Two Distinct Bacterial Biofilm Components Trigger Metamorphosis in the Colonial Hydrozoan Hydractinia echinata.
In marine environments, the bacterially induced metamorphosis of larvae is a widespread cross-kingdom communication phenomenon that is critical for the persistence of many marine invertebrates. However, the majority of inducing bacterial signals and underlying cellular mechanisms remain enigmatic. The marine hydroid Hydractinia echinata is a well-known model system for investigating bacterially stimulated larval metamorphosis, as larvae transform into the colonial adult stage within 24 h of signal detection. Although H. echinata has served as a cell biological model system for decades, the identity and influence of bacterial signals on the morphogenic transition remained largely unexplored. Using a bioassay-guided analysis, we first determined that specific bacterial (lyso)phospholipids, naturally present in bacterial membranes and vesicles, elicit metamorphosis in Hydractinia larvae in a dose-response manner. Lysophospholipids, as single compounds or in combination (50 μM), induced metamorphosis in up to 50% of all larvae within 48 h. Using fluorescence-labeled bacterial phospholipids, we demonstrated that phospholipids are incorporated into the larval membranes, where interactions with internal signaling cascades are proposed to occur. Second, we identified two structurally distinct exopolysaccharides of bacterial biofilms, the new Rha-Man polysaccharide from Pseudoalteromonas sp. strain P1-9 and curdlan from Alcaligenes faecalis, to induce metamorphosis in up to 75% of tested larvae. We also found that combinations of (lyso)phospholipids and curdlan induced transformation within 24 h, thereby exceeding the morphogenic activity observed for single compounds and bacterial biofilms. Our results demonstrate that two structurally distinct, bacterium-derived metabolites converge to induce high transformation rates of Hydractinia larvae and thus may help ensure optimal habitat selection. IMPORTANCE Bacterial biofilms profoundly influence the recruitment and settlement of marine invertebrates, critical steps for diverse marine processes such as the formation of coral reefs, the maintenance of marine fisheries, and the fouling of submerged surfaces. However, the complex composition of biofilms often makes the characterization of individual signals and regulatory mechanisms challenging. Developing tractable model systems to characterize these coevolved interactions is the key to understanding fundamental processes in evolutionary biology. Here, we characterized two types of bacterial signaling molecules, phospholipids and polysaccharides, that induce the morphogenic transition. We then analyzed their abundance and combinatorial activity. This study highlights the general importance of multiple bacterial signal converging activity in development-related cross-kingdom signaling and poses the question of whether complex lipids and polysaccharides are general metamorphic cues for cnidarian larvae.


Glycogen metabolism in psoriatic epidermis and in regenerating epidermis.
The observation that the glycogen content of epidermis from psoriatic lesions and from regenerating wound epithelium is increased has been confirmed by quantitative estimation. In epidermis from psoriatic lesions, although the proportion of glycogen synthase in the I form is only about 5% of the total and similar to control values, total glycogen synthase activity is increased approximately 4-fold and hence glycogen synthase I activity is increased to the same extent. In contrast, total phosphorylase activity is only slightly increased and, since the proportion of the enzyme in the a form is reduced, phosphorylase a activity is similar to control values. In epidermis from psoriatic lesions, the concentration of UDP-glucose is approximately doubled, and the concentrations of fructose 1,6-bisphosphate and of 6-phosphogluconate are increased approximately 5-fold. It is concluded that rates of glycogen synthesis, of glycolysis and of the pentose phosphate pathway are all enhanced in vivo and in consequence the rate of glucose uptake by psoriatic epidermis must be increased. In the non-involved epidermis of psoriatic patients the glycogen content is within normal limits, and although total glycogen synthase activity is increased the ratio of glycogen synthase I to phosphorylase a is maintained at normal levels by the appropriate phosphorylation of both enzymes. In regenerating wound epithelium in the pig, the changes in enzyme activity and in metabolite concentration closely resemble those found in epithelium from psoriatic lesions except that in wound epithelium the proportion of phosphorylase in the a form is increased relative to normal epithelium.


Outcomes research: the history, debate, and implications for the field of occupational therapy.
The field of occupational therapy, as with all facets of health care, has been profoundly affected by the changing climate of health care delivery. The combination of cost-effectiveness and quality of care has become the benchmark for and consequent drive behind the rise of managed health care delivery systems. The spawning of outcomes research is in direct response to the need for comparative databases to provide results of effectiveness in health care treatment protocols, evaluations of health-related quality of life, and cost containment measures. Outcomes management is the application of outcomes research data by all levels of health care providers. The challenges facing occupational therapists include proving our value in an economic trend of downsizing, competing within the medical profession, developing and affiliating with new payer sources, and reengineering our careers to meet the needs of the new, nontraditional health care marketplace.


Transesophageal Speckle-Tracking Echocardiography Improves Right Ventricular Systolic Function Assessment in the Perioperative Setting.
Perioperative evaluation of right ventricular (RV) systolic function is important to follow intraoperative changes, but it is often not possible to assess with transthoracic echocardiographic (TTE) imaging, because of surgical field constraints. Echocardiographic RV quantification is most commonly performed using tricuspid annular plane systolic excursion (TAPSE), but it is not clear whether this method works with transesophageal echocardiographic (TEE) imaging. This study was performed to evaluate the relationship between TTE and TEE TAPSE distances measured with M-mode imaging and in comparison with speckle-tracking TTE and TEE measurements. Prospective observational TTE and TEE imaging was performed during elective cardiac surgical procedures in 100 subjects. Speckle-tracking echocardiographic TAPSE distances were determined and compared with the TTE M-mode TAPSE standard. Both an experienced and an inexperienced user of the speckle-tracking echocardiographic software evaluated the images, to enable interobserver assessment in 84 subjects. The comparison between TTE M-mode TAPSE and TEE M-mode TAPSE demonstrated significant variability, with a Spearman correlation of 0.5 and a mean variance in measurement of 6.5 mm. There was equivalence within data pairs and correlations between TTE M-mode TAPSE and both speckle-tracking TTE and speckle-tracking TEE TAPSE, with Spearman correlations of 0.65 and 0.65, respectively. The average variance in measurement was 0.6 mm for speckle-tracking TTE TAPSE and 1.5 mm for speckle-tracking TEE TAPSE. Using TTE M-mode TAPSE as a control, TEE M-mode TAPSE results are not accurate and should not be used clinically to evaluate RV systolic function. The relationship between speckle-tracking echocardiographic TAPSE and TTE M-mode TAPSE suggests that in the perioperative setting, speckle-tracking TEE TAPSE might be used to quantitatively evaluate RV systolic function in the absence of TTE imaging.


Additional value of FDG-PET to contrast enhanced-computed tomography (CT) for the diagnosis of mediastinal lymph node metastasis in non-small cell lung cancer: a Japanese multicenter clinical study.
This study was a controlled multicenter clinical study to verify the diagnostic effects of additional FDG-PET to contrast-enhanced CT for mediastinal lymph node metastasis in patients with operable non-small cell lung cancer (NSCLC). NSCLC patients with enlarged mediastinal lymph nodes (short diameter, 7-20 mm), confirmed using contrast-enhanced CT, were examined using FDG-PET to detect metastases prior to surgery. The primary endpoint was the accuracy for concomitantly used CT and FDG-PET showing the additional effects of FDG, compared with CT alone. The secondary endpoints were the clinical impact of FDG-PET on therapeutic decisions and adverse reaction from FDG administration. The images were interpreted by investigators at each institution. Moreover, blinded readings were performed by an image interpretation committee independent of the institutions. The gold standard was the pathological diagnosis determined by surgery or biopsy after PET, and patients in whom a pathological diagnosis was not obtained were excluded from the analysis. Among 99 subjects, the results for 81 subjects eligible for analysis showed that the accuracy improved from 69.1% (56/81) for CT alone to 75.3% (61/81) for CT + PET (p = 0.404). These findings contributed to treatment decisions in 63.0% (51/81) of the cases, mainly with regard to the selection of the operative procedure. The results of the image interpretation committee showed that the accuracy improved from 64.2% (52/81) (95% CI 52.8-74.6) for CT to 75.3% (61/81) (95% CI 64.5-84.2) for CT + PET. The accuracy for 106 mediastinal lymph nodes improved significantly from 62.3% (66/106) (95% CI 52.3-71.5) for CT to 79.2% (84/106) (95% CI 70.3-86.5) for CT + PET (p < 0.05). We found that no serious adverse drug reactions appeared in any of the 99 patients who received FDG, except for transient mild outliers in the laboratory data for two patients. The addition of FDG-PET to contrast-enhanced CT imaging for the staging of NSCLC improved the diagnostic accuracy for mediastinal lymph node metastasis. FDG-PET improved the precision of the staging of NSCLC and contributed to the surgical decisions.


The Current Situation Regarding Long-Acting Insulin Analogues Including Biosimilars Among African, Asian, European, and South American Countries; Findings and Implications for the Future.
Background: Diabetes mellitus rates continue to rise, which coupled with increasing costs of associated complications has appreciably increased global expenditure in recent years. The risk of complications are enhanced by poor glycaemic control including hypoglycaemia. Long-acting insulin analogues were developed to reduce hypoglycaemia and improve adherence. Their considerably higher costs though have impacted their funding and use. Biosimilars can help reduce medicine costs. However, their introduction has been affected by a number of factors. These include the originator company dropping its price as well as promoting patented higher strength 300 IU/ml insulin glargine. There can also be concerns with different devices between the manufacturers. Objective: To assess current utilisation rates for insulins, especially long-acting insulin analogues, and the rationale for patterns seen, across multiple countries to inform strategies to enhance future utilisation of long-acting insulin analogue biosimilars to benefit all key stakeholders. Our approach: Multiple approaches including assessing the utilisation, expenditure and prices of insulins, including biosimilar insulin glargine, across multiple continents and countries. Results: There was considerable variation in the use of long-acting insulin analogues as a percentage of all insulins prescribed and dispensed across countries and continents. This ranged from limited use of long-acting insulin analogues among African countries compared to routine funding and use across Europe in view of their perceived benefits. Increasing use was also seen among Asian countries including Bangladesh and India for similar reasons. However, concerns with costs and value limited their use across Africa, Brazil and Pakistan. There was though limited use of biosimilar insulin glargine 100 IU/ml compared with other recent biosimilars especially among European countries and Korea. This was principally driven by small price differences in reality between the originator and biosimilars coupled with increasing use of the patented 300 IU/ml formulation. A number of activities were identified to enhance future biosimilar use. These included only reimbursing biosimilar long-acting insulin analogues, introducing prescribing targets and increasing competition among manufacturers including stimulating local production. Conclusions: There are concerns with the availability and use of insulin glargine biosimilars despite lower costs. This can be addressed by multiple activities.


