Complementary catalysis and analysis within solid state additively manufactured metal micro flow reactors.

Additive Manufacturing is transforming how researchers and industrialists look to design and manufacture chemical devices to meet their specific needs. In this work, we report the first example of a flow reactor formed via the solid-state metal sheet lamination technique, Ultrasonic Additive Manufacturing (UAM), with directly integrated catalytic sections and sensing elements. The UAM technology not only overcomes many of the current limitations associated with the additive manufacturing of chemical reactionware but it also significantly increases the functionality of such devices. A range of biologically important 1, 4-disubstituted 1, 2, 3-triazole compounds were successfully synthesised and optimised in-flow through a Cu mediated Huisgen 1, 3-dipolar cycloaddition using the UAM chemical device. By exploiting the unique properties of UAM and continuous flow processing, the device was able to catalyse the proceeding reactions whilst also providing real-time feedback for reaction monitoring and optimisation.

In-situ time resolved spectrographic measurement using an additively manufactured metallic micro-fluidic analysis platform.

INTRODUCTION: Microfluidic reactionware allows small volumes of reagents to be utilized for highly controlled flow chemistry applications. By integrating these microreactors with onboard analytical systems, the devices change from passive ones to active ones, increasing their functionality and usefulness. A pressing application for these active microreactors is the monitoring of reaction progress and intermediaries with respect to time, shedding light on important information about these real-time synthetic processes. OBJECTIVE: In this multi-disciplinary study the objective was to utilise advanced digital fabrication to research metallic, active microreactors with integrated fibre optics for reaction progress monitoring of solvent based liquids, incompatible with previously researched polymer devices, in combination with on-board Ultraviolet-visible spectroscopy for real-time reaction monitoring. METHOD: A solid-state, metal-based additive manufactured system (Ultrasonic Additive Manufacturing) combined with focussed ion beam milling, that permitted the accurate embedment of delicate sensory elements directly at the point of need within aluminium layers, was researched as a method to create active, metallic, flow reactors with on-board sensing. This outcome was then used to characterise and correctly identify concentrations of UV-active water-soluble B-vitamin nicotinamide and fluorescein. A dilution series was formed from 0.01-1.75 mM; which was pumped through the research device and monitored using UV-vis spectroscopy. RESULTS: The results uniquely showed the in-situ ion milling of ultrasonically embedded optical fibres resulted in a metallic microfluidic reaction and monitoring device capable of measuring solvent solutions from 18 µM to 18 mM of nicotinamide and fluorescein, in real time. This level of accuracy highlights that the researched device and methods are capable of real-time spectrographic analysis of a range of chemical reactions outside of those possible with polymer devices.

Review article: novel oral-targeted therapies in inflammatory bowel disease.

BACKGROUND: There is a great unmet clinical need for efficacious, tolerable, economical and orally administrated drugs for the treatment of inflammatory bowel disease (IBD). New therapeutic avenues have become possible including the development of medications that target specific genetic pathways found to be relevant in other immune mediated diseases. AIMS: To provide an overview of recent clinical trials for new generation oral targeted medications that may have a future role in IBD management. METHODS: Pubmed and Medline searches were performed up to 1 March 2018 using keywords: "IBD", "UC", "CD", "inflammatory bowel disease" "ulcerative colitis", "Crohn's disease" in combination with "phase", "study", "trial" and "oral". A manual search of the clinical trial register, article reference lists, abstracts from meetings of Digestive Disease Week, United European Gastroenterology Week and ECCO congress were also conducted. RESULTS: In randomised controlled trials primary efficacy endpoints were met for tofacitinib (JAK 1/3 inhibitor-phase III), upadacitinib (JAK 1 inhibitor-phase II) and AJM300 (α4-integrin antagonist-phase II) in ulcerative colitis. Ozanimod (S1P receptor agonist-phase II) also demonstrated clinical remission. For Crohn's disease, filgotinib (JAK1 inhibitor-phase II) met primary endpoints and laquinimod (quinolone-3-carboxide small molecule-phase II) was also efficacious. Trials using mongersen (SMAD7 inhibitor) and vidofludimus (dihydroorotate dehydrogenase inhibitor) have been halted. CONCLUSIONS: This is potentially the start of an exciting new era in which multiple therapeutic options are at the disposal of physicians to treat IBD on an individualised basis. Head-to-head studies with existing treatments and longer term safety data are needed for this to be possible.

