Estimations of the effective Young's modulus of specimens prepared by fused filament fabrication.

The elastic response of homogeneous isotropic materials is most commonly represented by their Young's modulus (E), but geometric variability associated with additive manufacturing results in materials that are neither homogeneous nor isotropic. Here we investigated methods to estimate the effective elastic modulus (Eeff) of samples fabricated by fused filament fabrication. We conducted finite element analysis (FEA) on printed samples based on material properties and CT-scanned geometries. The analysis revealed how the layer structure of a specimen altered the internal stress distribution and the resulting Eeff. We also investigated different empirical methods to estimate Eeff as guides. We envision the findings from our study can provide guidelines for modulus estimation of as-printed specimens, with the potential of applying to other extrusion-based additive manufacturing technologies.

Temporary faecal diversion in ileocolic resection for Crohn's disease: is there an impact on long-term surgical recurrence?

AIM: Temporary faecal diversion after ileocolic resection (ICR) for Crohn's disease reduces postoperative anastomotic complications in high-risk patients. The aim of this study was to assess if this approach also reduces long-term surgical recurrence. METHOD: This was a multicentre retrospective review of prospectively maintained databases. Patient demographics, medical and surgical details were collected by three specialist centres. All patients had undergone an ICR between 2000 and 2012. The primary end-point was surgical recurrence. RESULTS: Three hundred and twelve patients (80%) underwent an ICR without covering ileostomy (one stage). Seventy-seven (20%) had undergone an ICR with end ileostomy/double-barrel ileostomy/enterocolostomy followed by closure (two stage). The median follow-up was 105 months [interquartile range (IQR) 76-136 months]. The median time to ileostomy closure was 9 months (IQR 5-12 months). There was no significant difference in surgical recurrence between the one- and two-stage groups (18% vs 16%, P = 0.94). We noted that smokers (20% vs 34%, P = 0.01) and patients with penetrating disease (28% vs 52%, P < 0.01) were more likely to be defunctioned. A reduced recurrence rate was observed in the small high-risk group of patients who were smokers with penetrating disease behaviour treated with a two-stage strategy (0/10 vs 4/7, P = 0.12). CONCLUSION: Despite having higher baseline risk factors, the results in terms of rate of surgical recurrence over 9 years are similar for patients having a two-stage compared with a one-stage procedure.

Randomized clinical trial comparing collagen plug and advancement flap for trans-sphincteric anal fistula.

BACKGROUND: The role of a collagen plug for treating anal fistula is not well established. A randomized prospective multicentre non-inferiority study of surgical treatment of trans-sphincteric cryptogenic fistulas was undertaken, comparing the anal fistula plug with the mucosal advancement flap with regard to fistula recurrence rate and functional outcome. METHODS: Patients with an anal fistula were evaluated for eligibility in three centres, and randomized to either mucosal advancement flap surgery or collagen plug, with clinical follow-up at 3 and 12 months. The primary outcome was the fistula recurrence rate. Anal pain (visual analogue scale), anal incontinence (St Mark's score) and quality of life (Short Form 36 questionnaire) were also reported. RESULTS: Ninety-four patients were included; 48 were allocated to the plug procedure and 46 to advancement flap surgery. The median follow-up was 12 (range 9-24) months. The recurrence rate at 12 months was 66 per cent (27 of 41 patients) in the plug group and 38 per cent (15 of 40) in the flap group (P = 0·006). Anal pain was reduced after operation in both groups. Anal incontinence did not change in the follow-up period. Patients reported an increased quality of life after 3 months. There were no differences between the groups with regard to pain, incontinence or quality of life. CONCLUSION: There was a considerably higher recurrence rate after the anal fistula plug procedure than following advancement flap repair. Registration number: NCT01021774 (http://www.clinicaltrials.gov).

The Regulatory Challenges of Placing Dietary Ingredients on the European and US Market.

