Correlation of Clinico-pathological Classification of Hansen's Disease in a South Indian City.
Hansen's Disease (HD) presents itself in different forms depending on the individual's immune status, and based on this Ridley-Jopling classified the disease into five sub-groups. The aim of this study was to evaluate the role of histopathology and bacteriological index (BI) in accurate staging of HD with clinical correlation. Fifty HD patients with clinical diagnosis confirmed by histopathology were included. Patients in reaction and on treatment were excluded. Case records and histopathological slides were viewed and BI was recorded. In 10/50 cases, a diagnosis of HD was made or suspected, but were not clinically classified. In these, histopathology proved useful in diagnosis and classification. Indeterminate HD was the most common histopathological diagnosis (6 cases). The remaining 40 patients, were clinically classified using the Ridley-Jopling classification, as Indeterminate Leprosy (IL) in 10/40 (25%), Tuberculoid Leprosy (TT) 5/40 (12.5%), Borderline Tuberculoid (BT) 16/40 (40%), Borderline Lepromatous (BL) 4/40 (10%) and Lepromatous Leprosy (LL) 5 (12.5%). HD was common in males with male to female ratio of 1.66:1 and affected the younger individuals (maximum in 21 to 30 years). On histopathology BT was the most common type (40%) followed by IL (27.5%), BL (12.5%), TT (10%) and LL (10%). No case of Mid-Borderline (BB) type was diagnosed clinically or histopathologically. Overall concordance between clinical and histopathological diagnosis was 65% (26/40 cases) and for each type was IL = 80%, TT = 20%, BT = 75%, BL = 50% and LL = 60%. Where classification seemed difficult as in cases of BT and BL, II played an important role. The overall concordance between clinical classification and histopathological diagnosis of DO is 65% in this study. Th *discordance that is observed is between BT and TT, the paucibacillary type and BL andILL the multibacillary type and hence the treatment is not affected. Overall, IL was a common diagnosis on histopathology in this study'(11/40 cases and 6/10 cases). If clinically warranted, a repeat deeper punch of skin biopsyrmay be required for a proper categorization of the cases.


Using a cross-contextual reciprocal learning approach in a multisite implementation research project to improve self-management for type 2 diabetes.
This paper reports on the use of reciprocal learning for identifying, adopting and adapting a type 2 diabetes self-management support intervention in a multisite implementation trial conducted in a rural setting in a low-income country (Uganda), a periurban township in a middle-income country (South Africa) and socioeconomically disadvantaged suburbs in a high-income country (Sweden). The learning process was guided by a framework for knowledge translation and structured into three learning cycles, allowing for a balance between evidence, stakeholder interaction and contextual adaptation. Key factors included commitment, common goals, leadership and partnerships. Synergistic outcomes were the cocreation of knowledge, interventions and implementation methods, including reverse innovations such as adaption of community-linked models of care. Contextualisation was achieved by cross-site exchanges and local stakeholder interaction to balance intervention fidelity with local adaptation. Interdisciplinary and cross-site collaboration resulted in the establishment of learning networks. Limitations of reciprocal learning relate to the complexity of the process with unpredictable outcomes and the limited generalisability of results.


Blood Pressure and Angiogenic Markers in Pregnancy: Contributors to Pregnancy-Induced Hypertension and Offspring Cardiovascular Risk.
Pregnancy-induced hypertension is a severe pregnancy complication, increasing risk of long-term cardiovascular disease in mothers and offspring. We hypothesized that maternal blood pressure in pregnancy associated with offspring blood pressure; that the associations were sex-specific; and that maternal circulating placental angiogenic markers (PlGF [placental growth factor] and sFlt-1 [soluble fms-like tyrosine kinase-1]) mediated this relationship. We analyzed data from 2434 women and 2217 children from the Odense Child Cohort, a prospective Danish cohort study. Offspring blood pressure trajectory from 4 months to 5 years was highly associated to maternal first, second, and third trimester blood pressure, and mean blood pressure in pregnancy, independent of maternal and offspring covariates. There were offspring sex-specific associations: Girls from mothers in the highest quartile of first and third trimester blood pressure had significantly higher systolic blood pressure at 5 years than the rest of the cohort (mean difference±SEM: 1.81±0.59 and 2.11±0.59 mm Hg, respectively, all P<0.01); whereas boys had significantly higher diastolic blood pressure at 5 years (mean difference±SEM: 1.11±0.45 and 1.03±0.45, respectively, all P<0.05). Concentrations of PlGF at gestational week 28 correlated inversely to maternal gestational blood pressure trajectory, independent of the diagnosis of pregnancy-induced hypertension, adjusted β coefficients (95% CI) for predicting systolic blood pressure (SBP): -3.18 (-4.66 to -1.70) mm Hg, for predicting diastolic blood pressure (DBP): -2.48 (-3.57 to -1.40) mm Hg. In conclusion, maternal gestational blood pressure predicted offspring blood pressure trajectory until 5 years in a sex-differential manner. Furthermore, subtle alterations in blood pressure in early pregnancy preceded hypertension or preeclampsia, and PlGF was a mediator of cardiovascular health in pregnancy.


Antibody repertoire analysis in polygenic autoimmune diseases.
High-throughput sequencing of the DNA/RNA encoding antibody heavy- and light-chains is rapidly transforming the field of adaptive immunity. It can address key questions, including: (i) how the B-cell repertoire differs in health and disease; and (ii) if it does differ, the point(s) in B-cell development at which this occurs. The advent of technologies, such as whole-genome sequencing, offers the chance to link abnormalities in the B-cell antibody repertoire to specific genomic variants and polymorphisms. Here, we discuss the current research using B-cell antibody repertoire sequencing in three polygenic autoimmune diseases where there is good evidence for a pathological role for B-cells, namely systemic lupus erythematosus, multiple sclerosis and rheumatoid arthritis. These autoimmune diseases exhibit significantly skewed B-cell receptor repertoires compared with healthy controls. Interestingly, some common repertoire defects are shared between diseases, such as elevated IGHV4-34 gene usage. B-cell clones have effectively been characterized and tracked between different tissues and blood in autoimmune disease. It has been hypothesized that these differences may signify differences in B-cell tolerance; however, the mechanisms and implications of these defects are not clear.


c-Fos Protects Neurons Through a Noncanonical Mechanism Involving HDAC3 Interaction: Identification of a 21-Amino Acid Fragment with Neuroprotective Activity.
Proteins belonging to the AP-1 family of transcription factors are known to be involved in the regulation of neuronal viability. While strides have been made to elucidate the mechanisms of how individual members regulate cell death, much remains unknown. We find that the expression of one AP-1 member, c-Fos, is reduced in cerebellar granule neurons (CGNs) induced to die by low potassium (LK) treatment. Restoration and increase of this expression protect CGNs against LK-induced death, whereas knockdown induces death of otherwise healthy neurons. Furthermore, forced expression can protect cortical neurons against homocysteic acid (HCA)-induced toxicity. Taken together, this suggests that c-Fos is necessary for neuronal survival and that elevating c-Fos expression has a neuroprotective effect. Consistent with this idea is the finding that c-Fos expression is reduced selectively in the striatum in two separate mouse models of Huntington's disease and forced expression protects against neuronal death resulting from mutant huntingtin (mut-Htt) expression. Interestingly, neuroprotection by c-Fos does not require its DNA-binding, transcriptional, or heteromerization domains. However, this protective activity can be inhibited by pharmacological inhibition of c-Abl, CK-I, and MEK-ERK signaling. Additionally, expression of point mutant forms of this protein has identified that mutation of a tyrosine residue, Tyr345, can convert c-Fos from neuroprotective to neurotoxic. We show that c-Fos interacts with histone deacetylase-3 (HDAC3), a protein that contributes to mut-Htt neurotoxicity and whose overexpression is sufficient to promote neuronal death. When co-expressed, c-Fos can protect against HDAC3 neurotoxicity. Finally, our study identifies a 21-amino acid region at the C-terminus of c-Fos that is sufficient to protect neurons against death induced by LK, HCA treatment, or mut-Htt expression when expressed via a plasmid transfection or as a cell-permeable peptide. This cell-permeable peptide, designated as Fos-CTF, could have potential as a therapeutic agent for neurodegenerative diseases.


Use of polymerase chain reaction for early identification of Mycobacterium tuberculosis in positive cultures.
To develop a readily applicable polymerase chain reaction (PCR) based technique which would permit the identification of Mycobacterium tuberculosis complex isolates from Bactec phials at an earlier stage than currently available methods. Mycobacterial cells cultured in Bactec 12B medium were harvested by centrifugation. The cells were lysed by heating in distilled water. Oligonucleotide primers based on the sequence of the gene coding for the immunogenic protein MPB64 were then used to amplify a 240 base pair fragment of DNA directly from the crude cell lysate. The PCR product was visualised under ultraviolet light following electrophoresis of an aliquot in an agarose gel containing ethidium bromide. The sensitivity of the PCR was adjusted so that about 600 cfu of M tuberculosis gave a positive result. The lowest growth index at which this method of identification might be applied to Bactec phials was determined and a number of routine cultures giving a positive growth index examined. M tuberculosis was positively identified at the lowest growth index, as determined by the Bactec system. Of 45 routine cultures examined, with growth indexes ranging from 6 to 999, the 15 confirmed by conventional means to contain M tuberculosis were correctly identified from 1 ml of culture medium. The method described can be used to identify M tuberculosis isolates cultured in the Bactec system at the earliest detectable rise in growth index. It may therefore allow cultured mycobacteria to be identified at an earlier stage than conventional methods or the commercially available DNA probes adapted for use with the Bactec system.


Composition based strategies for controlling radii in lipid nanotubes.
Nature routinely carries out small-scale chemistry within lipid bound cells and organelles. Liposome-lipid nanotube networks are being developed by many researchers in attempt to imitate these membrane enclosed environments, with the goal to perform small-scale chemical studies. These systems are well characterized in terms of the diameter of the giant unilamellar vesicles they are constructed from and the length of the nanotubes connecting them. Here we evaluate two methods based on intrinsic curvature for adjusting the diameter of the nanotube, an aspect of the network that has not previously been controllable. This was done by altering the lipid composition of the network membrane with two different approaches. In the first, the composition of the membrane was altered via lipid incubation of exogenous lipids; either with the addition of the low intrinsic curvature lipid soy phosphatidylcholine (soy-PC) or the high intrinsic curvature lipid soy phosphatidylethanolamine (soy-PE). In the second approach, exogenous lipids were added to the total lipid composition during liposome formation. Here we show that for both lipid augmentation methods, we observed a decrease in nanotube diameter following soy-PE additions but no significant change in size following the addition of soy-PC. Our results demonstrate that the effect of soy-PE on nanotube diameter is independent of the method of addition and suggests that high curvature soy-PE molecules facilitate tube membrane curvature.