Protective antibodies against Clostridium difficile are present in intravenous immunoglobulin and are retained in humans following its administration.

The prevalence of serum antibodies against Clostridium difficile (CD) toxins A and B in healthy populations have prompted interest in evaluating the therapeutic activity of intravenous immunoglobulin (IVIg) in individuals experiencing severe or recurrent C. difficile infection (CDI). Despite some promising case reports, a definitive clinical role for IVIg in CDI remains unclear. Contradictory results may be attributed to a lack of consensus regarding optimal dose, timing of administration and patient selection as well as variability in specific antibody content between commercial preparations. The purpose of this study was to investigate retrospectively the efficacy of three commercial preparations of IVIg for treating severe or recurrent CDI. In subsequent mechanistic studies using protein microarray and toxin neutralization assays, all IVIg preparations were analysed for specific binding and neutralizing antibodies (NAb) to CD antigens in vitro and the presence of anti-toxin NAbs in vivo following IVIg infusion. A therapeutic response to IVIg was observed in 41% (10 of 17) of the CDI patients. Significant variability in multi-isotype specific antibodies to a 7-plex panel of CD antigens and toxin neutralization efficacies were observed between IVIg preparations and also in patient sera before and after IVIg administration. These results extend our current understanding of population immunity to CD and support the inclusion of surface layer proteins and binary toxin antigens in CD vaccines. Future strategies could enhance IVIg treatment response rates by using protein microarray to preselect donor plasma/serum with the highest levels of anti-CD antibodies and/or anti-toxin neutralizing capacities prior to fractionation.

Customisable 3D printed microfluidics for integrated analysis and optimisation.

The formation of smart Lab-on-a-Chip (LOC) devices featuring integrated sensing optics is currently hindered by convoluted and expensive manufacturing procedures. In this work, a series of 3D-printed LOC devices were designed and manufactured via stereolithography (SL) in a matter of hours. The spectroscopic performance of a variety of optical fibre combinations were tested, and the optimum path length for performing Ultraviolet-visible (UV-vis) spectroscopy determined. The information gained in these trials was then used in a reaction optimisation for the formation of carvone semicarbazone. The production of high resolution surface channels (100-500 µm) means that these devices were capable of handling a wide range of concentrations (9 µM-38 mM), and are ideally suited to both analyte detection and process optimisation. This ability to tailor the chip design and its integrated features as a direct result of the reaction being assessed, at such a low time and cost penalty greatly increases the user's ability to optimise both their device and reaction. As a result of the information gained in this investigation, we are able to report the first instance of a 3D-printed LOC device with fully integrated, in-line monitoring capabilities via the use of embedded optical fibres capable of performing UV-vis spectroscopy directly inside micro channels.

Potential of lactoferrin to prevent antibiotic-induced Clostridium difficile infection.

OBJECTIVES: Clostridium difficile infection (CDI) is a global healthcare problem. Recent evidence suggests that the availability of iron may be important for C. difficile growth. This study evaluated the comparative effects of iron-depleted (1% Fe(3+) saturated) bovine apo-lactoferrin (apo-bLf) and iron-saturated (85% Fe(3+) saturated) bovine holo-lactoferrin (holo-bLf) in a human in vitro gut model that simulates CDI. METHODS: Two parallel triple-stage chemostat gut models were inoculated with pooled human faeces and spiked with C. difficile spores (strain 027 210, PCR ribotype 027). Holo- or apo-bLf was instilled (5 mg/mL, once daily) for 35 days. After 7 days, clindamycin was instilled (33.9 mg/L, four times daily) to induce simulated CDI. Indigenous microflora populations, C. difficile total counts and spores, cytotoxin titres, short chain fatty acid concentrations, biometal concentrations, lactoferrin concentration and iron content of lactoferrin were monitored daily. RESULTS: In the apo-bLf model, germination of C. difficile spores occurred 6 days post instillation of clindamycin, followed by rapid vegetative cell proliferation and detectable toxin production. By contrast, in the holo-bLf model, only a modest vegetative cell population was observed until 16 days post antibiotic administration. Notably, no toxin was detected in this model. In separate batch culture experiments, holo-bLf prevented C. difficile vegetative cell growth and toxin production, whereas apo-bLf and iron alone did not. CONCLUSIONS: Holo-bLf, but not apo-bLf, delayed C. difficile growth and prevented toxin production in a human gut model of CDI. This inhibitory effect may be iron independent. These observations suggest that bLf in its iron-saturated state could be used as a novel preventative or treatment strategy for CDI.

