The influence of fasting on the disposition of warfarin in rats.
A study was undertaken to investigate the influence of fasting on the disposition of warfarin in rats. Fasting consisted of withholding solid food, but not water, immediately following warfarin (3 or 10 mg/kg s.c.) administration until sacrifice at different time intervals (3, 6, 12, 24 and 30 hours). Control animals were fed ad libitum. Total and unbound warfarin concentrations were measured in plasma and liver supernatant as well as unchanged warfarin and its metabolites in urine. The disposition of unbound warfarin was found to be markedly affected by fasting, especially at the 3 mg/kg dose: the disappearance rate of unbound warfarin from plasma was accelerated in fasted animals in contrast to that of total warfarin. In addition, unbound warfarin was cleared from plasma at a more rapid rate than total warfarin in both control and fasted animals. At the 10 mg/kg dose, the disposition of total and unbound warfarin was little affected by fasting. The concentration of unbound warfarin in the liver supernatant of fasted rats given warfarin, 3 mg/kg, was significantly increased at 6 and 24 hours of fasting. Plasma free fatty acids were significantly elevated starting at 6 hours, but no such difference was noticed with liver homogenate, except at 24 hours. The 24-hour urinary excretion of unchanged warfarin was higher in fasted rats, but fasting failed to produce any change in the excretion of warfarin metabolites. The results of the present investigation indicate that short periods of fasting influence the disposition of unbound warfarin without apparently modifying its biotransformation and further show the importance of plasma protein binding on the pharmacokinetics of warfarin.


Use of model organism and disease databases to support matchmaking for human disease gene discovery.
The Matchmaker Exchange application programming interface (API) allows searching a patient's genotypic or phenotypic profiles across clinical sites, for the purposes of cohort discovery and variant disease causal validation. This API can be used not only to search for matching patients, but also to match against public disease and model organism data. This public disease data enable matching known diseases and variant-phenotype associations using phenotype semantic similarity algorithms developed by the Monarch Initiative. The model data can provide additional evidence to aid diagnosis, suggest relevant models for disease mechanism and treatment exploration, and identify collaborators across the translational divide. The Monarch Initiative provides an implementation of this API for searching multiple integrated sources of data that contextualize the knowledge about any given patient or patient family into the greater biomedical knowledge landscape. While this corpus of data can aid diagnosis, it is also the beginning of research to improve understanding of rare human diseases.


A membrane protein required for dislocation of misfolded proteins from the ER.
After insertion into the endoplasmic reticulum (ER), proteins that fail to fold there are destroyed. Through a process termed dislocation such misfolded proteins arrive in the cytosol, where ubiquitination, deglycosylation and finally proteasomal proteolysis dispense with the unwanted polypeptides. The machinery involved in the extraction of misfolded proteins from the ER is poorly defined. The human cytomegalovirus-encoded glycoproteins US2 and US11 catalyse the dislocation of class I major histocompatibility complex (MHC) products, resulting in their rapid degradation. Here we show that US11 uses its transmembrane domain to recruit class I MHC products to a human homologue of yeast Der1p, a protein essential for the degradation of a subset of misfolded ER proteins. We show that this protein, Derlin-1, is essential for the degradation of class I MHC molecules catalysed by US11, but not by US2. We conclude that Derlin-1 is an important factor for the extraction of certain aberrantly folded proteins from the mammalian ER.


B-cell memory in malaria: Myths and realities.
B-cell and antibody responses to Plasmodium spp., the parasite that causes malaria, are critical for control of parasitemia and associated immunopathology. Antibodies also provide protection to reinfection. Long-lasting B-cell memory has been shown to occur in response to Plasmodium spp. in experimental model infections, and in human malaria. However, there are reports that antibody responses to several malaria antigens in young children living with malaria are not similarly long-lived, suggesting a dysfunction in the maintenance of circulating antibodies. Some studies attribute this to the expansion of atypical memory B cells (AMB), which express multiple inhibitory receptors and activation markers, and are hyporesponsive to B-cell receptor (BCR) restimulation in vitro. AMB are also expanded in other chronic infections such as tuberculosis, hepatitis B and C, and HIV, as well as in autoimmunity and old age, highlighting the importance of understanding their role in immunity. Whether AMB are dysfunctional remains controversial, as there are also studies in other infections showing that AMB can produce isotype-switched antibodies and in mouse can contribute to protection against infection. In light of these controversies, we review the most recent literature on either side of the debate and challenge some of the currently held views regarding B-cell responses to Plasmodium infections.


Rapid Cycling Genomic Selection in a Multiparental Tropical Maize Population.
Genomic selection (GS) increases genetic gain by reducing the length of the selection cycle, as has been exemplified in maize using rapid cycling recombination of biparental populations. However, no results of GS applied to maize multi-parental populations have been reported so far. This study is the first to show realized genetic gains of rapid cycling genomic selection (RCGS) for four recombination cycles in a multi-parental tropical maize population. Eighteen elite tropical maize lines were intercrossed twice, and self-pollinated once, to form the cycle 0 (C0) training population. A total of 1000 ear-to-row C0 families was genotyped with 955,690 genotyping-by-sequencing SNP markers; their testcrosses were phenotyped at four optimal locations in Mexico to form the training population. Individuals from families with the best plant types, maturity, and grain yield were selected and intermated to form RCGS cycle 1 (C1). Predictions of the genotyped individuals forming cycle C1 were made, and the best predicted grain yielders were selected as parents of C2; this was repeated for more cycles (C2, C3, and C4), thereby achieving two cycles per year. Multi-environment trials of individuals from populations C0, C1, C2, C3, and C4, together with four benchmark checks were evaluated at two locations in Mexico. Results indicated that realized grain yield from C1 to C4 reached 0.225 ton ha-1 per cycle, which is equivalent to 0.100 ton ha-1 yr-1 over a 4.5-yr breeding period from the initial cross to the last cycle. Compared with the original 18 parents used to form cycle 0 (C0), genetic diversity narrowed only slightly during the last GS cycles (C3 and C4). Results indicate that, in tropical maize multi-parental breeding populations, RCGS can be an effective breeding strategy for simultaneously conserving genetic diversity and achieving high genetic gains in a short period of time.