Maternal Health Care Utilization Among Syrian Refugees in Lebanon and Jordan.
Purpose The influx of Syrian refugees into Jordan and Lebanon over the last 5 years presents an immense burden to national health systems. This study was undertaken to assess utilization of maternal health services among Syrian refugees in both countries. Description A cross-sectional survey of Syrian refugees living in urban and rural (non-camp) settings was conducted using a two-stage cluster survey design with probability proportional to size sampling in 2014-2015. Eighty-six percent of surveyed households in Lebanon and 88% of surveyed households in Jordan included women with a live birth in the last year. Information from women in this sub-set of households was analyzed to understand antenatal and intrapartum health service utilization. Assessment A majority of respondents reported seeking antenatal care, 82% and 89% in Jordan and Lebanon, respectively. Women had an average of at least six antenatal care visits. Nearly all births (98% in Jordan and 94% in Lebanon) took place in a health facility. Cesarean rates were similar in both countries; approximately one-third of all births were cesarean deliveries. A substantial proportion of women incurred costs for intrapartum care; 33% of Syrian women in Jordan and 94% of Syrian women in Lebanon reported paying out of pocket for their deliveries. The proportion of women incurring costs for intrapartum care was higher in Jordan both countries for women with cesarean deliveries compared to those with vaginal deliveries; however, this difference was not statistically significant in either country (Jordan p-value = 0.203; Lebanon p-value = 0.099). Conclusion Syrian refugees living in Jordan and Lebanon had similar levels of utilization of maternal health services, despite different health systems and humanitarian assistance provisions. As expected, a substantial proportion of households incurred out-of-pocket costs for essential maternal and newborn health services, making cost a major factor in care-seeking decisions and locations. As health financing policies shift to account for the continued burden of refugee hosting on the health system, sustained attention to the availability and quality of essential maternal and newborn health services is needed to protect both refugee and host populations women's rights to health and health care during pregnancy, childbirth, and the postpartum period.


A systematic review of the research evidence for the benefits of teledentistry.
Objective This review is designed to inform future decisions about the benefits of integrating teledentistry into routine health services, by presenting an overview of the evidence for the effectiveness and economic impact of teledentistry. Methods Two reviewers searched PubMed, EMBASE and CINAHL databases through November 2016 to identify published peer-reviewed studies in English. Teledentistry studies were included if they were; (a) controlled (randomised or non-randomised) assessment studies; and (b) compared outcomes of a teledentistry intervention in terms of clinical or economic evaluation with the outcomes of traditional clinical alternatives. The quality of the studies was evaluated using a quality appraisal tool that considered study performance and design. Results This review identified 385 publications, of which 217 full-text articles were retrieved for further inspection. Of these, only 11 articles met the inclusion criteria. Nine of the included articles showed some clinical outcomes; the other two were primarily economic analyses. The balance of these studies assesed the efficacy of teledentistry interventions rather than their effectiveness. Four studies (36%) achieved higher quality scores and have greater potential to influence health-care decision-making. To date, the most convincing published evidence regarding the efficacy of teledentistry was provided by studies on paediatric dentistry, orthodontics and oral medicine. The economic analysis referred only to cost-minimisation, suggesting that the use of teleconsultation in dentistry can be cost-saving when compared to a conventional consultation. However, high-quality economic studies on teledentistry are rare. Conclusion There is emerging evidence supporting the efficacy of teledentistry. However, there is not yet enough conclusive evidence, particularly for its effectiveness, cost-effectiveness and long-term use, to make evidence-based policy decisions on teledentistry.


Lengthened Cutaneous Silent Period in Fibromyalgia Suggesting Central Sensitization as a Pathogenesis.
The pathogenesis of fibromyalgia (FM) has not been clearly elucidated, but central sensitization, which plays an important role in the development of neuropathic pain, is considered to be the main mechanism. The cutaneous silent period (CSP), which is a spinal reflex mediated by A-delta cutaneous afferents, is useful for the evaluation of sensorimotor integration at the spinal and supraspinal levels. To understand the pathophysiology of FM, we compared CSP patterns between patients with FM and normal healthy subjects. Twenty-four patients with FM diagnosed in accordance with the 1990 American College of Rheumatology classification system and 24 age- and sex-matched healthy volunteers were recruited. The CSP was measured from the abductor pollicis brevis muscle. Demographic data, number of tender points, and visual analog scale and FM impact questionnaire scores were collected. The measured CSP and clinical parameters of the patient and control groups were compared. In addition, possible correlations between the CSP parameters and the other clinical characteristics were analyzed. Mean CSP latencies did not differ between patients (55.50 ± 10.97 ms) and healthy controls (60.23 ± 11.87 ms; p = 0.158), although the mean CSP duration was significantly longer in patients (73.75 ± 15.67 ms) than in controls (63.50 ± 14.05 ms; p = 0.021). CSP variables did not correlate with any clinical variables. The significantly longer CSP duration in FM patients suggests central dysregulation at the spinal and supraspinal levels, rather than peripheral small fiber dysfunction.


Novel Benzodiazepine-Like Ligands with Various Anxiolytic, Antidepressant, or Pro-Cognitive Profiles.
Altered gamma-aminobutyric acid (GABA) function is consistently reported in psychiatric disorders, normal aging, and neurodegenerative disorders and reduced function of GABA interneurons is associated with both mood and cognitive symptoms. Benzodiazepines (BZ) have broad anxiolytic, but also sedative, anticonvulsant and amnesic effects, due to nonspecific GABA-A receptor (GABAA-R) targeting. Varying the profile of activity of BZs at GABAA-Rs is predicted to uncover additional therapeutic potential. We synthesized four novel imidazobenzodiazepine (IBZD) amide ligands and tested them for positive allosteric modulation at multiple α-GABAA-R (α-positive allosteric modulators), pharmacokinetic properties, as well as anxiolytic and antidepressant activities in adult mice. Efficacy at reversing stress-induced or age-related working memory deficits was assessed using a spontaneous alternation task. Diazepam (DZP) was used as a control. Three ligands (GL-II-73, GL-II-74, and GL-II-75) demonstrated adequate brain penetration and showed predictive anxiolytic and antidepressant efficacies. GL-II-73 and GL-II-75 significantly reversed stress-induced and age-related working memory deficits. In contrast, DZP displayed anxiolytic but no antidepressant effects or effects on working memory. We demonstrate distinct profiles of anxiolytic, antidepressant, and/or pro-cognitive activities of newly designed IBZD amide ligands, suggesting novel therapeutic potential for IBZD derivatives in depression and aging.


The endocrinology of the brain.
The brain hosts a vast and diverse repertoire of neuropeptides, a class of signalling molecules often described as neurotransmitters. Here I argue that this description entails a catalogue of misperceptions, misperceptions that feed into a narrative in which information processing in the brain can be understood only through mapping neuronal connectivity and by studying the transmission of electrically conducted signals through chemical synapses. I argue that neuropeptide signalling in the brain involves primarily autocrine, paracrine and neurohormonal mechanisms that do not depend on synaptic connectivity and that it is not solely dependent on electrical activity but on mechanisms analogous to secretion from classical endocrine cells. As in classical endocrine systems, to understand the role of neuropeptides in the brain, we must understand not only how their release is regulated, but also how their synthesis is regulated and how the sensitivity of their targets is regulated. We must also understand the full diversity of effects of neuropeptides on those targets, including their effects on gene expression.


Laparoscopic pyloroplasty is a safe and effective first-line surgical therapy for refractory gastroparesis.
Surgical options for symptomatic delayed gastric emptying include gastric stimulator implantation, subtotal gastrectomy, and pyloroplasty. Pyloroplasty has been shown to improve gastric emptying yet is seldom described as a primary treatment for gastroparesis. We present a single-institution experience of laparoscopic Heineke-Mikulicz pyloroplasty (LP) as treatment for gastroparesis. A prospective foregut surgery database was queried for LP over a 5-year period. Charts were reviewed for indications, complications, symptom score, and outcomes. Gastroparesis was defined by (1) abnormal gastric emptying study, (2) endoscopic visualization of retained food after prolonged NPO status, or (3) clinical symptoms suspicious of vagal nerve injury following complex re-operative foregut surgery. Results were analyzed using a paired T test and single-factor ANOVA. One hundred and seventy-seven LP patients were identified and reviewed. One hundred and five had a concurrent fundoplication for objective reflux. There were no intraoperative complications or conversions to laparotomy. Overall morbidity rate was 6.8% with four return to OR and two confirmed leaks (1.1% leak rate). Average length of stay was 3.5 days, and readmission rate was 7%. Eighty-six percent had improvement in GES with normalization in 77%. Gastric emptying half-time decreased from 175 ± 94 to 91 ± 45 min. Nineteen patients (10.7%) had subsequent surgical interventions: gastric stimulator implantation (12), feeding jejunostomy and/or gastrostomy tube (6), or subtotal gastrectomy (4). Symptom severity scores for nausea, vomiting, bloating, abdominal pain, and early satiety decreased significantly at 3 months. Laparoscopic pyloroplasty improves or normalizes gastric emptying in nearly 90% of gastroparesis patients with very low morbidity. It significantly improves symptoms of nausea, vomiting, bloating, and abdominal pain. Some patients may go on to another surgical treatment for GP, but it remains a safe and less invasive alternative to a subtotal gastrectomy in these clinically challenging patients.


Implementation of health-focused interventions in vulnerable populations: protocol for a scoping review.
Vulnerable populations face significant challenges in navigating the care continuum, ranging from diagnosis of illness to linkage and retention in healthcare. Understanding how best to move individuals within these vulnerable populations across the care continuum is critical to improving their health. A large body of literature has focused on evaluation of implementation of various health-focused interventions in this population. However, we do not fully understand the unique challenges to implementing healthcare interventions for vulnerable populations. This study aims to examine the literature describing implementation of health service interventions among vulnerable populations to identify how implementations using the Consolidated Framework for Implementation Research are adapted. Findings from this review will be useful to implementation scientists to identify gaps in evidence and for adapting similar interventions in unique settings. This study protocol outlines a scoping review of the peer-reviewed and grey literature, using established approaches delineated in Arksey and O'Malley's scoping review framework and the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews Checklist. Search strategies will be developed and refined by a medical librarian in collaboration with the research team. Searches will be conducted in electronic databases (CINAHL, Cochrane, PsychINFO, PubMed, Social Services Abstracts, Web of Science, Google and Google Scholar) and limited to studies published between 1 August 2009 and 1 June 2020. Additionally, hand searches will be conducted in three relevant journals-Implementation Science, Systematic Reviews and BMJ Open. English-language studies and reports meeting inclusion criteria will be screened independently by two reviewers and the final list will be abstracted and charted in duplicate. This is a review of the literature; ethics approval is not indicated. We will disseminate findings from this study in peer-reviewed journals as well as presentations to relevant stakeholders and conferences.