Differential binding and internalization of Clostridium difficile toxin A by human peripheral blood monocytes, neutrophils and lymphocytes.

Colitis due to Clostridium difficile infection is mediated by secreted toxins A and B and is characterized by infiltration by cells from the systemic circulation. The aim of our study was to investigate interactions between fluorescently labelled toxin A and peripheral blood monocytes, neutrophils and lymphocytes. Purified toxin A was labelled with Alexa Fluor® 488 (toxin A(488)) and incubated with isolated human peripheral blood mononuclear cells or washed whole blood cells for varying time intervals at either 37 or 4 °C/ice. The ability of trypan blue to quench cell surface-associated (but not cytoplasmic) fluorescence was also investigated. At 37 °C, toxin A(488) -associated fluorescence in monocytes peaked at 1 h (majority internalized), with subsequent loss associated with cell death. In contrast to monocytes, binding of toxin A(488) in neutrophils was greater on ice than at 37 °C. Studies using trypan blue suggested that over 3 h at 37 °C, most of the toxin A(488)-associated fluorescence in neutrophils remained at the cell surface. Over 48 h (37 °C and ice/4 °C), there was minimal toxin A(488)-associated fluorescence in lymphocytes. These studies suggest major differences in interactions between toxin A and circulating cells that infiltrate the mucosa during colonic inflammation in C. difficile infection.

Liaison neurologists facilitate accurate neurological diagnosis and management, resulting in substantial savings in the cost of inpatient care.

BACKGROUND: Despite understaffing of neurology services in Ireland, the demand for liaison neurologist input into the care of hospital inpatients is increasing. This aspect of the workload of the neurologist is often under recognised. AIMS/METHODS: We prospectively recorded data on referral and service delivery patterns to a liaison neurology service, the neurological conditions encountered, and the impact of neurology input on patient care. RESULTS: Over a 13-month period, 669 consults were audited. Of these, 79% of patients were seen within 48 h and 86% of patients were assessed by a consultant neurologist before discharge. Management was changed in 69% cases, and discharge from hospital expedited in 50%. If adequate resources for neurological assessment had been available, 28% could have been seen as outpatients, with projected savings of 857 bed days. CONCLUSIONS: Investment in neurology services would facilitate early accurate diagnosis, efficient patient and bed management, with substantial savings.

The clinical spectrum of Lyme neuroborreliosis.

Lyme disease is a multisystem infectious disease, endemic in parts of Europe, including the West of Ireland. Neurological manifestions (neuroborreliosis) are variable. Presenting neurological syndromes include meningitis, cranial neuropathies, myeloradiculitis and mononeuritis multiplex. A lack of specificity in serological diagnosis may add to diagnostic confusion. We reviewed thirty cases of acute Lyme disease in the West of Ireland and found neurological syndromes in 15 (50%), with painful radiculopathy (12 patients; 80%) and cranial neuropathy (7 patients; 46%) occurring frequently. Neuroborreliosis needs to be considered in the differential diagnosis of these neurological syndromes in the appropriate clinical context.

Recent advances in Clostridium difficile-associated disease.

The main purpose of this article is to review recent developments in the management of acute and recurrent Clostridium difficile-associated disease, with consideration of existing and new antibiotic and non-antibiotic agents for treatment. Details of the current developmental stage of new agents are provided and the role of surgery in the management of severe disease is discussed. Infection control measures considered comprise prudent use of antimicrobials, prevention of cross-infection and surveillance. Other topics that are covered include the recent emergence of an epidemic hypervirulent strain, pathogenesis, clinical presentation and approaches to rapid diagnosis and assessment of the colonic disease.