The DSHEA is 30 years old and its place in providing legitimate protections for public health through relevant agency oversight has created a patchwork of legal and scientific requirements. In contrast, the European Union has rules on supplements and permitted ingredients. Given the context of a global supply chain for food ingredients any conflict between the legality of ingredients between the U.S/EU can inhibit the economic viability of international trade. The purpose of this review is to contrast these different systems of legislative oversight. The analysis of both markets demonstrates a fragmentation in what are considered legal food ingredients between country wide harmonization and state rules and related interpretation. There are many commonalities in this regard between the U.S/EU, from borderline medicinal classifications to their resultant preclusion from food use. However, the codified legal system existing within the EU and excessive guidance can be viewed as time consuming and inflexible, especially for placing new ingredients on the market. The US in contrast is in a holding pattern for legislative interpretation regarding NDIs, GRAS and possible drug preclusion laws. As we hit the anniversary of the DSHEA recent commentary from U.S./EU central authorities point to increased international co-operation in ingredient safety assessments but whether this results in friction-free access between markets is to be determined.

AMB2018-03: Benchmark Physical Property Measurements for Material Extrusion Additive Manufacturing of Polycarbonate.

Material extrusion (MatEx) is finding increasing applications in additive manufacturing of thermoplastics due to the ease of use and the ability to process disparate polymers. Since part strength is anisotropic and frequently deviates negatively with respect to parts produced by injection molding, an urgent challenge is to predict final properties of parts made through this method. A nascent effort is underway to develop theoretical and computational models of MatEx part properties, but these efforts require comprehensive experimental data for guidance and validation. As part of the AM-Bench framework, we provide here a thorough set of measurements on a model system: polycarbonate printed in a simple rectangular shape. For the precursor material (as-received filament), we perform rheology, gel permeation chromatography, and dynamical mechanical analysis, to ascertain critical material parameters such as molar mass distribution, glass transition, and shear thinning. Following processing, we conduct X-ray computed tomography, scanning electron microscopy, depth sensing indentation, and atomic force microscopy modulus mapping. These measurements provide information related to pores, method of failure, and local modulus variations. Finally, we conduct tensile testing to assess strength and degree of anisotropy of mechanical properties. We find several effects that lead to degradation of tensile properties including the presence of pore networks, poor interfacial bonding, variations in interfacial mechanical behavior between rasters, and variable interaction of the neighboring builds within the melt state. The results provide insight into the processing-structure-property relationships and should serve as benchmarks for the development of mechanical models.

Oligomerization-dependent regulation of motility and morphogenesis by the collagen XVIII NC1/endostatin domain.

Collagen XVIII (c18) is a triple helical endothelial/epithelial basement membrane protein whose noncollagenous (NC)1 region trimerizes a COOH-terminal endostatin (ES) domain conserved in vertebrates, Caenorhabditis elegans and Drosophila. Here, the c18 NC1 domain functioned as a motility-inducing factor regulating the extracellular matrix (ECM)-dependent morphogenesis of endothelial and other cell types. This motogenic activity required ES domain oligomerization, was dependent on rac, cdc42, and mitogen-activated protein kinase, and exhibited functional distinction from the archetypal motogenic scatter factors hepatocyte growth factor and macrophage stimulatory protein. The motility-inducing and mitogen-activated protein kinase-stimulating activities of c18 NC1 were blocked by its physiologic cleavage product ES monomer, consistent with a proteolysis-dependent negative feedback mechanism. These data indicate that the collagen XVIII NC1 region encodes a motogen strictly requiring ES domain oligomerization and suggest a previously unsuspected mechanism for ECM regulation of motility and morphogenesis.

Nerve growth factor acts through cAMP-dependent protein kinase to increase the number of sodium channels in PC12 cells.

cAMP-dependent protein kinase (PKA) and phospholipid-dependent protein kinase (PKC) play a role in nerve growth factor (NGF)-mediated differentiation. In PC12 cells, NGF causes neurite outgrowth and increases the number of voltage-gated Na+ channels. Neurite outgrowth involves in part activation of PKC. How NGF regulates Na+ channel number is unknown. Using patch-clamp techniques, we find that agents activating PKC, including phorbol esters and a ras oncogene product (p21) that induces neurites, caused little increase in channel number. In contrast, agents increasing intracellular cAMP were as effective as NGF. A specific protein inhibitor of the PKA catalytic subunit blocked increases by NGF or cAMP. Thus, NGF increases Na+ channel number in PC12 cells in part by activating PKA but apparently not PKC.

Domains of E1A that bind p105Rb, p130, and p300 are required to block nerve growth factor-induced neurite growth in PC12 cells.