Diffusion of digital breast tomosynthesis among women in primary care: associations with insurance type.
Digital breast tomosynthesis (DBT) has shown potential to improve breast cancer screening and diagnosis compared to digital mammography (DM). The FDA approved DBT use in conjunction with conventional DM in 2011, but coverage was approved by CMS recently in 2015. Given changes in coverage policies, it is important to monitor diffusion of DBT by insurance type. This study examined DBT trends and estimated associations with insurance type. From June 2011 to September 2014, DBT use in 22 primary care centers in the Dartmouth -Brigham and Women's Hospital Population-based Research Optimizing Screening through Personalized Regimens research center (PROSPR) was examined among women aged 40-89. A longitudinal repeated measures analysis estimated the proportion of DBT performed for screening or diagnostic indications over time and by insurance type. During the study period, 93,182 mammograms were performed on 48,234 women. Of these exams, 16,506 DBT tests were performed for screening (18.1%) and 2537 were performed for diagnosis (15.7%). Between 2011 and 2014, DBT utilization increased in all insurance groups. However, by the latest observed period, screening DBT was used more frequently under private insurance (43.4%) than Medicaid (36.2%), Medicare (37.8%), other (38.6%), or no insurance (32.9%; P < 0.0001). No sustained differences in use of DBT for diagnostic testing were seen by insurance type. DBT is increasingly used for breast cancer screening and diagnosis. Use of screening DBT may be associated with insurance type. Surveillance is required to ensure that disparities in breast cancer screening are minimized as DBT becomes more widely available.


Human Hepatocyte-Derived Induced Pluripotent Stem Cells: MYC Expression, Similarities to Human Germ Cell Tumors, and Safety Issues.
Induced pluripotent stem cells (iPSC) are a most promising approach to the development of a hepatocyte transplantable mass sufficient to induce long-term correction of inherited liver metabolic diseases, thus avoiding liver transplantation. Their intrinsic self-renewal ability and potential to differentiate into any of the three germ layers identify iPSC as the most promising cell-based therapeutics, but also as drivers of tumor development. Teratoma development currently represents the gold standard to assess iPSC pluripotency. We analyzed the tumorigenic potential of iPSC generated from human hepatocytes (HEP-iPSC) and compared their immunohistochemical profiles to that of tumors developed from fibroblast and hematopoietic stem cell-derived iPSC. HEP-iPSC generated tumors significantly presented more malignant morphological features than reprogrammed fibroblasts or CD34+ iPSC. Moreover, the protooncogene myc showed the strongest expression in HEP-iPSC, compared to only faint expression in the other cell subsets. Random integration of transgenes and the use of potent protooncogenes such as myc might be a risk factor for malignant tumor development if hepatocytes are used for reprogramming. Nonviral vector delivery systems or reprogramming of cells obtained from less invasive harvesting methods would represent interesting options for future developments in stem cell-based approaches for liver metabolic diseases.


Curcumin suppresses growth of mesothelioma cells in vitro and in vivo, in part, by stimulating apoptosis.
Malignant pleural mesothelioma (MPM) is an aggressive, asbestos-related malignancy of the thoracic pleura. Although, platinum-based agents are the first line of therapy, there is an urgent need for second-line therapies to treat the drug-resistant MPM. Cell cycle as well as apoptosis pathways are frequently altered in MPM and thus remain attractive targets for intervention strategies. Curcumin, the major component in the spice turmeric, alone or in combination with other chemotherapeutics has been under investigation for a number of cancers. In this study, we investigated the biological and molecular responses of MPM cells to curcumin treatments and the mechanisms involved. Flow-cytometric analyses coupled with western immunoblotting and gene-array analyses were conducted to determine mechanisms of curcumin-dependent growth suppression of human (H2373, H2452, H2461, and H226) and murine (AB12) MPM cells. Curcumin inhibited MPM cell growth in a dose- and time-dependent manner while pretreatment of MPM cells with curcumin enhanced cisplatin efficacy. Curcumin activated the stress-activated p38 kinase, caspases 9 and 3, caused elevated levels of proapoptotic proteins Bax, stimulated PARP cleavage, and apoptosis. In addition, curcumin treatments stimulated expression of novel transducers of cell growth suppression such as CARP-1, XAF1, and SULF1 proteins. Oral administration of curcumin inhibited growth of murine MPM cell-derived tumors in vivo in part by stimulating apoptosis. Thus, curcumin targets cell cycle and promotes apoptosis to suppress MPM growth in vitro and in vivo. Our studies provide a proof-of-principle rationale for further in-depth analysis of MPM growth suppression mechanisms and their future exploitation in effective management of resistant MPM.


Neurogenic detrusor overactivity in patients with spinal cord injury: evaluation and management.
Lower urinary tract dysfunction can have a significant impact on patients with spinal cord injury. Over the years, many treatment options have become available. This article reviews the assessment and management of neurogenic detrusor overactivity, with a particular focus on articles from the recent literature. Recent guidelines on the subject will be discussed. Management options include antimuscarinics and bladder emptying measures, botulinum toxin A, and neuromodulation in refractory cases and surgery for intractable cases. Recent and relevant publications in these areas will be summarized and discussed.


Abnormal axon reflex-mediated sweating correlates with high state of anxiety in atopic dermatitis.
Sweating plays a key role in skin homeostasis, including antimicrobial and moisturizing effects, and regulation of skin surface pH. Impaired axon reflex-mediated (AXR) sweating has been observed in patients with atopic dermatitis (AD). However, the mechanism of such abnormal sudomotor axon reflex remains to be revealed. To investigate this mechanism, sudomotor function was analyzed using a quantitative sudomotor axon reflex test (acetylcholine iontophoresis) in patients with AD (n = 26) and healthy volunteers (n = 12). Correlation between sudomotor function and certain background factors, including Spielberger State Trait Anxiety Inventory score, Severity Scoring of Atopic Dermatitis (SCORAD) score, number of circulating eosinophils, and serum concentrations of thymus and activation-regulated chemokine and immunoglobulin E radioimmunosorbent test, was validated. Latency time was significantly prolonged in AD (p = 0.0352), and AXR sweating volume (mg/0-5 min) was significantly lower in AD patients than in healthy controls (p = 0.0441). Direct sweating volume (mg/0-5 min) was comparable in AD patients and healthy controls. A significant correlation between the evaluation results of quantitative sudomotor axon reflex tests and certain background factors was not observed. The latency time in non-lesioned and lesioned areas for AD patients versus continuous anxiety value in the Spielberger State Trait Anxiety Inventory and the AXR versus SCORAD showed significant correlations (p = 0.0424, p = 0.0169, and p = 0.0523, respectively). Although the number of study subjects was little, abnormal AXR sweating in patients with AD was observed. Correlative analysis suggests possible involvement of continuous anxiety and the immune system in such abnormal sudomotor function.


