CD38 Deficiency Protects Heart from High Fat Diet-Induced Oxidative Stress Via Activating Sirt3/FOXO3 Pathway.
Previous studies showed that CD38 deficiency protected heart from ischemia/reperfusion injury and high fat diet (HFD)-induced obesity in mice. However, the role of CD38 in HFD-induced heart injury remains unclear. In the present study, we have investigated the effects and mechanisms of CD38 deficiency on HFD-induced heart injury. The metabolites in heart from wild type (WT) and CD38 knockout (CD38-/-) mice were examined using metabolomics analysis. Cell viability, lactate hydrogenase (LDH) release, super oxide dismutase (SOD) activity, reactive oxygen species (ROS) production, triglyceride concentration and gene expression were examined by biochemical analysis and QPCR. Our results revealed that CD38 deficiency significantly elevated the intracellular glutathione (GSH) concentration and GSH/GSSG ratio, decreased the contents of free fatty acids and increased intracellular NAD+ level in heart from CD38-/- mice fed with HFD. In addition, in vitro knockdown of CD38 significantly attenuated OA-induced cellular injury, ROS production and lipid synthesis. Furthermore, the expression of mitochondrial deacetylase Sirt3 as well as its target genes FOXO3 and SOD2 were markedly upregulated in the H9C2 cell lines after OA stimulation. In contrast, the expressions of NOX2 and NOX4 were significantly decreased in the cells after OA stimulation. Our results demonstrated that CD38 deficiency protected heart from HFD-induced oxidative stress via activating Sirt3/FOXO3-mediated anti-oxidative stress pathway.


The effects of evening bright light exposure on subsequent morning exercise performance.
We investigated the effects of evening bright light on the circadian timing of core temperature and morning exercise performance under conditions of high thermal stress. At 20:00 h, 8 males were exposed to a standardised light protocol and thereafter to either polychromatic bright light (2,500 lux at 50 cm, BL) or no light (0 lux, NL) for 30 min. The following morning, intermittent cycling exercise was undertaken followed by a 10 km time-trial in an environmental chamber set to 35°C and 60% relative humidity. Core body temperature was measured throughout. Data were analysed using a within-subjects model and presented as mean±SD. Time of the sleep-trough in core temperature occurred ~1.75 h later following BL (P=0.07). Prior to time-trial, core temperature was 0.27±0.42°C lower in BL (95%CI: -0.02 to 0.57, P=0.07). The time-trial was completed 1.43±0.63 min (0.98-1.87) faster in BL (P=0.001). Post time-trial, intestinal temperature was 38.21±0.56°C (37.84-38.57) in BL compared to 38.64±0.42°C (38.34-38.93) in NL (P=0.10). These data provide the first evidence that a 30-min exposure to bright light prior to sleep can influence exercise performance under hot conditions during the subsequent early morning.


Submitogenic concentrations of anti-CD3 monoclonal antibody exert antiapoptotic effects in preactivated CD4+ but not CD8+ human T cells.
Immunological memory has been ascribed to the presence of long-lived memory cells. The mechanisms underlying their generation are not completely understood, but dependence on antigen persistence has been discussed in this regard. However, in spite of in vivo evidence favoring this model, studies on TCR/CD3 stimulation of T cell lines or unseparated peripheral blood T cell in vitro have failed to demonstrate prolonged survival of preactivated cells. We have examined the dose-dependent effect of TCR/CD3 engagement mimicked by immobilized anti-CD3 antibody. To this end, well-defined populations of CD4+ and CD8+ lymphoblasts isolated from bulk cultures of preactivated PBMCs by flow sorting were examined. These cells were restimulated with immobilized anti-CD3 in the presence or absence of various costimulatory factors, and were analyzed for their viability state, as well as their apoptotic and proliferative behavior. We have shown that inhibition of apoptosis following CD3 stimulation occurs at submitogenic concentrations, while activation-driven apoptosis requires high-density TCR/CD3 activation. Prevention of apoptosis by submitogenic CD3 stimulation was, however, observed only when CD4+ but not when CD8+ cells were investigated, and was not readily influenced by other costimulatory factors present in cultures. This observation points to the importance of antigen persistence in regulating survival of memory CD4+ but not CD8+ cells.


2-Hydroxyglutarate Inhibits ATP Synthase and mTOR Signaling.
We discovered recently that the central metabolite α-ketoglutarate (α-KG) extends the lifespan of C. elegans through inhibition of ATP synthase and TOR signaling. Here we find, unexpectedly, that (R)-2-hydroxyglutarate ((R)-2HG), an oncometabolite that interferes with various α-KG-mediated processes, similarly extends worm lifespan. (R)-2HG accumulates in human cancers carrying neomorphic mutations in the isocitrate dehydrogenase (IDH) 1 and 2 genes. We show that, like α-KG, both (R)-2HG and (S)-2HG bind and inhibit ATP synthase and inhibit mTOR signaling. These effects are mirrored in IDH1 mutant cells, suggesting a growth-suppressive function of (R)-2HG. Consistently, inhibition of ATP synthase by 2-HG or α-KG in glioblastoma cells is sufficient for growth arrest and tumor cell killing under conditions of glucose limitation, e.g., when ketone bodies (instead of glucose) are supplied for energy. These findings inform therapeutic strategies and open avenues for investigating the roles of 2-HG and metabolites in biology and disease.


Nonevent stress contributes to mental health disparities based on sexual orientation: evidence from a personal projects analysis.
This study examined the role of nonevent stress--in the form of frustrated personal project pursuits in the arenas of relationships and work--as a contributing factor to mental health disparities between heterosexual and lesbian, gay, and bisexual (LGB) populations. A purposive sample of 431 LGB (55%) and heterosexually identified (45%) individuals living in the United States and Canada completed the Personal Project Inventory by describing and rating core personal projects they were pursuing. The intensity of perceived barriers to the achievement of relationship- and work-related personal projects served as indicators nonevent stress. Hierarchical linear regression models tested the hypothesis that nonevent stress contributes to the association between sexual orientation and two indicators of mental health: depressive symptoms and psychological well-being. LGB individuals had significantly more depressive symptoms and lower levels of psychological well-being than heterosexuals. Indicators of nonevent stress were significantly associated with mental health outcomes and their inclusion in models attenuated sexual orientation differences in mental health. The critical indirect pathway leading from sexual minority status to mental health occurred via barriers to relationship projects from interpersonal sources. This research suggests that nonevent stress because of structural and interpersonal stigma may contribute to mental health disparities between LGB and heterosexual individuals. The findings have important implications for policy reform around same-sex relationship recognition and workplace discrimination. Future research and clinical work will benefit by expanding existing foci on stress to include nonevent stressors to better understand and address mental health problems, particularly in LGB populations.


Revealing the environmental pollution of two estuaries through histopathological biomarkers in five fishes from different trophic guilds of northeastern Brazil.
Estuaries in Brazil are mostly anthropically affected due to the discharge of industrial and domestic effluents. In two of them, the Santa Cruz Channel Estuary (ITAP) and Sirinhaém River Estuary (SIR), historically affected by mercury pollution and sugarcane industry in Northeast Brazil, we assessed environmental pollution using liver and gill histopathological biomarkers in fish from different trophic levels. Liver samples exhibited serious damages such as hepatic steatosis, necrosis, and infiltration. The gills showed moderate to severe changes, such as lifting of epithelial cells, lamellar aneurysm, and rupture of lamellar epithelium. Most of the changes in the liver and gills were reported for species Centropomus undecimalis and the Gobionellus stomatus, which were considered as good sentinels of pollution. The combination of biomarker methodologies was efficient in diagnosing the serious damage to the species, reinforcing the need for monitoring the health of the ecosystems evaluated.


Collagen-rich stroma in aggressive colon tumors induces mesenchymal gene expression and tumor cell invasion.
Gene expression-based classification systems have identified an aggressive colon cancer subtype with mesenchymal features, possibly reflecting epithelial-to-mesenchymal transition (EMT) of tumor cells. However, stromal fibroblasts contribute extensively to the mesenchymal phenotype of aggressive colon tumors, challenging the notion of tumor EMT. To separately study the neoplastic and stromal compartments of colon tumors, we have generated a stroma gene filter (SGF). Comparative analysis of stromahigh and stromalow tumors shows that the neoplastic cells in stromahigh tumors express specific EMT drivers (ZEB2, TWIST1, TWIST2) and that 98% of differentially expressed genes are strongly correlated with them. Analysis of differential gene expression between mesenchymal and epithelial cancer cell lines revealed that hepatocyte nuclear factor 4α (HNF4α), a transcriptional activator of intestinal (epithelial) differentiation, and its target genes are highly expressed in epithelial cancer cell lines. However, mesenchymal-type cancer cell lines expressed only part of the mesenchymal genes expressed by tumor-derived neoplastic cells, suggesting that external cues were lacking. We found that collagen-I dominates the extracellular matrix in aggressive colon cancer. Mimicking the tumor microenvironment by replacing laminin-rich Matrigel with collagen-I was sufficient to induce tumor-specific mesenchymal gene expression, suppression of HNF4α and its target genes, and collective tumor cell invasion of patient-derived colon tumor organoids. The data connect collagen-rich stroma to mesenchymal gene expression in neoplastic cells and to collective tumor cell invasion. Targeting the tumor-collagen interface may therefore be explored as a novel strategy in the treatment of aggressive colon cancer.


Female odours promote the activation of sheltered kissing bug Rhodnius prolixus males and modulate their orientation.
The existence of a pheromone emitted during copulation has been reported for Rhodnius prolixus (Hemiptera: Reduviidae) Stål. Adults possess one pair of metasternal glands (MGs) from which female R. prolixus release volatiles mainly at night. We investigated whether these volatiles emitted by adult R. prolixus can modulate sexual-related behaviours of opposite and/or same sex individuals. We first used a shelter bioassay to test if adult activity patterns can be affected by chemical signals emitted by opposite sex conspecifics. We observed that males left the shelter more frequently in the presence of females, showing higher activity and an increase of intrasexual copulation attempts. Females showed no shelter-related activation in the presence of male odours. Second, we used a locomotion compensator device to investigate whether females or males show oriented responses to odours emitted by adults. We found that males oriented towards air currents carrying female odours but females did not orient towards odours emitted by adults. Finally, we observed that males oriented towards the female MG compounds. Thus, a volatile chemical signal emitted by females from their MGs promoted the activation of sheltered males and modulated orientation to air currents.