Nerve growth factor (NGF) causes PC12 cells to cease division and undergo sympathetic neuron-like differentiation, including neurite outgrowth. We have tested whether differentiation and division share overlapping control mechanisms in these cells. To do this, we have perturbed the activity of proteins known to participate in cell-cycle regulation by introducing the E1A oncogene or its mutant forms via microinjection into PC12 cells. The E1A protein binds to several putative cell cycle control proteins, including p105Rb (the product of the retinoblastoma susceptibility gene), as well as others of unknown function such as p130, p107, and p300. Similar to previous results, we find that wild-type E1A abrogates NGF-induced neurite extension. However, NGF does cause neurite outgrowth in the presence of E1A mutants known to have greatly reduced binding to either p105Rb and p130 or p300. Our experiments suggest that p105Rb, p130, and p300 may participate either in E1A-mediated inhibition of differentiation or in the NGF signal transduction pathway. We also report here that NGF affects phosphorylation of p105Rb, suggesting that Rb mediates at least some of NGF's effects. Our results raise the possibility that putative cell-cycle control proteins may participate not only in NGF-induced cessation of division but also in differentiation.

Ras family GTPases control growth of astrocyte processes.

Astrocytes in neuron-free cultures typically lack processes, although they are highly process-bearing in vivo. We show that basic fibroblast growth factor (bFGF) induces cultured astrocytes to grow processes and that Ras family GTPases mediate these morphological changes. Activated alleles of rac1 and rhoA blocked and reversed bFGF effects when introduced into astrocytes in dissociated culture and in brain slices using recombinant adenoviruses. By contrast, dominant negative (DN) alleles of both GTPases mimicked bFGF effects. A DN allele of Ha-ras blocked bFGF effects but not those of Rac1-DN or RhoA-DN. Our results show that bFGF acting through c-Ha-Ras inhibits endogenous Rac1 and RhoA GTPases thereby triggering astrocyte process growth, and they provide evidence for the regulation of this cascade in vivo by a yet undetermined neuron-derived factor.

Micronutrient assay for cancer prevention clinical trials: serum retinol, retinyl palmitate, alpha-carotene, and beta-carotene with the use of high-performance liquid chromatography.

Assay of serum levels of retinol, retinyl palmitate, alpha-carotene, and beta-carotene to assess nutritional status, to trials of retinol and/or beta-carotene to assess nutritional status, to monitor compliance with medication schedules, and to conduct toxicity surveillance. The optimal assay method for clinical trial use represents a balance between analytical power and speed/simplicity. Three such methods were evaluated by means of shared samples between two laboratories. Each method required less than 15 minutes per assay and detected all of the analytes of interest. Careful evaluation of calibration materials and procedures permitted different laboratories using different methods to produce results with an interlaboratory variability smaller than the within-laboratory variability for each separate method. Typical precisions for the analytes in serum samples are: retinol, 0.06 relative standard deviation (RSD; standard deviation divided by mean value); retinyl palmitate, 0.08 RSD; alpha-carotene, 0.15 RSD; and beta-carotene, 0.11 RSD. Application of these methods to several hundred samples indicated that retinyl palmitate and beta-carotene levels were indicative of administered retinol and beta-carotene, whereas retinol itself was not. Population variability in pretreatment serum levels of these micronutrients expressed as RSD (retinol, 0.24; alpha-carotene, 1.11; and beta-carotene, 0.98) far exceeded the analytical imprecision in these determinations, confirming that the present assays could meet the needs of current clinical intervention trials.

Infrared thermography of welding zones produced by polymer extrusion additive manufacturing.

In common thermoplastic additive manufacturing (AM) processes, a solid polymer filament is melted, extruded though a rastering nozzle, welded onto neighboring layers and solidified. The temperature of the polymer at each of these stages is the key parameter governing these non-equilibrium processes, but due to its strong spatial and temporal variations, it is difficult to measure accurately. Here we utilize infrared (IR) imaging - in conjunction with necessary reflection corrections and calibration procedures - to measure these temperature profiles of a model polymer during 3D printing. From the temperature profiles of the printed layer (road) and sublayers, the temporal profile of the crucially important weld temperatures can be obtained. Under typical printing conditions, the weld temperature decreases at a rate of approximately 100 °C/s and remains above the glass transition temperature for approximately 1 s. These measurement methods are a first step in the development of strategies to control and model the printing processes and in the ability to develop models that correlate critical part strength with material and processing parameters.

The Safety, Pharmacokinetics, and Nervous System Effects of Two Natural Sources of Caffeine in Healthy Adult Males.