ER stress induces epithelial differentiation in the mouse oesophagus.
Stress in the endoplasmic reticulum (ER) leads to activation of the unfolded protein response (UPR). Xbp1, a key component of the UPR has recently been linked to the risk of developing oesophageal squamous cell carcinoma, suggesting an important role for the UPR in the oesophageal epithelium. Here we examined the role of ER stress and the UPR in oesophageal epithelial homoeostasis. We examined the expression of components of the UPR in the oesophageal epithelium. We used a pharmacological approach and a genetic approach to examine the effects of ER stress in vivo in the mouse oesophagus. The oesophagus of these mice was examined using immunohistochemistry and real-time reverse transcription (RT)-PCR. Components of the UPR were heterogeneously expressed in the basal layer of the epithelium. Induction of ER stress by 24-h treatment with thapsigargin resulted in depletion of proliferating cells in the basal layer of the oesophagus and induced differentiation. We next activated the UPR by inducible deletion of the major ER chaperone Grp78 in Ah1Cre-Rosa26-LacZ-Grp78(-/-) mice in which mutant cells could be traced by expression of LacZ. In these mice LacZ-positive mutant cells in the basal layer lost their proliferative capacity, migrated towards the oesophageal lumen and were replaced by LacZ-negative non-mutant cells. We observed no apoptosis in mutant cells. These results show that ER stress induces epithelial differentiation in precursor cells in the oesophageal epithelium. This UPR induced differentiation may serve as a quality control mechanism that protects against oesophageal cancer development.


Pharmacologic suppression of JAK1/2 by JAK1/2 inhibitor AZD1480 potently inhibits IL-6-induced experimental prostate cancer metastases formation.
Metastatic prostate cancer is lethal and lacks effective strategies for prevention or treatment, requiring novel therapeutic approaches. Interleukin-6 (IL-6) is a cytokine that has been linked with prostate cancer pathogenesis by multiple studies. However, the direct functional roles of IL-6 in prostate cancer growth and progression have been unclear. In the present study, we show that IL-6 is produced in distant metastases of clinical prostate cancers. IL-6-activated signaling pathways in prostate cancer cells induced a robust 7-fold increase in metastases formation in nude mice. We further show that IL-6 promoted migratory prostate cancer cell phenotype, including increased prostate cancer cell migration, microtubule reorganization, and heterotypic adhesion of prostate cancer cells to endothelial cells. IL-6-driven metastasis was predominantly mediated by Stat3 and to lesser extent by ERK1/2. Most importantly, pharmacologic inhibition of Jak1/2 by AZD1480 suppressed IL-6-induced signaling, migratory prostate cancer cell phenotypes, and metastatic dissemination of prostate cancer in vivo in nude mice. In conclusion, we demonstrate that the cytokine IL-6 directly promotes prostate cancer metastasis in vitro and in vivo via Jak-Stat3 signaling pathway, and that IL-6-driven metastasis can be effectively suppressed by pharmacologic targeting of Jak1/2 using Jak1/2 inhibitor AZD1480. Our results therefore provide a strong rationale for further development of Jak1/2 inhibitors as therapy for metastatic prostate cancer.


Absorption of amino acids from the intestine and their net flux across the mesenteric- and portal-drained viscera of lambs.
Experiments were conducted to compare the rates of apparent absorption (disappearance) of individual essential amino acids (EAA) from the small intestine with their net fluxes across the mesenteric- (MDV) and portal- (PDV) drained viscera in sheep given a pelleted alfalfa diet at two levels of intake. Disappearances of individual EAA across the region of the small intestine drained by the mesenteric arcade (jejunum to ileum) were similar to those across the whole of the small intestine (duodenum to ileum). The net MDV flux of each EAA was similar to its rate of disappearance, but, with the exception of threonine on the low intake level, the net PDV flux was lower (P < .05). Increasing the intake of alfalfa from 800 to 1,200 g/d increased the rate of disappearance of individual EAA between the duodenum and ileum by .56 (range .43 to .65) and between the jejunum and ileum by .51 (range .45 to .60). The MDV and PDV blood flows increased by .35 and .39, respectively, and, with the exception of valine, net fluxes of individual EAA increased by .39 (range .20 to .50) across the MDV and by .44 (range .21 to .71) across the PDV. When net fluxes across the MDV and PDV were measured simultaneously, the ratio of PDV: MDV flux for each EAA was less than (P < .05) unity (mean .61, range .55 to .69), even though all MDV blood enters the PDV, contributing approximately .45 of the total portal flow. This observation suggests that, in regions of the tract not drained by the MDV, extraction of arterial blood EAA for tissue and secretory protein synthesis must exceed the release of protein degradation products into the venous drainage. The results are discussed in terms of endogenous protein secretions into the lumen of the tract anterior to the small intestine.


Melatonin reverses morphine tolerance by inhibiting microglia activation and HSP27 expression.
Melatonin has been reported to attenuate opioid tolerance. In this study, we explored the possible mechanism of melatonin in diminishing morphine tolerance. Two intrathecal (i.t.) catheters were implanted in male Wistar rats for drug delivery. One was linked to a mini-osmotic pump for morphine or saline infusion. On the seventh day, 50μg of melatonin or vehicle was injected through the other catheter instantly after discontinuation of morphine or saline infusion; 3h later, 15μg of morphine or saline was injected. The antinociceptive response was then measured using the tail-flick test every 30min for 120min. The results showed that chronic morphine infusion elicited antinociceptive tolerance and upregulated heat shock protein 27 (HSP27) expression in the dorsal horn of the rat spinal cord. Melatonin pretreatment partially restored morphine's antinociceptive effect in morphine-tolerant rats and reversed morphine-induced HSP27 upregulation. In addition, chronic morphine infusion induced microglial cell activation and was reversed by melatonin treatment. The present study provides evidence that melatonin, acting via inhibiting morphine-induced neuroinflammation, can be useful as a therapeutic adjuvant for patients under long-term opioid treatment for pain relief.


Disruption of MBD5 contributes to a spectrum of psychopathology and neurodevelopmental abnormalities.
Microdeletions of chromosomal region 2q23.1 that disrupt MBD5 (methyl-CpG-binding domain protein 5) contribute to a spectrum of neurodevelopmental phenotypes; however, the impact of this locus on human psychopathology has not been fully explored. To characterize the structural variation landscape of MBD5 disruptions and the associated human psychopathology, 22 individuals with genomic disruption of MBD5 (translocation, point mutation and deletion) were identified through whole-genome sequencing or cytogenomic microarray at 11 molecular diagnostic centers. The genomic impact ranged from a single base pair to 5.4 Mb. Parents were available for 11 cases, all of which confirmed that the rearrangement arose de novo. Phenotypes were largely indistinguishable between patients with full-segment 2q23.1 deletions and those with intragenic MBD5 rearrangements, including alterations confined entirely to the 5'-untranslated region, confirming the critical impact of non-coding sequence at this locus. We identified heterogeneous, multisystem pathogenic effects of MBD5 disruption and characterized the associated spectrum of psychopathology, including the novel finding of anxiety and bipolar disorder in multiple patients. Importantly, one of the unique features of the oldest known patient was behavioral regression. Analyses also revealed phenotypes that distinguish MBD5 disruptions from seven well-established syndromes with significant diagnostic overlap. This study demonstrates that haploinsufficiency of MBD5 causes diverse phenotypes, yields insight into the spectrum of resulting neurodevelopmental and behavioral psychopathology and provides clinical context for interpretation of MBD5 structural variations. Empirical evidence also indicates that disruption of non-coding MBD5 regulatory regions is sufficient for clinical manifestation, highlighting the limitations of exon-focused assessments. These results suggest an ongoing perturbation of neurological function throughout the lifespan, including risks for neurobehavioral regression.


Early outcomes from the CLASP IID trial roll-in cohort for prohibitive risk patients with degenerative mitral regurgitation.
We report the 30-day outcomes from the roll-in cohort of the CLASP IID trial, representing the first procedures performed by each site. The currently enrolling CLASP IID/IIF pivotal trial is a multicenter, prospective, randomized trial assessing the safety and effectiveness of the PASCAL transcatheter valve repair system in patients with clinically significant MR. The trial allows for up to three roll-in patients per site. Eligibility criteria were: DMR ≥3+, prohibitive surgical risk, and deemed suitable for transcatheter repair by the local heart team. Trial oversight included a central screening committee and echocardiographic core laboratory. The primary safety endpoint was a 30-day composite MAE: cardiovascular mortality, stroke, myocardial infarction (MI), new need for renal replacement therapy, severe bleeding, and non-elective mitral valve re-intervention, adjudicated by an independent clinical events committee. Thirty-day echocardiographic, functional, and quality of life outcomes were assessed. A total of 45 roll-in patients with mean age of 83 years and 69% in NYHA class III/IV were treated. Successful implantation was achieved in 100%. The 30-day composite MAE rate was 8.9% including one cardiovascular death (2.2%) due to severe bleeding from a hemorrhagic stroke, one MI, and no need for re-intervention. MR≤1+ was achieved in 73% and ≤2+ in 98% of patients. 89% of patients were in NYHA class I/II (p < .001) with improvements in 6MWD (30 m; p = .054) and KCCQ (17 points; p < .001). Early results representing sites with first experience with the PASCAL repair system showed favorable 30-day outcomes in patients with DMR≥3+ at prohibitive surgical risk.