Collaborative text-annotation resource for disease-centered relation extraction from biomedical text.

Agglomerating results from studies of individual biological components has shown the potential to produce biomedical discovery and the promise of therapeutic development. Such knowledge integration could be tremendously facilitated by automated text mining for relation extraction in the biomedical literature. Relation extraction systems cannot be developed without substantial datasets annotated with ground truth for benchmarking and training. The creation of such datasets is hampered by the absence of a resource for launching a distributed annotation effort, as well as by the lack of a standardized annotation schema. We have developed an annotation schema and an annotation tool which can be widely adopted so that the resulting annotated corpora from a multitude of disease studies could be assembled into a unified benchmark dataset. The contribution of this paper is threefold. First, we provide an overview of available benchmark corpora and derive a simple annotation schema for specific binary relation extraction problems such as protein-protein and gene-disease relation extraction. Second, we present BioNotate: an open source annotation resource for the distributed creation of a large corpus. Third, we present and make available the results of a pilot annotation effort of the autism disease network.

Comparative analysis of neurological disorders focuses genome-wide search for autism genes.

The behaviors of autism overlap with a diverse array of other neurological disorders, suggesting common molecular mechanisms. We conducted a large comparative analysis of the network of genes linked to autism with those of 432 other neurological diseases to circumscribe a multi-disorder subcomponent of autism. We leveraged the biological process and interaction properties of these multi-disorder autism genes to overcome the across-the-board multiple hypothesis corrections that a purely data-driven approach requires. Using prior knowledge of biological process, we identified 154 genes not previously linked to autism of which 42% were significantly differentially expressed in autistic individuals. Then, using prior knowledge from interaction networks of disorders related to autism, we uncovered 334 new genes that interact with published autism genes, of which 87% were significantly differentially regulated in autistic individuals. Our analysis provided a novel picture of autism from the perspective of related neurological disorders and suggested a model by which prior knowledge of interaction networks can inform and focus genome-scale studies of complex neurological disorders.

Neurotrophic actions of PACAP-38 and LIF on human neuroblastoma SH-SY5Y cells.

The neurotrophic actions of pituitary adenylate cyclase-activating polypeptide (PACAP)-38 and leukemia inhibitory factor (LIF) were investigated in human neuroblastoma SH-SY5Y cells. Effects on differentiation were assessed through monitoring morphological changes and Western blot analysis of the expression of neuronal marker proteins. In contrast to PACAP-38, which induced a 5.5-fold increase in the number of neurite-bearing cells, LIF had no significant effect on cell morphology compared to control cells over the 4-day time course. Cells co-treated with PACAP-38+LIF showed a similar increase in neurite-bearing cells compared to those treated with PACAP-38 alone. Cell morphology was similar for PACAP-38-treated and PACAP-38+LIF-co-treated cells, with the formation of bipolar neuron-like cells with long thin neurites, topped by growth cone-like structures and varicosities. SH-SY5Y cells express tyrosine hydroxylase (TH) but only low levels of the neuronal marker proteins: Bcl-2, GAP-43 and choline acetyltransferase (ChAT). Treatment of cells with PACAP-38 induced the expression of Bcl-2, GAP-43, and ChAT but did not appear to alter the expression of TH. LIF failed to induce the expression of GAP-43 and had little effect on the expression of TH, but did induce the expression of Bcl-2 and upregulated the expression of ChAT. Co-treatment with LIF had no effect on PACAP-38-induced expression of Bcl-2, GAP-43, and ChAT. Cells differentiated for 4 days with PACAP-38 or treated with LIF also displayed increased resistance to hypoxic conditions and to treatment with H2O2 and TNFalpha. The increased resistance to hypoxic conditions for PACAP-differentiated cells was blocked by the p38 MAP kinase inhibitor, SB203580, but not by the MEK1 inhibitor, PD98059. Additionally, cell proliferation assays show that LIF, but not PACAP-38, stimulates proliferation of SH-SY5Y cells, and this observed increase by LIF is not attenuated by co-treatment with PACAP. Further investigation of the intracellular signaling pathways mediating the neurotrophic effects of PACAP on SH-SY5Y cells indicate that neither phospholipase C activation nor Ca2+/calmodulin-dependent kinase II (CAMKII) are involved.