Endogenous circadian rhythms in pigment composition induce changes in photochemical efficiency in plant canopies.
There is increasing evidence that the circadian clock is a significant driver of photosynthesis that becomes apparent when environmental cues are experimentally held constant. We studied whether the composition of photosynthetic pigments is under circadian regulation, and whether pigment oscillations lead to rhythmic changes in photochemical efficiency. To address these questions, we maintained canopies of bean and cotton, after an entrainment phase, under constant (light or darkness) conditions for 30-48 h. Photosynthesis and quantum yield peaked at subjective noon, and non-photochemical quenching peaked at night. These oscillations were not associated with parallel changes in carbohydrate content or xanthophyll cycle activity. We observed robust oscillations of Chl a/b during constant light in both species, and also under constant darkness in bean, peaking when it would have been night during the entrainment (subjective nights). These oscillations could be attributed to the synthesis and/or degradation of trimeric light-harvesting complex II (reflected by the rhythmic changes in Chl a/b), with the antenna size minimal at night and maximal around subjective noon. Considering together the oscillations of pigments and photochemistry, the observed pattern of changes is counterintuitive if we assume that the plant strategy is to avoid photodamage, but consistent with a strategy where non-stressed plants maximize photosynthesis.


Appetitive-based learning in rats: lack of effect of chronic exposure to fluoride.
Chronic ingestion of optimally fluoridated water (ca. 1.0 mg/L) has not been associated with any adverse health effects. Possible effects on the nervous system, however, have received little attention. One study with rats given high doses of fluoride reported subtle behavioral changes. The authors suggested that the ability of humans to learn might be reduced and recommended further study with humans and rats. The present study was done to provide data with which to assess this suggestion. Weanling, female rats (n=32) were provided with water containing graded doses of fluoride (0, 2.9, 5.7, 11.5 mg/kg body weight/day) for eight months. While under restricted food access they were tested for their ability to learn an operant response for food and to adjust their responding under schedules of reinforcement requiring high rates of responding (5 days) and then low rates of responding (10 days). Bone, plasma and seven regions of brain were analyzed for fluoride. There were no significant differences among the groups in learning or performance of the operant tasks. Tissue fluoride concentrations were directly related to the levels of exposure. In the 11.5 mg/kg/day group the bone, plasma and brain concentrations were 99, 305 and 221 times higher, respectively, than those in the control group. The average brain-to-plasma fluoride concentration ratios in each of the seven brain sections fell within a narrow range and did not exceed 0.40. There was no consistent evidence for the preferential uptake of fluoride by any given brain section. Chronic ingestion of fluoride at levels up to 230 times more than that experienced by humans whose main source of fluoride is fluoridated water had no significant effect on appetitive-based learning.


Butyrylcholinesterase K variant on chromosome 3 q is associated with Type II diabetes in white Caucasian subjects.
To determine the association of three genes associated with Alzheimer's disease--butyrylcholinesterase (BcHE) on chromosome 3 q, alpha2 macroglobulin (alpha2M) on chromosome 12 p and apolipoprotein E (ApoE) on chromosome 19 q--with Type II (non-insulin-dependent) diabetes mellitus. Frequencies of BcHE K variant, alpha2M insertion and/or deletion polymorphism, the ApoE common polymorphisms and promoter variants at ApoE-491 and -291, were examined by fluorescent RFLP in DNA from 276 United Kingdom Prospective Diabetes Study Type II diabetic subjects and 351 non-diabetic subjects from the Diabetes In Families study. Genetic data in diabetic subjects was analysed in relation to clinical characteristics and islet function as assessed by the requirement for insulin therapy 6 years after randomisation. The BcHE K variant allele was more common among Type II diabetic subjects (D) than non-diabetic subjects (ND) (22.8 % D vs 15.8 % ND; p = 0.00 017). Subjects homozygous for the variant were more frequent in the diabetic group (5.8 % D vs 2.6 % ND: p = 0.00 056). The K variant allele frequency was not associated with a requirement for insulin therapy (29.0 % insulin-requiring vs 21.8 % non-insulin-requiring; p = 0.121). There were no associations of alpha2M and ApoE polymorphisms or ApoE promoter variants with clinical characteristics or insulin requirement in diabetic subjects. There were differences in total cholesterol (p = 0.0005) and LDL-cholesterol (p = 0.0009) among non-diabetic subjects in relation to ApoE-491 genotypes. The association of the BcHE gene (3q26) with Type II diabetes could be related to an identified susceptibility locus on chromosome 3q27 but appears to be independent of islet function. The absence of diabetes-specific associations with alpha2M, ApoE or ApoE promoter variants suggest that these are not important in the onset of hyperglycaemia.


Connexins are expressed in primary brain tumors and enhance the bystander effect in gene therapy.
Experimental brain tumor gene therapy with the herpes simplex virus thymidine kinase (HSV-tk) gene has demonstrated that not only HSV-tk transduced but surrounding non-HSV-tk transduced cells are killed when given ganciclovir. This so-called bystander effect has recently been shown to be dependent on connexin-mediated intercellular communication. To assess potential susceptibility to the bystander effect, we examined levels of connexin-26 and connexin-43 expression in a series of primary brain tumors. Connexin-26 expression has not previously been studied in primary brain tumors and connexin-43 expression has not been studied in nonastrocytic primary brain tumors. We also attempted to enhance the bystander effect in vitro by overexpressing connexin in tumor cells with high basal levels of connexin expression. Western blot analysis and immunohistochemistry were used to determine levels of connexin-26 and connexin-43 expression in a series of primary brain tumors. Wild-type 9L gliosarcoma cells were transfected in vitro with the connexin-43 gene and the HSV-tk gene or the HSV-tk gene alone. The bystander effect of each transfectant was then assessed and compared. Most of the primary brain tumors tested, including low-grade astrocytomas, anaplastic astrocytomas, glioblastomas, oligodendrogliomas, gangliogliomas, meningiomas, and medulloblastomas, showed connexin-26 and connexin-43 expression. Bystander experiments revealed a significant enhancement of the bystander effect in the gliosarcoma cells transfected with connexin-43 and HSV-tk, as compared with gliosarcoma cells transfected with HSV-tk alone. Most primary brain tumors express connexin-26 and connexin-43. This suggests that most primary brain tumors may be susceptible to the bystander effect of HSV-tk gene therapy. The bystander effect can be enhanced in vitro by overexpression of connexin-43 in a cell line with a high basal level of connexin-43 expression.