Massive localized malignant pleural mesothelioma (LMPM): manifestations on computed tomography in 6 cases.
Our study analyzed the clinical symptoms and computed tomography (CT) manifestations of massive localized malignant pleural mesothelioma (LMPM) patients to improve the knowledge and diagnosis of this disease. Our study collected 6 massive LMPM patients pathologically confirmed by CT in the department of Radiology of the People's Hospital of Yuyao, Zhejiang Province, from January, 2007 to June, 2013; data of patients were also collected. The clinical symptoms, clinicopathological characteristics, CT manifestations, treatments and prognosis of enrolled patients were analyzed. Our study enrolled 6 LMPM patients (2 males; 4 females) classified to epitheliated type (n = 4) and sarcomatous type (n = 2) with mean age of 62.7 ± 7.4, and 5 of them had a history of asbestos exposure. CT manifestations revealed that large soft-tissue mass close to pleura, which was smooth and lobulated, was discovered in all patients with maximum diameter of 10~15 cm and mean diameter of 13.67 ± 1.15 cm; The mean value of CT was 36.29 ± 2.62 HU; after enhancement, the mean value was increased to 76.36 ± 7.73 HU; patients showed zones of small patchy necrosis and large patchy necrosis. The following presentations were founded: enlargement of tumor vessel which showed arborization (2 patients), mass wrap around the descending aorta in left lower chest (1 patient), strips of fat density in mediastinum superior (1 patient), pleural tail sign (3 patients). Among 6 patients, pleural effusion (n = 4), mediastinal lymph node enlargement (n = 3), invasion and destruction of local ribs (n = 2). Median survival time of patients were 20 months (2 cases conducted operation), 24 (2 cases chose combined radiotherapy and chemotherapy) and less than 6 months (2 cases underwent chemotherapy). To sum up, CT showed important diagnostic values on massive LMPM patients; patients with a history of asbestos exposure, large soft-tissue mass of pleura with an abundant blood supply and wrap around large vessels might increase the risk of massive LMPM.


Recurrent malignant melanoma: effect of adjuvant immunotherapy on survival.
Twenty-nine patients referred consecutively to a cancer clinic because of recurrent metastatic malignant melanoma were given 5 mg of Connaught Laboratories bacillus Calmette-Guérin (BCG) by multiple cutaneous puncture at weekly and later at monthly intervals. Eight were also treated with autologous tumour vaccine and three with intralesional BCG. This group was compared with a retrospective control group of 54 patients treated with surgery and radiotherapy alone after recurrence. Prognostic features such as site of primary and of first metastasis, disease-free interval, age and sex were similar in the two groups. However, the median survival from the time of first recurrence was 12 months in the control group but 21 months in the BCG-treated group. The major improvement was in patients with disease limited to the regional lymph nodes: the median survival was 16 months in the control group but over 32 months in the BCG-treated group. Autologous tumour vaccine appeared to have no effect on survival. Serial testing of immunocompetence did not offer any prognostic advantage, although the results of some tests correleated well with extent of disease.


Identification of asparagine-linked oligosaccharides involved in tumor cell adhesion to laminin and type IV collagen.
MDW4, a wheat germ agglutinin-resistant nonmetastatic mutant of the highly metastatic murine tumor cell line called MDAY-D2 has previously been shown to attach to fibronectin and type IV collagen, whereas MDAY-D2 and phenotypic revertants of MDW4 attached poorly to these substrates. The increased adhesiveness of the mutant cells appeared to be closely related to a lesion in cell surface carbohydrate structures. In an effort to identify the carbohydrates involved in cell attachment, glycopeptides isolated from mutant and wild-type cells as well as from purified glycoproteins were tested for their ability to inhibit the attachment of MDW4 cells to plastic surfaces coated with fibronectin, laminin, or type IV collagen. The addition of mannose-terminating glycopeptide to the adhesion assay inhibited MDW4 cell attachment to type IV collagen. In contrast, a sialylated poly N-acetyllactosamine-containing glycopeptide, isolated from wheat germ agglutinin-sensitive MDAY-D2 cells but absent in MDW4 cells, inhibited MDW4 attachment to laminin. None of the glycopeptides used in this study inhibited attachment of MDW4 cells to fibronectin-coated plastic. Peptide N-glycosidase treatment of the cells to remove surface asparagine-linked oligosaccharides inhibited MDW4 adhesion to type IV collagen, but not to laminin, and the same treatment of the wheat germ agglutinin-sensitive cells enhanced attachment to laminin. Tumor cell attachment to, and detachment from, the sublaminal matrix protein laminin and type IV collagen are thought to be important events in the metastatic process. Our results indicate that tumor cell attachment to these proteins may be partially modulated by the expression of specific oligosaccharide structures associated with the cell surface.


Collective soul: the spirituality of an interdisciplinary palliative care team.
Although spirituality as it relates to patients is gaining increasing attention, less is known about how health care professionals (HCP) experience spirituality personally or collectively in the workplace. This study explores the collective spirituality of an interdisciplinary palliative care team, by studying how individuals felt about their own spirituality, whether there was a shared sense of a team spirituality, how spirituality related to the care the team provided to patients and whether they felt that they provided spiritual care. A qualitative autoethnographic approach was used. The study was conducted in a 10-bed Tertiary Palliative Care Unit (TPCU) in a large acute-care referral hospital and cancer center. Interdisciplinary team members of the TPCU were invited to participate in one-to-one interviews and/or focus groups. Five interviews and three focus groups were conducted with a total of 20 participants. Initially participants struggled to define spirituality. Concepts of spirituality relating to integrity, wholeness, meaning, and personal journeying emerged. For many, spirituality is inherently relational. Others acknowledged transcendence as an element of spirituality. Spirituality was described as being wrapped in caring and often manifests in small daily acts of kindness and of love, embedded within routine acts of caring. Palliative care served as a catalyst for team members' own spiritual journeys. For some participants, palliative care represented a spiritual calling. A collective spirituality stemming from common goals, values, and belonging surfaced. This was the first known study that focused specifically on the exploration of a collective spirituality. The culture of palliative care seems to foster spiritual reflection among health care professionals both as individuals and as a whole. While spirituality was difficult to describe, it was a shared experience often tangibly present in the provision of care on all levels.


Porcupine-mediated lipidation is required for Wnt recognition by Wls.
Wnt proteins are members of a conserved family of secreted signaling ligands and play crucial roles during development and in tissue homeostasis. There is increasing evidence that aberrant Wnt production is an underlying cause of dysregulated Wnt signaling, however little is known about this process. One protein known to play a role in secretion is the transmembrane protein Wntless (Wls). However, the mechanism by which Wls promotes Wnt secretion is a riddle. It is not known which Wnt family members require Wls and what the structural requirements are that make some of them reliant on Wls for secretion. Here we present a systematic analysis of all known Drosophila Wnt family members with respect to their dependence on Wls function for secretion. We first show that the glycosylation status of Wg at conserved sites does not determine its dependence on Wls. Moreover, in apparent contrast to murine wls, Drosophila wls is not a target gene of canonical Wnt signaling. We then show that all Wnts, with the exception of WntD, require Wls for secretion. All Wnts, with the exception of WntD, also contain a conserved Serine residue (in Wg S239), which we show to be essential for their functional and physical interaction with Wls. Finally, all Wnts, with the exception of WntD, require the acyltransferase Porcupine for activity and for functionally interacting with Wls. Together, these findings indicate that Por-mediated lipidation of the S239-equivalent residue is essential for the interaction with, and secretion by, Wls.


Apoptotic cell death triggered by nitric oxide in pancreatic beta-cells.
Nitric oxide (NO) is believed to be an effector molecule that mediates interleukin (IL)-1 beta-induced destruction and dysfunction of pancreatic beta-cells. We have demonstrated that both exogenous NO and NO generated endogenously by IL-1 beta brought about apoptosis of isolated rat pancreatic islet cells as well as pancreatic beta-cell tumor-derived cell line HIT. This apoptosis was characterized by cleavage of DNA into nucleosomal fragments of 180-200 bp and morphologically by nuclear shrinkage, chromatic condensation, and apoptotic body formation. The IL-1 beta-induced internucleosomal DNA cleavage occurred in a time- and dose-dependent manner. Actinomycin D, cycloheximide, and nitric oxide synthase inhibitors inhibited the DNA cleavage, which was correlated with the amount of NO produced, indicating that NO produced by HIT cells themselves could mediate the apoptosis. Furthermore, in the presence of tumor necrosis factor (TNF)-alpha, large amounts of NO were produced by IL-1 beta and DNA cleavage occurred more noticeably, although TNF-alpha alone did not generate NO. Streptozotocin (STZ), a diabetogenic reagent containing a nitroso moiety, also released NO and induced internucleosomal DNA cleavage in HIT cells. These results suggest that NO-induced internucleosomal DNA cleavage is an important initial step in the destruction and dysfunction of pancreatic beta-cells induced by inflammatory stimulation or treatment with STZ.


Selective intratumoral amplification of an antiangiogenic vector by an oncolytic virus produces enhanced antivascular and anti-tumor efficacy.
The development of effective cancer therapy will require the simultaneous targeting of multiple steps in tumor development. We have previously described an antiangiogenic gene therapy vector, Ad Flk1-Fc, which expresses a soluble VEGF receptor capable of inhibiting tumor angiogenesis and growth. We have also described an oncolytic virus, dl922/947, whose replication and subsequent cytotoxicity are restricted to cancer cells with a loss of the G1-S cell cycle checkpoint. Here we have optimized methods for combining these therapies, yielding significantly greater anti-tumor effects than the respective monotherapies. In cultured tumor lines, co-infection with both Ad Flk1-Fc and dl922/947 allowed replication and repackaging of the replication-deficient Ad Flk1-Fc and enhanced soluble VEGF receptor expression. Similar repackaging and increased gene expression were demonstrated in vivo using bioluminescence imaging studies. Finally, coadministration of these therapeutic viral therapies in vivo produced significantly enhanced anti-tumor effects in colon HCT 116 and prostate PC-3 xenografts in mice. This increased therapeutic benefit correlated with replication of Ad Flk1-Fc viral genomes, increased intratumoral levels of Flk1-Fc protein, and decreased microvessel density, consistent with enhanced antiangiogenic activity.


