounced antibacterial activity against Gram-negative and Gram-positive pathogenic bacteria. We expect our results will contribute to the growing understanding of using sterilisation methods for antibacterial hydrogels that have the potential for wider tissue engineering applications.
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Dec 2023
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I22-Small angle scattering & Diffraction
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Diamond Proposal Number(s):
[14126]
Open Access
Abstract: Echinoderms, such as sea cucumbers, have the remarkable property of changing the stiffness of their dermis according to the surrounding chemical environments. When sea cucumber dermal specimens are constantly strained, stress decays exponentially with time. Such stress relaxation is a hallmark of visco-elastic mechanical behavior. In this paper, in contrast, we attempted to interpret stress relaxation from the chemoelasticity viewpoint. We used a finite element model for the microstructure of the sea cucumber dermis. We varied stiffness over time and framed such changes against the first-order reactions of the interfibrillar matrix. Within this hypothetical scenario, we found that stress relaxation would then occur primarily due to fast crosslink splitting between the chains and a much slower macro-chain scission, with characteristic reaction times compatible with relaxation times measured experimentally. A byproduct of the model is that the concentration of undamaged macro-chains in the softened state is low, less than 10%
, which tallies with physical intuition. Although this study is far from being conclusive, we believe it opens an alternative route worthy of further investigation.
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Dec 2023
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I12-JEEP: Joint Engineering, Environmental and Processing
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Open Access
Abstract: Infection is the leading cause of biomedical implant failure, making the incorporation of antibacterial elements highly desirable. However, new alloys must also possess a low elastic modulus, to prevent stress shielding and bone resorption. Achieving this is challenging, with current alloys containing antibacterial elements being too stiff. Here, we report a novel Ti-Nb-Au alloy that contains appreciable concentrations of an antibacterial element, is free from the deleterious omega phase, has an exceptionally low elastic modulus (38.4 GPa) and high strain recoverability. These results indicate that the Ti-Nb-Au system has promise for biomedical applications, warranting further investigation and development.
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Dec 2023
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DIAD-Dual Imaging and Diffraction Beamline
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Abstract: Purpose / Aim: Dentine hypersensitivity (DH) is a commonly occurring dental condition where sharp pain derived from exposed dentine in response to stimuli that cannot be ascribed to any other dental diseases. It is highly prevalent affecting up to 69% of the UK population and has a significant impact on the quality of life. Bioactive glasses that degrade in oral environment and form apatite are thought to be beneficial in occluding the exposed open dentine tubules and have been introduced to toothpastes, e.g. Novamin® (45S5 Bioglass) for Sensodyne by GSK, fluoride containing bioactive glass for BioMin® F by BioMin Technologies Ltd. Post-mortem characterisations evidenced tubule occlusion (Fig.1) but failed in providing dynamic history. Therefore, this study aimed to monitor dentine tubule occlusion with bioactive glasses using an operando time-lapse X-ray diffraction tomography experiment. Materials & Methods: Disinfected Teeth (collected under REC reference 16/SW/0220) were sectioned mesio-distally into discs approximately 500 μm thick using a precision diamond saw, polished down to 300 μm manually. Matchstick specimens (5 mm length x 3 mm width) prepared were brushed for 2 mins with bioactive glasses pastes, housed in a modified Eppendorf tube and positioned on the tomography stage of the Dual Imaging and Diffraction (DIAD) beamline at Diamond Light Source (UK’s national synchrotron). A baseline X-ray tomography (pink beam, 0-180°, detector exposure of 0.01 and 5,000 projections) and X-ray diffraction mapping (matrix scan with 10x10 points, 20 s exposure) were collected before artificial saliva was introduced. Time-lapse X-ray tomography and X-ray diffraction mapping using the same parameters as baseline scan were carried out consecutively for 8 h allowing the visualisation of tubule occlusion and changes of mineral density as well as monitor the phase evolution from glass to apatite. Artificial saliva was manually replenished. Monochromatic beam with an energy of 20 keV was used and calibrations were performed. Results: The collected tomography data allow visualisation of dentine tubule occlusion showing improved occlusion with time. 2D XRD data provide qualitative and quantitative information relates to glass dissolution and apatite formation as a function of time. Conclusions: The Dual Imaging and Diffraction (DIAD) beamline correlates X-ray tomography and X-ray diffraction mapping offers the opportunity to study dentine occlusion by bioactive glasses in a time evolving manner that is not available via other techniques. Although in vitro but clinically relevant. The results will potentially provide a guidance for optimising and designing products for dentine tubule occlusion/treating dentine hypersensitivity – one of the most prevalent global diseases with healthy ageing.