Fabrication of fibrin based electrospun multiscale composite scaffold for tissue engineering applications.
Fabricating scaffolds mimicking the native extracellular matrix (ECM) in both structure and function is a key challenge in the field of tissue engineering. Previously we have demonstrated a novel electrospinnig method for the fabrication of fibrin nanofibers using Poly(vinyl alcohol) (PVA) as an 'electrospinning-driving' polymer. Here we demonstrate the fabrication and characterization of a multiscale fibrin based composite scaffold with polycaprolactone (PCL) by sequential electrospinning of PCL microfibers and fibrin nanofibers. This multiscale scaffold has great potential for tissue engineering applications due to the combined benefits of biological nanofibers such as cell attachment and proliferation and that of microfibers such as open structure, larger pore size and adequate mechanical strength. Physico chemical characterization of the electrospun scaffold was done by Scanning Electron Microscopy (SEM), Contact angle analysis, fibrin specific Phosphotungstic acid haematoxyllin (PTAH) staining and evaluation of mechanical properties. SEM data revealed the formation of bead free nanofibers of fibrin with a fiber diameter ranging from 50-500 nm and microfibers of PCL in the size range of 1 microns to 2.5 microns. These dimensions mimic the hierarchical structure of ECM found in native tissues. Cell attachment and viability studies using human mesenchymal stem cells (hMSC) revealed that the scaffold is non toxic and supports cell attachment, spreading and proliferation. In addition, we examined the inflammatory potential of the scaffold to demonstrate its usefulness in tissue engineering applications.


Enlargement of halloysite clay nanotube lumen by selective etching of aluminum oxide.
Halloysite clay tubes have 50 nm diameter and chemically different inner and outer walls (inner surface of aluminum oxide and outer surface of silica). Due to this different chemistry, the selective etching of alumina from inside the tube was realized, while preserving their external diameter (lumen diameter changed from 15 to 25 nm). This increases 2-3 times the tube lumen capacity for loading and further sustained release of active chemical agents such as metals, corrosion inhibitors, and drugs. In particular, halloysite loading efficiency for the benzotriazole increased 4 times by selective etching of 60% alumina within the tubes' lumens. Specific surface area of the tubes increased over 6 times, from 40 to 250 m(2)/g, upon acid treatment.


Crystallization and preliminary X-ray characterization of tobacco streak virus and a proteolytically modified form of the capsid protein.
Isolated tobacco streak virus strain mild (TSV-M) top nucleoprotein component (TV) was crystallized by vapor diffusion with polyethylene glycol (PEG) and methyl pentanediol. The morphology of the crystal suggested a hexagonal space group, however, the crystals were disordered as analyzed by X-ray diffraction. Capsid protein from TSV (strains M and white clover) was treated with trypsin to remove 87 amino terminal residues. The modified capsid protein was purified by ion exchange chromatography and crystallized in both hexagonal and triclinic forms. Hexagonal crystals grown by vapor diffusion in PEG diffract X rays to 4.5 A. The crystals, in the space group P6(2) or its enantiomorph, possess unit cell dimensions of a = 98.3 A, c = 108.7 A and probably contain 12 dimers/unit cell. Triclinic crystals grown by vapor diffusion in ammonium sulfate diffracted X rays to 2.4 A resolution when exposed to X-ray synchrotron radiation. The crystals have unit cell dimensions of a = 60.1 A, b = 77.5 A, c = 73.9 A with alpha = 67.8 degrees, beta = 130 degrees, and gamma = 106 degrees and probably contain 4 dimers/unit cell.


Potential benefit from using the alpha(s1)-casein genotype information in a selection scheme for dairy goats.
A stochastic approach is proposed to predict responses to selection when using alpha(s1)-casein genotype information in a selection scheme of a Spanish breed of dairy goats. Two independent selection objectives were considered: protein yield (PY), where the major additive gene CSN1S1, which codes for alpha(s1)-casein, has a small effect, and protein content (P%), where this gene has a large effect on performances. Significant differences in response between using and ignoring information on the major gene were observed only when the major gene has a large effect. The main result was in the case of P%, the total genetic gain obtained in the early generations of selection was maintained in the long-term. Taking account of genotype information either in the evaluation model or in the selection criteria leads to a faster fixation of the favourable allele and a reduction of the total genetic variance over generations. The inbreeding rates varied across generations, the highest rates observed in later generations of selection and when the major gene has a large effect and its genotype was included in the genetic evaluation procedure. It is concluded that inclusion of the casein genotype as an additional selection criteria will improve gains for protein traits, in particular P%. Recommendations are also given in order to optimize the use of this molecular information in dairy goat selection programs.


Inhibition of carcinoma cell motility by epoxyeicosatrienoic acid (EET) antagonists.
Cytochrome P450 (CYP) epoxygenases, CYP2C8, 2C9 and 2J2 mRNA and proteins, were expressed in prostate carcinoma (PC-3, DU-145 and LNCaP) cells. 11,12-Epoxyeicosatrienoic acid (11,12-EET) was the major arachidonic acid metabolite in these cells. Blocking EET synthesis by a selective CYP epoxygenase inhibitor (N-methylsulfonyl-6-(2-propargyloxyphenyl)hexanamide [MS-PPOH]) inhibited tonic (basal) invasion and migration (motility) while exogenously added EET induced cell motility in a concentration-dependent manner. An epidermal growth factor receptor (EGFR) kinase inhibitor (AG494) or a PI3 kinase inhibitor (LY294002) inhibited cell migration and reduced 11,12-EET-induced cell migration. Importantly, synthetic EET antagonists (14,15-epoxyeicosa-5(Z)-enoic acid [14,15-EEZE], 14,15-epoxyeicosa-5(Z)-enoic acid 2-[2-(3-hydroxy-propoxy)-ethoxy]-ethyl ester [14,15-EEZE-PEG] and 14,15-epoxyeicosa-5(Z)-enoic-methylsulfonylimide [14,15-EEZE-mSI]) inhibited EET-induced cell invasion and migration. 11,12-EET induced cell stretching and myosin-actin microfilament formation as well as increased phosphorylation of EGFR and Akt (Ser473), while 14,15-EEZE inhibited these effects. These results suggest that EET induce and EET antagonists inhibit cell motility, possibly by putative EET receptor-mediated EGFR and PI3K/Akt pathways, and suggest that EET antagonists are potential therapeutic agents for prostate cancer.


Effect of chronic hypoxia on glucose transporters in heart and skeletal muscle of immature and adult rats.
Glucose transporter (GLUT) modulation can be an important mechanism that contributes to adaptation to hypoxic stress, but little is known about GLUT modulation in heart and skeletal muscle with prolonged hypoxia. In this work, the effect of chronic hypoxia on GLUT-4 and GLUT-1 mRNA and protein was studied in these two tissues in the adult and during development. Hypoxia (fractional inspired O2 = 9 +/- 0.5%) was administered to two groups, i.e., an immature group exposed from 3 to 30 days of age and an adult group exposed from 90 to 120 days of age. Rats were then killed and their heart and skeletal muscles were sampled for measurements of GLUT mRNA and protein with Northern and Western blots. In the adult, chronic hypoxia significantly decreased cardiac GLUT mRNA level by > 25% of control (P < 0.05), but had little effect on GLUT protein. A very different hypoxic effect was seen in the immature rat heart with a major increase in protein and no appreciable change in mRNA density. Adult skeletal muscle had no change in GLUT mRNA level but GLUT protein increased (15-20%, P < 0.05) while both GLUT mRNA and protein were significantly increased in the immature skeletal muscles (60-90% over control). We conclude that during chronic O2 deprivation, GLUT-1 and GLUT-4 expressions show a similar pattern but greatly depend on tissue type and age. These differences in GLUT regulation may be due to different strategies for coping with prolonged O2 deprivation in both immature and adult animals.


Knowledge, motivations, expectations, and traits of an African, African-American, and Afro-Caribbean sequencing cohort and comparisons to the original ClinSeq® cohort.
Racial minority populations are underrepresented in genomics research. This study enrolled African-descended individuals in a sequencing study and reported their characteristics. We purposively recruited 467 individuals self-identified as African, African American, or Afro-Caribbean to the ClinSeq® study and surveyed them about knowledge, motivations, expectations, and traits. Summary statistics were calculated and compared with data from the study's original cohort, which was primarily White and self-referred. Recruitment took five years and 83% of enrollees completed the survey. Participants had modest knowledge about benefits and limitations of sequencing (x̅s = 5.1, ranges: 0-10), and less than the original cohort (x̅ = 7.5 and 7.7, respectively). Common motivations to enroll were learning information relevant to personal health (49%) or family members' health (33%), and most had realistic expectations of sequencing. Like the original cohort, they had high levels of optimism, openness, and resilience. Early adopters may have relatively consistent personality traits irrespective of majority/minority status and recruitment methods, but high levels of genomics knowledge are not universal. Research should determine whether recruitment and consent procedures provide adequate education to promote informed choices and realistic expectations, which are vital to ethical research and increasing genomics research participation in underrepresented communities.


Preoperative Radiographic Parameters to Predict a Higher Pseudarthrosis Rate After Anterior Cervical Discectomy and Fusion.
Retrospective study. To determine whether postoperative pseudarthrosis can be predicted from specific preoperative radiograph measurements. Various factors reportedly influence the occurrence of pseudarthrosis after anterior cervical discectomy and fusion (ACDF). However, to our knowledge, there are no reports on the relationships between preoperative radiographic parameters and pseudarthrosis. We analyzed 84 consecutive patients (45 males, 39 females, mean age, 58.9 ± 11.2 yrs) who underwent ACDF. In all patients, allografts filled with local chip bone were inserted after discectomy and anterior plating was performed. On preoperative plain radiographs, we analyzed C2-C7 sagittal vertical axis, T1 sagittal slope, segmental motion, global cervical motion, and location of fusion segments. Pseudarthrosis was diagnosed as interspinous motion >1 mm with superjacent interspinous motion ≥4 mm on magnified dynamic lateral radiographs. Multivariate logistic regression was used to analyze the risk factors for pseudarthrosis and the receiver operating characteristic (ROC) curve was used to define a cutoff value. One hundred and twenty-five segments from 84 patients were included. The pseudarthrosis rate was 29% based on number of patients (24/84) and 20% based on number of segments (25/125). Multilevel surgery and segments at the lowest levels showed higher pseudarthrosis rates (P = 0.01). Per multivariate logistic regression analysis, greater preoperative segmental motion, greater preoperative T1 sagittal slope, and C6-7 segments were associated with a higher risk of pseudarthrosis (all P < 0.05). A segmental motion cutoff value of 12° demonstrated pseudarthrosis with sensitivity of 87%, specificity of 84%, and area under the curve of 0.899, indicating moderate accuracy. Greater preoperative segmental motion, greater preoperative T1 sagittal slope, and lower fusion levels could be risk factors for pseudarthrosis following ACDF. Preoperative segmental motion >12° is likely to be an important indicator of the development of pseudarthrosis. 3.


