Optimization of SDS exposure on preservation of ECM characteristics in whole organ decellularization of rat kidneys.

Renal transplantation is well established as the optimal form of renal replacement therapy but is restricted by the limited pool of organs available for transplantation. The whole organ decellularisation approach is leading the way for a regenerative medicine solution towards bioengineered organ replacements. However, systematic preoptimization of both decellularization and recellularization parameters is essential prior to any potential clinical application and should be the next stage in the evolution of whole organ decellularization as a potential strategy for bioengineered organ replacements. Here we have systematically assessed two fundamental parameters (concentration and duration of perfusion) with regards to the effects of differing exposure to the most commonly used single decellularizing agent (sodium dodecyl sulphate/SDS) in the perfusion decellularization process for whole rat kidney ECM bioscaffolds, with findings showing improved preservation of both structural and functional components of the whole kidney ECM bioscaffold. Whole kidney bioscaffolds based on our enhanced protocol were successfully recellularized with rat primary renal cells and mesenchymal stromal cells. These findings should be widely applicable to decellularized whole organ bioscaffolds and their optimization in the development of regenerated organ replacements for transplantation. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1352-1360, 2017.

Surface effects of vapour-liquid-solid driven Bi surface droplets formed during molecular-beam-epitaxy of GaAsBi.

Herein we investigate a (001)-oriented GaAs1-xBix/GaAs structure possessing Bi surface droplets capable of catalysing the formation of nanostructures during Bi-rich growth, through the vapour-liquid-solid mechanism. Specifically, self-aligned "nanotracks" are found to exist trailing the Bi droplets on the sample surface. Through cross-sectional high-resolution transmission electron microscopy the nanotracks are revealed to in fact be elevated above surface by the formation of a subsurface planar nanowire, a structure initiated mid-way through the molecular-beam-epitaxy growth and embedded into the epilayer, via epitaxial overgrowth. Electron microscopy studies also yield the morphological, structural, and chemical properties of the nanostructures. Through a combination of Bi determination methods the compositional profile of the film is shown to be graded and inhomogeneous. Furthermore, the coherent and pure zincblende phase property of the film is detailed. Optical characterisation of features on the sample surface is carried out using polarised micro-Raman and micro-photoluminescence spectroscopies. The important light producing properties of the surface nanostructures are investigated through pump intensity-dependent micro-PL measurements, whereby relatively large local inhomogeneities are revealed to exist on the epitaxial surface for important optical parameters. We conclude that such surface effects must be considered when designing and fabricating optical devices based on GaAsBi alloys.

Mechanism of periodic height variations along self-aligned VLS-grown planar nanostructures.

In this study we report in-plane nanotracks produced by molecular-beam-epitaxy (MBE) exhibiting lateral self-assembly and unusual periodic and out-of-phase height variations across their growth axes. The nanotracks are synthesized using bismuth segregation on the GaAsBi epitaxial surface, which results in metallic liquid droplets capable of catalyzing GaAsBi nanotrack growth via the vapor-liquid-solid (VLS) mechanism. A detailed examination of the nanotrack morphologies is carried out employing a combination of scanning electron and atomic force microscopy and, based on the findings, a geometric model of nanotrack growth during MBE is developed. Our results indicate diffusion and shadowing effects play significant roles in defining the interesting nanotrack shape. The unique periodicity of our lateral nanotracks originates from a rotating nucleation "hot spot" at the edge of the liquid-solid interface, a feature caused by the relative periodic circling of the non-normal ion beam flux incident on the sample surface, inside the MBE chamber. We point out that such a concept is divergent from current models of crawling mode growth kinetics and conclude that these effects may be utilized in the design and assembly of planar nanostructures with controlled non-monotonous structure.

Raman scattering reveals strong LO-phonon-hole-plasmon coupling in nominally undoped GaAsBi: optical determination of carrier concentration.

We report room-temperature Raman scattering studies of nominally undoped (100) GaAs1−xBix epitaxial layers exhibiting Bi-induced (p-type) longitudinal-optical-plasmon-coupled (LOPC) modes for 0.018 ≤ x ≤ 0.048. Redshifts in the GaAs-like optical modes due to alloying are evaluated and are paralleled by strong damping of the LOPC. The relative integrated Raman intensities of LO(Γ) and LOPC ALO/ALOPC are characteristic of heavily doped p-GaAs, with a remarkable near total screening of the LO(Γ) phonon (ALO/ALOPC → 0) for larger Bi concentrations. A method of spectral analysis is set out which yields estimates of hole concentrations in excess of 5×1017cm−3 and correlates with the Bi molar fraction. These findings are in general agreement with recent electrical transport measurements performed on the alloy, and while the absolute size of the hole concentrations differ, likely origins for the discrepancy are discussed. We conclude that the damped LO-phonon-hole-plasmon coupling phenomena plays a dominant role in Raman scattering from unpassivated nominally undoped GaAsBi.

Therapeutic cell encapsulation techniques and applications in diabetes.