This double-blind crossover clinical trial randomized 12 adult males to receive 200 mg of caffeine from a green coffee extract, a guayusa leaf extract, and a synthetic control to compare their safety, absorption, and effect on neurotransmitters. The results showed no statistically significant changes in blood pressure or heart rate from baseline to 120 min postdose of each natural source compared with changes from baseline in the control (0.094 < = P < = 0.910). The ratios of Cmax , AUC0-4 , and AUC0-∞ of each natural source to the control were bioequivalent by US Food and Drug Administration standards (90% CI within 80-125%). The guayusa leaf extract stimulated a significantly lower increase in epinephrine compared with the control (+0.5 vs. +2.78 µg/gCr, P = 0.04), while the green coffee extract provoked an increase in epinephrine similar to the control (+3.21 vs. +2.78 µg/gCr, P = 0.569). Implications for future clinical research are discussed.

STANDARD REFERENCE MATERIALS FOR THE POLYMERS INDUSTRY.

The National Institute of Standards and Technology (NIST) provides science, industry, and government with a central source of well-characterized materials certified for chemical composition or for some chemical or physical property. These materials are designated Standard Reference Materials® (SRMs) and are used to calibrate measuring instruments, to evaluate methods and systems, or to produce scientific data that can be referred readily to a common base. In this paper, we discuss the history of polymer based SRMs, their current status, and challenges and opportunities to develop new standards to address industrial measurement challenges.

Calcium-dependent inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells.

The inactivation of calcium channels in mammalian pituitary tumor cells (GH3) was studied with patch electrodes under voltage clamp in cell-free membrane patches and in dialyzed cells. The calcium current elicited by depolarization from a holding potential of -40 mV passed predominantly through one class of channels previously shown to be modulated by dihydropyridines and cAMP-dependent phosphorylation (Armstrong and Eckert, 1987). When exogenous calcium buffers were omitted from the pipette solution, the macroscopic calcium current through those channels inactivated with a half time of approximately 10 ms to a steady state level 40-75% smaller than the peak. Inactivation was also measured as the reduction in peak current during a test pulse that closely followed a prepulse. Inactivation was largely reduced or eliminated by (a) buffering free calcium in the pipette solution to less than 10(-8) M; (b) replacing extracellular calcium with barium; (c) increasing the prepulse voltage from +10 to +60 mV; or (d) increasing the intracellular concentration of cAMP, either 'directly' with dibutyryl-cAMP or indirectly by activating adenylate cyclase with forskolin or vasoactive intestinal peptide. Thus, inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells only occurs when membrane depolarization leads to calcium ion entry and intracellular accumulation.

Micronuclei in exfoliated bladder cells among individuals chronically exposed to arsenic in drinking water.

Inorganic arsenic is an established cause of lung and skin cancer. Epidemiological evidence from Taiwan suggests that arsenic causes more fatal internal cancers, with the highest relative risks reported for bladder cancer. We conducted a cross-sectional biomarker study in a Chilean male population chronically exposed to high (70 subjects) and low (55 subjects) arsenic levels in their drinking water (average concentrations, 600 and 15 micrograms As/liter, respectively). A fluorescent version of the exfoliated bladder cell micronucleus (MN) assay was used employing fluorescence in situ hybridization with a centromeric probe to identify the presence (MN+) or absence (MN-) of whole chromosomes within micronuclei, thereby determining the mechanism of arsenic-induced genotoxicity in vivo. We divided the study population into quintiles by urinary arsenic levels and found an exposure-dependent increase in micronucleated cell prevalence in quintiles 2-4 (urinary arsenic, 54-729 micrograms/liter). The largest increase appeared when quintile 4 was compared to quintile 1 [prevalence ratio, 3.0; 95% confidence interval (CI), 1.9-4.6]. The prevalence of MN+ increased to 3.1-fold in quintile 4 (95% CI, 1.4-6.6), and the prevalence of MN-increased to 7.5-fold in quintile 3 (95% CI, 2.8-20.3), suggesting that chromosome breakage was the major cause of MN formation. Prevalences of total MN, MN+, and MN- returned to baseline levels in quintile 5 (urinary arsenic, 729-1894 micrograms/liter), perhaps due to cytostasis or cytotoxicity. These results add additional weight to the hypothesis that ingesting arsenic-contaminated water enhances bladder cancer risk and suggest that arsenic induces genetic damage to bladder cells at drinking water levels close to the current United States Maximum Contaminant Level of 50 micrograms/liter for arsenic.