Preparation of Whole Bone Marrow for Mass Cytometry Analysis of Neutrophil-lineage Cells.
In this article, we present a protocol that is optimized to preserve neutrophil-lineage cells in fresh BM for whole BM CyTOF analysis. We utilized a myeloid-biased 39-antibody CyTOF panel to evaluate the hematopoietic system with a focus on the neutrophil-lineage cells by using this protocol. The CyTOF result was analyzed with an open-resource dimensional reduction algorithm, viSNE, and the data was presented to demonstrate the outcome of this protocol. We have discovered new neutrophil-lineage cell populations based on this protocol. This protocol of fresh whole BM preparation may be used for 1), CyTOF analysis to discover unidentified cell populations from whole BM, 2), investigating whole BM defects for patients with blood disorders such as leukemia, 3), assisting optimization of fluorescence-activated flow cytometry protocols that utilize fresh whole BM.


Structure of purine-purine mispairs during misincorporation and extension by Escherichia coli DNA polymerase I.
The mechanism by which purine-purine mispairs are formed and extended was examined with the high-fidelity Klenow fragment of Escherichia coli DNA polymerase I with the proofreading exonuclease activity inactivated. The structures of the purine-purine mispairs were examined by comparing the kinetics of mispair formation with adenine versus 7-deazaadenine and guanine versus 7-deazaguanine at four positions in the DNA, the incoming dNTP, the template base, and both positions of the terminal base pair. A decrease in rate associated with a 7-deazapurine substitution would suggest that the nucleotide is in a syn conformation in a Hoogsteen base pair with the opposite base. During mispair formation, the k(pol)/K(d) values for the insertion of dATP opposite A (dATP/A) as well as dATP/G and dGTP/G were decreased greater than 10-fold with the deazapurine in the dNTP. These results suggest that during mispair formation the newly forming base pair is in a Hoogsteen geometry with the incoming dNTP in the syn conformation and the template base in the anti conformation. During mispair extension, the only decrease in k(pol)/K(d) was associated with the G/G base pair in which 7-deazaguanine was in the template strand. These results as well as previous results [McCain et al. (2005) Biochemistry 44, 5647-5659] in which a hydrogen bond was found between the 3-position of guanine at the primer terminus and Arg668 during G/A and G/G mispair extension indicate that the conformation of the purine at the primer terminus is in the anti conformation during mispair extension. These results suggest that purine-purine mispairs are formed via a Hoogsteen geometry in which the dNTP is in the syn conformation and the template is in the anti conformation. During extension, however, the conformation of the primer terminus changes to an anti configuration while the template base may be in either the syn or anti conformations.


Recommendations for Clinical Trial Development in Mantle Cell Lymphoma.
Mantle cell lymphoma (MCL) comprises around 6% of all non-Hodgkin's lymphoma (NHL) diagnoses. In younger patients, age less than 60 to 65 years, aggressive induction often followed by consolidation with autologous stem cell transplant has suggested improved outcomes in this population. Less intensive therapies in older patients often followed by maintenance have been studied or are under active investigation. However, despite recent advances, MCL remains incurable, with a median overall survival of around five years. Patients with high-risk disease have particularly poor outcomes. Treatment varies widely across institutions, and to date no randomized trials comparing intensive vs less intensive approaches have been reported. Although recent data have highlighted the heterogeneity of MCL outcomes, patient assessment for treatment selection has largely been driven by patient age with little regard to fitness, disease biology, or disease risk. One critical advance is the finding that minimal residual disease status (MRD) after induction correlates with long-term outcomes. As such, its use as a potential end point could inform clinical trial design. In order to more rapidly improve the outcomes of MCL patients, clinical trials are needed that prospectively stratify patients on the basis of MCL biology and disease risk, incorporate novel agents, and use MRD to guide the need for additional therapy.


The Old and the New: Can Physics-Informed Deep-Learning Replace Traditional Linear Solvers?
Physics-Informed Neural Networks (PINN) are neural networks encoding the problem governing equations, such as Partial Differential Equations (PDE), as a part of the neural network. PINNs have emerged as a new essential tool to solve various challenging problems, including computing linear systems arising from PDEs, a task for which several traditional methods exist. In this work, we focus first on evaluating the potential of PINNs as linear solvers in the case of the Poisson equation, an omnipresent equation in scientific computing. We characterize PINN linear solvers in terms of accuracy and performance under different network configurations (depth, activation functions, input data set distribution). We highlight the critical role of transfer learning. Our results show that low-frequency components of the solution converge quickly as an effect of the F-principle. In contrast, an accurate solution of the high frequencies requires an exceedingly long time. To address this limitation, we propose integrating PINNs into traditional linear solvers. We show that this integration leads to the development of new solvers whose performance is on par with other high-performance solvers, such as PETSc conjugate gradient linear solvers, in terms of performance and accuracy. Overall, while the accuracy and computational performance are still a limiting factor for the direct use of PINN linear solvers, hybrid strategies combining old traditional linear solver approaches with new emerging deep-learning techniques are among the most promising methods for developing a new class of linear solvers.


Radioimmunotherapy with (111)In/(90)Y-2IT-BAD-m170 for metastatic prostate cancer.
Over 31,000 Americans die of androgen-independent metastatic prostate cancer each year. New strategies that do not involve hormonal manipulation but instead recognize the biochemical and molecular characteristics of prostate cancer are needed. Radioimmunotherapy (RIT) uses a tumor-specific monoclonal antibody to deliver systemic, targeted radiation to cancer. The objectives of this Phase I study of (111)In-2IT-BAD-m170 (for imaging) and (90)Y-2IT-BAD-m170 (for therapy) were to determine the toxicity and maximum tolerated dose (MTD), the specificity for targeting metastatic prostate cancer, and the efficacy for palliation of pain. M170 is a mouse monoclonal antibody that targets adenocarcinomas. Patients with adequate renal and liver function, rising prostate-specific antigen, and androgen-independent metastatic prostate cancer were eligible. After estimation of dosimetry and pharmacokinetics with (111)In-2IT-BAD-m170, a single dose of (90)Y-2IT-BAD-m170 (0.185, 0.370, 0.555, or 0.740 GBq/m(2)) was administered to cohorts of three patients. Pain was assessed objectively by questionnaires before and for 8 weeks after RIT; weekly prostate-specific antigen levels were obtained for 2 months after RIT. The MTD of (90)Y-2IT-BAD-m170 was 0.740 GBq/m(2) for patients that had up to 10% of the axial skeleton involved with prostate cancer. Toxicity was almost exclusively confined to reversible myelosuppression. Metastatic prostate cancer was targeted by (111)In-2IT-BAD-m170 in all 17 patients. The mean radiation dose delivered to 39 bone and 18 nodal metastases by (90)Y-2IT-BAD-m170 was 10.5 Gy/GBq (range 2.8-25.1). Thirteen of 17 patients reported pain before (90)Y-2IT-BAD-m170; 7 of these 13 had a partial or complete resolution of pain that lasted an average of 4.3 weeks. This study determined the MTD of (111)In/(90)Y-2IT-BAD-m170 in patients with metastatic prostate cancer. The drugs were well tolerated, targeted metastases, and temporarily palliated pain.


Impact of Enteral Feeding on Vasoactive Support in Septic Shock: A Retrospective Observational Study.
Introducing enteral nutrition (EN) during hemodynamic instability may induce the splanchnic steal phenomenon, which may worsen systemic oxygen delivery and increase vasopressor dose. We aimed to determine the change in vasopressor dose in septic shock patients who received concomitant EN. We hypothesize that EN delivery is not associated with worsening hemodynamic instability, as defined by an increase in vasopressor dose ≥50% at 24 hours. This is a retrospective observational cohort study of adult patients with septic shock who were admitted to the intensive care unit from January 2015 to June 2015 and received EN. Vasopressor and EN parameters were collected at 6-hour intervals for the first 24 hours. Data were available for 28 consecutive patients. The mean age was 60 years (SD = 18), and 54% were females. Norepinephrine (NE) was used in 100%. EN and vasopressor overlap totaled 36 hours (interquartile range [IQR], 27-69). Median NE dose when starting EN was 5.9 μg/min (IQR, 3.88). Median change in dose from 0 to 6 hours was 0.85 μg/min (95% CI, 0.681.06; P = 0.136), corresponding to a median increase of 14.5%. Total NE duration was 60.5 hours (IQR, 47.5-75.5). No serious complications occurred. The median vasopressor dose did not increase by ≥50% during the first 24 hours of EN. This suggests early EN delivered during septic shock is not associated with worsening hemodynamic instability. Limitations include a small sample size and residual confounding. Prospective data are needed.


Specific rearrangement reactions of acetylated lysine containing peptide bn (n = 4-7) ion series.
Characterization of ε-N-acetylated lysine containing peptides, one of the most prominent post-translational modifications of proteins, is an important goal for tandem mass spectrometry experiments. A systematic study for the fragmentation reactions of b ions derived from ε-N-acetyllysine containing model octapeptides (KAc YAGFLVG and YAKAc GFLVG) has been examined in detail. Collision-induced dissociation (CID) mass spectra of bn (n = 4-7) fragments of ε-N-acetylated lysine containing peptides are compared with those of N-terminal acetylated and doubly acetylated (both ε-N and N-terminal) peptides, as well as acetyl-free peptides. Both direct and nondirect fragments are observed for acetyl-free and singly acetylated (ε-N or N-terminal) peptides. In the case of ε-N-acetylated lysine containing peptides, however, specific fragment ions (m/z 309, 456, 569 and 668) are observed in CID mass spectra of bn (n = 4-7) ions. The CID mass spectra of these four ions are shown to be identical to those of selected protonated C-terminal amidated peptides. On this basis, a new type of rearrangement chemistry is proposed to account for the formation of these fragment ions, which are specific for ε-N-acetylated lysine containing peptides. Consistent with the observation of nondirect fragments, it is proposed that the b ions undergo head-to-tail macrocyclization followed by ring opening. The proposed reaction pathway assumes that bn (n = 4-7) of ε-N-acetylated lysine containing peptides has a tendency to place the KAc residue at the C-terminal position after macrocyclization/reopening mechanism. Then, following the loss of CO, it is proposed that the marker ions are the result of the loss of an acetyllysine imine as a neutral fragment.