Three-dimensional co-culture of primary human liver cells in bioreactors for in vitro drug studies: effects of the initial cell quality on the long-term maintenance of hepatocyte-specific functions.
In vitro culture models that employ human liver cells could be potent tools for predictive studies on drug toxicity and metabolism in the pharmaceutical industry. A bioreactor culture model was developed that permits the three-dimensional co-culture of liver cells under continuous medium perfusion with decentralised mass exchange and integral oxygenation. We tested the ability of the system to support the long-term maintenance and differentiation of primary human liver cells. The effects of the initial cell quality were investigated by comparing cultures from resected, non-preserved liver with cultures from liver graft tissue damaged by long-term preservation. In cultures originating from non-preserved liver, protein and urea synthesis, glucose metabolism, and cytochrome (CYP450) activities were stable over the 2-week culture period, with maximal activities at the end of the first week in culture. Enzyme induction led to increased 7-ethoxyresorufin O-deethylase activities of up to 20 times the basal value. In cultures from preservation-damaged liver, recovery of metabolic activities was detected during bioreactor culture. After two weeks, most biochemical parameters approached those of cultures from non-preserved human liver. Light microscopy demonstrated the three-dimensional reorganisation of hepatocytes and non-parenchymal cells in co-culture. Long-term maintenance, and even the regeneration of specific functional activities of human liver cells, can be achieved in the bioreactor. This could facilitate the introduction into the pharmaceutical industry of in vitro drug testing with primary human liver cells.


A novel scoring system for assessing Chiari malformation type I treatment outcomes.
Outcome assessment for the management of Chiari malformation type 1 is difficult because of the lack of a reliable and specific surgical outcome assessment scale. Such a scale could reliably correlate postoperative outcomes with preoperative symptoms. We developed a novel scoring system and applied it retrospectively to 146 patients treated at our institution in order to create and verify a simple and quantifiable assessment of Chiari outcomes. The Chicago Chiari Outcome Scale (CCOS) uses 4 postoperative outcome categories (pain, nonpain symptoms, functionality, and complications) graded 1 to 4 for a total possible score of 16. As a comparison with current Chiari outcome methodology, each patient was also placed into a gestalt outcome group of "improved," "unchanged," or "worse" (I/U/W). Patients were stratified by CCOS scores and by I/U/W group. Stratifying patients by total CCOS scores showed that patients who achieved CCOS scores between 13 and 16 were predominantly in the I/U/W improved group (n = 101, 69%); scores between 9 and 12 were predominantly I/U/W unchanged (n = 39, 27%), and scores between 4 and 8 were I/U/W worse (n = 6, 4%). Symptom subscore results provided insight into the specifics of the overall outcome in addition to the more quantitative nature of the 16-point scale. We describe a CCOS that assigns higher scores to patients judged improved by gestalt I/U/W ratings and lower scores to those who were unchanged or worse while defining outcome in 4 specific subcategories. As such, this CCOS should allow for a more unified and quantifiable outcome assessment after Chiari surgery.


Pattern of neurotoxicity of n-hexane, methyl n-butyl ketone, 2,5-hexanediol, and 2,5-hexanedione alone and in combination with O-ethyl O-4-nitrophenyl phenylphosphonothioate in hens.
This investigation was designed to study the neurotoxicity produced in hens by the aliphatic hexacarbons n-hexane, methyl n-butyl ketone (MnBK), 2,5-hexanediol (2,5-HDOH), and 2,5-hexanedione (2,5-HD) following daily dermal application of each chemical alone and in combination with O-ethyl O-4-nitrophenyl phenylphosphonothioate (EPN). Dermal application was carried out on the unprotected back of the neck. To assess whether the joint neurotoxic action of various chemicals is caused by the enhancement of absorption through the skin or by interaction at the molecular level, two additional experiments were performed. In the first experiment, EPN was dissolved in each of the aliphatic hydrocarbons prior to their topical application. In the second experiment, EPN was dissolved in acetone and applied at a different location from that of the aliphatic hexacarbons. Dermal application was carried out for 90 d followed by a 30-d observation period. The results show that hens treated with EPN developed severe ataxia followed by improvement during the observation period; n-hexane produced leg weakness with subsequent recovery, whereas the same dose of MnBK, 2,5-HDOH, or 2,5-HD produced clinical signs of neurotoxicity characterized by gross ataxia; concurrent dermal application of EPN with n-hexane or 2,5-HDOH at the same site or at different sites produced an additive neurotoxic action; simultaneous dermal application of EPN and MnBK at different sites resulted in an additive effect, whereas it caused potentiation when applied at the same site; and concurrent topical application of EPN and 2,5-HD produced a potentiating neurotoxic effect. While no histopathologic lesion was produced at the end of the observation period when any test chemical was applied alone, binary treatments of EPN and aliphatic hexacarbons resulted in histopathologic changes in some hens, with morphology and distribution characteristic of EPN neurotoxicity. The joint potentiating or additive action of aliphatic hexacarbons on EPN neurotoxicity was: 2,5-HD greater than MnBK greater than 2,5-HDOH greater than n-hexane. The mechanism of this joint action seems to be related both to enhancing skin absorption of EPN and/or its metabolic activation by n-hexane and its related chemicals.


Soft ionization of thermally evaporated hypergolic ionic liquid aerosols.
Isolated ion pairs of a conventional ionic liquid, 1-Ethyl-3-Methyl-Imidazolium Bis(trifluoromethylsulfonyl)imide ([Emim(+)][Tf(2)N(-)]), and a reactive hypergolic ionic liquid, 1-Butyl-3-Methyl-Imidazolium Dicyanamide ([Bmim(+)][Dca(-)]), are generated by vaporizing ionic liquid submicrometer aerosol particles for the first time; the vaporized species are investigated by dissociative ionization with tunable vacuum ultraviolet (VUV) light, exhibiting clear intact cations, Emim(+) and Bmim(+), presumably originating from intact ion pairs. Mass spectra of ion pair vapor from an effusive source of the hypergolic ionic liquid show substantial reactive decomposition due to the internal energy of the molecules emanating from the source. Photoionization efficiency curves in the near threshold ionization region of isolated ion pairs of [Emim(+)][Tf(2)N(-)] ionic liquid vapor are compared for an aerosol source and an effusive source, revealing changes in the appearance energy due to the amount of internal energy in the ion pairs. The aerosol source has a shift to higher threshold energy (∼0.3 eV), attributed to reduced internal energy of the isolated ion pairs. The method of ionic liquid submicrometer aerosol particle vaporization, for reactive ionic liquids such as hypergolic species, is a convenient, thermally "cooler" source of isolated intact ion pairs in the gas phase compared to effusive sources.


Use of Cannabis to Relieve Pain and Promote Sleep by Customers at an Adult Use Dispensary.
Medical cannabis patients consistently report using cannabis as a substitute for prescription medications; however, little is known about individuals accessing cannabis through adult-use markets. A survey at two retail stores was conducted in Colorado, United States. Between August 2016 and October 2016, store staff asked customers if they wanted to participate and, if so, provided an electronic survey link. All customers reporting medical certification were excluded. Of 1,000 adult-use only customer respondents, 65% reported taking cannabis to relieve pain and 74% reported taking cannabis to promote sleep. Among respondents taking cannabis for pain, 80% reported that it was very or extremely helpful, and most of those taking over-the-counter pain medications (82%) or opioid analgesics (88%) reported reducing or stopping use of those medications. Among respondents taking cannabis for sleep, 84% found it very or extremely helpful, and most of those taking over-the-counter (87%) or prescription sleep aids (83%) reported reducing or stopping use of those medications. De facto medical use of cannabis for symptom relief was common among adult-use dispensary customers and the majority reported that cannabis decreased their medication use. Adult use cannabis laws may broaden access to cannabis for the purpose of symptom relief.


Sectional Impressions and 'Every Other Tooth' Technique in FPD.
Gingival displacement procedure is an important component of fixed prosthodontics. It helps in delivery of the impression material subgingivally to record the finish line as well as the tooth contour apical to it, so as to produce restorations with closely adapted margins in harmony with the tooth anatomy and gingiva. While displacing gingiva with retraction cords, the health of the tissues must be maintained. In case of multiple anterior tooth preparations with close root proximity, placing the retraction cord simultaneously around all teeth will result in strangulation of interdental papillae. This can impair gingival health as well as cause black inter-dental triangles. To avoid this, 'every other tooth' technique of gingival displacement in combination with sectional impressions may be used to provide a better master cast in multiple tooth preparations. The present case report describes the use of this simple combination procedure for the restoration of lower six anterior teeth as a part of full mouth rehabilitation.


Long-term desensitization of nicotinic acetylcholine receptors is regulated via protein kinase A-mediated phosphorylation.
During prolonged application of transmitter, ligand-gated ion channels enter a nonconducting desensitized state. Studies on Torpedo electroplax nicotinic acetylcholine (ACh) receptors have shown that entry into the desensitized state is accelerated by protein kinase A-dependent (PKA) receptor phosphorylation. To examine the effects of phosphorylation on desensitization of muscle-type ACh receptors, we expressed the frog embryonic receptor type in Xenopus oocytes. Treatment of embryonic muscle ACh receptors with 8-Br cAMP had no measurable effect on the rate of entry into a desensitized state, but it greatly accelerated the recovery from desensitization. Three complementary approaches to reduce the levels of receptor phosphorylation provided additional evidence for a role of PKA-dependent phosphorylation in rescuing receptors from long-term desensitization. Inactivation of the endogenous PKA activity by coexpression of an inhibitor protein, treatment of receptors with phosphatase, and removal of phosphorylation sites by site-specific subunit mutation all resulted in slowed recovery. Our findings point to the existence of two distinct desensitized states: one requiring several seconds for full recovery and a second state from which recovery requires minutes. Receptors lacking PKA phosphorylation sites exhibit a pronounced increase in the slowly recovering component of desensitization, suggesting that receptor phosphorylation speeds overall recovery by reducing the entry into a deep desensitized state. This newly described effect of phosphorylation on ACh receptor function may serve as an important modulator of postsynaptic receptor sensitivity.


Production of multilineage hemopoietic growth-stimulating activities by a human melanoma cell line.
In this study, we report that conditioned medium (CM) from the human melanoma cell line Hs0294 contains growth-stimulating activity for human and murine hemopoietic progenitor cells of multiple lineages. Crude CM was assayed for hemopoietic progenitor cell growth-stimulating activity (HP-GSA) in plasma clot cultures of 10(5) C57B1/6J mouse bone marrow cells or 10(5) light-density, nonadherent human peripheral blood mononuclear cells. CM concentrations as low as 0.2% had demonstrable HP-GSA for murine and human hemopoietic progenitor cells, although peak activity for all hemopoietic progenitor cell types assayed (murine CFU-Mix, BFU-E, CFU-GM, and CFU-M and human BFU-E) was observed with 5%-10% Hs0294-CM. Growth-stimulating activity for human erythroid progenitors (BFU-E) was comparable to that produced by phytohemagglutinin-stimulated human mononuclear cell CM. Stimulation of murine hemopoietic progenitors by Hs0294-CM was comparable to the activity in pokeweed mitogen-stimulated mouse spleen cell CM. The Hs0294-CM did not contain detectable quantities of erythropoietin or interleukin 3. Human burst-promoting activity was stable through sequential freezing and thawing, heating to 50 degrees C for 1 h or 100 degrees C for 5 min, and incubation at pH 9.8 for 1 h, but lost 70% of its activity when incubated at pH 2.2 for 1 h. More than 95% of the HP-GSA was recovered in the 50%-80% fraction by (NH4)2SO4 precipitation. The HP-GSA had an estimated molecular weight of approximately 21,000. In conclusion, we have shown that a human melanoma cell line, Hs0294, produces multilineage HP-GSA.