Kangaroo care: national survey of practice, knowledge, barriers, and perceptions.
A national survey was conducted to assess practice, knowledge, barriers, and perceptions regarding Kangaroo Care (KC)--the holding of diaper-clad preterm infants skin-to-skin, chest-to-chest by parents. A descriptive survey was conducted. Kangaroo Care Questionnaires (KCQs), developed for the study, were sent to nurse managers in all hospitals in the United States that were identified as providing neonatal intensive care services (N = 1,133), and were to be completed by the nurse most familiar with the practice of KC in that unit. A second KCQ was sent to non-respondents. Descriptive statistics were used to summarize the data. A response rate of 59% (N = 537) was achieved. Over 82% of the respondents reported practicing KC in their neonatal intensive care units (NICUs). Nurses were knowledgeable about KC. Major barriers to practicing KC for certain types of infants were infant safety concerns, as well as reluctance by nurses, physicians, and families to initiate or participate in KC. Many NICUs do not permit KC for certain types of infants (e.g., those on vasopressors or high-frequency ventilation). Over 60% of respondents agreed that low gestational age or weight were not contraindications. Respondents from NICUs in which KC is practiced were more positive in their perceptions than respondents from NICUs that do not practice KC. The findings suggest that in order to overcome barriers to the practice of KC, nurses need educational offerings highlighting the knowledge and skills needed to provide KC safely and effectively. These educational offerings should also emphasize the value of KC to infants and parents. In addition, knowledgeable practitioners need to develop evidence-based policies and procedures that will lead to successful KC.


Anatomical Atlas of the Quail's Ear (Coturnix coturnix).
This study aims to enhance the anatomical knowledge of the ear of the adult quail (Coturnix coturnix) through the creation of a scaled 3D model utilizing data from micro-CT images. In addition, 17 annotated histological sections of the quail's ear are aligned to their 3D position in the model. The resulting anatomical atlas provides an intuitive insight into the 3D anatomy and can be used for medical education. The model also allows measuring anatomical structures and can thus serve as reference for the quail's auricular anatomy and as a basis to evaluate clinical diagnostic imaging results.


Functional analysis of three CYP1A2 variants found in a Japanese population.
Human cytochrome P450 1A2 (CYP1A2) catalyzes the metabolism of many important drugs and environmental chemicals. We previously reported three naturally occurring genetic polymorphisms (125C>G, Pro42Arg, CYP1A2*15; 1130G>A, Arg377Gln, *16; and 1367G>A, Arg456His, *8) found in a Japanese population. In this study, these variant enzymes were expressed in Chinese hamster V79 cells, and their mRNA and protein expression levels as well as catalytic activities were determined. All three variant enzymes showed reduced protein expression levels (66% for Pro42Arg and approximately 30% for Arg377Gln and Arg456His) compared with that of the wild type (WT) without any change in mRNA expression levels. Kinetic analysis for 7-ethoxyresorufin O-deethylation revealed that V(max) and V(max)/K(m) of all three variants were less than 3 and 1% of the WT, respectively, although the K(m) value was significantly increased only in the Arg377Gln variant (approximately a 9-fold increase). Markedly reduced activities of the three variants were also observed for phenacetin O-deethylation. In the reduced CO difference spectral analysis using recombinant proteins produced in the Sf21/baculovirus system, the peak at 450 nm seen in the WT protein was hardly observed in the three variants, suggesting marked reductions in their hemoprotein formation. These results suggest that Pro42, Arg377, and Arg456 are critical amino acids for the production of catalytically active CYP1A2 holoenzyme.


Nitric oxide induces the synthesis of vascular endothelial growth factor by rat vascular smooth muscle cells.
Vascular endothelial growth factor (VEGF) is known to induce the release of nitric oxide (NO) from endothelial cells. However, the effect of NO on VEGF synthesis is not clear. Accordingly, the effect of endogenous and exogenous NO on VEGF synthesis by rat vascular smooth muscle cells (VSMCs) was investigated. Two in vitro models were used: (1) VSMCs stimulated to produce NO by treatment with interleukin (IL)-1beta (10 ng/mL) and (2) VSMCs lipotransfected with pKecNOS plasmid, containing the endothelial constitutive NO synthase (ecNOS) cDNA. The synthesis of NO was inhibited by N(omega)-nitro-L-arginine methyl ester (L-NAME, 2 to 5 mmol/L) or diaminohydroxypyrimidine (DAHP, 2.5 to 5 mmol/L), inhibitors of NOS and GTP cyclohydrolase I, respectively. Some cells treated with L-NAME or DAHP were supplemented with L-arginine (10 mmol/L) or tetrahydrobiopterin (BH(4); 100 micromol/L), respectively. In addition, we studied the effect of sodium nitroprusside (SNP; 10 and 100 micromol/L) and chemically related compounds, potassium ferrocyanide and ferricyanide, on VEGF generation. IL-1beta induced iNOS expression and NO generation and significantly upregulated VEGF mRNA expression and protein synthesis. L-NAME and DAHP totally inhibited NO generation and decreased the IL-1beta-upregulated VEGF synthesis by 30% to 40%. Supplementation with L-arginine or BH(4) increased NO generation by L-NAME- or DAHP-treated cells, and VEGF synthesis was augmented by addition of BH(4). The cells generating NO after pKecNOS transfection released significantly higher amounts of VEGF than cells transfected with control plasmids. Inhibition of NO generation by L-NAME decreased VEGF synthesis. In contrast to the effect of endogenous NO, we observed the inhibition of VEGF synthesis in the presence of high (10 or 100 micromol/L) concentrations of SNP. This effect was mimicked by chemically related ferricyanide and ferrocyanide compounds, suggesting that the inhibitory effect of sodium nitroprusside may be mediated by an NO-independent mechanism. The results indicate that endogenous NO enhances VEGF synthesis. The positive interaction between endogenous NO and VEGF may have implications for endothelial regeneration after balloon angioplasty and for angiogenesis.