Recent advances in Clostridium difficile-associated disease.

The main purpose of this article is to review recent developments in the management of acute and recurrent Clostridium difficile-associated disease, with consideration of existing and new antibiotic and non-antibiotic agents for treatment. Details of the current developmental stage of new agents are provided and the role of surgery in the management of severe disease is discussed. Infection control measures considered comprise prudent use of antimicrobials, prevention of cross-infection and surveillance. Other topics that are covered include the recent emergence of an epidemic hypervirulent strain, pathogenesis, clinical presentation and approaches to rapid diagnosis and assessment of the colonic disease.

PACAP-38 induces neuronal differentiation of human SH-SY5Y neuroblastoma cells via cAMP-mediated activation of ERK and p38 MAP kinases.

The intracellular signaling pathways mediating the neurotrophic actions of pituitary adenylate cyclase-activating polypeptide (PACAP) were investigated in human neuroblastoma SH-SY5Y cells. Previously, we showed that SH-SY5Y cells express the PAC(1) and VIP/PACAP receptor type 2 (VPAC(2)) receptors, and that the robust cAMP production in response to PACAP and vasoactive intestinal peptide (VIP) was mediated by PAC(1) receptors (Lutz et al. 2006). Here, we investigated the ability of PACAP-38 to differentiate SH-SY5Y cells by measuring morphological changes and the expression of neuronal markers. PACAP-38 caused a concentration-dependent increase in the number of neurite-bearing cells and an up-regulation in the expression of the neuronal proteins Bcl-2, growth-associated protein-43 (GAP-43) and choline acetyltransferase: VIP was less effective than PACAP-38 and the VPAC(2) receptor-specific agonist, Ro 25-1553, had no effect. The effects of PACAP-38 and VIP were blocked by the PAC(1) receptor antagonist, PACAP6-38. As observed with PACAP-38, the adenylyl cyclase activator, forskolin, also induced an increase in the number of neurite-bearing cells and an up-regulation in the expression of Bcl-2 and GAP-43. PACAP-induced differentiation was prevented by the adenylyl cyclase inhibitor, 2',5'-dideoxyadenosine (DDA), but not the protein kinase A (PKA) inhibitor, H89, or by siRNA-mediated knock-down of the PKA catalytic subunit. PACAP-38 and forskolin stimulated the activation of extracellular signal-regulated kinase (ERK), mitogen-activated protein kinase (MAP; p38 MAP kinase) and c-Jun N-terminal kinase (JNK). PACAP-induced neuritogenesis was blocked by the MEK1 inhibitor PD98059 and partially by the p38 MAP kinase inhibitor SB203580. Activation of exchange protein directly activated by cAMP (Epac) partially mimicked the effects of PACAP-38, and led to the phosphorylation of ERK but not p38 MAP kinase. These results provide evidence that the neurotrophic effects of PACAP-38 on human SH-SY5Y neuroblastoma cells are mediated by the PAC(1) receptor through a cAMP-dependent but PKA-independent mechanism, and furthermore suggest that this involves Epac-dependent activation of ERK as well as activation of the p38 MAP kinase signaling pathway.

Simultaneous bilateral obturator nerve stimulation during transurethral electrovaporization of the prostate.

Transurethral vaporization of the prostate is a new method of electrosurgery used for treating benign prostatic hypertrophy. We observed simultaneous obturator nerve stimulation during spinal anesthesia while the middle lobe of the prostate was being resected. A discussion of why electrovaporization is different from traditional electrocautery devices is presented. Due to the increased power required during electrovaporization, and the arcing at the electrode, a large amount of demodulated, low-frequency current is produced that can stimulate the obturator nerve. This can occur even if the electrode is not close to the lateral bladder wall. The solution to the problem of obturator nerve stimulation, including bilateral obturator nerve blocks using a lower power setting during resection and converting from a regional to general anesthetic with neuromuscular blockade, is discussed.