Laboratory screening of thrombophilia. Evaluation of the diagnostic efficacy of a global test to detect congenital deficiencies of the protein C anticoagulant pathway.
Clinical laboratories are at present confronted with increasing demands for thrombophilia work-up, which may seriously overwhelm their capacity. Recently, methods able to investigate the protein C anticoagulant pathway globally have been proposed. In this study we investigated the reliability of one such method for its ability to detect patients with known defects of the pathway by testing plasmas from patients with the FVQ506 mutation, with congenital protein C, protein S or antithrombin deficiencies, and patients with previous history of thrombosis, but no identifiable defects. The results show that the new global test fulfils the requirements for congenital protein C deficiency and activated protein C resistance associated with the FVQ506 mutation, which account for more than half of the congenital defects found in thrombophilia. However, congenital protein S deficiency very often remains undetected by this test. Improvement of sensitivity toward this component of the protein C anticoagulant pathway would enroll the global test as a suitable candidate to explore the pathway. Since antithrombin, which also remains undetected by this test, is an additional important risk factor for venous thrombosis, devoting time and effort to developing global tests able to detect defects in both the antithrombin and protein C pathways is warranted.


Lynch syndrome (hereditary nonpolyposis colorectal cancer) diagnostics.
Preventive programs for individuals who have high lifetime risks of colorectal cancer may reduce disease morbidity and mortality. Thus, it is important to identify the factors that are associated with hereditary colorectal cancer and to monitor the effects of tailored surveillance. In particular, patients with Lynch syndrome, hereditary nonpolyposis colorectal cancer (HNPCC), have an increased risk to develop colorectal cancer at an early age. The syndrome is explained by germline mutations in DNA mismatch repair (MMR) genes, and there is a need for diagnostic tools to preselect patients for genetic testing to diagnose those with HNPCC. Patients (n = 112) from 285 families who were counseled between 1990 and 2005 at a clinic for patients at high risk for HNPCC were selected for screening to detect mutations in MMR genes MLH1, MSH2, MSH6, and PMS2 based on family history, microsatellite instability (MSI), and immunohistochemical analysis of MMR protein expression. Tumors were also screened for BRAF V600E mutations; patients with the mutation were considered as non-HNPCC. Among the 112 patients who were selected for screening, 69 had germline MMR mutations (58 pathogenic and 11 of unknown biologic relevance). Sixteen of the 69 mutations (23%) were missense mutations. Among patients with MSI-positive tumors, pathogenic MMR mutations were found in 38 of 43 (88%) of patients in families who met Amsterdam criteria and in 13 of 22 (59%) of patients in families who did not. Among patients with MSI-negative tumors, pathogenic MMR mutations were found in 5 of 17 (29%) of families meeting Amsterdam criteria and in 1 of 30 (3%) of non-Amsterdam families with one patient younger than age 50 years. In three patients with MSI-negative tumors who had pathogenic mutations in MLH1 or MSH6, immunohistochemistry showed loss of the mutated protein. Our findings suggest that missense MMR gene mutations are common in HNPCC and that germline MMR mutations are also found in patients with MSI-negative tumors.


Genetic complexity and quantitative trait loci mapping of yeast morphological traits.
Functional genomics relies on two essential parameters: the sensitivity of phenotypic measures and the power to detect genomic perturbations that cause phenotypic variations. In model organisms, two types of perturbations are widely used. Artificial mutations can be introduced in virtually any gene and allow the systematic analysis of gene function via mutants fitness. Alternatively, natural genetic variations can be associated to particular phenotypes via genetic mapping. However, the access to genome manipulation and breeding provided by model organisms is sometimes counterbalanced by phenotyping limitations. Here we investigated the natural genetic diversity of Saccharomyces cerevisiae cellular morphology using a very sensitive high-throughput imaging platform. We quantified 501 morphological parameters in over 50,000 yeast cells from a cross between two wild-type divergent backgrounds. Extensive morphological differences were found between these backgrounds. The genetic architecture of the traits was complex, with evidence of both epistasis and transgressive segregation. We mapped quantitative trait loci (QTL) for 67 traits and discovered 364 correlations between traits segregation and inheritance of gene expression levels. We validated one QTL by the replacement of a single base in the genome. This study illustrates the natural diversity and complexity of cellular traits among natural yeast strains and provides an ideal framework for a genetical genomics dissection of multiple traits. Our results did not overlap with results previously obtained from systematic deletion strains, showing that both approaches are necessary for the functional exploration of genomes.


Non-invasive evaluation of the equine gastrointestinal mucosal transcriptome.
Evaluating the health and function of the gastrointestinal tract can be challenging in all species, but is especially difficult in horses due to their size and length of the gastrointestinal (GI) tract. Isolation of mRNA of cells exfoliated from the GI mucosa into feces (i.e., the exfoliome) offers a novel means of non-invasively examining the gene expression profile of the GI mucosa. This approach has been utilized in people with colorectal cancer. Moreover, we have utilized this approach in a murine model of GI inflammation and demonstrated that the exfoliome reflects the tissue transcriptome. The ability of the equine exfoliome to provide non-invasive information regarding the health and function of the GI tract is not known. The objective of this study was to characterize the gene expression profile found in exfoliated intestinal epithelial cells from normal horses and compare the exfoliome data with the tissue mucosal transcriptome. Mucosal samples were collected from standardized locations along the GI tract (i.e. ileum, cecum, right dorsal colon, and rectum) from four healthy horses immediately following euthanasia. Voided feces were also collected. RNA isolation, library preparation, and RNA sequencing was performed on fecal and intestinal mucosal samples. Comparison of gene expression profiles from the tissue and exfoliome revealed correlation of gene expression. Moreover, the exfoliome contained reads representing the diverse array of cell types found in the GI mucosa suggesting the equine exfoliome serves as a non-invasive means of examining the global gene expression pattern of the equine GI tract.


Geometric alignment and chromatic calibration of serial radiographic images.
To develop software for automated registration and intensity calibration of serial dental radiographs for the analysis of longitudinal changes in bone density. Serial dental radiographs were acquired using a positioning device designed to minimize projection divergence. Each radiograph included an image of a standardized aluminium wedge. The radiographs were scanned on a flatbed scanner (AGFA Duo Scan) with a spatial resolution of 300 dpi, and pixel intensity coded in 16-bit grey scale. The intensity was calibrated using serial images of selected areas with defined thickness of the aluminium wedge. A robust B-splines multiresolution registration algorithm was implemented to overcome the acquisition misalignment. Radiographs, taken before and after periodontal therapy, were subtracted to assess bone density evolution. The intensity calibration decreased the maximum intensity variations between serial radiographs from 30+/-17% to 1+/-1% (mean+/-standard deviation), and improved the visual comparison between the radiographs. The registration stage allowed correcting the misalignment of the radiographs on the scanner screen and superimposing the radiography contents. The observed residual motion was about 0.02+/-0.01 mm. Very user-friendly software was developed. The manipulator needs to scan the radiographs only one time. The software performs all subsequent processing steps.


Examining the crossmodal consequences of viewing the Müller-Lyer illusion.
For many years, the Müller-Lyer illusion was studied as a purely "visual" illusion, but like many other optical illusions, the evidence now shows that it also occurs when stimuli are presented tactually. In the present study, we investigated whether the visual perception of the illusion would have any crossmodal consequences for haptic perception. The wings-in and wings-out parts of the Müller-Lyer illusion were placed end-to-end, sharing a central fin. This Brentano version of the illusion was presented visually on a screen in front of the participants, who had to compare the "felt" length of two sticks placed on the back of the screen, one behind either part of the illusion. Our results show that the presentation of the visual illusion modified the felt lengths of the sticks presented directly behind the illusion. In particular, the stick presented on the side of space perceived visually as being shorter (behind the wings-in part of the display) was perceived as longer, and vice versa for the stick mounted behind the space perceived visually as longer (behind the wings-out part of the display). These results highlight the crossmodal consequences of the visual perception of the Müller-Lyer illusion for the haptic perception of line length.


Stereoisomers of tetrahydrothiamin pyrophosphate, potent inhibitors of the pyruvate dehydrogenase multienzyme complex from Escherichia coli.
Tetrahydrothiamin pyrophosphate, an analogue of thiamin pyrophosphate (TPP) in which the thiazolium ring has been reduced to a thiazolidine ring, was prepared by borohydride reduction of TPP. It consists of four stereoisomers, comprising two diastereomers each of which is a racemic mixture, generated by the introduction of two chiral centers on the thiazolidine ring. The major and minor diastereomers were separated and inferred to be of the cis and trans configurations, respectively, from a study of the nuclear Overhauser effects in the 1H NMR spectrum of the simpler tetrahydrothiamin. Tetrahydro-TPP behaves as a mixture of potent inhibitors of the pyruvate decarboxylase (E1) component of the pyruvate dehydrogenase complex from Escherichia coli. The site of binding is probably the TPP-binding site on E1, and the Kd for each of the four stereoisomers was estimated. The cis isomers of tetrahydro-TPP bind more tightly than does TPP, whereas the trans isomers appear to bind with about the same Kd as TPP. Sodium borohydride caused a rapid inhibition of E1 activity in the presence of TPP, believed to be due to reaction of borohydride with enzyme-bound TPP. The experiments are consistent with an enhancement of the reactivity of the thiazole ring of TPP when bound to the catalytic site of E1, which could be due to polarization of the greater than +N=C bond near a hydrophobic or positively charged region of the protein. A spontaneous reactivation occurred after the initial inhibition by borohydride, attributable to a weakly binding inhibitor, not tetrahydro-TPP, being formed at the catalytic site.