Stroke Incidence by Sex Across the Lifespan.
We evaluated the influence of age on the association between sex and the incidence of stroke or transient ischemic attack (TIA) using a population-based cohort from Ontario, Canada. We followed a cohort of adults (≥18 years) without prior stroke from January 1, 2003 (cohort start date) to March 31, 2018, to identify incident events. We calculated hazard ratios (HRs), in women compared to men, of incident stroke or TIA, adjusted for demographics and comorbidities, overall and stratified by stroke type. We calculated piecewise adjusted HRs for each decade of age to evaluate the effect of age on sex differences in stroke incidence. We followed 9.2 million adults for a median of 15 years and observed 280,197 incident stroke or TIA events. Compared with men, women had an overall lower adjusted hazard of stroke or TIA (HR, 0.82 [95% CI, 0.82-0.83]), with similar findings across all stroke types except for subarachnoid hemorrhage (HR, 1.29 [95% CI, 1.24-1.33]). We found a U-shaped association between age and sex differences in the incidence of stroke or TIA: compared with men, the hazard of stroke was higher in women among those aged ≤30 years (HR, 1.26 [95% CI, 1.10-1.45]), lower among those between ages 40 and 80 years (eg, age 50-59, HR, 0.69 [95% CI, 0.68-0.70]), and similar among those aged ≥80 years (HR, 0.99 [95% CI, 0.98-1.01]). Overall, women have a lower hazard of stroke than men, but this association varies by age and across stroke types. Recognition of age-sex variations in stroke incidence can help guide prevention efforts to reduce stroke incidence in both men and women.


Vegan diets: practical advice for athletes and exercisers.
With the growth of social media as a platform to share information, veganism is becoming more visible, and could be becoming more accepted in sports and in the health and fitness industry. However, to date, there appears to be a lack of literature that discusses how to manage vegan diets for athletic purposes. This article attempted to review literature in order to provide recommendations for how to construct a vegan diet for athletes and exercisers. While little data could be found in the sports nutrition literature specifically, it was revealed elsewhere that veganism creates challenges that need to be accounted for when designing a nutritious diet. This included the sufficiency of energy and protein; the adequacy of vitamin B12, iron, zinc, calcium, iodine and vitamin D; and the lack of the long-chain n-3 fatty acids EPA and DHA in most plant-based sources. However, via the strategic management of food and appropriate supplementation, it is the contention of this article that a nutritive vegan diet can be designed to achieve the dietary needs of most athletes satisfactorily. Further, it was suggested here that creatine and β-alanine supplementation might be of particular use to vegan athletes, owing to vegetarian diets promoting lower muscle creatine and lower muscle carnosine levels in consumers. Empirical research is needed to examine the effects of vegan diets in athletic populations however, especially if this movement grows in popularity, to ensure that the health and performance of athletic vegans is optimised in accordance with developments in sports nutrition knowledge.


Hexachlorobenzene promotes angiogenesis in vivo, in a breast cancer model and neovasculogenesis in vitro, in the human microvascular endothelial cell line HMEC-1.
Exposure to environmental pollutants may alter proangiogenic ability and promotes tumor growth. Hexachlorobenzene (HCB) is an organochlorine pesticide found in maternal milk and in lipid foods, and a weak ligand of the aryl hydrocarbon receptor (AhR). HCB induces migration and invasion in human breast cancer cells, as well as tumor growth and metastasis in vivo. In this study, we examined HCB action on angiogenesis in mammary carcinogenesis. HCB stimulates angiogenesis and increases vascular endothelial growth factor (VEGF) expression in a xenograft model with the human breast cancer cell line MDA-MB-231. Human microvascular endothelial cells HMEC-1 exposed to HCB (0.005, 0.05, 0.5 and 5μM) showed an increase in cyclooxygenase-2 (COX-2) and VEGF protein expression involving AhR. In addition, we found that HCB enhances VEGF-Receptor 2 (VEGFR2) expression, and activates its downstream pathways p38 and ERK1/2. HCB induces cell migration and neovasculogenesis in a dose-dependent manner. Cells pretreatment with AhR, COX-2 and VEGFR2 selective inhibitors, suppressed these effects. In conclusion, our results show that HCB promotes angiogenesis in vivo and in vitro. HCB-induced cell migration and tubulogenesis are mediated by AhR, COX-2 and VEGFR2 in HMEC-1. These findings may help to understand the association among HCB exposure, angiogenesis and mammary carcinogenesis.


Trocar-assisted intrascleral sutureless fixation of a dislocated three-piece sulcus intraocular lens.
Trocar-assisted intraocular lens (IOL) reposition surgery using a scleral fixation method was performed for a patient with a dislocated sulcus IOL. Two 3-mm-long scleral tunnels 2 mm from and parallel to the limbus were formed using a 23-gauge vitrectomy trocar transconjunctivally entering the sclera at an angle of approximately 10°. Haptics were collected using a 23-gauge serrated retinal forceps entering from the trocar cannula and externalized from the scleral tunnels together with the trocar. The same procedure was applied for the other haptic. Both haptics were pushed into the scleral tunnel and a transconjunctival secure 10-0 nylon suture was placed at the scleral tunnel entry site around the haptic. Sutures were removed 1 week later. No complications occurred intraoperatively or postoperatively. At a 10-month follow-up, IOL was stabilized. IOL reposition surgery using the trocar-assisted IOL scleral fixation method is a viable alternative to intrascleral fixation surgery.