Reversal of compensatory flow in severe intrauterine growth restriction: middle cerebral artery and intracardiac volume flow modifications.
The aim of the study was to investigate cerebral and cardiac volume flow modifications in a group of preterm severely growth restricted (GR) fetuses with absent end diastolic flow (AEDF) in the umbilical artery. Doppler indices of GR fetuses born at less than 32 weeks of gestation with AEDF in umbilical artery were longitudinally analyzed. Ductus venosus S/A, umbilical artery PI, middle cerebral artery (MCA) pulsatility index (PI), MCA volume flow and intracardiac flows were checked daily. The value of volume flow was determined by multiplying the vessel area obtained by means of a power angiography by the integral of the velocimetric curve. In Group 1 the time of delivery was decided when abnormal Doppler index in ductus venosus (S/A>3) was detected. In Group 2, decisions about delivery were based on maternal indications or on various cardiotocographic (CTG) abnormalities. The division in the two groups was made with a retrospective method. Fifteen GR fetuses were studied: seven cases entered Group 1 and eight cases Group 2. In Group 2 velocimetric (PI) and quantitative (volume flow) values remained stable until delivery. In all cases in Group 1 ductus venosus anomalies (S/A>3) were preceded by 24 hours by cerebral volume flow indices indicating the loss of MCA vasodilatation and by changes in intracardiac volume flow indices, while the PI increase in MCA was present only 24 hours after the changes detected by the volume flow study and were simultaneous to ductus venosus modifications. The decrease in volume flow through the middle cerebral artery occurs at least 24 hours before the increase of middle cerebral artery PI and before the ductus venosus anomalies. Flow through the four cardiac valves show modification in agreement to the changes of the middle cerebral artery flow.


[Field survey of the anticancer drug contamination in the preparation environment. Usefulness of the 5-FU monitoring by the coupon method].
For medical professionals involved in cancer chemotherapy, occupational exposure of anticancer agents is considered a health risk. Education about the handling of anticancer drugs and proper use of protective equipment are important for reducing occupational exposure of anticancer drugs. Furthermore, monitoring of the contamination level of anticancer drugs is important for determining the propriety of anti-contamination methods. Cyclophosphamide (CPA) has been used as the standard drug of the contamination level; however, it is rarely detected because of the disparity between drug preparation frequency and consumption, and use of a closed system. Therefore, we chose 5-fluorouracil (5-FU) as the standard drug and attempted to monitor its contamination levels by sampling using drug absorption sheets (the coupon method). We measured contamination levels inside a biological safety cabinet (BSC) and at its lower floor, and at a preparation worktable, nurses' station worktable, its lower floor and floor of the hospital room in a chemotherapy room for outpatients of the Iwate Medical University Hospital for 3 time periods. 5-FU was detected in 72% of the coupons (n=108), while CPA was detected in only 7% of the coupons (n=108). Monitoring of 5-FU contamination levels by using the coupon method was considered useful for evaluating anti-contamination method and the contamination process.


Upon impact: the fate of adhering Pseudomonas fluorescens cells during nanofiltration.
Nanofiltration (NF) is a high-pressure membrane filtration process increasingly applied in drinking water treatment and water reuse processes. NF typically rejects divalent salts, organic matter, and micropollutants. However, the efficiency of NF is adversely affected by membrane biofouling, during which microorganisms adhere to the membrane and proliferate to create a biofilm. Here we show that adhered Pseudomonas fluorescens cells under high permeate flux conditions are met with high fluid shear and convective fluxes at the membrane-liquid interface, resulting in their structural damage and collapse. These results were confirmed by fluorescent staining, flow cytometry, and scanning electron microscopy. This present study offers a "first-glimpse" of cell damage and death during the initial phases of bacterial adhesion to NF membranes and raises a key question about the role of this observed phenomena during early-stage biofilm formation under permeate flux and cross-flow conditions.


An alcohol-focused intervention versus a healthy living intervention for problem drinkers identified in a general hospital setting (ADAPTA): study protocol for a randomized, controlled pilot trial.
Alcohol misuse is a major cause of premature mortality and ill health. Although there is a high prevalence of alcohol problems among patients presenting to general hospital, many of these people are not help seekers and do not engage in specialist treatment. Hospital admission is an opportunity to steer people towards specialist treatment, which can reduce health-care utilization and costs to the public sector and produce substantial individual health and social benefits. Alcohol misuse is associated with other lifestyle problems, which are amenable to intervention. It has been suggested that the development of a healthy or balanced lifestyle is potentially beneficial for reducing or abstaining from alcohol use, and relapse prevention. The aim of the study is to test whether or not the offer of a choice of health-related lifestyle interventions is more acceptable, and therefore able to engage more problem drinkers in treatment, than an alcohol-focused intervention. This is a pragmatic, randomized, controlled, open pilot study in a UK general hospital setting with concurrent economic evaluation and a qualitative component. Potential participants are those admitted to hospital with a diagnosis likely to be responsive to addiction interventions who score equal to or more than 16 on the Alcohol Use Disorders Identification Test (AUDIT). The main purpose of this pilot study is to evaluate the acceptability of two sorts of interventions (healthy living related versus alcohol focused) to the participants and to assess the components and processes of the design. Qualitative research will be undertaken to explore acceptability and the impact of the approach, assessment, recruitment and intervention on trial participants and non-participants. The effectiveness of the two treatments will be compared at 6 months using AUDIT scores as the primary outcome measure. There will be additional economic, qualitative and secondary outcome measurements. Development of the study was a collaboration between academics, commissioners and clinicians in general hospital and addiction services, made possible by the Collaboration in Leadership in Applied Health Research and Care (CLAHRC) program of research. CLAHRC was a necessary vehicle for overcoming the barriers to answering an important NHS question--how better to engage problem drinkers in a hospital setting. ISRCTN47728072.


Monitored home-based with or without face-to-face exercise for maternal mental health during the COVID-19 pandemic.
Despite the known beneficial effects of exercise, most pregnant women do not exercise regularly. Most studies on exercise have been conducted on supervised exercise and there is limited evidence regarding the adherence and effect of other exercise programs on pregnancy outcomes. Therefore, we aimed to investigate adherence to a face-to-face plus monitored home exercise program versus a monitored home-based exercise program on its own during pregnancy. In addition, effects of these two exercise programs on women's mental health during pregnancy and postpartum (primary outcomes) and on some other maternal and neonatal outcomes (secondary outcomes) will be assessed. In this superiority trial with three parallel arms, 150 women at 12-18 weeks of gestation will be randomised equally into three groups (face-to-face plus monitored home exercise, only monitored home-based exercise, and control). The exercise programs will be performed up to the 38th week of gestation during which participants will be assessed at specific intervals during the pregnancy, and post-partum and followed up until six months after childbirth. The exercise diary will be used to assess the adherence. The Edinburgh Depression Scale and the Positive and Negative Affect Schedule will be used to assess prenatal and postnatal depression and affect, respectively. This study reflects the feasibility and acceptance of two exercise programs for pregnant women and their effects on important outcomes. If these programs are followed properly and effectively, pregnant women's health can be improved using these methods at a lower cost compared to the conventional supervised exercise program.


Outcome after Elective Percutaneous Coronary Intervention Depends on Age in Patients with Stable Coronary Artery Disease - An Analysis of Relative Survival in a Multicenter Cohort and an OCT Substudy.
Age is a strong predictor of survival in patients with coronary artery disease. In elder patients with increasing co-morbidities percutaneous coronary intervention (PCI) is associated with more complications and worse outcome. The calculation of relative survival rates adjusts for the "background" mortality in the general population by correcting for age and gender. We analyzed if elder patients after elective PCI have a worse relative survival compared to younger patient groups. A total of 8,342 patients who underwent elective PCI at two high volume centers between 1998 and 2009 were analyzed. The survival of our patients after PCI (observed survival) was slightly lower compared to the general population (expected survival) resulting in a slightly decreasing relative survival curve. In a multivariate Cox regression model age amongst others was a strong predictor of survival. Stratifying patients according to their age the relative survival curves of younger patients (Quartile 1: <58 years; 2,046 patients), elder patients (Quartile 3: 66-73 years; 2,090 patients) and very old patients (Quartile 4: >73 years; 2,307 patients) were similar. The relative survival of mid-aged patients (Quartile 2: 58-65 years; 1,899 patients) was better than that of all other patient groups. The profile of cardiovascular risk factors differs between the various groups resulting in different composition and burden of coronary plaques in an optical coherence tomography sub-study. Patients after elective PCI have a slightly worse long-term survival compared to the age- and sex-matched general population. This is also true for different groups of age except for mid-aged patients between 58 and 63 years. Elder patients between 66 and 73 years and above 73 years have a similar relative survival compared to younger patients below 58 years, and might therefore have similar benefit from elective PCI.


High and low inhibitor soybean meals affect human duodenal proteinase activity differently: in vivo comparison with bovine serum albumin.
The objective of this study was to investigate the effects of high and low inhibitor soybean meals on the duodenal enzyme activities and on the possible regulatory role of gastrointestinal hormones in the pancreatic response. After an overnight fast, 11 healthy volunteers received an intraduodenal infusion of saline for 60 min. This was followed by infusion of either of three test meals: extract of raw soybeans (RS), a low inhibitor soy protein isolate (SPI) or bovine serum albumin (BSA), 10 g/h for 60 min. Then saline was again given intraduodenally for 30 min. Gastric juice was collected continuously and duodenal juice and peripheral blood samples were collected every 10 min. Duodenal chymotryptic activity was severely inhibited by RS, whereas SPI and BSA increased the chymotryptic activity. Tryptic activity showed a transient reduction (55%) during RS infusion, whereas BSA and in particular SPI increased the tryptic activity. No change was seen in amylase activity. The lack of total inhibition of tryptic activity has been studied further and is the subject of the accompanying paper. The peripheral plasma levels of cholecystokinin (CCK) increased significantly during BSA but not during SPI or RS infusions. Thus, CCK levels were not increased by the inhibition of the proteolytic activity by RS in duodenal juice.


Blood levels of oxytocin and vasopressin during suckling in the rabbit and the problem of their independent release.
1. Oxytocin and vasopressin were assayed in samples of blood collected from conscious rabbits during suckling. The milk yields were determined from the weight gain of the litters.2. With normal milk yields (35-155 g), relatively high concentrations (31-375 muu./ml.) of oxytocin were present in the blood but vasopressin was detected in only one out of eight samples. It is concluded that the release of oxytocin by the milk-ejection reflex is essential for normal milk removal in the rabbit and that this hormone is released independently of vasopressin.3. When the milk yield did not exceed 15 g, an amount which could be removed by passive withdrawal from the mammary gland, vasopressin was detected in four out of five blood samples but none contained a detectable concentration of oxytocin. This result can be explained by the intervention of emotional stress which is known to block the milk-ejection reflex but which acts as a stimulus for the release of vasopressin.4. Emotional stress could inhibit the milk-ejection reflex either centrally by blocking the release of oxytocin, or peripherally by blocking the response of the mammary gland to circulating oxytocin. The absence of the hormone in blood during ineffectual suckling suggests that the block is central.