Indication and timing of thyroid surgery for patients with hereditary medullary thyroid cancer syndromes.
Hereditary medullary thyroid cancer syndromes comprise familial medullary thyroid cancer (FMTC) and multiple endocrine neoplasia types 2A and 2B. Hereditary medullary thyroid cancers have an autosomal dominant pattern of inheritance and are caused by activating germline point mutations in the RET proto-oncogene. Evaluation of the onset, extent, and progression of hereditary medullary thyroid cancer associated with specific RET mutations has enabled clinicians to treat patients based on the level of risk associated with their specific mutation. Children identified by RET screening to be at risk for the development of medullary thyroid cancer can be treated with prophylactic thyroidectomy before developing the disease. This review covers the diagnosis, evaluation, timing of surgical management, and optimal follow-up of patients with hereditary medullary thyroid cancer syndromes.


The Opioid Hydra: Understanding Overdose Mortality Epidemics and Syndemics Across the Rural-Urban Continuum.
The rapid increase of fatal opioid overdoses over the past two decades is a major U.S. public health problem, especially in non-metropolitan communities. The crisis has transitioned from pharmaceuticals to illicit synthetic opioids and street mixtures, especially in urban areas. Using latent profile analysis, we classify n = 3,079 counties into distinct classes using CDC fatal overdose rates for specific opioids in 2002-2004, 2008-2012, and 2014-2016. We identify three distinct epidemics (prescription opioids, heroin, and prescription-synthetic opioid mixtures) and one syndemic involving all opioids. We find that prescription-related epidemic counties, whether rural or urban, have been "left behind" the rest of the nation. These communities are less populated and more remote, older and mostly white, have a history of drug abuse, and are former farm and factory communities that have been in decline since the 1990s. Overdoses in these places exemplify the "deaths of despair" narrative. By contrast, heroin and opioid syndemic counties tend to be more urban, connected to interstates, ethnically diverse, and in general more economically secure. The urban opioid crisis follows the path of previous drug epidemics, affecting a disadvantaged subpopulation that has been left behind rather than the entire community. County data on opioid epidemic class membership are provided.


Mutations in the Drosophila melanogaster gene three rows permit aspects of mitosis to continue in the absence of chromatid segregation.
We have cloned the three rows (thr) gene, by a combination of chromosome microdissection and P element tagging. We describe phenotypes of embryos homozygous for mutations at the thr locus. Maternal mRNA and protein appear to be sufficient to allow 14 rounds of mitosis in embryos homozygous for thr mutations. However, a small percentage of cells in syncytial blastoderm stage thr embryos sink into the interior of the embryo as if they have failed to divide properly. Following cellularisation all cells complete mitosis 14 normally. All cells become delayed at mitosis 15 with their chromosomes remaining aligned on the spindle in a metaphase-like configuration, even though both cyclins A and B have both been degraded. As cyclin B degradation occurs at the metaphase-anaphase transition, subsequent to the microtubule integrity checkpoint, the delay induced by mutations at the thr locus defines a later point in mitotic progression. Chromosomes in the cells of thr embryos do not undertake anaphase separation, but remain at the metaphase plate. Subsequently they decondense. A subset of nuclei go on to replicate their DNA but there is no further mitotic division.


Purinergic signalling: what is missing and needed next? The use of transgenic mice, crystallographic analysis and MicroRNA.
While ATP is recognized as an intracellular energy source for many biochemical reactions, it is now recognised it is also an important extracellular signalling molecule. ATP is involved in both physiological and pathological events in most cell types, and receptor subtypes have been cloned and characterised. An important goal of purinergic research today is to annotate the human genome with functional information regarding the role of genes for purinergic receptors, ectonucleotidases and transporters, in brain physiology and pathology. Insights into these roles have been gained also from studies of the various purinergic knockouts, and here we report on the generation of these purinergic receptor/ectonucleotidase-null mice. Recent X-ray structures of purinergic ligand-activated receptors provide promising templates to understand the molecular mechanism of receptor actions at the atomic level, and to deploy X-ray structures to be used for structure-based drug design. In the present work we also summarize recent findings about X-ray structures of ionotropic and metabotropic purinergic receptors and ectonucleotidases. A novel and prominent role as modulators of signal propagation in animal cells is played by microRNAs. By acting as genetic switches, they might become stringent regulators of the variety of cellular responses triggered by the dynamic interactions between purinergic receptors, nucleotides/nucleosides, transporters and ectonucleotidases. In this review we highlight data on the regulation of purinergic mechanisms by microRNAs. Finally, we would like to illustrate what information is still missing or needed for the acquisition of a more complete knowledge of purinergic signalling.


Recovery of Palm Oil and Valuable Material from Oil Palm Empty Fruit Bunch by Sub-critical Water.
Oil palm empty fruit bunch (EFB) is one of the solid wastes produced in huge volume by palm oil mill. Whilst it still contains valuable oil, approximately 22.6 million tons is generated annually and treated as solid waste. In this work, sub-critical water (sub-cw) was used to extract oil, sugar and tar from spikelet of EFB. The spikelet was treated with sub-cw between 180-280°C and a reaction time of 2 and 5 minutes. The highest yield of oil was 0.075 g-oil/g-dry EFB, obtained at 240°C and reaction time of 5 minutes. Astonishingly, oil that was extracted through this method was 84.5% of that obtained through Soxhlet method using hexane. Yield of oil extracted was strongly affected by the reaction temperature and time. Higher reaction temperature induces the dielectric constant of water towards the non-polar properties of solvent; thus increases the oil extraction capability. Meanwhile, the highest yield of sugar was 0.20 g-sugar/g-dry EFB obtained at 220°C. At this temperature, the ion product of water is high enough to enable maximum sub-critical water hydrolysis reaction. This study showed that oil and other valuable material can be recovered using water at sub-critical condition, and most attractive without the use of harmful organic solvent.


Interactive conference voting. The OMERACT II Committee. Outcome Measures in Rheumatoid Arthritis Clinical Trial Conference.
We describe and analyze opinion polling results from interactive voting procedures undertaken before and after presentations during the Outcome Measures in Rheumatoid Arthritis Clinical Trials Conference (OMERACT II) in Ottawa, Canada, June 30-July 2, 1994. The scoring procedure was a matched voting design; when a participant used the same keypad at the beginning and end of voting, change within a participant could be estimated. Participants, experienced in the rheumatic diseases included clinicians, researchers, methodologists, regulators, and representatives of the pharmaceutical industry. Patients under consideration were those with any rheumatic diseases. Questions were constructed to evaluate the change in voting behavior expected from the content of the presentation. Statistically significant and substantively important changes were evident in most questions.


IgA cross-reactivity between a nuclear autoantigen and wheat proteins suggests molecular mimicry as a possible pathomechanism in celiac disease.
Celiac disease patients display IgA antibody reactivity to wheat as well as to human proteins. We used serum IgA from celiac patients and, for control purposes, from patients with Crohn's disease, ulcerative colitis and from healthy individuals to identify celiac disease-specific IgA autoantigens in nitrocellulose-blotted extracts from various human cell types (epithelial, endothelial, intestinal cells, fibroblasts). The pattern, recognition intensity and time course of IgA autoreactivity was monitored using serial serum samples obtained from celiac children before and under gluten-free diet. By immunoblot inhibition and subcellular (cytosolic, nuclear) cell fractionation we identified a 55 kDa nuclear autoantigen expressed in intestinal, endothelial cells and in fibroblasts which was recognized by IgA antibodies of approximately half of the celiac disease patients and cross-reacted with wheat proteins. IgA reactivity to the 55 kDa autoantigen disappeared during gluten-free diet and was inhibited after pre-absorption of sera with wheat proteins but not with tissue transglutaminase, previously reported as the unique celiac disease-specific autoantigen. In conclusion, we defined a novel 55 kDa celiac disease-specific nuclear IgA autoantigen which shares epitopes with wheat proteins and which is different from tissue transglutaminase and calreticulin. Although the newly defined autoantigen was recognized much less frequently than tissue transglutaminase, our data suggest molecular mimicry between wheat and human proteins as a possible pathomechanism for the induction and/or maintenance of mucosal tissue damage in celiac disease.


Influence of attended repetition trials on negative priming in younger and older adults.
A lengthened response time when a distractor becomes a target, called negative priming, is an undisputed phenomenon in selective attention, yet just what the underlying mechanism responsible for negative priming is has not been resolved. In this study, the proportion of attended repetition trials was manipulated in order to test the predictions of three theories that have been proposed for explaining spatial negative priming: distractor suppression (e.g., Tipper, 1985), episodic memory retrieval (e.g., Neill, Valdes, & Terry, 1995), and novelty bias (e.g., Milliken, Tipper, Houghton, & Lupiáñez, 2000). The results supported the proposal that a novelty bias, which is flexible and can be overridden, is the primary mechanism responsible for priming in spatial tasks. Memory retrieval obscured the novelty bias for target processing, was more selective in older adults, and did not affect distractor processing. Novelty bias and distractor suppression may share the same inhibitory attentional mechanism.


MOC-PS(SM) CME article: self-assessment and performance in practice: the carpal tunnel.
After studying the article, the participant should be able to: 1. Conduct an appropriate history and physical examination for a patient suspected of having carpal tunnel syndrome. 2. Understand the role of provocative and other diagnostic tests pertinent to the diagnosis of carpal tunnel syndrome. 3. Understand the goals of the surgical treatment of carpal tunnel syndrome and how to obtain these. 4. Appreciate the common complications of carpal tunnel surgery and their management. The purpose of this article is to review important aspects of the history, physical examination, diagnosis, and management of carpal tunnel syndrome. Associated diseases, predisposing factors, and prognostic features are explored. The significance of diagnostic studies and the variety of anesthetic techniques with which to perform the surgery are reviewed. Evidence regarding the different surgical approaches, such as the open versus the endoscopic, is examined. Postoperative care issues such as therapy and splinting are examined. Finally, complications of carpal tunnel surgery and their management are outlined.