Patient derived organoids to model rare prostate cancer phenotypes.
A major hurdle in the study of rare tumors is a lack of existing preclinical models. Neuroendocrine prostate cancer is an uncommon and aggressive histologic variant of prostate cancer that may arise de novo or as a mechanism of treatment resistance in patients with pre-existing castration-resistant prostate cancer. There are few available models to study neuroendocrine prostate cancer. Here, we report the generation and characterization of tumor organoids derived from needle biopsies of metastatic lesions from four patients. We demonstrate genomic, transcriptomic, and epigenomic concordance between organoids and their corresponding patient tumors. We utilize these organoids to understand the biologic role of the epigenetic modifier EZH2 in driving molecular programs associated with neuroendocrine prostate cancer progression. High-throughput organoid drug screening nominated single agents and drug combinations suggesting repurposing opportunities. This proof of principle study represents a strategy for the study of rare cancer phenotypes.


Modeling the impact of alternative strategies for rapid molecular diagnosis of tuberculosis in Southeast Asia.
Novel diagnostic tests hold promise for improving tuberculosis (TB) control, but their epidemiologic impact remains uncertain. Using data from the World Health Organization (2011-2012), we developed a transmission model to evaluate the deployment of 3 hypothetical TB diagnostic tests in Southeast Asia under idealized scenarios of implementation. We defined diagnostics by their sensitivity for smear-negative TB and proportion of patients testing positive who initiate therapy ("point-of-care amenability"), with tests of increasing point-of-care amenability having lower sensitivity. Implemented in the public sector (35% of care-seeking attempts), each novel test reduced TB incidence by 7%-9% (95% uncertainty range: 4%-13%) and mortality by 20%-22% (95% uncertainty range: 14%-27%) after 10 years. If also deployed in the private sector (65% of attempts), these tests reduced incidence by 13%-16%, whereas a perfect test (100% sensitivity and treatment initiation) reduced incidence by 20%. Annually detecting 20% of prevalent TB cases through targeted screening (70% smear-negative sensitivity, 85% treatment initiation) also reduced incidence by 19%. Sensitivity and point-of-care amenability are equally important considerations when developing novel diagnostic tests for TB. Novel diagnostics can substantially reduce TB incidence and mortality in Southeast Asia but are unlikely to transform TB control unless they are deployed actively and in the private sector.


Golf courses and wetland fauna.
Golf courses are often considered to be chemical-intensive ecosystems with negative impacts on fauna. Here we provide evidence that golf courses can contribute to the support and conservation of wetland fauna, i.e., amphibians and macroinvertebrates. Comparisons of amphibian occurrence, diversity of macroinvetebrates, and occurrence of species of conservation concern were made between permanent freshwater ponds surveyed on golf courses around Sweden's capital city, Stockholm, and off-course ponds in nature-protected areas and residential parklands. A total of 71 macroinvertebrate species were recorded in the field study, with no significant difference between golf course ponds and off-course ponds at the species, genus, or family levels. A within-group similarities test showed that golf course ponds have a more homogenous species composition than ponds in nature-protected areas and ponds in residential parkland. Within the macroinvertebrate group, a total of 11 species of odonates were identified, with no difference detected between the categories of ponds, nor any spatial autocorrelation. Significant differences were found between pond categories in the occurrence of five species of amphibians, although anuran occurrence did not differ between ponds. The great crested newt (Triturus cristatus) was significantly associated with golf course ponds, but the smooth newt (Triturus vulgaris) was not. We found no evidence of any correlation between pond size and occurrence of amphibians. Among the taxa of conservation concern included in the sample, all amphibians are nationally protected in Sweden, with the internationally threatened T. cristatus more frequently found in golf course ponds. Among macroinveterbrates of conservation status, the large white-faced darter dragonfly (Leucorrhinia pectoralis) was only detected in golf course ponds, and Tricholeiochiton fagesi (Trichoptera) was only found in one off-course pond. GIS results revealed that golf courses provide over a quarter of all available permanent, freshwater ponds in central greater Stockholm. We assert that golf courses have the potential to contribute to wetland fauna support, particularly in urban settings where they may significantly contribute to wetland creation. We propose a greater involvement of ecologists in the design of golf courses to further bolster this potential.


A household-based survey of knowledge, attitudes and practices towards dengue fever among local urban communities in Taiz Governorate, Yemen.
Yemen has witnessed several dengue fever outbreaks coincident with the social unrest and war in the country. The aim of the present study was to describe the knowledge, attitudes and practices (KAPs) of at-risk urban populations residing in Taiz, southwest of Yemen. In addition, factors possibly associated with poor preventive practices were investigated. A household-based, cross-sectional survey was conducted in three urban districts encompassing 383 households. Data on the socio-demographic characteristics and KAPs of the participating household heads were collected using a pre-designed, structured questionnaire. The association of socio-demographic characteristics, knowledge and attitudes of the population with poor preventive practices against dengue fever was then analyzed using logistic regression. More than 90.0 % of respondent household heads had correct knowledge about fever, headache and joint pain as common signs and symptoms of dengue fever. Moreover, muscular pain and bleeding were perceived by more than 80.0 % of the respondents as being associated with dengue fever; however, only 65.0 % of the respondents reported skin rash as a sign of dengue fever. More than 95.0 % of respondents agreed about the seriousness and possible transmission of dengue fever; however, negative attitudes regarding the facts of being at risk of the disease and that the infection is preventable were expressed by 15.0 % of respondents. Despite the good level of knowledge and attitudes of the respondent population, poor preventive practices were common. Bivariate analysis identified poor knowledge of dengue signs and symptoms (OR = 2.1, 95 % CI = 1.24-3.68; P = 0.005) and its vector (OR = 2.1, 95 % CI = 1.14-3.84; P = 0.016) as factors significantly associated with poor preventive practices. However, multivariable analysis showed that poor knowledge of the vector is an independent predictor of poor preventive practices of the population (adjusted OR = 2.1, 95 % CI = 1.14-3.84; P = 0.018). The majority of people in urban communities of Taiz have a clear understanding of most signs/symptoms of dengue fever as well as positive attitudes towards the seriousness and possible transmissibility of dengue fever. However, negative attitudes regarding their perception of the risk and possible prevention of the infection are prevailing among a small proportion of the population and need to be targeted by educational campaigns. It appears that the good level of the population knowledge of the signs/symptoms of dengue fever and the factors contributing to the spread and control of its vectors did not translate into good practices.