Congenital generalized lipodystrophy, type 4 (CGL4) associated with myopathy due to novel PTRF mutations.
Congenital generalized lipodystrophy (CGL) is a rare autosomal recessive disorder characterized by near total absence of body fat since birth with predisposition to insulin resistance, diabetes, hypertriglyceridemia, and hepatic steatosis. Three CGL loci, AGPAT2, BSCL2, and CAV1, have been identified previously. Recently, mutations in polymerase I and transcript release factor (PTRF) were reported in five Japanese patients presenting with myopathy and CGL (CGL4). We report novel PTRF mutations and detailed phenotypes of two male and three female patients with CGL4 belonging to two pedigrees of Mexican origin (CGL7100 and CGL178) and one pedigree of Turkish origin (CGL180). All patients had near total loss of body fat and congenital myopathy manifesting as weakness, percussion-induced muscle mounding, and high serum creatine kinase levels. Four of them had hypertriglyceridemia. Three of them had atlantoaxial instability. Two patients belonging to CGL178 pedigree required surgery for pyloric stenosis in the first month of life. None of them had prolonged QT interval on electrocardiography but both siblings belonging to CGL7100 had exercise-induced ventricular arrhythmias. Three of them had mild acanthosis nigricans but had normal glucose tolerance. Two of them had hepatic steatosis. All patients had novel null mutations in PTRF gene. In conclusion, mutations in PTRF result in a novel phenotype that includes generalized lipodystrophy with mild metabolic derangements, myopathy, cardiac arrhythmias, atlantoaxial instability, and pyloric stenosis. It is unclear how mutations in PTRF, which plays an essential role in formation of caveolae, affect a wide variety of tissues resulting in a variable phenotype.


[Hypertrophic cardiomyopathy during ACTH treatment].
We report on two infants (two and six month old) with infantile myoclonic seizures, who developed signs of hypertrophic cardiomyopathy (HCM), while receiving ACTH treatment (Tetracosactid=Synacthen Depot). The diagnosis of HCM was established by echocardiography. The first patient (R.M. female). showed signs of cardiac insufficiency and was treated with a beta-blocking agent (Propranolol=Dociton). This led to resolution of the clinical symptoms and reversal of myocardial muscle thickness as determined by echocardiography. The second patient (R.S. female) did not develop cardiac symptoms. A correlation between cardiomyopathy and ACTH treatment is discussed. Based on the various biological effects of ACTH different hypothetical explanations for this correlation are proposed: increased deposition of glycogen, enhanced protein synthesis, oedema of the myocardial tissue and systemic hypertension. Because of the correlation observed between ACTH treatment and the development of cardiomyopathy we recommend regular physical and echocardiographic examinations to detect cardiac involvement during treatment with ACTH.


Osteoporosis and the Potential of Cell-Based Therapeutic Strategies.
Osteoporosis, the most common chronic metabolic bone disease, is characterized by low bone mass and increased bone fragility. Nowadays more than 200 million individuals are suffering from osteoporosis and still the number of affected people is dramatically increasing due to an aging population and longer life, representing a major public health problem. Current osteoporosis treatments are mainly designed to decrease bone resorption, presenting serious adverse effects that limit their safety for long-term use. Numerous studies with mesenchymal stem cells (MSCs) have helped to increase the knowledge regarding the mechanisms that underlie the progression of osteoporosis. Emerging clinical and molecular evidence suggests that inflammation exerts a significant influence on bone turnover, thereby on osteoporosis. In this regard, MSCs have proven to possess broad immunoregulatory capabilities, modulating both adaptive and innate immunity. Here, we will discuss the role that MSCs play in the etiopathology of osteoporosis and their potential use for the treatment of this disease.


In vitro passage selects for Chlamydia muridarum with enhanced infectivity in cultured cells but attenuated pathogenicity in mouse upper genital tract.
Although modern Chlamydia muridarum has been passaged for decades, there are no reports on the consequences of serial passage with strong selection pressure on its fitness. In order to explore the potential for Pasteurian selection to induce genomic and phenotypic perturbations to C. muridarum, a starter population was passaged in cultured cells for 28 generations without standard infection assistance. The resultant population, designated CMG28, displays markedly reduced in vitro dependence on centrifugation for infection and low incidence and severity of upper genital tract pathology following intravaginal inoculation into mice compared to the parental C. muridarum population, CMG0. Deep sequencing of CMG0 and CMG28 revealed novel protein variants in the hypothetical genes TC0237 (Q117E) and TC0668 (G322R). In vitro attachment assays of isogenic plaque clone pairs with mutations in either TC0237 and TC0668 or only TC0237 reveal that TC0237(Q117E) is solely responsible for enhanced adherence to host cells. Paradoxically, double mutants, but not TC0237(Q117E) single mutants, display severely attenuated in vivo pathogenicity. These findings implicate TC0237 and TC0668 as novel genetic factors involved in chlamydial attachment and pathogenicity, respectively, and show that serial passage under selection pressure remains an effective tool for studying Chlamydia pathogenicity.


Okara: A Nutritionally Valuable By-product Able to Stabilize Lactobacillus plantarum during Freeze-drying, Spray-drying, and Storage.
Okara is a nutritionally valuable by-product produced in large quantities as result of soymilk elaboration. This work proposes its use as both culture and dehydration medium during freeze-drying, spray-drying, and storage of Lactobacillus plantarum CIDCA 83114. Whole and defatted okara were employed as culture media for L. plantarum CIDCA 83114. The growth kinetics were followed by plate counting and compared with those of bacteria grown in MRS broth (control). No significant differences in plate counting were observed in the three media. The fatty acid composition of bacteria grown in whole and defatted okara showed a noticeable increase in the unsaturated/saturated (U/S) fatty acid ratio, with regard to bacteria grown in MRS. This change was mainly due to the increase in polyunsaturated fatty acids, namely C18:2. For dehydration assays, cultures in the stationary phase were neutralized and freeze-dried (with or without the addition of 250 mM sucrose) or spray-dried. Bacteria were plate counted immediately after freeze-drying or spray-drying and during storage at 4°C for 90 days. Freeze-drying in whole okara conducted to the highest bacterial recovery. Regarding storage, spray-dried bacteria previously grown in whole and defatted okara showed higher plate counts than those grown in MRS. On the contrary, freeze-dried bacteria previously grown in all the three culture media were those with the lowest plate counts. The addition of sucrose to the dehydration media improved their recovery. The higher recovery of microorganisms grown in okara after freeze-drying and spray-drying processes and during storage was ascribed to both the presence of fiber and proteins in the dehydration media, and the increase in U/S fatty acids ratio in bacterial membranes. The obtained results support for the first time the use of okara as an innovative matrix to deliver L. plantarum. Considering that okara is an agro-waste obtained in large quantities, these results represent an innovative strategy to add it value, providing a symbiotic ingredient with promising industrial applications in the development of novel functional foods and feeds.


Genotypic and phenotypic spectrum of pyridoxine-dependent epilepsy (ALDH7A1 deficiency).
Pyridoxine-dependent epilepsy was recently shown to be due to mutations in the ALDH7A1 gene, which encodes antiquitin, an enzyme that catalyses the nicotinamide adenine dinucleotide-dependent dehydrogenation of l-alpha-aminoadipic semialdehyde/L-Delta1-piperideine 6-carboxylate. However, whilst this is a highly treatable disorder, there is general uncertainty about when to consider this diagnosis and how to test for it. This study aimed to evaluate the use of measurement of urine L-alpha-aminoadipic semialdehyde/creatinine ratio and mutation analysis of ALDH7A1 (antiquitin) in investigation of patients with suspected or clinically proven pyridoxine-dependent epilepsy and to characterize further the phenotypic spectrum of antiquitin deficiency. Urinary L-alpha-aminoadipic semialdehyde concentration was determined by liquid chromatography tandem mass spectrometry. When this was above the normal range, DNA sequencing of the ALDH7A1 gene was performed. Clinicians were asked to complete questionnaires on clinical, biochemical, magnetic resonance imaging and electroencephalography features of patients. The clinical spectrum of antiquitin deficiency extended from ventriculomegaly detected on foetal ultrasound, through abnormal foetal movements and a multisystem neonatal disorder, to the onset of seizures and autistic features after the first year of life. Our relatively large series suggested that clinical diagnosis of pyridoxine dependent epilepsy can be challenging because: (i) there may be some response to antiepileptic drugs; (ii) in infants with multisystem pathology, the response to pyridoxine may not be instant and obvious; and (iii) structural brain abnormalities may co-exist and be considered sufficient cause of epilepsy, whereas the fits may be a consequence of antiquitin deficiency and are then responsive to pyridoxine. These findings support the use of biochemical and DNA tests for antiquitin deficiency and a clinical trial of pyridoxine in infants and children with epilepsy across a broad range of clinical scenarios.


The absence of macrophage Nrf2 promotes early atherogenesis.
The loss of nuclear factor E2-related factor 2 (Nrf2) has been shown to protect against atherogenesis in apoE-deficient mice. The mechanism by which Nrf2 deficiency affords atheroprotection in this model is currently unknown, but combined systemic and local vascular effects on lesion macrophages have been proposed. We investigated the effect of bone marrow-specific loss of Nrf2 on early atherogenesis in low-density lipoprotein (LDL) receptor-deficient (LDLR(-/-)) mice, and assessed the effect of Nrf2 on cellular accumulation of modified LDLs and the expression of inflammatory markers in macrophages. The effect of bone marrow-specific loss of Nrf2 on atherogenesis was studied using bone marrow transplantation of wild-type (WT) or Nrf2(-/-) bone marrow to LDLR(-/-) mice. Mice transplanted with Nrf2(-/-) bone marrow and fed a high-fat diet for 6 weeks exhibited significantly larger atherosclerotic lesions than WT bone marrow transplanted mice. Moreover, in thioglycollate-elicited Nrf2(-/-) macrophages, the uptake of acetylated and malondialdehyde-modified LDLs was increased in comparison with WT controls, with the concomitant increase in the expression of scavenger receptor A and toll-like receptor 4. In addition, the expression of pro-inflammatory monocyte chemoattractant protein-1 and interleukin-6 were increased in Nrf2(-/-) vs. WT macrophages. Nrf2 deficiency specific to bone marrow-derived cells aggravates atherosclerosis in LDLR(-/-) mice. Furthermore, the loss of Nrf2 in macrophages enhances foam cell formation and promotes the pro-inflammatory phenotype.