Immunomodulatory properties of Brucella melitensis lipopolysaccharide determinants on mouse dendritic cells in vitro and in vivo.
The lipopolysaccharide (LPS) is a major virulence factor of Brucella, a facultative intracellular pathogenic Gram-negative bacterium. Brucella LPS exhibits a low toxicity and its atypical structure was postulated to delay the host immune response, favouring the establishment of chronic disease. Here we carried out an in-depth in vitro and in vivo characterisation of the immunomodulatory effects of Brucella LPS on different dendritic cell (DC) subpopulations. By using LPSs from bacteria that share some of Brucella LPS structural features, we demonstrated that the core component of B. melitensis wild-type (Bm-wt) LPS accounts for the low activation potential of Brucella LPS in mouse GM-CSF-derived (GM-) DCs. Contrary to the accepted dogma considering Brucella LPS a poor TLR4 agonist and DC activator, Bm-wt LPS selectively induced expression of surface activation markers and cytokine secretion from Flt3-Ligand-derived (FL-) DCs in a TLR4-dependent manner. It also primed in vitro T cell proliferation by FL-DCs. In contrast, modified LPS with a defective core purified from Brucella carrying a mutated wadC gene (Bm-wadC), efficiently potentiated mouse and human DC activation and T cell proliferation in vitro. In vivo, Bm-wt LPS promoted scant activation of splenic DC subsets and limited recruitment of monocyte- DC like cells in the spleen, conversely to Bm-wadC LPS. Bm-wadC live bacteria drove high cytokine secretion levels in sera of infected mice. Altogether, these results illustrate the immunomodulatory properties of Brucella LPS and the enhanced DC activation ability of the wadC mutation with potential for vaccine development targeting Brucella core LPS structure.


Expression of cholecystokinin A receptors in neurons innervating the rat stomach and intestine.
Two distinct receptors, cholecystokinin (CCK)-A and CCK-B, mediate CCK effects in the digestive system. The aim of this study was to elucidate the cellular sites of expression of CCK-A receptor in the rat stomach and small intestine. We developed and characterized an antibody to the N-terminal region (LDQPQPSKEWQSA) of rat CCK-A receptor and used it for localization studies with immunohistochemistry. Specificity of the antiserum was demonstrated by (1) detection of a broad band at 85-95 kilodaltons in Western blots of membranes from CCK-A receptor CHO-transfected cells; (2) cell surface staining of CCK-A receptor-transfected cells, (3) translocation of CCK-A receptor immunostaining in CCK-A receptor-transfected cells after exposure to CCK; and (4) abolition of tissue immunostaining by preadsorbtion of the antibody with the peptide used for immunization. CCK-A receptor immunoreactivity was localized to myenteric neurons and to fibers in the muscle and mucosa. In the stomach, myenteric neurons and mucosal fibers were abundant. Many CCK-A receptor myenteric neurons contained the inhibitory transmitter vasoactive intestinal polypeptide, and some were immunoreactive for the excitatory transmitter substance P. Subdiaphragmatic vagotomy reduced the density of CCK-A receptor fibers in the gastric mucosa by approximately 50%, whereas celiac/superior mesenteric ganglionectomy had no detectable effect on fiber density. CCK-A receptor is expressed in functionally distinct neurons of the gastrointestinal tract. CCK-A receptor may mediate reflexes stimulated by CCK through the release of other transmitters from neurons bearing the receptor.


Elevated trefoil factor 3 plasma levels in critically ill patients with abdominal sepsis or non-infectious abdominal illness.
Trefoil factor 3 (TFF3) is a small peptide secreted mainly by goblet cells in the gut, where it plays a key role in gastrointestinal defence and repair. Plasma TFF3 has been reported as a biomarker of intestinal injury and as such it has been evaluated as a marker of disease activity in colitis. Impaired gut barrier function has been postulated as the "motor" of critical illness. We here sought to determine the temporal dynamics of plasma TFF3 in adult patients admitted to intensive care unit with abdominal sepsis or after major abdominal surgery for a non-infectious condition (post-op GI patients). TFF3 was measured in plasma obtained from 143 patients with abdominal sepsis and 98 post-op GI patients on admission to the intensive care (day 0) and at days 2 and 4 thereafter. Abdominal sepsis patients showed sustained elevated plasma TFF3 levels from day 0 to 4 relative to healthy control values, while in post-op GI patients admission TFF3 levels were not increased, only rising at day 2 and 4. In both patient groups, the presence of shock was associated with higher TFF3 levels. Moreover, patients with 3 or more organs failing had higher plasma TFF3 concentrations. While plasma TFF3 was higher in sepsis patients who did not survive until day 30, TFF3 levels were not independently associated with 30-day mortality in a Cox regression analysis. These results could support the theory that intestinal injury contributes to the pathogenesis of critical illness. Future studies are needed to elucidate whether the proposed gut dysfunction precedes or supersedes organ dysfunction in time.


Multisite monitoring of choline using biosensor microprobe arrays in combination with CMOS circuitry.
A miniature device enabling parallel in vivo detection of the neurotransmitter choline in multiple brain regions of freely behaving rodents is presented. This is achieved by combining a biosensor microprobe array with a custom-developed CMOS chip. Each silicon microprobe comprises multiple platinum electrodes that are coated with an enzymatic membrane and a permselective layer for selective detection of choline. The biosensors, based on the principle of amperometric detection, exhibit a sensitivity of 157±35 µA mM(-1) cm(-2), a limit of detection of below 1 µM, and a response time in the range of 1 s. With on-chip digitalization and multiplexing, parallel recordings can be performed at a high signal-to-noise ratio with minimal space requirements and with substantial reduction of external signal interference. The layout of the integrated circuitry allows for versatile configuration of the current range and can, therefore, also be used for functionalization of the electrodes before use. The result is a compact, highly integrated system, very convenient for on-site measurements.


[Cavinton in the treatment of ischemic cerebral stroke. Clinical and computerized-tomographic evaluation].
Intravenous Cavinton (ethyl ester of apovincaminic acid) was given to 27 patients with acute ischaemic stroke, while 30 patients were given aminophylline twice daily during 10 days. The results of this treatment was compared on the basis of clinical manifestations and changes in computerized tomograms (CT) before and after the treatment. It was found that Cavinton gave similar results as aminophylline and other vasoactive preparations studied by other authors. In the group of severe strokes better results were observed after Cavinton, --out of 14 patients 2 died and 12 improved. In the aminophylline group 5 out of 10 patients died, and in 2 no improvement was observed. Clinical improvement was not correlated with CT improvement, while a significant correlation was observed between the state of the patient during admission and the size of the focus in CT. Cavinton had a significantly greater effect on the regression of CT changes.


Effect of walnut consumption on markers of blood glucose control: a systematic review and meta-analysis.
Type 2 diabetes mellitus is a chronic disease increasing in global prevalence. Although habitual consumption of walnuts is associated with reduced risk of CVD, there is inconsistent evidence for the impact of walnut consumption on markers of glycaemic control. This systematic review and meta-analysis aimed to examine the effect of walnut consumption on markers of blood glucose control. A systematic search of Medline, PubMed, CINAHL and Cochrane databases (to 2 March 2019) was conducted. Inclusion criteria were randomised controlled trials conducted with adults which assessed the effect of walnut consumption on fasting blood glucose and insulin, glycated Hb and homeostatic model assessment of insulin resistance. Random effects meta-analyses were conducted to assess the weighted mean differences (WMD) for each outcome. Risk of bias in studies was assessed using the Cochrane Risk of Bias tool 2.0. Sixteen studies providing eighteen effect sizes were included in the review. Consumption of walnuts did not result in significant changes in fasting blood glucose levels (WMD: 0·331 mg/dl; 95 % CI -0·817, 1·479) or other outcome measures. Studies were determined to have either 'some concerns' or be at 'high risk' of bias. There was no evidence of an effect of walnut consumption on markers of blood glucose control. These findings suggest that the known favourable effects of walnut intake on CVD are not mediated via improvements in glycaemic control. Given the high risk of bias observed in the current evidence base, there is a need for further high-quality randomised controlled trials.


Establishment of cell lines using a doxycycline-inducible gene expression system to regulate expression of hepatitis B virus X protein.
The hepatitis B virus (HBV) X gene plays an important role in HBV-associated pathogenesis, especially hepatocarcinogenesis. Establishment of a stable and regulable HBx expression system will allow study of the function of this gene. Here, we describe the development of a doxycycline-inducible recombinant plasmid (pBPSTR3-FlagX) with the full-length HBV X gene and all components of the tetracycline-on ("Tet-on") gene expression system. This vector exhibited dose-dependent doxycycline-dependent induction of the Flag-HBx protein in HepG2 and Hep3B cells. We also observed dose-dependent doxycycline transactivation of HBx in HepG2 cells. After transfecting HepG2 cells with the pBPSTR3-FlagX plasmid, we isolated five puromycin-resistant cell clones with stable HBx expression, two of which exhibited stable and tight control of HBx expression by doxycycline. This new system has great potential for functional studies of the HBV X gene.


Ferritin and LDL-cholesterol as biomarkers of fat-free mass loss in ALS.
The availability of longitudinal clinical and biological data led us to wonder whether these parameters could be used to predict disturbances in body composition during ALS progression. Bioelectrical impedance analysis (BIA), as well as clinical and biological parameters (blood lipids and ferritin), were collected one year after diagnosis in ALS patients. The correlations were evaluated by the Spearman test. Performances to predict the evolution of BIA parameters during ALS evolution were evaluated by ROC analysis. Forty-two ALS patients were enrolled. Variations in FFM over one year were correlated to the variations in LDL-cholesterol (r = 0.53, p = 0.002) and ferritin (r = -0.58, p = 0.0002). To predict FFM loss, an increase in ferritin over 9 µg/L had a sensitivity of 90.0% and a specificity of 80.0% (p < 0.0001). Ferritine evolution would allow to easily follow the FFM without BIA during ALS. In addition, an adapted nutritional treatment based on this biological parameter might slow down ALS progression.


Regulation of iron homeostasis mediated by the heme-binding protein Dap1 (damage resistance protein 1) via the P450 protein Erg11/Cyp51.
Fungal infections arise frequently in immunocompromised patients, and sterol synthesis is a primary pathway targeted by antifungal drugs. In particular, the P450 protein Erg11/Cyp51 catalyzes a critical step in ergosterol synthesis, and the azole class of antifungal drugs inhibits Erg11. Dap1 is a heme-binding protein related to cytochrome b5 that activates Erg11, so that cells lacking Dap1 accumulate the Erg11 substrate and are hypersensitive to Erg11 inhibitors. Heme binding by Dap1 is crucial for its function, and point mutants in its heme-binding domain render Dap1 inactive for sterol biosynthesis and DNA damage resistance. Like Dap1, the human homologue, PGRMC1/Hpr6, also regulates sterol synthesis and DNA damage resistance. In the present study, we demonstrate that the Dap1 heme-1 domain is required for growth under conditions of low iron availability. Loss of Dap1 is suppressed by elevated levels of Erg11 but not by increased heme biosynthesis. Dap1 localizes to punctate cytoplasmic structures that co-fractionate with endosomes, and Dap1 contributes to the integrity of the vacuole. The results suggest that Saccharomyces cerevisiae Dap1 stimulates a P450-catalyzed step in sterol synthesis via a distinct localization from its homologues in Schizosaccharomyces pombe and mammals and that this function regulates iron metabolism.