Structural insights into the evolution of a non-biological protein: importance of surface residues in protein fold optimization.
Phylogenetic profiling of amino acid substitution patterns in proteins has led many to conclude that most structural information is carried by interior core residues that are solvent inaccessible. This conclusion is based on the observation that buried residues generally tolerate only conserved sequence changes, while surface residues allow more diverse chemical substitutions. This notion is now changing as it has become apparent that both core and surface residues play important roles in protein folding and stability. Unfortunately, the ability to identify specific mutations that will lead to enhanced stability remains a challenging problem. Here we discuss two mutations that emerged from an in vitro selection experiment designed to improve the folding stability of a non-biological ATP binding protein. These mutations alter two solvent accessible residues, and dramatically enhance the expression, solubility, thermal stability, and ligand binding affinity of the protein. The significance of both mutations was investigated individually and together, and the X-ray crystal structures of the parent sequence and double mutant protein were solved to a resolution limit of 2.8 and 1.65 A, respectively. Comparative structural analysis of the evolved protein to proteins found in nature reveals that our non-biological protein evolved certain structural features shared by many thermophilic proteins. This experimental result suggests that protein fold optimization by in vitro selection offers a viable approach to generating stable variants of many naturally occurring proteins whose structures and functions are otherwise difficult to study.


Benefits of pomegranate (Punica granatum Linn) fruit extracts to weight changes, total protein, and uric acid in white rats (Rattus norvegicus) as an animal model of acute renal failure.
The occurrence of acute renal failure (ARF) cases continues to increase every year. Some of the cases are due to nephrotoxic effect caused by overdose of antibiotic consumption or abuse of the drug, gentamicin. An antibiotic therapy that can be used to overcome in such a case is the pomegranate extracts. However, until now, studies using pomegranate for cases of ARF have not been done. This study aims to determine changes in body weight, the levels of total protein (TP), and the levels of serum uric acid (UA) as a result of the pomegranate extract consumption. A total number of 32 rats (Rattus norvegicus) were divided into four groups randomly. One group was assigned as the control group (P0) and given intraperitoneal (i.p.) saline and 0.3% carboxy methyl cellulose sodium (CMC) Na; P1 was provided with 80 mg/kg bw/i.p. gentamicin and 0.3% CMC Na orally, P2 was supplied with 80 mg/kg bw/i.p. gentamicin and ellagic acid in 0.3% CMC Na, and P3 was given 80 mg/kg bw/i.p. gentamicin and 150 mg/kg bw pomegranate extract in 0.3% CMC Na. The provision of treatment was carried out in 8 days and followed by making the overthrow of body weight and blood sampling for the examination of study variables. The results taken by doing the analysis of variance method for the four treatment groups show that the control group (P0) has significant differences from P1, P2, and P3 (p<0.05), but there are no significant differences among the other three treatment groups. Meanwhile, the average values of serum UA levels among P1, P2, and P3 indicate significant differences. In conclusion, the administration of pomegranate extracts in the treatment of nephrotoxicity toward rats is effective to maintain normal body weight, normal TP levels, and the UA blood serum of the rats. As this study is a preventive therapy, it needs further researches about the effective dose as a curative therapy, its level of effectiveness and its long-term side effects.


In vitro tests of the validity of singlet oxygen luminescence measurements as a dose metric in photodynamic therapy.
Singlet oxygen ((1)O(2)) is widely believed to be the major cytotoxic agent involved in photodynamic therapy (PDT). We showed recently that measurement of the weak near infrared luminescence of (1)O(2) is possible in cells in vitro and tissues in vivo. Here, we investigated the relationship between the integrated luminescence signal and the in vitro PDT response of AML5 leukemia cells sensitized with aminolevulinic acid-induced protoporphyrin IX (PpIX). Sensitized cell suspensions were irradiated with pulsed 523 nm laser light at average fluence rates of 10, 25, or 50 mWcm(-2) and, (1)O(2) luminescence measurements were made throughout the treatment. Cell survival was measured with either propidium iodide-labeled flow cytometry or colony-forming assay. The PpIX concentration in the cells, the photobleaching, and the pO(2) in the cell suspensions were also monitored. There were large variations in cell survival and (1)O(2) generation in different experiments due to different controlled treatment parameters (fluence and fluence rate) and other uncontrolled factors (PpIX synthesis and oxygenation). However, in all of the cases, cell kill correlated strongly with the cumulative (1)O(2) luminescence and allowed direct estimation of the (1)O(2) per cell required to achieve a specific level of cell kill. This study supports the validity and potential utility of (1)O(2) luminescence measurement as a dosimetric tool for PDT, as well as confirming the likely role of (1)O(2) in porphyrin-based PDT.


Copper deficiency myelopathy: A report of two cases.
Copper deficiency myelopathy represents an often underdiagnosed, acquired neurological syndrome, clinically characterized by posterior column dysfunction. The main causes of copper deficiency are bariatric surgery, increased consumption of zinc, and malabsorption. However, even after a careful history taking and extensive laboratory researches, the etiology of copper deficiency remains undetermined in a significant percentage of cases. Patients affected by copper deficiency myelopathy usually present with sensory ataxia due to dorsal column dysfunction and sometimes with mild leg spasticity. In such patients, spinal cord magnetic resonance imaging (MRI) may show hyperintense lesions in T2-weighted sequences involving the posterior columns of cervical and thoracic cord. These MRI findings are not distinguishable from those of subacute combined degeneration associated with vitamin B12 deficiency. Here, we describe two patients with gait ataxia and sensory symptoms in which a diagnosis of copper deficiency myelopathy was made. Both patients showed a significant clinical, neuroradiological, and neurophysiological improvement after proper supplementation therapy. The patients herein described underline the importance to include serum copper and ceruloplasmin levels as part of the myelopathy diagnostic workup, especially in the cases of otherwise unexplained subacute myelopathy involving the posterior columns. Since copper deficiency myelopathy is a progressive syndrome, early diagnosis is mandatory in order to promptly provide a proper supplementation therapy and, thus, prevent an irreversible neurological damage.