Mutations in tetratricopeptide repeat domain 7A result in a severe form of very early onset inflammatory bowel disease.
Very early onset inflammatory bowel diseases (VEOIBD), including infant disorders, are a diverse group of diseases found in children younger than 6 years of age. They have been associated with several gene variants. Our aim was to identify the genes that cause VEOIBD. We performed whole exome sequencing of DNA from 1 infant with severe enterocolitis and her parents. Candidate gene mutations were validated in 40 pediatric patients and functional studies were carried out using intestinal samples and human intestinal cell lines. We identified compound heterozygote mutations in the Tetratricopeptide repeat domain 7 (TTC7A) gene in an infant from non-consanguineous parents with severe exfoliative apoptotic enterocolitis; we also detected TTC7A mutations in 2 unrelated families, each with 2 affected siblings. TTC7A interacts with EFR3 homolog B to regulate phosphatidylinositol 4-kinase at the plasma membrane. Functional studies demonstrated that TTC7A is expressed in human enterocytes. The mutations we identified in TTC7A result in either mislocalization or reduced expression of TTC7A. Phosphatidylinositol 4-kinase was found to co-immunoprecipitate with TTC7A; the identified TTC7A mutations reduced this binding. Knockdown of TTC7A in human intestinal-like cell lines reduced their adhesion, increased apoptosis, and decreased production of phosphatidylinositol 4-phosphate. In a genetic analysis, we identified loss of function mutations in TTC7A in 5 infants with VEOIBD. Functional studies demonstrated that the mutations cause defects in enterocytes and T cells that lead to severe apoptotic enterocolitis. Defects in the phosphatidylinositol 4-kinase-TTC7A-EFR3 homolog B pathway are involved in the pathogenesis of VEOIBD.


Optimization and comparison of two different 3D culture methods to prepare cell aggregates as a bioink for organ printing.
The ultimate goal of tissue engineering is to design and fabricate functional human tissues that are similar to natural cells and are capable of regeneration. Preparation of cell aggregates is one of the important steps in 3D tissue engineering technology, particularly in organ printing. Two simple methods, hanging drop (HD) and conical tube (CT) were utilized to prepare cell aggregates. The size and viability of the aggregates obtained at different initial cell densities and pre-culture duration were compared. The proliferative ability of the cell aggregates and their ability to spread in culture plates were also investigated. In both methods, the optimum average size of the aggregates was less than 500 microm. CT aggregates were smaller than HD aggregates. 5,000 cells per drop HD aggregates showed a marked ability to attach and spread on the culture surface. The proliferative ability reduced when the initial cell density was increased. Comparing these methods, we found that the HD method having better size controlling ability as well as enhanced ability to maintain higher rates of viability, spreading, and proliferation. In conclusion, smaller HD aggregates might be a suitable choice as building blocks for making bioink particles in bioprinting technique.


Acetylcholine receptors loss and postsynaptic damage in MuSK antibody-positive myasthenia gravis.
Muscle-specific tyrosine kinase (MuSK) antibodies are found in some patients with "seronegative" myasthenia gravis (MG), but how they cause myasthenic symptoms is not clear. We visualized acetylcholine receptors (AChRs) and complement component 3 (C3) in muscle biopsies from 10 Japanese MG patients with MuSK antibodies, compared with 42 with AChR antibodies. The AChR density was not significantly decreased in MuSK antibody (Ab)-positive end-plates compared with AChR antibody-positive end-plates, and C3 was detected in only two of eight MuSK Ab-positive patients. MuSK antibodies do not appear to cause substantial AChR loss, complement deposition, or morphological damage. Effects on MuSK function need to be explored.


Nutritional and Pharmacological Targeting of the Calcium-Sensing Receptor Influences Chemically Induced Colitis in Mice.
The calcium-sensing receptor (CaSR) is the main regulator of extracellular Ca2+ homeostasis. It has diverse functions in different tissues, including the intestines. Intestine-specific knockout of the CaSR renders mice more susceptible to dextran sulphate sodium (DSS)-induced colitis. To test our hypothesis that the CaSR reduces intestinal inflammation, we assessed the effects of nutritional and pharmacological agonists of the CaSR in a colitis model. We treated female Balb/C mice with dietary calcium and protein (nutritional agonists of the CaSR) or pharmacological CaSR modulators (the agonists cinacalcet and GSK3004774, and the antagonist NPS-2143; 10 mg/kg), then induced colitis with DSS. The high-protein diet had a strong pro-inflammatory effect-it shortened the colons (5.3 ± 0.1 cm vs. 6.1 ± 0.2 cm normal diet, p < 0.05), lowered mucin expression and upregulated pro-inflammatory cytokines, such as interferon-γ, (4.2-fold, p < 0.05) compared with the normal diet. Cinacalcet reduced mucin expression, which coincided with an increase in tumor necrosis factor-α (4.4-fold, p < 0.05) and IL-6 (4.9-fold, p < 0.05) in the plasma, compared with vehicle. The CaSR antagonist, NPS-2143, significantly reduced the cumulative inflammation score compared with the vehicle control (35.3 ± 19.1 vs. 21.9 ± 14.3 area under the curve, p < 0.05) and reduced infiltration of inflammatory cells. While dietary modulation of the CaSR had no beneficial effects, pharmacological inhibition of the CaSR may have the potential of a novel add-on therapy in the treatment of inflammatory bowel diseases.


Hemodynamic and neural mechanisms of action of thyrotropin-releasing hormone in the rat.
The mechanisms mediating the effects of thyrotropin-releasing hormone (TRH) on the cardiovascular system were studied in the conscious rat. Intracerebroventricular (i.c.v.) injection of TRH (8 pmol-80 nmol/kg) induced dose-dependent increases in mean arterial pressure, heart rate, and cardiac index. Hindquarter blood flow increased due to vasodilation, while an increase in renal and mesenteric vascular resistance caused a decrease in blood flow in the respective organs. The plasma levels of norepinephrine and epinephrine were increased by TRH, while there was no change in plasma renin activity or vasopressin. The cardiovascular actions of i.c.v. TRH were not influenced by blockade of the renin-angiotensin system or vasopressin receptors. The ganglion blocker chlorisondamine and the alpha 1- and alpha 2-adrenoreceptor antagonist phentolamine (2 mg/kg i.v.) abolished the increase in blood pressure and mesenteric vasoconstriction after i.c.v. TRH. Propranolol (2 mg/kg i.v.) blocked the TRH-induced increase in cardiac index, heart rate, and hindquarter blood flow. The hindquarter vasodilation induced by TRH was also blocked by the selective beta 2-adrenoceptor antagonist ICI 188,551 (1 or 2 mg/kg i.v.), while the beta 1-adrenoceptor blocker practolol (10 mg/kg i.v.) had no effect on the hindquarter vasodilation produced by TRH but totally blocked the increase in cardiac index. In adrenal demedullated rats, the systemic hemodynamic effects of i.c.v. TRH were diminished along with the decrease in renal blood flow and increase in renal vascular resistance; however, the increase in hindquarter blood flow was attenuated only in adrenal demedullated rats pretreated with the sympathetic blocker bretylium. The renal vasoconstriction induced by i.c.v. TRH was not abolished by renal denervation. In sinoaortic debuffered rats, the pressor, tachycardic, and mesenteric vasoconstrictor responses to centrally administered TRH were significantly potentiated. Taken together, these data suggest that the putative neurotransmitter TRH may play a role in central regulation of cardiac functions and organ blood flow distribution through both the sympathetic nerves and the adrenal medulla. A pivotal role for beta 2-adrenoceptors in mediation of hindquarter vasodilation is also demonstrated.


Photodynamic therapy for non-melanoma skin cancer.
Photodynamic therapy (PDT) involves the administration of a photosensitizing drug and its subsequent activation by light at wavelengths matching the absorption spectrum of the photosensitizer. Because the skin is readily accessible to light-based therapies, PDT with systemic and particularly with topical agents has become important in treating cutaneous disorders. Topical PDT is indicated for treating actinic keratosis, superficial or thin non-melanoma skin cancer, including some cases of nodular basal cell carcinoma, and some cutaneous lymphomas. Advantages of aminolevulinic acid/methyl aminolevulinate PDT include the possibility of simultaneous treatment of multiple tumors and large surface areas, good cosmesis, and minimal morbidity, such as bleeding, scarring, or infection.


Two-photon induced collagen cross-linking in bioartificial cardiac tissue.
Cardiac tissue engineering is a promising strategy for regenerative therapies to overcome the shortage of donor organs for transplantation. Besides contractile function, the stiffness of tissue engineered constructs is crucial to generate transplantable tissue surrogates with sufficient mechanical stability to withstand the high pressure present in the heart. Although several collagen cross-linking techniques have proven to be efficient in stabilizing biomaterials, they cannot be applied to cardiac tissue engineering, as cell death occurs in the treated area. Here, we present a novel method using femtosecond (fs) laser pulses to increase the stiffness of collagen-based tissue constructs without impairing cell viability. Raster scanning of the fs laser beam over riboflavin-treated tissue induced collagen cross-linking by two-photon photosensitized singlet oxygen production. One day post-irradiation, stress-strain measurements revealed increased tissue stiffness by around 40% being dependent on the fibroblast content in the tissue. At the same time, cells remained viable and fully functional as demonstrated by fluorescence imaging of cardiomyocyte mitochondrial activity and preservation of active contraction force. Our results indicate that two-photon induced collagen cross-linking has great potential for studying and improving artificially engineered tissue for regenerative therapies.


Diversity, Prevalence, and Longitudinal Occurrence of Type II Toxin-Antitoxin Systems of Pseudomonas aeruginosa Infecting Cystic Fibrosis Lungs.
Type II toxin-antitoxin (TA) systems are most commonly composed of two genes encoding a stable toxin, which harms the cell, and an unstable antitoxin that can inactivate it. TA systems were initially characterized as selfish elements, but have recently gained attention for regulating general stress responses responsible for pathogen virulence, formation of drug-tolerant persister cells and biofilms-all implicated in causing recalcitrant chronic infections. We use a bioinformatics approach to explore the distribution and evolution of type II TA loci of the opportunistic pathogen, Pseudomonas aeruginosa, across longitudinally sampled isolates from cystic fibrosis lungs. We identify their location in the genome, mutations, and gain/loss during infection to elucidate their function(s) in stabilizing selfish elements and pathogenesis. We found (1) 26 distinct TA systems, where all isolates harbor four in their core genome and a variable number of the remaining 22 on genomic islands; (2) limited mutations in core genome TA loci, suggesting they are not under negative selection; (3) no evidence for horizontal transmission of elements with TA systems between clone types within patients, despite their ability to mobilize; (4) no gain and limited loss of TA-bearing genomic islands, and of those elements partially lost, the remnant regions carry the TA systems supporting their role in genomic stabilization; (5) no significant correlation between frequency of TA systems and strain ability to establish as chronic infection, but those with a particular TA, are more successful in establishing a chronic infection.