The effect of mucolytic agents on gene transfer across a CF sputum barrier in vitro.
Trials of gene transfer for cystic fibrosis (CF) are currently underway. However, direct application to the airways may be impeded by the presence of airway secretions. We have therefore assessed the effect of CF sputum on the expression of the reporter gene beta-galactosidase complexed with the cationic liposome DC-Chol/DOPE in a number of cell lines in vitro. Transfection was markedly inhibited in the presence of sputum; the effect was concentration dependent and was only partially ameliorated by removal of sputum with phosphate-buffered saline (PBS) washing before gene transfer. However, treatment of the sputum-covered cells with recombinant human DNase (rhDNase, 50 micrograms/ml) but not with N-acetylcysteine, Nacystelyn, lysine (all 20 mM) or recombinant alginase (0.5 U/ml) significantly (P < 0.005) improved gene transfer. Adenovirus-mediated gene transfer efficiency in the presence of sputum was similarly inhibited, and again, treatment with rhDNase before transfection significantly improved gene transfer (P < 0.005). Transfection of Cos 7 cells in the presence of exogenous genomic DNA alone demonstrated similar inhibition to that observed with sputum and was also ameliorated by pre-treatment of DNA-covered cells with rhDNase. In a separate series of experiments performed in the absence of added sputum or genomic DNA, increasing concentrations of rhDNase resulted in a concentration-related decline in transfection efficiency. However, even at the highest concentration (500 micrograms/ml of rhDNase), transfection efficiency remained more than 50% of control. Thus, pre-treatment of CF airways with rhDNase may be appropriate before liposome or adenovirus-mediated gene therapy.


Genetic delineation between and within the widespread coccolithophore morpho-species Emiliania huxleyi and Gephyrocapsa oceanica (Haptophyta).
Emiliania huxleyi and Gephyrocapsa oceanica are abundant coccolithophore morpho-species that play key roles in ocean carbon cycling due to their importance as both primary producers and cal-cifiers. Global change processes such as ocean acidification impact these key calcifying species. The physiology of E. huxleyi, a developing model species, has been widely studied, but its genetic delineation from G. oceanica remains unclear due to a lack of resolution in classical genetic markers. Using nuclear (18S rDNA and 28S rDNA), mitochondrial (cox1, cox2, cox3, rpl16, and dam), and plastidial (16S rDNA, rbcL, tufA, and petA) DNA markers from 99 E. huxleyi and 44 G. oceanica strains, we conducted a multigene/multistrain survey to compare the suitability of different markers for resolving phylogenetic patterns within and between these two morpho-species. The nuclear genes tested did not provide sufficient resolution to discriminate between the two morpho-species that diverged only 291Kya. Typical patterns of incomplete lineage sorting were generated in phylogenetic analyses using plastidial genes. In contrast, full morpho-species delineation was achieved with mitochondrial markers and common intra-morpho-species phylogenetic patterns were observed despite differing rates of DNA substitution. Mitochondrial genes are thus promising barcodes for distinguishing these coccolithophore morpho-species, in particular in the context of environmental monitoring.


Modeling behavioral and neuronal symptoms of Alzheimer's disease in mice: a role for intraneuronal amyloid.
The amyloid Abeta-peptide (Abeta) is suspected to play a critical role in the cascade leading to AD as the pathogen that causes neuronal and synaptic dysfunction and, eventually, cell death. Therefore, it has been the subject of a huge number of clinical and basic research studies on this disease. Abeta is typically found aggregated in extracellular amyloid plaques that occur in specific brain regions enriched in nAChRs in Alzheimer's disease (AD) and Down syndrome (DS) brains. Advances in the genetics of its familiar and sporadic forms, together with those in gene transfer technology, have provided valuable animal models that complement the traditional cholinergic approaches, although modeling the neuronal and behavioral deficits of AD in these models has been challenging. More recently, emerging evidence indicates that intraneuronal accumulation of Abeta may also contribute to the cascade of neurodegenerative events and strongly suggest that it is an early, pathological biomarker for the onset of AD and associated cognitive and other behavioral deficits. The present review covers these studies in humans, in in vitro and in transgenic models, also providing more evidence that adult 3xTg-AD mice harboring PS1M146V, APPSwe, tauP301L transgenes, and mimicking many critical hallmarks of AD, show cognitive deficits and other behavioral alterations at ages when overt neuropathology is not yet observed, but when intraneuronal Abeta, synaptic and cholinergic deficits can already be described.


Comparison of the incidence, nature and cause of injuries sustained on grass and new generation artificial turf by male and female football players. Part 1: match injuries.
To compare the incidence, nature, severity and cause of match injuries sustained on grass and new generation artificial turf by male and female footballers. The National Collegiate Athletic Association Injury Surveillance System was used for a two-season (August to December) prospective study of American college and university football teams (2005 season: men 52 teams, women 64 teams; 2006 season: men 54 teams, women 72 teams). Injury definitions and recording procedures were compliant with the international consensus statement for epidemiological studies of injuries in football. Athletic trainers recorded details of the playing surface and the location, diagnosis, severity and cause of all match injuries. The number of days lost from training and match play was used to define the severity of an injury. Match exposures (player hours) were recorded on a team basis. The overall incidence of match injuries for men was 25.43 injuries/1000 player hours on artificial turf and 23.92 on grass (incidence ratio 1.06; p = 0.46) and for women was 19.15 injuries/1000 player hours on artificial turf and 21.79 on grass (incidence ratio = 0.88; p = 0.16). For men, the mean severity of non-season ending injuries was 7.1 days (median 5) on artificial turf and 8.4 days (median 5) on grass and, for women, 11.2 days (median 5) on artificial turf and 8.9 days (median 5) on grass. Joint (non-bone)/ligament/cartilage and contusion injuries to the lower limbs were the most common general categories of match injury on artificial turf and grass for both male and female players. Most injuries were acute (men: artificial turf 24.60, grass 22.91; p = 0.40; women: artificial turf 18.29, grass 20.64; p = 0.21) and resulted from player-to-player contact (men: artificial turf 14.73, grass 13.34; p = 0.37; women: artificial turf 10.72; grass 11.68; p = 0.50). There were no major differences in the incidence, severity, nature or cause of match injuries sustained on new generation artificial turf and grass by either male or female players.


Effect of P2X4 and P2X7 receptor antagonism on the pressure diuresis relationship in rats.
Reduced glomerular filtration, hypertension and renal microvascular injury are hallmarks of chronic kidney disease, which has a global prevalence of ~10%. We have shown previously that the Fischer (F344) rat has lower GFR than the Lewis rat, and is more susceptible to renal injury induced by hypertension. In the early stages this injury is limited to the pre-glomerular vasculature. We hypothesized that poor renal hemodynamic function and vulnerability to vascular injury are causally linked and genetically determined. In the present study, normotensive F344 rats had a blunted pressure diuresis relationship, compared with Lewis rats. A kidney microarray was then interrogated using the Endeavour enrichment tool to rank candidate genes for impaired blood pressure control. Two novel candidate genes, P2rx7 and P2rx4, were identified, having a 7- and 3- fold increased expression in F344 rats. Immunohistochemistry localized P2X4 and P2X7 receptor expression to the endothelium of the pre-glomerular vasculature. Expression of both receptors was also found in the renal tubule; however there was no difference in expression profile between strains. Brilliant Blue G (BBG), a relatively selective P2X7 antagonist suitable for use in vivo, was administered to both rat strains. In Lewis rats, BBG had no effect on blood pressure, but increased renal vascular resistance, consistent with inhibition of some basal vasodilatory tone. In F344 rats BBG caused a significant reduction in blood pressure and a decrease in renal vascular resistance, suggesting that P2X7 receptor activation may enhance vasoconstrictor tone in this rat strain. BBG also reduced the pressure diuresis threshold in F344 rats, but did not alter its slope. These preliminary findings suggest a physiological and potential pathophysiological role for P2X7 in controlling renal and/or systemic vascular function, which could in turn affect susceptibility to hypertension-related kidney damage.


Extrapleural pneumonectomy: still indicated?
The optimal treatment of malignant pleural mesothelioma (MPM) has not yet been established and is still under investigation. Surgery is one of the pillars in the multimodality approach with the purpose of removing as much as visible tumor as possible and to relieve symptoms. To date, two major surgical procedures are available for removal or debulking of MPM that is considered to be resectable: [extended (e)] pleurectomy/decortication (P/D) and extrapleural pneumonectomy (EPP). Historically, EPP was regarded as the only way to achieve a macroscopic complete resection. However, in the last years, there is a shift in literature towards (e)P/D as the preferred surgical procedure whenever possible as several retrospective studies and meta-analyses showed a similar or lower long-term survival and higher perioperative mortality and postoperative morbidity in patients who been treated with EPP. On the other hand, no randomized-controlled trials regarding surgical treatment with (e)P/D or EPP exist and therefore level A evidence favoring one surgical procedure is lacking. In this review we provide a nuanced and well-considered answer to the question whether EPP is still indicated in the surgical treatment of MPM.