Genome wide association study identifies two loci associated with cadmium in erythrocytes among never-smokers.
Cadmium is a non-essential toxic metal with multiple adverse health effects. Exposure in the general population occurs by smoking and diet. Cadmium in erythrocytes is a valid biomarker of exposure and body burden of cadmium. We aimed to identify genetic variants related to concentrations of cadmium in erythrocytes. Erythrocyte cadmium was analyzed in 4432 individuals (1728 never smokers) from the Swedish population-based Malmö Diet and Cancer cohort. Genotyping was performed using the Illumina HumanOmniExpressExome Bead chip with genome-wide coverage. Genome wide analyses were performed in the whole sample and in never smokers. No single nucleotide polymorphism (SNP) reached a genome-wide significant association with erythrocyte cadmium in the whole sample. However, in never smokers, 14 variants showed genome-wide significant relationships with erythrocyte cadmium after adjusting for age and sex. Thirteen variants were in linkage disequilibrium on chromosome 8q13.3 in the XKR9 and LACTB2 genes. The lead SNP on 8q13.3 was rs12681420 (minor allele G, minor allele frequency [MAF] = 0.46, β: -0.11, P = 3.48 × 10-11), an intron variant within the XKR9 gene. The other significant locus, rs17574271 (minor allele C, MAF = 0.09, β: 0.17, P = 6.18 × 10-9), was an intron variant within the DLGAP1 gene at chromosome 18p11.31. This genome-wide study of never smokers from the general population identified two independent regions related to erythrocyte cadmium. The strongest locus covers the XKR9 and LACTB2 genes, which both could have related functions in cadmium absorption and metabolism. Replication studies are needed to confirm the findings and mechanisms should be further investigated.


Predictors of smoking lapse during a 48-hour laboratory analogue smoking cessation attempt.
Many individuals who attempt to quit smoking experience a smoking lapse early on in the quitting process, with most lapses resulting in a return to regular smoking. Using a novel laboratory model, this study sought to investigate baseline predictors of smoking lapse during a brief, simulated smoking quit attempt. Self-report baseline measures were completed by 81 smokers, who also smoked a cigarette in the laboratory to equate recent smoke exposure. Participants were then given brief face-to-face smoking-cessation counseling along with monetary incentives to abstain from smoking for 48 hr (i.e., $40). Participants returned to the laboratory after 24 hr and 48 hr for assessment of smoking behavior. By 48 hr, 25 participants lapsed, with rates equivalent among men and women (31% vs. 31%). Higher rates of delay discounting and a preference for menthol cigarettes significantly predicted greater odds of lapsing. Shorter time to first cigarette after waking (TTFC) was associated with greater lapse risk at trend levels. No effects were observed for demographic variables, cigarettes per day, prequit abstinence self-efficacy, or depressive symptoms. Future research examining predictors of early lapse and underlying mechanisms is needed, and laboratory analogue models offer a controlled time- and cost-effective framework in which to investigate smoking cessation processes. (PsycINFO Database Record


Routine pre-operative focused ultrasonography by anesthesiologists in patients undergoing urgent surgical procedures.
Unexpected cardiopulmonary complications are well described during surgery and anesthesia. Pre-operative evaluation by focused cardiopulmonary ultrasonography may prevent such mishaps. The aim of this study was to determine the frequency of unexpected cardiopulmonary pathology with focused ultrasonography in patients undergoing urgent surgical procedures. We performed pre-operative focused cardiopulmonary ultrasonography in patients aged 18 years or above undergoing urgent surgical procedures at pre-defined study days. Known and unexpected cardiopulmonary pathology was recorded, and subsequent changes in the anesthesia technique or supportive actions were registered. A total of 112 patients scheduled for urgent surgical procedures were included. Their mean age (standard deviation) was 62 (21) years. Of these patients, 24% were American Society of Anesthesiologists (ASA) class 1, 39% were ASA class 2, 32% were ASA class 3, and 4% were ASA class 4. Unexpected cardiopulmonary pathology was disclosed in 27% [95% confidence interval (CI) 19-36] of the patients and led to a change in anesthesia technique or supportive actions in 43% (95% CI 25-63) of these. Unexpected pathology leading to changes in anesthesia technique or supportive actions was only disclosed in a group of patients above the age of 60 years and/or in ASA class ≥ 3. Focused cardiopulmonary ultrasonography disclosed unexpected pathology in patients undergoing urgent surgical procedures and induced changes in the anesthesia technique or supportive actions. Pre-operative focused ultrasonography seems feasible in patients above 60 year and/or with physical limitations but not in young, healthy individuals.


Ifenprodil attenuates the acquisition and expression of methamphetamine-induced behavioral sensitization and activation of Ras-ERK1/2 cascade in the caudate putamen.
Chronic discontinuous use of many psychomotor stimulants leads to behavioral sensitization and, owing to it shares common mechanisms with relapse, most researchers use its animal model to explore the neurobiological mechanisms of addiction. Recent studies have proved that N-methyl-d-aspartate receptors (NMDARs) are implicated in psychomotor stimulant-induced behavioral sensitization. However, the function of GluN2B-containing NMDARs and their potential downstream cascade(s) in the acquisition and expression of behavioral sensitization to methamphetamine (METH) have not been explored. In this study, 2.5, 5, and 10mg/kg ifenprodil, the specific inhibitor of GluN2B, was used to explore the function of these receptors in distinct phases of behavioral sensitization to METH in mice. Then, using western blot, Ras, pERK1/2/ERK1/2, and ΔFosB levels in the prefrontal cortex (PFc), nucleus accumbens (NAc), and caudate putamen (CPu) were detected. Behavioral results showed that low-dose ifenprodil attenuated the acquisition and expression of behavioral sensitization to METH significantly. Western blot analysis revealed that pre-injection of low-dose ifenprodil in the acquisition markedly attenuated METH-induced ascent of Ras, pERK1/2/ERK1/2, and ΔFosB protein levels in the CPu. However, pre-treatment in the expression only affected the alterations of Ras and pERK1/2/ERK1/2 levels in the CPu. Moreover, chronic METH administration increased pERK1/2/ERK1/2 level in the NAc. In conclusion, GluN2B-containing NMDARs contribute to both the acquisition and expression of behavioral sensitization to METH in mice. Furthermore, the acquisition phase might be mediated by the Ras-ERK1/2-ΔFosB cascade in the CPu while the expression phase may be regulated by the Ras-ERK1/2 cascade in the CPu.


Conventional morphological and functional evaluation of the microbial populations in a sequencing batch reactor performing EBPR.
To help confirm and interpret the Enhanced Biological Phosphorus Removal (EBPR) performance of the microbial populations in a laboratory-scale activated sludge (AS) system, conventional microscopic examinations were carried out. A lab-scale sequencing batch reactor (SBR), named ARC, was fed with acetate, as the sole carbon source, and operated for EBPR. Daily monitoring and cyclic behavior evaluation studies indicated that the system always worked for EBPR in the long run, with efficiencies depending on the influent characteristics and operational stability. Poly-P and PHB-staining experiments revealed that the enriched biomass of the reactor was quite diverse in terms of morphology, hosting populations of traditional rod-shaped PAOs, tetrad/sarcina-like cells (referred here as TFOs, rather than GAOs), diplococci-shaped cells, and staphylococci-like clustered populations, in addition to few filaments. Although the microscopic observations were qualitative, rather than quantitative, they seemed likely to correlate well to the biochemical performance of the reactor.


CRISPR-Cas12a trans-cleaves DNA G-quadruplexes.
We for the first time report that the activated CRISPR-Cas12a system trans-cleaves DNA G-quadruplexes (G4). The cleavage activity on human telomere G4 and TBA G4 was investigated and verified by FRET, CD, gel electrophoresis and NMR. We believe that this finding will pave a new avenue for advancing the applications of CRISPR-Cas12a and G4 in biosensing and biochemistry.


Exostoses and cavernous venous formation in the external auditory canal of the hooded seal as a functional physiological organ.
Exostoses of the external auditory canal (EAC) develop after protracted mechanical, chemical or thermal irritation in particular. This is a common disorder among aquatic sportsmen and has been considered unique to Man. We dissected and photodocumented the EACs of 5 newborn and 3 adult Hooded Seals (Cystophora cristata). Serial sections of the EACs were prepared for light microscopic evaluation after staining with haematoxylin-eosin or toluidine blue. All EACs exhibited a firm, broad-based. mountain peak-shaped exostosis on the floor of the meatus, lateral to the eardrum. In addition, the meatal skin of the bony EAC harboured large venous sinuses. The exostosis and venous sinuses of the seal EAC participate in the protection of the sensitive hearing apparatus, particularly the pars tensa portion of the drum, during divine.


In vitro toxicity tests of zinc phosphate cement.
BHK-21 (C-13) cells were exposed to freshly mixed zinc phosphate cement for 0.5, 1 and 2 h in a simulated cavity apparatus. The effect on the cells was evaluated by quantitative measurement of acid phosphatase and succinic dehydrogenase reaction product in order to determine organelle membrane permeability. After 0.5 h cell-material contact there was no significant effect. After 1 h cell-material contact, there was slight labilization of the lysosomal membranes, and a significant decrease in succinic dehydrogenase activity. The mean overall stain density in the experimental cells was 109% and 57% of the control values for acid phosphotase and succinic dehydrogenase respectively. After 2 h cell-material contact, there was significant labilization of the lysosomal membranes, and also labilization of the mitochondrial membranes. The mean overall stain densities were 114% and 75% of the control values for acid phosphatase and succinic dehydrogenase respectively. The relevance of these findings to the in vivo situation is discussed.


The role of platelet growth factors in cancer therapy.
The use of myeloid growth factors has markedly reduced the complications of chemotherapy-induced neutropenia, however, abrogation of severe thrombocytopenia remains a major clinical problem. Platelet transfusions remain the standard method of preventing or treating thrombocytopenia but are associated with a variety of complications and are a limited resource. A number of cytokines have been clinically investigated for their thrombopoietic activity, the most promising of which is the recently cloned ligand to the hematopoietic growth factor receptor, c-Mpl. The c-Mpl ligand, also referred to as thrombopoietin, megakaryocyte growth and development factor (MGDF) and megapoietin, is a potent lineage-specific agent that promotes growth and maturation of megakaryocytes and their progenitors. It holds promise for clinical use in the treatment of iatrogenic or disease-associated bone marrow failure states and possibly in syndromes of excessive platelet consumption. Early clinical trials assessing the safety and activity of recombinant human MGDF are now underway.