Circadian Disruption and Occupational Toxicants Exposure Affecting the Immunity of Shift Workers During SARS CoV-2 Pandemic.
In several regions of the world, the recent Coronavirus Disease-2019 (COVID-19) pandemic outbreak increased morbidity and mortality. The pandemic situation disrupted many workers' previously established lifestyles. The main aim of the present review was to describe the circadian disruption and occupational toxicant exposure affecting the immunity of shift workers during the SARS CoV-2 pandemic. We retrieved pertinent published literature from the Google Scholar, PubMed, and Scopus databases. In the present review, we discuss the circadian rhythm involving the hypothalamic-pituitary-adrenal (HPA) axis at the molecular level, its disruption, occupational toxicant exposure causing immunomodulatory effects, and the role of immunity during the SARS CoV-2 pandemic. The severity of the progression of the viral infection depends on multiple factors affecting immunity. Hence, shift workers may need to be aware of those factors such as circadian rhythm disruption as well as occupational toxicant exposure. The timing of shift workers' energy intake is also important concerning the shift of the workers. The information in the present review may be important for all workers who are at risk during the pandemic. In the absence of any published literature related to association of circadian rhythm disruption with occupational toxicant exposure, the present review may have greater importance.


Peak and average rectified EMG measures: which method of data reduction should be used for assessing core training exercises?
Core strengthening and stability exercises are fundamental for any conditioning training program. Although surface electromyography (sEMG) is used to quantify muscle activity there is a lack of research using this method to investigate the core musculature and core stability. Two types of data reduction are commonly used for sEMG; peak and average rectified EMG methods. Peak EMG has been infrequently reported in the literature with regard to the assessment of core training while even fewer studies have incorporated average rectified EMG data (ARV). The aim of the study was to establish the repeatability of peak and average rectified EMG data during core training exercises and their interrelationship. Ten male highly trained athletes (inter-subject repeatability group; age, 18 ± 1.2 years; height, 176.5 ± 3.2 cm; body mass, 71 ± 4.5 kg) and one female highly trained athlete (intra-subject repeatability group; age; 27 years old; height; 180 cm; weight; 53 kg) performed five maximal voluntary isometric contractions (MVIC) and five core exercises, chosen to represent a range of movement and muscle recruitment patterns. Peak EMG and ARV EMG were calculated for eight core muscles (rectus abdominis, RA; external oblique, EO; internal oblique, IO; multifidis, MF; latissimus dorsi, LD; longissimus, LG; gluteus maximus, GM; rectus femoris, RF) using sEMG. Average coefficient of variation (CV%) for peak EMG across all the exercises and muscles was 45%. This is in comparison to 35% for the ARV method, which was found to be a significant difference (P<0.05), therefore implying that the ARV method is the more reliable measure for these types of exercise. Analysis of the inter-subject and intra-subject CV% values suggest that these exercises and muscles are sufficiently repeatable using sEMG. Five muscles were highly correlated (R>0.70; RA, EO, MF, GM, LG) between peak and ARV EMG suggesting, that for these core muscles, the two methods provide a similar evaluation of muscle activity. However, for other muscles (IO, RF, LD) the relationship was found to range from poor to moderate (R=0.10-0.70). The relationship between peak and ARV EMG was also affected by exercise type. Dynamic low and high-threshold exercises and asymmetrical low-threshold exercises had a moderate correlation between the variables (R=0.74-0.81), while the static exercise showed a poor correlation (R=0.46). It can be concluded that there are similarities between the two EMG variables, however due to the effect of type of exercise and muscle on the EMG data, both methods should be included in any future EMG study on the core musculature and core stability exercises.


Reducing distress in mothers of children with autism and other disabilities: a randomized trial.
Compared with other parents, mothers of children with autism spectrum disorder or other neurodevelopmental disabilities experience more stress, illness, and psychiatric problems. Although the cumulative stress and disease burden of these mothers is exceptionally high, and associated with poorer outcomes in children, policies and practices primarily serve the identified child with disabilities. A total of 243 mothers of children with disabilities were consented and randomized into either Mindfulness-Based Stress Reduction (mindfulness practice) or Positive Adult Development (positive psychology practice). Well-trained, supervised peer mentors led 6 weeks of group treatments in 1.5-hour weekly sessions, assessing mothers 6 times before, during, and up to 6 months after treatment. Mothers had children with autism (65%) or other disabilities (35%). At baseline, 85% of this community sample had significantly elevated stress, 48% were clinically depressed, and 41% had anxiety disorders. Using slopes-as-outcomes, mixed random effects models, both treatments led to significant reductions in stress, depression, and anxiety, and improved sleep and life satisfaction, with large effects in depression and anxiety. Mothers in Mindfulness-Based Stress Reduction versus Positive Adult Development had greater improvements in anxiety, depression, sleep, and well-being. Mothers of children with autism spectrum disorder improved less in anxiety, but did not otherwise differ from their counterparts. Future studies are warranted on how trained mentors and professionals can address the unmet mental health needs of mothers of children with developmental disabilities. Doing so improves maternal well-being and furthers their long-term caregiving of children with complex developmental, physical, and behavioral needs.


Bacterial growth kinetics under a novel flexible methacrylate dressing serving as a drug delivery vehicle for antiseptics.
A flexible methacrylate powder dressing (Altrazeal®) transforms into a wound contour conforming matrix once in contact with wound exudate. We hypothesised that it may also serve as a drug delivery vehicle for antiseptics. The antimicrobial efficacy and influence on bacterial growth kinetics in combination with three antiseptics was investigated in an in vitro porcine wound model. Standardized in vitro wounds were contaminated with Staphylococcus aureus (MRSA; ATCC 33591) and divided into six groups: no dressing (negative control), methacrylate dressing alone, and combinations with application of 0.02% Polyhexamethylene Biguanide (PHMB), 0.4% PHMB, 0.1% PHMB + 0.1% betaine, 7.7 mg/mL Povidone-iodine (PVP-iodine), and 0.1% Octenidine-dihydrochloride (OCT) + 2% phenoxyethanol. Bacterial load per gram tissue was measured over five days. The highest reduction was observed with PVP-iodine at 24 h to log10 1.43 cfu/g, followed by OCT at 48 h to log10 2.41 cfu/g. Whilst 0.02% PHMB resulted in a stable bacterial load over 120 h to log10 4.00 cfu/g over 120 h, 0.1% PHMB + 0.1% betaine inhibited growth during the first 48 h, with slightly increasing bacterial numbers up to log10 5.38 cfu/g at 120 h. These results indicate that this flexible methacrylate dressing can be loaded with various antiseptics serving as drug delivery system. Depending on the selected combination, an individually shaped and controlled antibacterial effect may be achieved using the same type of wound dressing.


From video recordings to whisker stable isotopes: a critical evaluation of timescale in assessing individual foraging specialisation in Australian fur seals.
Estimating the degree of individual specialisation is likely to be sensitive to the methods used, as they record individuals' resource use over different time-periods. We combined animal-borne video cameras, GPS/TDR loggers and stable isotope values of plasma, red cells and sub-sampled whiskers to investigate individual foraging specialisation in female Australian fur seals (Arctocephalus pusillus doriferus) over various timescales. Combining these methods enabled us to (1) provide quantitative information on individuals' diet, allowing the identification of prey, (2) infer the temporal consistency of individual specialisation, and (3) assess how different methods and timescales affect our estimation of the degree of specialisation. Short-term inter-individual variation in diet was observed in the video data (mean pairwise overlap = 0.60), with the sampled population being composed of both generalist and specialist individuals (nested network). However, the brevity of the temporal window is likely to artificially increase the level of specialisation by not recording the entire diet of seals. Indeed, the correlation in isotopic values was tighter between the red cells and whiskers (mid- to long-term foraging ecology) than between plasma and red cells (short- to mid-term) (R(2) = 0.93-0.73 vs. 0.55-0.41). δ(13)C and δ(15)N values of whiskers confirmed the temporal consistency of individual specialisation. Variation in isotopic niche was consistent across seasons and years, indicating long-term habitat (WIC/TNW = 0.28) and dietary (WIC/TNW = 0.39) specialisation. The results also highlight time-averaging issues (under-estimation of the degree of specialisation) when calculating individual specialisation indices over long time-periods, so that no single timescale may provide a complete and accurate picture, emphasising the benefits of using complementary methods.


Coal combustion related fine particulate matter (PM2.5) induces toxicity in Caenorhabditis elegans by dysregulating microRNA expression.
We employed an in vivo assay system of Caenorhabditis elegans to determine if and which microRNAs (miRNAs) were dysregulated upon exposure to coal combustion related fine particulate matter (PM2.5) by profiling the miRNAs using SOLiD sequencing. From this, expression of 25 miRNAs was discovered to become dysregulated by exposure to PM2.5. Using the corresponding C. elegans deletion mutants, 5 miRNAs (mir-231, mir-232, mir-230, mir-251 and mir-35) were found to be involved in the control of PM2.5 toxicity. Furthermore, mutation of mir-231 or mir-232 induced a resistance to PM2.5 toxicity, whereas mutation of mir-230, mir-251, or mir-35 induced a susceptibility to PM2.5 toxicity. SMK-1, an ortholog of the mammalian SMEK protein, was identified as a molecular target for mir-231 in the regulation of PM2.5 toxicity. In addition, the genes of sod-3, sod-4 and ctl-3, which are necessary for protection against oxidative stress, were determined to be important downstream targets of smk-1 in the regulation of PM2.5 toxicity. The triggering of this mir-231-SMK-1-SOD-3/SOD-4/CTL-3 signaling pathway may be a critical molecular basis for the role of oxidative stress in the induction of coal combustion related PM2.5 toxicity.


Bacillus hisashii sp. nov., isolated from the caeca of gnotobiotic mice fed with thermophile-fermented compost.
A taxonomic study was performed on 15 bacterial isolates from the caeca of gnotobiotic mice that had been fed with thermophile-fermented compost. The 15 isolates were thermophilic, Gram-stain-positive, facultatively anaerobic, endospore-forming bacteria, and were most closely related to Bacillus thermoamylovorans CNCM I-1378T. The 16S rRNA gene sequence of strain N-11T, selected as representative of this new group, showed a similarity of 99.4 % with Bacillus thermoamylovorans CNCM I-1378T, 94.7 % with Bacillus thermolactis R-6488T, and 94.4 % with Bacillus kokeshiiformis MO-04T. The isolates were then classified into two distinct groups based on a (GTG)5-fingerprint analysis. Two isolates, N-11T and N-21, were the representatives of these two groups, respectively.` The N-11T and N-21 isolates showed 66-71 % DNA-DNA relatedness with one other, but had less than 37 % DNA-DNA relatedness with B. thermoamylovorans LMG 18084T. The other 13 isolates showed DNA-DNA relatedness values above 74 % with the N-11T isolate. All 15 isolates grew at 25-60 °C (optimum 50 °C), pH 6-8 (optimum pH 7) and were capable of growing on a medium containing 6 % (w/v) NaCl (optimum 0.5 %). The 15 isolates could be distinguished from B. thermoamylovorans LMG 18084T because they showed Tween 80 hydrolysis activity and did not produce acid from melibiose. The major fatty acids were anteiso-C15 : 0, C16 : 0, iso-C15 : 0, iso-C14 : 0 and iso-C16 : 0. The major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and several unidentified phospholipids. The diagnostic diamino acid in the cell-wall peptidoglycan was meso-diaminopimelic acid. The menaquinone was MK-7. The DNA G+C content was 37.9 mol%. Based on the phenotypic properties, the 15 strains represent a novel species of the genus Bacillus, for which the name Bacillus hisashii sp. nov. is proposed. The type strain is N-11T ( = NRBC 110226T = LMG 28201T).