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Oct 2023
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B21-High Throughput SAXS
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Diamond Proposal Number(s):
[26998]
Abstract: Topographical cues on materials can manipulate cellular fate, particularly for neural cells that respond well to such cues. Utilizing biomaterial surfaces with topographical features can effectively influence neuronal differentiation and promote neurite outgrowth. This is crucial for improving the regeneration of damaged neural tissue after injury. Here, we utilized groove patterns to create neural conduits that promote neural differentiation and axonal growth. We investigated the differentiation of human neural stem cells (NSCs) on silicon dioxide groove patterns with varying height-to-width/spacing ratios. We hypothesize that NSCs can sense the microgrooves with nanoscale depth on different aspect ratio substrates and exhibit different morphologies and differentiation fate. A comprehensive approach was employed, analyzing cell morphology, neurite length, and cell-specific markers. These aspects provided insights into the behavior of the investigated NSCs and their response to the topographical cues. Three groove-pattern models were designed with varying height-to-width/spacing ratios of 80, 42, and 30 for groove pattern widths of 1 μm, 5 μm, and 10 μm and nanoheights of 80 nm, 210 nm, and 280 nm. Smaller groove patterns led to longer neurites and more effective differentiation towards neurons, whereas larger patterns promoted multidimensional differentiation towards both neurons and glia. We transferred these cues onto patterned polycaprolactone (PCL) and PCL-graphene oxide (PCL-GO) composite ‘stamps’ using simple soft lithography and reproducible extrusion 3D printing methods. The patterned scaffolds elicited a response from NSCs comparable to that of silicon dioxide groove patterns. The smallest pattern stimulated the highest neurite outgrowth, while the middle-sized grooves of PCL-GO induced effective synaptogenesis. We demonstrated the potential for such structures to be wrapped into tubes and used as grafts for peripheral nerve regeneration. Grooved PCL and PCL-GO conduits could be a promising alternative to nerve grafting.
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Oct 2023
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Open Access
Abstract: Mesoporous glasses are a promising class of bioresorbable biomaterials characterized by high surface area and extended porosity in the range of 2 to 50 nm. These peculiar properties make them ideal materials for the controlled release of therapeutic ions and molecules. Whilst mesoporous silicate-based glasses (MSG) have been widely investigated, much less work has been done on mesoporous phosphate-based glasses (MPG). In the present study, MPG in the P2O5–CaO–Na2O system, undoped and doped with 1, 3, and 5 mol% of Cu ions were synthesized via a combination of the sol–gel method and supramolecular templating. The non-ionic triblock copolymer Pluronic P123 was used as a templating agent. The porous structure was studied via a combination of Scanning Electron Microscopy (SEM), Small-Angle X-ray Scattering (SAXS), and N2 adsorption–desorption analysis at 77 K. The structure of the phosphate network was investigated via solid state 31P Magic Angle Spinning Nuclear Magnetic Resonance (31P MAS-NMR) and Fourier Transform Infrared (FTIR) spectroscopy. Degradation studies, performed in water via Inductively Coupled Plasma-Optical Emission Spectroscopy (ICP-OES), showed that phosphates, Ca2+, Na+ and Cu ions are released in a controlled manner over a 7 days period. The controlled release of Cu, proportional to the copper loading, imbues antibacterial properties to MPG. A significant statistical reduction of Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) bacterial viability was observed over a 3 days period. E. coli appeared to be more resistant than S. aureus to the antibacterial effect of copper. This study shows that copper doped MPG have great potential as bioresorbable materials for controlled delivery of antibacterial ions.
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Jun 2023
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I22-Small angle scattering & Diffraction
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Minghui
Sun
,
Zheng
Dong
,
Liyuan
Wu
,
Haodong
Yao
,
Wenchao
Niu
,
Deting
Xu
,
Ping
Chen
,
Himadri S.
Gupta
,
Yi
Zhang
,
Yuhui
Dong
,
Chunying
Chen
,
Lina
Zhao
Open Access
Abstract: Structural disclosure of biological materials can help our understanding of design disciplines in nature and inspire research for artificial materials. Synchrotron microfocus X-ray diffraction is one of the main techniques for characterizing hierarchically structured biological materials, especially the 3D orientation distribution of their interpenetrating nanofiber networks. However, extraction of 3D fiber orientation from X-ray patterns is still carried out by iterative parametric fitting, with disadvantages of time consumption and demand for expertise and initial parameter estimates. When faced with high-throughput experiments, existing analysis methods cannot meet the real time analysis challenges. In this work, using the assumption that the X-ray illuminated volume is dominated by two groups of nanofibers in a gradient biological composite, a machine-learning based method is proposed for fast and automatic fiber orientation metrics prediction from synchrotron X-ray micro-focused diffraction data. The simulated data were corrupted in the training procedure to guarantee the prediction ability of the trained machine-learning algorithm in real-world experimental data predictions. Label transformation was used to resolve the jump discontinuity problem when predicting angle parameters. The proposed method shows promise for application in the automatic data-processing pipeline for fast analysis of the vast data generated from multiscale diffraction-based tomography characterization of textured biomaterials.