Chopper, a new death domain of the p75 neurotrophin receptor that mediates rapid neuronal cell death.
The cytoplasmic juxtamembrane region of the p75 neurotrophin receptor (p75(NTR)) has been found to be necessary and sufficient to initiate neural cell death. The region was named "Chopper" to distinguish it from CD95-like death domains. A 29-amino acid peptide corresponding to the Chopper region induced caspase- and calpain-mediated death in a variety of neural and non-neural cell types and was not inhibited by signaling through Trk (unlike killing by full-length p75(NTR)). Chopper triggered cell death only when bound to the plasma membrane by a lipid anchor, whereas non-anchored Chopper acted in a dominant-negative manner, blocking p75(NTR)-mediated death both in vitro and in vivo. Removal of the ectodomain of p75(NTR) increased the potency of Chopper activity, suggesting that it regulates the association of Chopper with downstream signaling proteins.


Perlecan is recruited by dystroglycan to nodes of Ranvier and binds the clustering molecule gliomedin.
Fast neural conduction requires accumulation of Na(+) channels at nodes of Ranvier. Dedicated adhesion molecules on myelinating cells and axons govern node organization. Among those, specific laminins and dystroglycan complexes contribute to Na(+) channel clustering at peripheral nodes by unknown mechanisms. We show that in addition to facing the basal lamina, dystroglycan is found near the nodal matrix around axons, binds matrix components, and participates in initial events of nodogenesis. We identify the dystroglycan-ligand perlecan as a novel nodal component and show that dystroglycan is required for the selective accumulation of perlecan at nodes. Perlecan binds the clustering molecule gliomedin and enhances clustering of node of Ranvier components. These data show that proteoglycans have specific roles in peripheral nodes and indicate that peripheral and central axons use similar strategies but different molecules to form nodes of Ranvier. Further, our data indicate that dystroglycan binds free matrix that is not organized in a basal lamina.


[Modification changes of the genetic material in Saccharomyces yeasts].
The problem of mating-type switches in heterothallic yeast cells was investigated. In selective system for cytoduction in alpha x alpha crosses alpha-cytoductants were predominantly obtained. Thus matings in alpha x alpha crosses can proceed through non-heritable changes (modifications) of the mating type alpha----a. The frequency of alpha-cytoductants after UV-irradiation of the recipient cells exceeded the control value 50-90 times. The extra copy of MAT alpha dramatically decreased the frequency of cytoductants in alpha x alpha crosses, either spontaneously or after UV-irradiation. The rad18 recipient defective in postreplication repair had 70-times increased level of mating-type modifications, as compared with isogenic Rad+ strain. An explanation consistent with these data is that mating-type modifications are due to phenotypic expression of primary lesions of MAT alpha locus. Such lesions might be expressed as transient a-mating type. After the mating event, these lesions can be repaired or turned to true mutations within the MAT locus. In fact, approximately half of non-mating cytoductants from alpha x alpha crosses had the phenotype of mat alpha 2 mutants.


[Karyologic research on murine B-cell hybridomas].
Karyotypes of 10 murine hybridomas producing monoclonal antibodies to microbal antigenes were examined using chromosome slides stained with Azur-eosine. Hybrid origin of all the cell clones was confirmed. The cultures differed from each other in modal chromosome numbers, in novel markers that were absent from cells of the parental myeloma X63.Ag8.653, in the frequency of metaphases with double minute chromosomes and in the level of cells with chromosome aberrations. The results obtained enable us to recommend a cytogenetic analysis for the identification of hybridomas. The following observations point out to a relative instability of the chromosomal apparatus of hybridomas: chromosome numbers varied significantly from cell to cell within one and the same clone; modal chromosome counts decreased in 3 of 5 hybridomas that were studied repeatedly within 1-2 months; in some hybridomas unstable chromosome aberrations were found in 18-38% of cells.


Control of membrane protein homeostasis by a chaperone-like glial cell adhesion molecule at multiple subcellular locations.
The significance of crosstalks among constituents of plasma membrane protein clusters/complexes in cellular proteostasis and protein quality control (PQC) remains incompletely understood. Examining the glial (enriched) cell adhesion molecule (CAM), we demonstrate its chaperone-like role in the biosynthetic processing of the megalencephalic leukoencephalopathy with subcortical cyst 1 (MLC1)-heteromeric regulatory membrane protein complex, as well as the function of the GlialCAM/MLC1 signalling complex. We show that in the absence of GlialCAM, newly synthesized MLC1 molecules remain unfolded and are susceptible to polyubiquitination-dependent proteasomal degradation at the endoplasmic reticulum. At the plasma membrane, GlialCAM regulates the diffusional partitioning and endocytic dynamics of cluster members, including the ClC-2 chloride channel and MLC1. Impaired folding and/or expression of GlialCAM or MLC1 in the presence of diseases causing mutations, as well as plasma membrane tethering compromise the functional expression of the cluster, leading to compromised endo-lysosomal organellar identity. In addition, the enlarged endo-lysosomal compartments display accelerated acidification, ubiquitinated cargo-sorting and impaired endosomal recycling. Jointly, these observations indicate an essential and previously unrecognized role for CAM, where GliaCAM functions as a PQC factor for the MLC1 signalling complex biogenesis and possess a permissive role in the membrane dynamic and cargo sorting functions with implications in modulations of receptor signalling.


Melaminivora jejuensis sp. nov., isolated from Swinery waste.
A sulfur-oxidizing bacterium, designated strain KBB12T, was isolated from swinery waste collected in Jeju, Republic of Korea. The cells were Gram-stain-negative, flagellated and rod-shaped. Growth occurred at 15-45 °C (optimum, 30-37 °C), at pH 6-9 (optimum, pH 7.0) and in the presence of 0-1 % (w/v) NaCl. The major cellular fatty acids were summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1 ω7c, C16 : 0 and C18 : 1ω7c. The polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phospholipid and an unidentified lipid. The major isoprenoid quinone was ubiquinone-8 (Q-8) and the DNA G+C content of the genomic DNA was 69.6 mol%. Phylogenetic analyses, based on 16S rRNA gene sequences, showed that the novel isolate belongs to the genus Melaminivora and was most closely related to Melaminivora alkalimesophila CY1T (97.2 % similarity). The DNA-DNA relatedness values between strain KBB12T and M. alkalimesophila DSM26005T was 43.4±2.7 %. On the basis of phylogenetic and phenotypic evidence, it is proposed that strain KBB12T represents a novel species of the genus Melaminivora, for which the name Melaminivora jejuensis sp. nov. is proposed. The type strain is KBB12T (=KCTC 32230T=JCM 18740T).


Aldosterone impairs bone marrow-derived progenitor cell formation.
Aldosterone has been suggested recently to cause vascular injury by directly acting on the vasculature, in addition to causing injury by raising the blood pressure. Bone marrow-derived endothelial progenitor cells (EPCs) have been shown to exert an important role in the repair of the endothelium. In addition, cell-based therapy using EPCs is emerging as a novel therapeutic strategy for myocardial and peripheral vascular diseases. However, impaired formation and function of EPCs has been observed in patients with risk factors for cardiovascular diseases. We evaluated the possible effects of aldosterone on EPCs by examining the progenitor cell formation from bone marrow mononuclear cells ex vivo. Aldosterone (10 to 1000 nmol/L) reduced the formation of progenitor cells in a concentration-dependent manner. This effect of aldosterone was attenuated by cotreatment with spironolactone. Aldosterone reduced the mRNA levels of vascular endothelial growth factor (VEGF) receptor (VEGFR) 2 without having any effect on the production of VEGF or mRNA levels of VEGF and hepatocyte growth factor in the progenitor cells. However, the expression of stromal-derived growth factor 1 mRNA was paradoxically increased. Consistent with the downregulation of VEGFR-2, VEGF-induced phosphorylation of Akt was abolished in the progenitor cells after aldosterone treatment. N-acetylcysteine, an antioxidant, attenuated the inhibitory effects of aldosterone. These data indicate that aldosterone inhibits the formation of bone marrow-derived progenitor cells, at least partly, by attenuating VEGFR-2 expression and the subsequent Akt signaling. Reduction of aldosterone levels, blockade of mineralocorticoid receptor, and/or cotreatment with antioxidants may, therefore, enhance vascular regeneration by EPCs.


Accuracy of self-collected vaginal dry swabs using the Xpert human papillomavirus assay.
Polymerase chain reaction-based Xpert human papillomavirus (HPV) assay is a rapid test that detects high-risk HPV (hrHPV) infection. This point-of-care test is usually performed by collecting a cervical specimen in a vial of PreservCyt® transport medium. We compared HPV test positivity and accuracy between self-collected sample with a dry swab (s-DRY) versus physician-collected cervical sampling using a broom like brush and immediate immersion in PreservCyt (dr-WET). In this cross-sectional study, we recruited 150 women ≥ 18 years old attending the colposcopy clinic in the University Hospital of Geneva. Each participant first self-collected a vaginal sample using a dry swab and then the physician collected a cervical specimen in PreservCyt. HPV analysis was performed with Xpert. Part of the PreservCyt-collected sample was used for hrHPV detection with the cobas® HPV test. HPV test positivity and performance of the two collection methods was compared. HPV positivity was 49.1% for s-DRY, 41.8% for dr-WET and 46.2% for cobas. Good agreement was found between s-DRY and dr-WET samples (kappa±Standard error (SE) = 0.64±0.09,), particularly for low-grade squamous intraepithelial lesions (LSIL+) (kappa±SE = 0.80±0.17). Excellent agreement was found between the two samples for HPV16 detection in general (kappa±SE = 0.91±0.09) and among LSIL+ lesions (kappa±SE = 1.00±0.17). Sensitivities and specificities were, respectively, 84.2% and 47.1%(s-DRY), 73.1% and 58.7%. (dr-WET) and 77.8% and 45.7% (cobas) for CIN2+ detection. The median delay between sampling and HPV analysis was 7 days for the Xpert HPV assay and 19 days for cobas. There were 36 (24.0%) invalid results among s-DRY samples and 4 (2.7%) among dr-WET (p = 0.001). Invalid results happened due to the long interval between collection and analysis. Self-collected vaginal dry swabs are a valid alternative to collecting cervical samples in PreservCyt solution for HPV testing with the Xpert HPV assay. HPV self-collection with dry cotton swabs might assist in the implementation of an effective screening strategy in developing countries. International Standard Randomized Controlled Trial Number Registry ISRCTN83050913.