Diversity and abundance of aromatic catabolic genes in lake sediments in response to temperature change.
The abundance and composition of genes involved in the catabolism of aromatic compounds provide important information on the biodegradation potential of organic pollutants and naturally occurring compounds in the environment. We studied catechol 2, 3 dioxygenase (C23O) and benzylsuccinate synthase (bssA) genes coding for key enzymes of aerobic and anaerobic degradation of aromatic compounds in experimental incubations with sediments from two contrasting lakes; humic lake Svarttjärn and eutrophic Vallentunasjön, respectively. Sediment cores from both lakes were incubated continuously for 5 months at constant temperatures ranging from 1.0 to 21.0 °C. The difference in C23O gene composition of the sediment analyzed at the end of the experiment was larger between lakes, than among temperature treatments within each lake. The abundance of C23O gene copies and measured respiration was positively correlated with temperature in Vallentunasjön, whereas putative C23O genes were present in lower concentrations in Svarttjärn sediments. Putative bssA genes were only detected in Svarttjärn. For both lakes, the two catabolic genes were most abundant in the surface sediment. The results emphasize the important role of temperature and nutrient availability in controlling the functional potential of sediment microorganisms and reveal differences between systems with contrasting trophic status. A better understanding of catabolic pathways and enzymes will enable more accurate forecasting of the functional properties of ecosystems under various scenarios of environmental change.


Identifying Core Components of EPA Implementation: A Path to Knowing if a Complex Intervention Is Being Implemented as Intended.
Competency-based assessment, using entrustable professional activities (EPAs), is rapidly being implemented worldwide without sufficient agreement on the essential elements of EPA-based assessment. The rapidity of implementation has left little time to understand what works in what circumstances and why or why not. The result is the attempted execution of a complex service intervention without a shared mental model for features needed to remain true to implementing an EPA assessment framework as intended. The purpose of this study was to identify the essential core components necessary to maintain integrity in the implementation of this intended intervention. A formal consensus-building technique, the Delphi process, was used to identify core components for implementing an EPA-based assessment framework. Twelve EPA experts from the United States, Canada, and the Netherlands participated in this process in February and March 2020. In each Delphi round, participants rated possible core components on a scale from 1 to 6, with 1 reflecting the worst fit and 6 the best fit for EPA-based assessment implementation. Predetermined automatic inclusion and exclusion criteria for candidate core components were set at ≥ 80% of participants assigning a value of 5 or 6 and ≥ 80% assigning a value of 1 or 2, respectively. After 3 rounds, participants prioritized 10 of 19 candidate core components for inclusion: performance prediction, shared local mental model, workplace assessment, high-stakes entrustment decisions, outcomes based, value of the collective, informed clinical competency committee members, construct alignment, qualitative data, and entrustment decision consequences. The study closed after 3 rounds on the basis of the rankings and comments. Using the core components identified in this study advances efforts to implement an EPA assessment framework intervention as intended, which mitigates the likelihood of making an incorrect judgment that the intervention demonstrates negative results.


Outcome after the single-stage, nonfenestrated Fontan procedure.
A completed Fontan circulation is the goal in the management of patients with single-ventricle physiology. To achieve this end, a two-stage rather than a single-stage approach is carried out routinely at many centers. Some groups have advocated baffle fenestration for virtually all patients to minimize post-Fontan complications. Other centers perform single-stage Fontan operations and do not fenestrate. Thus controversies have arisen regarding the indications for the staged procedure versus single stage and for fenestration versus no fenestration. The preoperative risk factors and postoperative course were characterized in 61 consecutive patients (median age, 3.3 years) undergoing a single-stage, nonfenestrated Fontan. The patients were followed for 3.5+/-1.9 years. The relationship between preoperative risk factors and mortality and morbidity was assessed. Preoperative risk factors assessed included age <2 years (n=18), branch pulmonary artery stenosis (n=20), elevated mean pulmonary artery pressure >15 mm Hg (n=16), atrioventricular valve regurgitation (n=5), and decreased ventricular function (n=2). Total caval pulmonary anastomosis was performed in 53 patients. Additional surgery was required at the time of the Fontan in 25 patients (41%). The median duration of mechanical ventilation was 1 day; median chest tube drainage was 5.5 days (range, 1 to 35). Oxygen saturation rose significantly postoperatively, from 83% to 95%. Early mortality was 4.9%; one patient died from pacemaker failure 9 months postoperatively, and one patient underwent successful heart transplant 4 months post-Fontan. One- and 5-year actuarial survival was 93%. No preoperative risk factor was associated with a failed Fontan or significant effusions. A single-stage, nonfenestrated Fontan was performed in a large group of patients with excellent surgical results and intermediate outcome. There is no evidence that a two-stage approach and/or baffle fenestration is required for a large cohort of patients who are candidates for a Fontan operation.


Haemodynamic simulation and the effect of early left ventricular unloading in pre-shock acute coronary syndrome.
Understanding the pathophysiological background on haemodynamic changes in acute myocardial infarction and during its interventional treatment is important to adequately use mechanical circulatory support. We describe haemodynamic simulations based on a real case scenario of infarct-related ischaemia with beginning haemodynamic compromise illustrating the advantage of active haemodynamic support. The patient case used for computer simulation is that of an acute coronary syndrome, slightly hypotonic. The right coronary artery is chronically occluded, and both left main and a saphenous vein graft to the left anterior descending coronary artery (LAD) show subtotal stenosis. In this scenario used for computer modelling of haemodynamics, we illustrate how unprotected percutaneous coronary intervention would limit coronary blood flow and constantly reduce myocardial contractility until cardiac arrest occurs. The simulation demonstrates how an intra-aortic balloon pump would delay but not prevent that compromise and how an Impella microaxial pump will actively support cardiac output and stabilize haemodynamics even when prolonged balloon inflations are performed, which will temporarily stop coronary perfusion. The simulation illustrates how temporary circulatory support with an Impella microaxial pump can stabilize haemodynamics and allow for a safe procedure in an unstable patient. Using computer simulation of haemodynamics to understand changes in haemodynamics when performing interventions in unstable patients might help to properly select a suitable support device if needed.


Impact of COVID-19 pandemic on treatment patterns in metastatic clear cell renal cell carcinoma.
The coronavirus pandemic has provoked discussions among healthcare providers how to manage cancer patients when faced with the threat of severe acute respiratory syndrome related coronavirus 2 (SARS-CoV-2) infection. Immune checkpoint inhibitor (ICI) containing regimens are standard of care in the majority of metastatic clear cell renal cell carcinoma (mccRCC) patients. It remains unclear whether therapies should be modified in response to the COVID-19 pandemic. We performed an online survey among physicians involved in the treatment of mccRCC, and 41 experts responded. Questions focused on criteria relevant for treatment decision outside the pandemic and the modifications of systemic therapy during COVID-19. For the majority of experts (73%), the combination of International metastatic renal cell carcinoma Database Consortium (IMDC) risk category and patient fitness are two important factors for decision-making. The main treatment choice in fit, favourable risk patients outside the pandemic is pembrolizumab/axitinib for 53%, avelumab/axitinib, sunitinib or pazopanib for 13% of experts each. During the pandemic, ICI-containing regimens are chosen less often in favour of a tyrosine kinase inhibitors (TKI) monotherapy, mainly sunitinib or pazopanib (35%).In fit, intermediate/poor-risk patients outside the pandemic, over 80% of experts choose ipilimumab/nivolumab, in contrast to only 41% of physicians during COVID-19, instead more TKI monotherapies are given. In patients responding to established therapies with ICI/ICI or ICI/TKI combinations, most participants modify treatment regimen by extending cycle length, holding one ICI or even both. mccRCC treatment modifications in light of the coronavirus pandemic are variable, with a shift from ICI/ICI to ICI/TKI or TKI monotherapy.


Use of urodynamics prior to surgery for urinary incontinence: How helpful is preoperative testing?
It has not yet been definitively demonstrated that preoperative evaluation of women with stress urinary incontinence with urodynamic testing enhances presurgical counseling, more effectively models patients' expectations or improves postoperative outcome. Nonetheless, urodynamic testing is frequently utilized in the assessment of women with stress urinary incontinence and clearly accomplishes a number of goals when utilized for this purpose. For example, there are data to suggest that the risk of voiding dysfunction can be mitigated by utilizing data obtained from urodynamic testing to identify women more likely to void ineffectively after conventional stress incontinence procedures. Furthermore, it has been suggested though not proven, that patients with more severe forms of stress incontinence as identified by urodynamic testing, might be less likely to improve after surgery compared to others with more modest degrees of incontinence. Since urodynamic testing is invasive, costly and not always available, it is imperative that the usefulness of such testing be carefully explored and its utility appropriately defined. In this review, we discuss urodynamic techniques to assess stress urinary incontinence, particularly focusing on the ability of leak point pressure testing and urethral pressure profilometry to predict which patients would most likely benefit from surgery and which might be more likely to experience adverse events following surgery.


Yellow Net of Triumffeta Is Caused by a Geminivirus: A First Report.
Triumffeta rhomboidiaceae Jacq. (Tiliaceae family) is an annual rainy season weed that is commonly found throughout India. For the last 3 years, during the rainy season, several plants of T. rhomboidiaceae in and around the gardens of the National Botanical Research Institute have been found with vein yellowing symptoms. The initial symptoms were vein clearing but in later stages the veins became yellow and thickened. In severe cases, the chlorosis extends into interveinal areas, resulting in complete yellowing of the leaves. In a few cases, green leafy or thorny enations could be seen on the dorsal side of the leaf. The disease was investigated to identify the causal agent. Vector transmission studies showed that the causal agent is transmitted by the whitefly, Bemisia tabaci, from infected to healthy seedlings of T. rhomdoidiaceae. Since whitefly transmission of the disease is consistent with a geminivirus as the causal agent, the role of such a virus was investigated. DNA isolated from Triumffeta plants (both from the infected plants in the field as well as from those inoculated experimentally in the greenhouse) showing above mentioned symptoms was amplified with two sets of degenerate primers, PAL1v1978/PAR1c496 (set 1) and PAL1v1978/PCRc1 (set 2), that have been shown to be specific for DNA-A of whitefly transmitted geminiviruses (WTGs), in polymerase chain reaction (1). We could amplify DNA-A fragments of approximately 1.2 kb from set 1 and 0.7 kb from set 2, as expected (1). DNA isolated from healthy seedlings gave no amplification of such fragments. Identification of the amplified DNA fragments (from infected samples) to be of geminiviral in nature was confirmed by Southern blot hybridization carried out under high stringency conditions. DNA-A of Indian tomato leaf curl virus (2) was used as a general probe for WTGs for the above hybridization experiment. Therefore, Triumffeta yellow net disease is caused by a geminivirus. A review of literature revealed that there is no record of a viral disease affecting this weed and, therefore, this is the first report of a viral disease affecting this plant. References: (1) M. R. Rojas et al. Plant Dis. 77:340, 1993. (2) K. M. Srivastava et al. J. Virol. Methods 51:297, 1995.