Genetic transformation of lime (Citrus aurantifolia Swing.): factors affecting transformation and regeneration.
We have previously developed procedures for the efficient production of sweet orange (Citrus sinensis L. Osbeck) and Carrizo citrange (C. sinensis L. Osbeck×Poncirus trifoliata L. Raf.) transgenic plants using an Agrobacterium tumefaciens-mediated transformation and shoot tip grafting in vitro regeneration system. We now report on the optimization of the cocultivation, regeneration and selection conditions for efficient and reliable production of transgenic lime (C. aurantifolia Swing.) plants. Improved transformation frequencies were obtained by cocultivating the explants with Agrobacterium on feeder plates. Optimum regeneration of transgenic shoots was obtained by exposing the explants to darkness for 2 weeks and by using kanamycin at 100 mg/l as selective agent. Attempts to use geneticin as selection antibiotic were not successful. Shoot tip grafting of regenerated shoots on Troyer citrange seedlings resulted in 100% successful production of transgenic plants. The presence and expression of the transferred genes in the regenerated plants was verified by β-glucuronidase histochemical and fluorimetric assays, neomycin phosphotransferase ELISA assays, PCR and Southern analyses.


Congenital Bednar's tumour.
Bednar's tumour (BT) is a rare variant of dermatofibrosarcoma protuberans (DFSP). In addition to the typical histological findings of DFSP, melanin-containing dendritic cells are found in BT. Its occurrence at birth is very rare and there have been only two case reports of congenital BT in the English literature. A 12-year-old boy presented with a single, dome-shaped erythematous nodule, 25 x 15 mm in size, on the lower back. The lesion was present as a depressed patch at birth. Subsequently the lesion grew gradually to a dome-shaped mass. Histopathological examination found a dense infiltration of spindle cells arranged in a storiform pattern in the dermis, with pigmented cells admixed with the spindle cells. Cell mitosis and atypia were rarely seen. On immunohistochemical study, the tumour cells were positive for CD34 and melanin-containing cells were positive for S-100. A diagnosis of BT was made and the mass was removed by wide local excision. There was no evidence of recurrence at follow-up 8 months later.


Transcriptional responses to complex mixtures: a review.
Exposure of people to hazardous compounds is primarily through complex environmental mixtures, those that occur through media such as air, soil, water, food, cigarette smoke, and combustion emissions. Microarray technology offers the ability to query the entire genome after exposure to such an array of compounds, permitting a characterization of the biological effects of such exposures. This review summarizes the published literature on the transcriptional profiles resulting from exposure of cells or organisms to complex environmental mixtures such as cigarette smoke, diesel emissions, urban air, motorcycle exhaust, carbon black, jet fuel, and metal ore and fumes. The majority of the mixtures generally up-regulate gene expression, with heme oxygenase 1 and CYP1A1 being up-regulated by all of the mixtures. Most of the mixtures altered the expression of genes involved in oxidative stress response (OH-1, metallothioneins), immune/inflammation response (IL-1b, protein kinase), xenobiotic metabolism (CYP1A1, CYP1B1), coagulation and fibrinolysis (plasminogen activator/inhibitor), proto-oncogenes (FUS1, JUN), heat-shock response (HSP60, HSP70), DNA repair (PCNA, GADD45), structural unit of condensed DNA (Crf15Orf16, DUSP 15), and extracellular matrix degradation (MMP1, 8, 9, 11, 12). Genes involved in aldehyde metabolism, such as ALDH3, appeared to be uniquely modulated by cigarette smoke. Cigarette smoke-exposed populations have been successfully distinguished from control nonexposed populations based on the expression pattern of a subset of genes, thereby demonstrating the utility of this approach in identifying biomarkers of exposure and susceptibility. The analysis of gene-expression data at the pathway and functional level, along with a systems biology approach, will provide a more comprehensive insight into the biological effects of complex mixtures and will improve risk assessment of the same. We suggest critical components of study design and reporting that will achieve this goal.


Facilitating the implementation of pharmacokinetic-guided dosing of prophylaxis in haemophilia care by discrete choice experiment.
Patients', parents' and providers' preferences with regard to medical innovations may have a major impact on their implementation. To evaluate barriers and facilitators for individualized pharmacokinetic (PK)-guided dosing of prophylaxis in haemophilia patients, parents of young patients, and treating professionals by discrete choice experiment (DCE) questionnaire. The study population consisted of patients with haemophilia currently or previously on prophylactic treatment with factor concentrate (n = 114), parents of patients aged 12-18 years (n = 19) and haemophilia professionals (n = 91). DCE data analysis was performed, taking preference heterogeneity into account. Overall, patients and parents, and especially professionals were inclined to opt for PK-guided dosing of prophylaxis. In addition, if bleeding was consequently reduced, more frequent infusions were acceptable. However, daily dosing remained an important barrier for all involved. 'Reduction of costs for society' was a facilitator for implementation in all groups. To achieve implementation of individualized PK-guided dosing of prophylaxis in haemophilia, reduction of bleeding risk and reduction of costs for society should be actively discussed as they are motivating for implementation; daily dosing is still reported to be a barrier for all groups. The knowledge of these preferences will enlarge support for this innovation, and aid in the drafting of implementable guidelines and information brochures for patients, parents and professionals.


Deleterious Outcomes in Long-Hauler COVID-19: The Effects of SARS-CoV-2 on the CNS in Chronic COVID Syndrome.
Amid our understanding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the mechanisms involved in the causation of acute-phase coronavirus disease (COVID-19), we have come across clinical cases that have been shown to run a protracted course of COVID-19 with complex clinical findings related to organ systems in general and the CNS in particular that deserve to be addressed in the COVID long-haulers, for which the more clinically-related term chronic COVID syndrome (CCS) has been coined recently. An in-depth understanding of the mechanism that forms the basis of CCS and neurological deficits in CCS is needed as this can help in determining the management of cases of neuro-COVID, which is emerging as a less lethal but more disabling disease state. This Viewpoint highlights this syndrome, the possible pathogenetic pathways involved, and the treatment approaches that can be taken to help manage COVID long-haulers in CCS.


Dermatoses caused by cultural practices: Therapeutic cultural practices.
With globalization and widespread immigration, physicians increasingly encounter patients from varying backgrounds and diverse customs. Although certain cultural practices are widely performed, there is limited medical literature describing their dermatologic and systemic effects and complications. Population diversity and sharing of traditions make it increasingly important for dermatologists to understand the role of cultural practices and recognize physiologic and pathologic sequelae. In addition, dermatologists are often adjured to assess skin findings that may be mistaken for abuse. Child abuse misdiagnosis can be traumatizing to all those involved, and immigrant families with limited English proficiency may have difficulty explaining their traditional practices. The first article of this 2-part continuing medical education series begins with a review of therapeutic cultural practices, including traditional Chinese medicine, Ayurveda, acupuncture, cupping, moxibustion, and coining, and the clinically relevant complications that may occur. Therapeutic practices can cause a range of complications, including contact dermatitis, heavy metal toxicity, and severe cutaneous adverse reactions.


A Multifaceted Intervention to Improve Patient Knowledge and Safe Use of Opioids: Results of the ED EMC2 Randomized Controlled Trial.
Despite increased focus on opioid prescribing, little is known about the influence of prescription opioid medication information given to patients in the emergency department (ED). The study objective was to evaluate the effect of an Electronic Medication Complete Communication (EMC2 ) Opioid Strategy on patients' safe use of opioids and knowledge about opioids. This was a three-arm prospective, randomized controlled pragmatic trial with randomization occurring at the physician level. Consecutive discharged patients at an urban academic ED (>88,000 visits) with new hydrocodone-acetaminophen prescriptions received one of three care pathways: 1) usual care, 2) EMC2 intervention, or 3) EMC2 + short message service (SMS) text messaging. The ED EMC2 intervention triggered two patient-facing educational tools (MedSheet, literacy-appropriate prescription wording [Take-Wait-Stop]) and three provider-facing reminders to counsel (directed to ED physician, dispensing pharmacist, follow-up physician). Patients in the EMC2 + SMS arm additionally received one text message/day for 1 week. Follow-up at 1 to 2 weeks assessed "demonstrated safe use" (primary outcome). Secondary outcomes including patient knowledge and actual safe use (via medication diaries) were assessed 2 to 4 days and 1 month following enrollment. Among the 652 enrolled, 343 completed follow-up (57% women; mean ± SD age = 42 ± 14.0 years). Demonstrated safe opioid use occurred more often in the EMC2 group (adjusted odds ratio [aOR] = 2.46, 95% confidence interval [CI] = 1.19 to 5.06), but not the EMC2 + SMS group (aOR = 1.87, 95% CI = 0.90 to 3.90) compared with usual care. Neither intervention arm improved medication safe use as measured by medication diary data. Medication knowledge, measured by a 10-point composite knowledge score, was greater in the EMC2 + SMS group (β = 0.57, 95% CI = 0.09 to 1.06) than usual care. The study found that the EMC2 tools improved demonstrated safe dosing, but these benefits did not translate into actual use based on medication dairies. The text-messaging intervention did result in improved patient knowledge.