The encapsulation of therapeutic cells permits the implantation of allogeneic and xenogeneic cells for the regulation of certain physiological processes damaged by the death or senescence of host tissues. The encapsulation of pancreatic cells for the treatment of diabetes is emphasized; however, many of the techniques are applicable to a wide array of mammalian cell applications. The summary of both established and novel encapsulation techniques, clinical trials, and commercial product developments highlights the metered but steady pace of therapeutic cell encapsulation towards implementation.

Laser-induced oxidation kinetics of bismuth surface microdroplets on GaAsBi studied in situ by Raman microprobe analysis.

We report the cw-laser-induced oxidation of molecular-beam-epitaxy grown GaAsBi bismuth surface microdroplets investigated in situ by micro-Raman spectroscopy under ambient conditions as a function of irradiation power and time. Our results reveal the surface droplets are high-purity crystalline bismuth and the resultant Bi2O3 transformation to be ß-phase and stable at room temperature. A detailed Raman study of Bi microdroplet oxidation kinetics yields insights into the laser-induced oxidation process and offers useful real-time diagnostics. The temporal evolution of new ß-Bi2O3 Raman modes is shown to be well described by Johnson-Mehl-Avrami-Kolmogorov kinetic transformation theory and while this study limits itself to the laser-induced oxidation of GaAsBi bismuth surface droplets, the results will find application within the wider context of bismuth laser-induced oxidation and direct Raman laser processing.

Combinatorial scaffold morphologies for zonal articular cartilage engineering.

Articular cartilage lesions are a particular challenge for regenerative medicine strategies as cartilage function stems from a complex depth-dependent organization. Tissue engineering scaffolds that vary in morphology and function offer a template for zone-specific cartilage extracellular matrix (ECM) production and mechanical properties. We fabricated multi-zone cartilage scaffolds by the electrostatic deposition of polymer microfibres onto particulate-templated scaffolds produced with 0.03 or 1.0mm(3) porogens. The scaffolds allowed ample space for chondrocyte ECM production within the bulk while also mimicking the structural organization and functional interface of cartilage's superficial zone. Addition of aligned fibre membranes enhanced the mechanical and surface properties of particulate-templated scaffolds. Zonal analysis of scaffolds demonstrated region-specific variations in chondrocyte number, sulfated GAG-rich ECM, and chondrocytic gene expression. Specifically, smaller porogens (0.03mm(3)) yielded significantly higher sGAG accumulation and aggrecan gene expression. Our results demonstrate that bilayered scaffolds mimic some key structural characteristics of native cartilage, support in vitro cartilage formation, and have superior features to homogeneous particulate-templated scaffolds. We propose that these scaffolds offer promise for regenerative medicine strategies to repair articular cartilage lesions.

Importance of phenylalanine 107 in agonist recognition by the 5-hydroxytryptamine(3A) receptor.

The 5-hydroxytryptamine (5-HT)(3) receptor is a member of the ligand-gated ion channel receptor family with significant homology to the nicotinic acetylcholine, gamma-aminobutyric acid(A), and glycine receptors. In this receptor class, the agonist binding site is formed by parts of the extracellular amino-terminal region. This study examines the effects of altering phenylalanine 107 (F107) of the 5-HT(3AL) subunit, obtained from NG108-15 cells, using site-directed mutagenesis. The wild-type (WT) and mutant receptors were expressed in HEK 293 cells and characterized using both whole-cell patch-clamp and radioligand binding. The tyrosine mutant F107Y exhibits a significantly lower affinity for the agonist 5-HT (K(i) = 203 versus 15.6 nM) and an increase of similar magnitude in the EC(50) value (10.6 versus 1.2 microM) compared with WT. The activation kinetics of the maximal currents generated by 5-HT with this mutant were markedly slower than those of the WT receptor, but application of supramaximal concentrations of the agonist markedly decreased the time to half-peak. The asparagine mutant F107N displayed a significantly higher affinity for 5-HT than the WT receptor (1.62 versus 15.6 nM), which was mirrored in direction and magnitude by changes in the EC(50) value for this agonist (0.2 versus 1.2 microM). In contrast to the WT receptor, the mutant F107N was activated by acetylcholine (EC(50) = 260 microM). The response to acetylcholine was blocked by the 5-HT(3) receptor antagonist renzapride with a similar IC(50) value as that determined against currents generated by 5-HT in the WT receptor. These data suggest that F107 is an important determinant of agonist recognition at the 5-HT(3) receptor.

Plastic bullet injuries in Northern Ireland: experiences during a week of civil disturbance.

BACKGROUND: Plastic bullets were introduced to Northern Ireland for riot-control purposes in 1973. Their use has been controversial, with a number of fatalities. In the week beginning July 7, 1996, some 8,000 plastic bullets were fired during widespread rioting. METHODS: Details of injuries attributed to plastic bullets were obtained retrospectively from patient notes for the period July 8 to 14, 1996, in six hospitals. A total of 172 injuries in 155 patients were recorded. RESULTS: Nineteen percent of injuries were to the face/head/neck, 20% were to the chest or abdomen, and 61% were to the limbs. Abbreviated Injury Scale scores ranged from I to 3. Forty-two patients were admitted for hospitalization, three to intensive care units. No fatalities occurred. CONCLUSION: Plastic bullet impact to the abdomen or above may cause life-threatening injuries. Below this site, major trauma is unlikely.