Decrease in bladder cell micronucleus prevalence after intervention to lower the concentration of arsenic in drinking water.

Epidemiological studies performed in Taiwan, Argentina, and Chile suggest that ingestion of arsenic (As) may cause bladder cancer. Because of these findings, we previously investigated the relationship between As ingestion and genetic damage to the urothelium in two cross-sectional biomarker studies, one in Nevada and one in Chile. In both studies, we found that increased levels of micronucleated cells (MNCs) in exfoliated bladder cells were associated with elevated concentrations of As in drinking water, suggesting that As induces genetic damage to bladder cells. To further investigate this relationship, we conducted an intervention study in a subset of highly exposed men (n = 34) from the cross-sectional study in Chile. Subjects whose usual source of water contained about 600 micrograms/liter As were supplied with water lower in As (45 micrograms/liter) for 8 weeks, allowing ample opportunity for renewal and exfoliation of bladder epithelial cells. Mean urinary As levels decreased during the intervention from 742 to 225 micrograms/liter. Bladder MNC prevalence also decreased from 2.63 MNCs/1000 cells preintervention to 1.79 MNCs/1000 cells postintervention (P < 0.05). When the analysis was limited to individuals previously having subcytotoxic urinary As levels (< 700 micrograms/liter), the change between pre- and postintervention MNC was more pronounced: the level decreased from 3.54 to 1.47 MNCs/1000 cells, respectively (P = 0.002). Among smokers, MNC prevalences decreased from 4.45 MNCs/1000 cells preintervention to 1.44 MNCs/1000 cells postintervention (P = 0.002). Among nonsmokers, the decrease was much smaller: 2.04 MNCs/1000 cells preintervention to 1.90 MNCs/1000 cells postintervention (P = 0.25), suggesting that smoker's bladder cells could be more susceptible to genotoxic damage caused by As. The reduction in bladder MNC prevalence with reduction in As intake provides further evidence that As is genotoxic to bladder cells.

Increased micronuclei in exfoliated bladder cells of individuals who chronically ingest arsenic-contaminated water in Nevada.

It is well established that inorganic arsenic is causally associated with lung cancer via inhalation and skin cancer via ingestion. Epidemiological evidence based on studies in Taiwan suggests that ingestion of inorganic arsenic may also cause other more fatal internal cancers, with the highest relative risks reported for bladder cancer. Here, we have used a biological marker of response, the micronucleus assay in exfoliated bladder cells, to evaluate the possible genotoxic effects of chronic arsenic ingestion on the bladder. The overall objective of this study was to compare the frequency of micronucleated cells in exfoliated bladder and buccal cells between a group of 18 individuals in Nevada who chronically ingested high levels of inorganic arsenic from their well water (average level, 1,312 micrograms/liter) and an individually matched control group with low exposure to arsenic (average level, 16 micrograms/liter). A 1.8-fold increase (90% confidence interval, 1.06-2.99) was observed in the weighted mean frequency of micronucleated bladder cells in the exposed group (2.79 per 1000 cells) compared with the unexposed group (1.57 per 1000 cells). In addition, the frequency of micronucleated bladder cells was positively associated with the urinary concentration of inorganic arsenic plus its methylated metabolites (Spearman correlation = 0.33; P = 0.03). In contrast, there was no increase in micronucleated buccal cells associated with arsenic ingestion (frequency ratio = 1.0; 90% confidence interval, 0.65-1.53). The results of this study provide evidence that chronic ingestion of high levels of inorganic arsenic in drinking water is associated with an increased frequency of micronucleated bladder cells. These findings are consistent with a genotoxic effect of arsenic on bladder cells, but a larger study is needed to confirm them.

Scientists in the courtroom: basic pointers for the expert scientific witness.

The need for expert and unbiased participation in legal proceedings by physicians, industrial hygienists, toxicologists, environmental scientists, regulators, and similar professionals is hampered by lack of familiarity with the requirements of expert testimony and lack of opportunities for professional training in this activity. Drawing on material developed in a continuing education course offered by the University of Washington, we describe the role and process of being an expert witness and provide basic information regarding good professional practices pertaining to the testifying expert role.