[AIDS and the eye].
The acquired immune deficiency syndrome (AIDS), is a recently described irreversible dysfunction of cell-mediated immunity in homosexuals, intravenous, drug abusers, and hemophiliacs, with subsequent development of potentially lethal opportunistic infections and/or unusual neoplasms, such as Kaposi's sarcoma. A prospective evaluation of ophthalmic findings in 14 patients with AIDS revealed that 8 patients had ophthalmoscopically and biomicroscopically significant ocular abnormalities, including peripapillary cotton-wool spots of changing frequency and diameter, retinal hemorrhages, progressive cytomegaly virus (CMV) retinitis, acute destructive retinal necrosis, periphlebitis, CMV conjunctivitis and keratitis, and in one patient a conjunctival Kaposi's sarcoma. All patients with AIDS and abnormal ocular findings carry a poor prognosis. Early detection of ocular manifestations is important, since most patients with AIDS are visually asymptomatic, and the ophthalmic presentation may be the primary one, and the initiating contact leading to diagnosis and permitting the prognosis to be assessed.


A best practice fall prevention exercise program to improve balance, strength / power, and psychosocial health in older adults: study protocol for a randomized controlled trial.
With increasing age neuromuscular deficits (e.g., sarcopenia) may result in impaired physical performance and an increased risk for falls. Prominent intrinsic fall-risk factors are age-related decreases in balance and strength / power performance as well as cognitive decline. Additional studies are needed to develop specifically tailored exercise programs for older adults that can easily be implemented into clinical practice. Thus, the objective of the present trial is to assess the effects of a fall prevention program that was developed by an interdisciplinary expert panel on measures of balance, strength / power, body composition, cognition, psychosocial well-being, and falls self-efficacy in healthy older adults. Additionally, the time-related effects of detraining are tested. Healthy old people (n = 54) between the age of 65 to 80 years will participate in this trial. The testing protocol comprises tests for the assessment of static / dynamic steady-state balance (i.e., Sharpened Romberg Test, instrumented gait analysis), proactive balance (i.e., Functional Reach Test; Timed Up and Go Test), reactive balance (i.e., perturbation test during bipedal stance; Push and Release Test), strength (i.e., hand grip strength test; Chair Stand Test), and power (i.e., Stair Climb Power Test; countermovement jump). Further, body composition will be analysed using a bioelectrical impedance analysis system. In addition, questionnaires for the assessment of psychosocial (i.e., World Health Organisation Quality of Life Assessment-Bref), cognitive (i.e., Mini Mental State Examination), and fall risk determinants (i.e., Fall Efficacy Scale - International) will be included in the study protocol. Participants will be randomized into two intervention groups or the control / waiting group. After baseline measures, participants in the intervention groups will conduct a 12-week balance and strength / power exercise intervention 3 times per week, with each training session lasting 30 min. (actual training time). One intervention group will complete an extensive supervised training program, while the other intervention group will complete a short version ('3 times 3') that is home-based and controlled by weekly phone calls. Post-tests will be conducted right after the intervention period. Additionally, detraining effects will be measured 12 weeks after program cessation. The control group / waiting group will not participate in any specific intervention during the experimental period, but will receive the extensive supervised program after the experimental period. It is expected that particularly the supervised combination of balance and strength / power training will improve performance in variables of balance, strength / power, body composition, cognitive function, psychosocial well-being, and falls self-efficacy of older adults. In addition, information regarding fall risk assessment, dose-response-relations, detraining effects, and supervision of training will be provided. Further, training-induced health-relevant changes, such as improved performance in activities of daily living, cognitive function, and quality of life, as well as a reduced risk for falls may help to lower costs in the health care system. Finally, practitioners, therapists, and instructors will be provided with a scientifically evaluated feasible, safe, and easy-to-administer exercise program for fall prevention.


Host ranges of gregarious muscoid fly parasitoids: Muscidifurax raptorellus (Hymenoptera: Pteromalidae), Tachinaephagus zealandicus (Hymenoptera: Encyrtidae), and Trichopria nigra (Hymenoptera: Diapriidae).
Attack rates, progeny production, sex ratios, and host utilization efficiency of Muscidifurax raptorellus (Kogan and Legner) (Hymenoptera: Pteromalidae), Tachinaephagus zealandicus Ashmead (Hymenoptera: Encyrtidae), and Trichopria nigra (Nees) (Hymenoptera: Diapriidae) were evaluated in laboratory bioassays with five dipteran hosts: house fly (Musca domestica L.), stable fly (Stomoxys calcitrans L.), horn fly (Hematobia irritans L.), black dump fly [Hydrotaea aenescens (Weidemann)] (Diptera: Muscidae), and a flesh fly (Sarcophaga bullata Parker) (Diptera: Sarcophagidae). M. raptorellus killed and successfully parasitized all five host species and produced an average 2.6 parasitoid progeny from each host. Host attack rates were highest on stable fly and lowest on horn fly; there were no differences among hosts in the total number of progeny produced. T. zealandicus killed larvae of all fly host species in similar numbers, but parasitism was most successful on H. aenescens and S. bullata and least successful on horn fly and house fly hosts. Significantly more parasitoid progeny emerged from S. bullata (10.2 parasitoids per host) than the other hosts; only 2.5 progeny were produced from parasitized horn fly hosts. Most of the killed puparia that produced neither adult flies nor parasitoids ("duds") contained dead parasitoids; in house fly, stable fly, and horn fly hosts, >30% of these dudded pupae contained adult wasps that failed to eclose. T. nigra successfully parasitized pupae of all host species except house fly and was most successful on stable fly. Significantly more parasitoid progeny emerged from S. bullata (30.6 parasitoids per host) than the other hosts; only 5.7 progeny were produced from horn fly hosts.


Effect of pectin on gastric emptying and gut hormone release in the dumping syndrome.
The effect of pectin on gastric emptying, gut hormone release, and symptoms was studied in four patients with dumping syndrome and in two healthy volunteers after ingestion of a hypertonic glucose meal with and without addition of pectin. The initial fraction emptied from the stomach was reduced in the patients, whose symptoms of dumping were abolished or alleviated by pectin. This change of the emptying seems to be caused by a prolonged stomach transit, probably due to the viscous nature of the pectin meal. Pectin had no effect on the gastric emptying of the volunteers. The motor activity of the stomach was not altered by pectin in either the patients or volunteers. In the patients insulin, enteroglucagon, neurotensin, and gastric inhibitory polypeptide rose to higher levels after the glucose meal than after the glucose-pectin meal. The individual differences in the hormone release were considered secondary to the altered gastric emptying produced by pectin.


No association with the ETM2 locus in Czech patients with familial essential tremor.
Essential tremor (ET) is one of the most common neurological movement disorders. In more than half of the cases, ET is inherited in an autosomal dominant manner, but no causative ET gene has been identified. However, several candidate loci have been reported, including the ETM2 locus that was originally found in a large American family of Czech descent. To ascertain the association with ETM2, we performed a genetic analysis of three polymorphic loci, etm1231, etm1234, and etm1240, located within the ETM2 candidate region in 61 Czech patients with a family history of ET and 68 healthy controls. The allele frequencies were not significantly different between the patients and the controls, and we did not observe any haplotype specifically associated with ET. This is the first report on the genetics of familial essential tremor in Czech patients. Our findings provide further evidence of genetic heterogeneity for ET.


Agent-based model provides insight into the mechanisms behind failed regeneration following volumetric muscle loss injury.
Skeletal muscle possesses a remarkable capacity for repair and regeneration following a variety of injuries. When successful, this highly orchestrated regenerative process requires the contribution of several muscle resident cell populations including satellite stem cells (SSCs), fibroblasts, macrophages and vascular cells. However, volumetric muscle loss injuries (VML) involve simultaneous destruction of multiple tissue components (e.g., as a result of battlefield injuries or vehicular accidents) and are so extensive that they exceed the intrinsic capability for scarless wound healing and result in permanent cosmetic and functional deficits. In this scenario, the regenerative process fails and is dominated by an unproductive inflammatory response and accompanying fibrosis. The failure of current regenerative therapeutics to completely restore functional muscle tissue is not surprising considering the incomplete understanding of the cellular mechanisms that drive the regeneration response in the setting of VML injury. To begin to address this profound knowledge gap, we developed an agent-based model to predict the tissue remodeling response following surgical creation of a VML injury. Once the model was able to recapitulate key aspects of the tissue remodeling response in the absence of repair, we validated the model by simulating the tissue remodeling response to VML injury following implantation of either a decellularized extracellular matrix scaffold or a minced muscle graft. The model suggested that the SSC microenvironment and absence of pro-differentiation SSC signals were the most important aspects of failed muscle regeneration in VML injuries. The major implication of this work is that agent-based models may provide a much-needed predictive tool to optimize the design of new therapies, and thereby, accelerate the clinical translation of regenerative therapeutics for VML injuries.


DSM-5 unspecified feeding and eating disorders in adolescents: what do they look like and are they clinically significant?
The recent DSM-5 categorization of eating disorders introduces a new category of eating disorders, Unspecified Feeding and Eating Disorders (UFED), where symptoms do not meet criteria for any other diagnostic category, but cause clinically significant distress or impairment. The aim of the current study was to explore what disorders in UFED might look like in an adolescent population. We examined a large cohort of adolescent female twins (N = 699) who were assessed on three occasions and who did not meet a DSM-5 eating disorder diagnosis but who reported threshold levels of either fasting and/or driven exercise (N = 33; 4.7%). This group of girls was compared to girls who reported no eating disorder over the three waves, and girls who met a diagnosis of either anorexia nervosa (AN) or atypical AN. The UFED group was characterized as being in the overweight range while striving to lose weight, and placing a high degree of importance on weight and shape in their self-evaluation. This group was indistinguishable from the two eating disorder groups on measures of global eating disorder severity, and demonstrated significantly elevated impairment and distress compared to the no eating disorder group commensurate with the eating disorder groups. Further research of this group is necessary to ensure that these individuals are not overlooked, and that treatment options are appropriate and available.