Analysis of the ligand binding site of the 5-HT3 receptor using site directed mutagenesis: importance of glutamate 106.

The 5-HT3 receptor is a ligand-gated ion channel with significant structural similarity to the nicotinic acetylcholine receptor. Several regions that form the ligand binding site in the nicotinic acetylcholine receptor are partially conserved in the 5-HT3 receptor, presumably reflecting the conserved signal transduction mechanism. Specific amino acid differences in these regions may account for their distinct ligand recognition properties. Using site-directed mutagenesis, we have replaced one of these residues, glutamate 106 (E106), with aspartate (D), asparagine (N), alanine (A) or glutamine (Q) and characterized the ligand-binding and electrophysiological properties of the mutant receptors after transient expression in HEK-293 cells. The affinity for the selective 5-HT3 receptor antagonist [3H]GR65630 was decreased 14-fold in the mutant E106D (Kd = 3.69 +/- 0.32 nM) when compared to wildtype (WT, E106) 5-HT3 receptor (0.27 +/- 0.03 nM), while the affinity for E106N was unchanged (0.42 +/- 0.07 nM, means +/- SEM, n = 3-10). Decreased affinities for both E106D and E106N were observed for the antagonists granisetron, ondansetron and renzapride and for the agonists 5-HT (130- and 30-fold) and 2-methyl-5-HT (250- and 20-fold), respectively. Both mutants still formed 5-HT-activatable ion channels, but the high Hill coefficient of the concentration effect curves in wildtype (2.0) was decreased to unity in both cases. The EC50 of 5-HT was increased seven-fold in E106N (8.7 microM) when compared to wildtype (1.2 microM), but unchanged in E106D, and the potency of the antagonist ondansetron for both mutants was decreased. E106A and E106Q expressed poorly preventing a detailed characterization. These data suggest that E106 contributes to the ligand-binding site of the 5-HT3 receptor and may form an ionic or hydrogen bond interaction with the primary ammonium group of 5-HT.

The accident and emergency department as a single portal of entry for the reassessment of all trauma patients transferred to specialist units.

127 patients who were transferred to a regional referral centre for specialist treatment within 48 h of serious injury were reassessed on arrival in the receiving accident and emergency department by a trauma team. 80 transferred patients (63%) required intervention in the accident and emergency department to complete assessment or resuscitation. In view of the well recognised difficulties in managing patients with multiple trauma and the possibility that initially occult injuries may become clinically significant during transport, transferred trauma patients should be reassessed in the accident and emergency department of the receiving hospital by a trauma team consisting of senior medical staff experienced in all aspects of trauma care.

The effects of first aid training on public awareness of the management of a seriously injured patient.

Members of the public were questioned regarding previous first aid training and whether they believed that they would be able to manage a seriously injured person adequately. They were then assessed as to their awareness of basic principles of dealing with such a person. It was found that, whilst confidence as an independent variable was not associated with better knowledge, those who had some form of training were significantly more confident about their own abilities and also performed better in the assessment. However there remained a minority who would be unwilling to become involved in the initial treatment of a seriously injured person and training did not appear to reduce this tendency.

Production of 15-HETE by cultured smooth muscle cells from cerebral artery.

Hydroperoxyeicosatetraenoic acids (HPETEs) are potent vasoconstrictors which have been implicated in the pathogenesis of the delayed cerebrovascular spasm which follows subarachnoid haemorrhage. We have previously shown that one of their stable breakdown products, 15-hydroxyeicosatetraenoic acid (15-HETE), is the major lipoxygenase product released from isolated cerebral arteries. To investigate the vascular cell type responsible for their production we have established the culture of smooth muscle cells from cerebral arteries and have measured their release of HETEs upon stimulation with arachidonic acid. The cultured cells isolated from bovine cerebral arteries expressed smooth muscle phenotype as they stained positively for smooth muscle alpha-actin and not factor VIII-related antigen. Furthermore, these cells possessed the calcium and potassium conductances characteristic of smooth muscle cells. Upon incubation with arachidonic acid, the major lipoxygenase product was 15-HETE; no leukotrienes were detected. The formation of 15-HETE was concentration-dependent and was attenuated by nordihydroguaiaretic acid, but not by indomethacin. Similar to intact cerebral artery, cultured cerebrovascular smooth muscle cells have lipoxygenase activity, the major product of which is 15-HETE. These results are consistent with a possible role of the HPETEs in the pathogenesis of cerebral vasospasm.

Angulated radial neck fractures in children. A prospective study of percutaneous reduction.

We made a prospective study of angulated radial neck fractures in children reduced by leverage with a percutaneous Kirschner wire. Of 36 consecutive cases with angulation of more than 30 degrees reduction was successful in 33. We obtained an excellent or good final result in 31 (94%).