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May 2023
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B21-High Throughput SAXS
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Diamond Proposal Number(s):
[32446]
Open Access
Abstract: Biomaterials for tissue regeneration must mimic the biophysical properties of the native physiological environment. A protein engineering approach allows the generation of protein hydrogels with specific and customised biophysical properties designed to suit a particular physiological environment. Herein, repetitive engineered proteins were successfully designed to form covalent molecular networks with defined physical characteristics able to sustain cell phenotype. Our hydrogel design was made possible by the incorporation of the SpyTag (ST) peptide and multiple repetitive units of the SpyCatcher (SC) protein that spontaneously formed covalent crosslinks upon mixing. Changing the ratios of the protein building blocks (ST:SC), allowed the viscoelastic properties and gelation speeds of the hydrogels to be altered and controlled. The physical properties of the hydrogels could readily be altered further to suit different environments by tuning the key features in the repetitive protein sequence. The resulting hydrogels were designed with a view to allow cell attachment and encapsulation of liver derived cells. Biocompatibility of the hydrogels was assayed using a HepG2 cell line constitutively expressing GFP. The cells remained viable and continued to express GFP whilst attached or encapsulated within the hydrogel. Our results demonstrate how this genetically encoded approach using repetitive proteins could be applied to bridge synthetic biology with nanotechnology creating a level of biomaterial customisation previously inaccessible.
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May 2023
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B21-High Throughput SAXS
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Diamond Proposal Number(s):
[28806, 28287]
Abstract: Bioinspired de novo self-assembling peptides have been widely used for the development of soft biomaterials for a wide variety of biomedical and pharmaceutical applications, such as cell scaffolding for tissue engineering1, controlled and localised drug delivery2, biosensing3, and many others. The meticulous control of peptide-based nanomaterial properties over the length scale, by molecular design, remains the main challenge for tailoring biomaterials properties to meet the application needs. In our group, we have recently adopted a minimalistic molecular engineering approach for the development of Ultrashort Ionic-complementary Constrained Peptides (UICPs), which were rationally designed to self-assemble into amphiphilic β-sheet nanofibers with unique hydrogelation properties and surface activity.4 We have previously demonstrated the crucial role played by aromatic stacking for the formation and thermodynamic stabilisation of UICP β-sheet structures. Herein, we will show how charge interactions can be manipulated for fine tuning molecular self-assembly, morphology and size of nanofibrous structures formation and viscoelasticity of UICP hydrogels.
A library of 18 peptide sequences (4-5 residues long) was developed to study the effect of the sequence net charge, charge density distribution, reversal of charge order and ionic self-complementarity on their propensity towards self-assembly and gelation. Interestingly, 12 of these peptides self-assembled into β-sheet nanofibrous structures forming hydrogels at pH 4.5-5, as confirmed by ATR-FTIR, SEM, TEM, SAXS and oscillatory rheology. Full control over β-sheet content (ranging from ~30-80%), fibre morphology (thin fibrils, thick straight fibre bundles, twisted helical nanofibres, flat nanoribbons and nanotubes) and sizes (~4-67 nm in diameter), as well as gelation (critical gelation concentrations ranging from <7.5 to >100 mM) and viscoelastic properties (storage moduli G’ ~0.1-100 KPa) was achieved by the careful positioning of both Glu and Lys residues at both C- and N-termini, in the sequence core and on both the hydrophilic and hydrophobic faces of the peptide chain. In essence, this design approach enabled/disabled lateral growth along the β-sheet ladder via electrostatic attraction (counter charge, anion-pi and cation-pi)/repulsion, hence controlling fibre thickness, morphology, entanglement, and the resulting viscoelasticity of the system. Our UICPs platform thus provides the flexibility in peptide molecular design for the manufacturing of soft biomaterials with versatile properties that can be in future tailored to the relevant biomedical application.
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Apr 2023
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I22-Small angle scattering & Diffraction
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Abstract: Cardiovascular diseases are the primary cause of death globally [1]. Bioresorbable vascular scaffolds (BVSs) were developed to overcome the limitations of permanent metallic stents. Poly-(l-lactic acid)(PLLA) is the preferred material choice but, with a tensile modulus lower than metal, the polymeric stents need to be ~2x as thick to achieve similar structural stiffness, hindering deployment and disrupting blood flow [2, 3].
Initially uniform PLLA is subjected to a series of thermo-mechanical processes to create a BVS, with properties strongly dependent on the thermal and strain histories experienced by the polymer — in particular, the crimping and expansion stages. There is a lack of consensus on the impact of crimping on the mechanical properties of the BVS [4]. U-bends have been highlighted as an area of interest and microcracks that form during crimping can indicate possible failure sites [5]. Crimping is thus key to creating a thinner scaffold.
Due to the cost of medical grade PLLA, it is far more feasible to carry out initial testing on a cheaper, packaging grade PLLA with a different molecular weight. This research aims to determine if the materials behave similarly enough during each phase of BVS processing for this to be a valid exercise.
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Apr 2023
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