The proapoptotic influenza A virus protein PB1-F2 forms a nonselective ion channel.
PB1-F2 is a proapoptotic influenza A virus protein of approximately 90 amino acids in length that is located in the nucleus, cytosol and in the mitochondria membrane of infected cells. Previous studies indicated that the molecule destabilizes planar lipid bilayers and has a strong inherent tendency for multimerization. This may be correlate with its capacity to induce mitochondrial membrane depolarization. Here, we investigated whether PB1-F2 is able to form ion channels within planar lipid bilayers and microsomes. For that purpose, a set of biologically active synthetic versions of PB1-F2 (sPB1-F2) derived from the IAV isolates A/Puerto Rico/8/34(H1N1) (IAV(PR8)), from A/Brevig Mission/1/1918(H1N1) (IAV(SF2)) or the H5N1 consensus sequence (IAV(BF2)) were used. Electrical and fluorimetric measurements show that all three peptides generate in planar lipid bilayers or in liposomes, respectively, a barely selective conductance that is associated with stochastic channel type fluctuations between a closed state and at least two defined open states. Unitary channel fluctuations were also generated when a truncated protein comprising only the 37 c-terminal amino acids of sPB1-F2 was reconstituted in bilayers. Experiments were complemented by extensive molecular dynamics simulations of the truncated fragment in a lipid bilayer. The results indicate that the c-terminal region exhibits a slightly bent helical fold, which is stable and remains embedded in the bilayer for over 180 ns. The data support the idea that PB1-F2 is able to form protein channel pores with no appreciable selectivity in membranes and that the c-terminus is important for this function. This information could be important for drug development.


Chemoimmunotherapy of metastatic large bowel cancer: nonspecific stimulation with BCG and levamisole.
The administration of two chemoimmunotherapy programs to 103 consecutive patients with metastatic colorectal cancer resulted in improved survival for patients who achieved either objective tumor regressions or disease stabilization for more than 8 weeks. Objective tumor regression was observed in 47% of patients treated with the Ftorafur-methyl-CCNU-methotrexate-Bacillus Calmette-Guerin (FTOR-MeM-BCG) program and in 34% of patients treated with the 5-fluorouracil-methotrexate-Baker's antifol (FU-M-BAF) +/- Levamisole program. The combinated median duration of survival for patients who achieved objective tumor regression and disease stabilization with FTOR-MeM-BCG was 13 months compared with 6 months for patients who had progression of disease (p = 0.001). The corresponding values for patients treated with FU-M-BAF +/- levamisole were 11 months and seven months, respectively (p = 0.001). While the role of BCG immunotherapy in these results remains speculative, the administration of levamisole immunotherapy did not appear to have influenced results significantly. Patients who presented at diagnosis with Dukes A, B and C lesions, and therefore had longer disease-free intervals, responded more frequently to chemoimmunotherapy and survived longer than patients who presented at diagnosis with Dukes D lesions. Similarly, greater antitumor effect was observed in patients with lower pretreatment plasma CEA levels evaluation of these pretreatment characteristics may have insignificant implications for the design of future clinical trials.


Lessons learned from cloning dogs.
The aim of this article is to review dog cloning research and to suggest its applications based on a discussion about the normality of cloned dogs. Somatic cell nuclear transfer was successfully used for production of viable cloned puppies despite limited understanding of in vitro dog embryo production. Cloned dogs have similar growth characteristics to those born from natural fertilization, with no evidence of serious adverse effects. The offspring of cloned dogs also have similar growth performance and health to those of naturally bred puppies. Therefore, cloning in domestic dogs can be applied as an assisted reproductive technique to conserve endangered species, to treat sterile canids or aged dogs, to improve reproductive performance of valuable individuals and to generate disease model animals.


Effectiveness of the introduction of a Chronic Care Model-based program for type 2 diabetes in Belgium.
During a four-year action research project (2003-2007), a program targeting all type 2 diabetes patients was implemented in a well-defined geographical region in Belgium. The implementation of the program resulted in an increase of the overall Assessment of Chronic Illness Care (ACIC) score from 1.45 in 2003 to 5.5 in 2007. The aim of the follow-up study in 2008 was to assess the effect of the implementation of Chronic Care Model (CCM) elements on the quality of diabetes care in a country where the efforts to adapt primary care to a more chronic care oriented system are still at a starting point. A quasi-experimental study design involving a control region with comparable geographical and socio-economic characteristics and health care facilities was used to evaluate the effect of the intervention in the region. In collaboration with the InterMutualistic Agency (IMA) and the laboratories from both regions a research database was set up. Study cohorts in both regions were defined by using administrative data from the Sickness Funds and selected from the research database. A set of nine quality indicators was defined based on current scientific evidence. Data were analysed by an institution experienced in longitudinal data analysis. In total 4,174 type 2 diabetes patients were selected from the research database; 2,425 patients (52.9% women) with a mean age of 67.5 from the intervention region and 1,749 patients (55.7% women) with a mean age of 67.4 from the control region. At the end of the intervention period, improvements were observed in five of the nine defined quality indicators in the intervention region, three of which (HbA1c assessment, statin therapy, cholesterol target) improved significantly more than in the control region. Mean HbA1c improved significantly in the intervention region (7.55 to 7.06%), but this evolution did not differ significantly (p = 0.4207) from the one in the control region (7.44 to 6.90%). The improvement in lipid control was significantly higher (p = 0.0021) in the intervention region (total cholesterol 199.07 to 173 mg/dl) than in the control region (199.44 to 180.60 mg/dl). The systematic assessment of long-term diabetes complications remained insufficient. In 2006 only 26% of the patients had their urine tested for micro-albuminuria and only 36% had consulted an ophthalmologist. Although the overall ACIC score increased from 1.45 to 5.5, the improvement in the quality of diabetes care was moderate. Further improvements are needed in the CCM components delivery system design and clinical information systems. The regional networks, as they are financed now by the National Institute for Health and Disability Insurance (NIHDI), are an opportunity to explore how this can be achieved in consultation with the GPs. But it is clear that, simultaneously, action is needed on the health system level to realize the installation of an accurate quality monitoring system and the necessary preconditions for chronic care delivery in primary care (patient registration, staff support, IT support). ClinicalTrials.gov Identifier: NCT00824499.


Coagulation behavior of Al(13) aggregates.
The coagulation behavior of Al(13) aggregates formed in coagulation of kaolin was investigated by small angle static light scattering (SASLS), solid-state (27)Al NMR and tapping mode atomic force microscope (TM-AFM). A kaolin suspension was coagulated by PACl containing high content of Al(13) polycation (PACl-Al(13)). The results indicated that Al(13) was predominant in destabilizing kaolin particles for PACl-Al(13) coagulation even though at alkaline pH (pH 10). At such high pH, Al(13) aggregates were observed when the dosage of PACl-Al(13) was increased. In addition, the mechanism of coagulation by PACl-Al(13) at alkaline pH was affected by dosage. When the dosage was insufficient, coagulation was caused by electrostatic patch, which led to compact flocs with high fractal dimension (D(f)). Interparticle bridging dominated the coagulation when the coagulant dosage approached the plateau of adsorption, which caused the looser flocs with low D(f). The in-situ AFM scanning in liquid system proved that the existence of linear Al(13) aggregates composed of a chain of coiled Al(13) in coagulation by PACl-Al(13) at a high dosage and alkaline pH. Meanwhile, several coiled Al(13) aggregates with various dimensions were observed at such condition.


[Significance of electron microscopy in differential diagnosis of small cell neoplasms].
The diagnosis of small cell anaplastic neoplasms is one of the difficult problems in routine pathology. In some cases, it is almost impossible by light microscopy (LM) to arrive at a correct diagnosis. Electron microscopy (EM) may contribute greatly in solving this problem. However, it is impractical to make EM examination as a routine in every difficult neoplasm. Therefore, we have reviewed, by EM specimens of 21 cases (9 cases of small cell carcinoma and 12 cases of lymphoma, previously diagnosed by LM in this hospital), and compared the light and ultrastructural features of these tumors. We found that the most important light microscopic feature to differentiate lymphoma from small cell carcinoma is the nuclear structure of tumor cells. According to the characteristics such as the shape of nuclei, the distribution of chromatin, the stereoscopic impression of nuclear section plane while focusing, the appearance of nucleoli, it is often possible to arrive at a correct diagnosis of lymphoma or small cell carcinoma by LM.


Validity of the executive function theory of attention-deficit/hyperactivity disorder: a meta-analytic review.
One of the most prominent neuropsychologic theories of attention-deficit/hyperactivity disorder (ADHD) suggests that its symptoms arise from a primary deficit in executive functions (EF), defined as neurocognitive processes that maintain an appropriate problem-solving set to attain a later goal. To examine the validity of the EF theory, we conducted a meta-analysis of 83 studies that administered EF measures to groups with ADHD (total N = 3734) and without ADHD (N = 2969). Groups with ADHD exhibited significant impairment on all EF tasks. Effect sizes for all measures fell in the medium range (.46-.69), but the strongest and most consistent effects were obtained on measures of response inhibition, vigilance, working memory, and planning. Weaknesses in EF were significant in both clinic-referred and community samples and were not explained by group differences in intelligence, academic achievement, or symptoms of other disorders. ADHD is associated with significant weaknesses in several key EF domains. However, moderate effect sizes and lack of universality of EF deficits among individuals with ADHD suggest that EF weaknesses are neither necessary nor sufficient to cause all cases of ADHD. Difficulties with EF appear to be one important component of the complex neuropsychology of ADHD.


Variable patterns of expression of luciferase in transgenic tobacco leaves.
A carboxyl-terminally modified firefly luciferase, encoded as a gene fusion to the neomycin phosphotransferase gene (which confers kanamycin resistance), was found to be enzymatically active for both enzymes when expressed in bacteria and in transgenic plants. A military-type starlight vision system was used to conveniently analyze the pattern of gene expression in transgenic tobacco plant leaves. Transgenic tobacco plants which expressed luciferase uniformly in all areas of the leaf, and assays for luciferin, demonstrated that luciferin rapidly penetrates all regions of a tobacco leaf in at least two dimensions. Depending on the test gene structure or, presumably, on the transferred DNA (T-DNA) insertional context, other transgenic plants were obtained that expressed luciferase with a wide range of nonuniform patterns from nominally the same cauliflower mosaic virus 35S promoter. For instance, the veins can be dark, while only the interveinal regions of the leaf lamina glow, or only the small capillary veins glow, or only the major veins glow. Local and/or systemic induction in response to wounding was also demonstrated.


Low-bandwidth, low-cost telemedicine consultations in rural family practice.
Telemedicine, based on the use of interactive video consultations, is being used more commonly in rural settings. This development is potentially important to rural patients because there are fewer physicians, particularly specialist physicians, in rural areas. Declining costs of telemedicine equipment and transmission have created increased access to these technologies for rural family physicians and their patients. This study considers satisfaction levels of rural family physicians, academic-based specialists, and rural patients in 130 consultations between rural physicians, rural patients, and urban academic specialists. To increase the practicability for rural use, low-cost equipment and low-bandwidth digital telephone transmission lines were utilized. Data were collected using questionnaires that were completed by patients, family physicians, and specialist consultants after each consultation. All categories of participants noted very high levels of satisfaction. Telemedicine-based consultations are well accepted by rural patients, rural family physicians, and urban academic specialist consultants. This approach could offer a useful adjunct to rural health care.