The effects of Red Bull energy drink compared with caffeine on cycling time-trial performance.
This study investigated the ergogenic effects of a commercial energy drink (Red Bull) or an equivalent dose of anhydrous caffeine in comparison with a noncaffeinated control beverage on cycling performance. Eleven trained male cyclists (31.7 ± 5.9 y 82.3 ± 6.1 kg, VO2max = 60.3 ± 7.8 mL · kg-1 · min-1) participated in a double-blind, placebo-controlled, crossover-design study involving 3 experimental conditions. Participants were randomly administered Red Bull (9.4 mL/kg body mass [BM] containing 3 mg/kg BM caffeine), anhydrous caffeine (3 mg/kg BM given in capsule form), or a placebo 90 min before commencing a time trial equivalent to 1 h cycling at 75% peak power output. Carbohydrate and fluid volumes were matched across all trials. Performance improved by 109 ± 153 s (2.8%, P = .039) after Red Bull compared with placebo and by 120 ± 172 s (3.1%, P = .043) after caffeine compared with placebo. No significant difference (P > .05) in performance time was detected between Red Bull and caffeine treatments. There was no significant difference (P > .05) in mean heart rate or rating of perceived exertion among the 3 treatments. This study demonstrated that a moderate dose of caffeine consumed as either Red Bull or in anhydrous form enhanced cycling time-trial performance. The ergogenic benefits of Red Bull energy drink are therefore most likely due to the effects of caffeine, with the other ingredients not likely to offer additional benefit.


Renal function is the same 6 months after robot-assisted partial nephrectomy regardless of clamp technique: analysis of outcomes for off-clamp, selective arterial clamp and main artery clamp techniques, with a minimum follow-up of 1 year.
To compare the renal functional outcomes, with >1 year of follow-up, of patients who underwent robot-assisted partial nephrectomy (RAPN) performed with different clamping techniques. The peri-operative data of patients undergoing RAPN performed with different clamping techniques were retrospectively analysed (group 1: off-clamp, n = 23; group 2: selective clamp, n = 25; group 3: main artery clamp, n = 114). The main outcome measures were postoperative serum creatinine level, estimated glomerular filtration rate (eGFR) and percentage change in eGFR, the data for which were collected at periodic intervals during the first 12 months and annually thereafter, in addition to late eGFR value. Only patients with >1 year of follow-up were included in the analysis. The baseline characteristics of groups 2 and 3 were similar, while patients in group 1 had smaller sized tumours and lower tumour complexity. The median follow-up periods were 45 (group 1), 20 (group 2) and 47 (group 3) months. The median clamping times were 24.8 min in the main artery clamp and 18 min in the selective artery clamp groups. Group 2 had greater median blood loss volume (100 vs 500 vs 200 mL for groups 1, 2 and 3, respectively; P < 0.01) and a longer length of hospital stay (3 vs 4 vs 3 days for groups 1, 2 and 3, respectively; P = 0.02). No significant differences were found among the groups with regard to transfusion rates, positive surgical margin rates, complications, recurrence or mortality rates. Groups 1 and 2 had significantly less deterioration of postoperative renal function during the first 3 months after surgery (P = 0.04; percent change in eGFR -1.5, -2 and -8% for groups 1, 2 and 3, respectively), but this beneficial outcome was not observed after 6 months or for the latest eGFR measurement (P = 0.48; latest percent change in eGFR -3, -6 and -3.5% for groups 1, 2 and 3, respectively). In regression analysis, baseline eGFR, type of clamp procedure and tumour complexity score were predictive of normal renal function 7 days after surgery, while only baseline eGFR and age could predict it 1 year postoperatively. Off-clamp and selective artery clamp techniques result in superior short-term renal functional outcomes compared with the main artery clamp approach; however, after the 6th postoperative month, there were no significant differences regarding the functional outcome among the above surgical techniques, as long as the warm ischaemia time was 20-30 min.


Deep Proteomics Network and Machine Learning Analysis of Human Cerebrospinal Fluid in Japanese Encephalitis Virus Infection.
Japanese encephalitis virus is a leading cause of neurological infection in the Asia-Pacific region with no means of detection in more remote areas. We aimed to test the hypothesis of a Japanese encephalitis (JE) protein signature in human cerebrospinal fluid (CSF) that could be harnessed in a rapid diagnostic test (RDT), contribute to understanding the host response and predict outcome during infection. Liquid chromatography and tandem mass spectrometry (LC-MS/MS), using extensive offline fractionation and tandem mass tag labeling (TMT), enabled comparison of the deep CSF proteome in JE vs other confirmed neurological infections (non-JE). Verification was performed using data-independent acquisition (DIA) LC-MS/MS. 5,070 proteins were identified, including 4,805 human proteins and 265 pathogen proteins. Feature selection and predictive modeling using TMT analysis of 147 patient samples enabled the development of a nine-protein JE diagnostic signature. This was tested using DIA analysis of an independent group of 16 patient samples, demonstrating 82% accuracy. Ultimately, validation in a larger group of patients and different locations could help refine the list to 2-3 proteins for an RDT. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD034789 and 10.6019/PXD034789.


The "GEnomics of Musculo Skeletal Traits TranslatiOnal NEtwork": Origins, Rationale, Organization, and Prospects.
Musculoskeletal research has been enriched in the past ten years with a great wealth of new discoveries arising from genome wide association studies (GWAS). In addition to the novel factors identified by GWAS, the advent of whole-genome and whole-exome sequencing efforts in family based studies has also identified new genes and pathways. However, the function and the mechanisms by which such genes influence clinical traits remain largely unknown. There is imperative need to bring multidisciplinary expertise together that will allow translating these genomic discoveries into useful clinical applications with the potential of improving patient care. Therefore "GEnomics of MusculoSkeletal traits TranslatiOnal NEtwork" (GEMSTONE) aims to set the ground for the: 1) functional characterization of discovered genes and pathways; 2) understanding of the correspondence between molecular and clinical assessments; and 3) implementation of novel methodological approaches. This research network is funded by The European Cooperation in Science and Technology (COST). GEMSTONE includes six working groups (WG), each with specific objectives: WG1-Study populations and expertise groups: creating, maintaining and updating an inventory of experts and resources (studies and datasets) participating in the network, helping to assemble focus groups defined by phenotype, functional and methodological expertise. WG2-Phenotyping: describe ways to decompose the phenotypes of the different functional studies into meaningful components that will aid the interpretation of identified biological pathways. WG3 Monogenic conditions - human KO models: makes an inventory of genes underlying musculoskeletal monogenic conditions that aids the assignment of genes to GWAS signals and prioritizing GWAS genes as candidates responsible for monogenic presentations, through biological plausibility. WG4 Functional investigations: creating a roadmap of genes and pathways to be prioritized for functional assessment in cell and organism models of the musculoskeletal system. WG5 Bioinformatics seeks the integration of the knowledge derived from the distinct efforts, with particular emphasis on systems biology and artificial intelligence applications. Finally, WG6 Translational outreach: makes a synopsis of the knowledge derived from the distinct efforts, allowing to prioritize factors within biological pathways, use refined disease trait definitions and/or improve study design of future investigations in a potential therapeutic context (e.g. clinical trials) for musculoskeletal diseases.


Dexamethasone inhibits receptor-activated phosphoinositide breakdown in rat basophilic leukemia (RBL-2H3) cells.
The activation of rat basophilic leukemia cells for histamine release is accompanied by Ca2+ influx and arachidonic acid release. IgE receptor but not A23187 ionophore stimulation of these cells also resulted in phosphoinositide breakdown. In these experiments, the culture of these cells with dexamethasone inhibited IgE- and ionophore-mediated histamine release. The concentration for 50% of maximal inhibition was 12 nM, and prolonged exposure to the drug was required, with maximal effect observed in 8 to 15 hr. The inhibitory effect of dexamethasone was reversible (t1/2 for recovery was 16 hr). Dexamethasone blocked the IgE-mediated 45Ca2+ influx and the release of [14C]-arachidonic acid (IC50 of 1 nM and 10 nM respectively). Dexamethasone inhibited the IgE receptor-mediated phosphoinositide breakdown (IC50 of 5 nM). It also decreased arachidonic acid release after A23187 stimulation demonstrating an effect on phospholipase A2. Therefore, exposure of the cells to dexamethasone results in the inhibition of both phospholipase A2 and phospholipase C pathways of arachidonic acid generation.


Effects of Paraquat Ban on Herbicide Poisoning-Related Mortality.
In Korea, registration of paraquat-containing herbicides was canceled in November 2011, and sales thereof were completely banned in November 2012. We evaluated the effect of the paraquat ban on the epidemiology and mortality of herbicide-induced poisoning. This retrospective study analyzed patients treated for herbicide poisoning at 17 emergency departments in South Korea between January 2010 and December 2014. The overall and paraquat mortality rates were compared pre- and post-ban. Factors associated with herbicide mortality were evaluated using logistic analysis. To determine if there were any changes in the mortality rates before and after the paraquat sales ban and the time point of any such significant changes in mortality, R software, version 3.0.3 (package, bcp) was used to perform a Bayesian change point analysis. We enrolled 2257 patients treated for herbicide poisoning (paraquat=46.8%). The overall and paraquat poisoning mortality rates were 40.6% and 73.0%, respectively. The decreased paraquat poisoning mortality rate (before, 75% vs. after, 67%, p=0.014) might be associated with increased intentionality. The multivariable logistic analysis revealed the paraquat ban as an independent predictor that decreased herbicide poisoning mortality (p=0.035). There were two major change points in herbicide mortality rates, approximately 3 months after the initial paraquat ban and 1 year after complete sales ban. This study suggests that the paraquat ban decreased intentional herbicide ingestion and contributed to lowering herbicide poisoning-associated mortality. The change point analysis suggests a certain timeframe was required for the manifestation of regulatory measures outcomes.


Insulinaemic and glycaemic responses to three forages in ponies.
Reduction of the hyperinsulinaemic response to feeding is central to the management of insulin dysregulation (ID). The aim of this study was to compare insulinaemic and glycaemic responses to soaked hay, dry hay and haylage in ponies. Twelve ponies of mixed breeds were maintained under identical management conditions. A randomised four-way crossover trial was conducted, in which fasted animals were fed a meal of 0.25% body weight as dry matter intake soaked hay, dry hay or haylage, or administered an oral glucose test (OGT). Blood glucose and serum insulin concentrations were measured before and at 2h following OGT, and regularly for 5h following forage meals. Median and interquartile range (IQR) area under the curve (AUC) for insulin (AUCi) was greater for haylage (median 6495; IQR 17352) vs. dry hay (2932; IQR 5937; P=0.019) and soaked hay (1066; IQR 1753; P=0.002), and greater for dry hay vs. soaked hay (P=0.002). The AUC for glucose (AUCg) was lower for soaked hay (1021; IQR 99) vs. dry hay (1075; IQR 105; P=0.002) and haylage (1107; IQR 221; P=0.003). Six ponies were classified as having ID based on the OGT. AUCi was greater in ID vs. non-ID ponies after all forages. In contrast, there was no detectable effect of ID status on AUCg. On an equivalent dry matter basis, soaked hay produced the lowest insulinaemic and glycaemic responses to feeding, while haylage produced the highest responses. The insulinaemic effects of all forages were greater in ponies with ID. These data support the practice of soaking hay with water to reduce postprandial insulinaemic responses in ponies.