Large polypeptides of 10S DNA polymerase alpha from calf thymus: rapid isolation using monoclonal antibody and tryptic peptide mapping analysis.
The polypeptides recognized by a monoclonal antibody against calf thymus DNA polymerase alpha (secreted from a hybridoma CL22 -2- 42B , Nucleic Acids Res. (1982) 10, 4703-4713) were identified by the immunoblot method as the large polypeptides of the partially-purified 10S DNA polymerase alpha fraction. Using an immunoprecipitation technique with the monoclonal antibody, a rapid immunological isolation of the polypeptides has been achieved. By this method, the large polypeptides with Mr = 140,000, 145,000, and 150,000 were isolated from a partially-purified preparation of 10S DNA polymerase alpha. On the other hand, the polypeptides with Mr = 150,000, 180,000, and 240,000 were obtained from a crude extract of calf thymus. Tryptic peptide maps showed that the large polypeptides with Mr = 150,000, and 180,000 were very similar in primary structure and that the structures of Mr = 180,000 and 240,000 polypeptides contained partially common sequences. Among these polypeptides, the Mr = 150,000 polypeptide was shown to correlate with the enzyme activity. These results suggest that the large polypeptide of 10S DNA polymerase alpha is initially synthesized as Mr = 180,000 or larger polypeptide, then converted to the form with Mr = 150,000. The Mr = 140,000 and 145,000 polypeptides in the purified preparation may be artificial products formed during purification.


Influence of stimulus parameters on amplitude-modulated stimulus frequency otoacoustic emissions.
The present study evaluated the influence of suppressor frequency (fs) and level (Ls) on stimulus-frequency otoacoustic emissions (SFOAEs) recorded using the amplitude-modulated (AM) suppressor technique described by Neely et al. [J. Acoust. Soc. Am. 118, 2124-2127 (2005a)]. Data were collected in normal-hearing subjects, with data collection occurring in two phases. In phase 1, SFOAEs were recorded with probe frequency (fp) = 1, 2, and 4 kHz and probe levels (Lp) ranging from 0 to 60 dB sound pressure level (SPL). At each fp, Ls ranged from Ls = Lp to Ls = Lp + 30 dB. Additionally, nine relationships between fs and fp were evaluated, ranging from fs/fp = 0.80 to fs/fp = 1.16. Results indicated that for low suppressor levels, suppressors higher in frequency than fp (fs > fp) resulted in higher AM-SFOAE levels than suppressors lower in frequency than fp (fs < fp). At higher suppressor levels, suppressors both higher and lower in frequency than fp produced similar AM-SFOAE levels, and, in many cases, low-frequency suppressors produced the largest response. Recommendations for stimulus parameters that maximize AM-SFOAE level were derived from these data. In phase 2, AM-SFOAEs were recorded using these parameters for fp = 0.7-8 kHz and Lp = 20-60 dB SPL. Robust AM-SFOAE responses were recorded in this group of subjects using the parameters developed in phase 1.


Attenuation of the effects of punishment by ethanol: comparisons with chlordiazepoxide.
Ethanol (ETOH), like chlordiazepoxide (CDZ), significantly attenuated the suppressive effect of punishment on licking behavior in water-deprived rats and mice. In rats, the greatest effects of ETOH (1.5 g/kg) were observed between 30 and 60 min following IP administration. tert-Butanol also attenuated the effects of punishment, suggesting that acetaldehyde was not contributing to this effect of ETOH. Since a dose of ETOH that increased punished drinking did not increase unpunished drinking, alteration in thirst motivation would not appear to be responsible for its antipunishment action. However, doses of ETOH or CDZ that significantly increased punished responding increased jump thresholds to aversive shock, suggesting that decreased sensitivity to aversive stimulation may contribute to the anti-punishment action of both agents. In addition to these similarities between ethanol and CDZ, several differences were noted in their effects. For example, CDZ decreased serum corticosterone concentration, whereas ETOH did not. Further, ETOH impaired aerial righting reflex and reduced rectal temperature, whereas CDZ had no effect on these parameters at doses that had anti-punishment activity. Finally, specific binding of [3H]flunitrazepam to crude brain cortical membranes was decreased by CDZ, but not ETOH. Although ETOH and CDZ similarly alter punished behavior, results suggest that ETOH does not act through a direct interaction with a benzodiazepine binding site.


Regulation of cell surface beta 1 integrin levels during keratinocyte terminal differentiation.
Integrins of the beta 1 family play a central role in controlling adhesion and terminal differentiation within the epidermis. When human epidermal keratinocytes undergo terminal differentiation, intracellular transport of newly synthesized integrins is inhibited, and mature receptors are lost from the cell surface. We have examined the mechanisms underlying these processes, using an experimental model in which keratinocytes are placed in suspension to induce terminal differentiation. The block in intracellular transport was keratinocyte- and integrin-specific since it was not observed when fibroblasts were placed in suspension and did not affect E-cadherin synthesis in suspended keratinocytes. Newly synthesized beta 1 integrins associated with an endoplasmic reticulum resident protein, calnexin; the association was prolonged when keratinocytes were placed in suspension, suggesting a role for calnexin in the inhibition of transport. After 24 h, the level of beta 1 integrin mRNA declines in suspended keratinocytes, reflecting inhibition of gene transcription, but in fibroblasts, the level remained constant. Transport of integrins could be blocked in both adherent keratinocytes and fibroblasts by inhibiting total protein synthesis, raising the possibility that transport is coupled to de novo integrin synthesis. The fate of receptors on the surface of keratinocytes was followed by confocal immunofluorescence microscopy, immunoelectron microscopy, and biochemical analysis: with the onset of terminal differentiation, endocytosed receptors were transported to the lysosomes. These experiments reveal novel mechanisms by which integrin levels can be controlled. Together with our earlier evidence for transcriptional regulation and affinity modulation of integrins, they highlight the complexity of the mechanisms which ensure that the onset of terminal differentiation is linked to detachment of keratinocytes from the underlying basement membrane.


Manipulation of polarization-dependent effects by magnetostrictive stress on silicon-on-insulator rib waveguides.
Polarization dependent loss (PDL) can cause deterioration in optical network performance owing to the resultant fluctuation in received power because of random changes in fiber polarization. We describe the use of a magnetostrictive layer integrated on a planar light-wave circuit that can offer the functionality of modifying the variable PDL or differential group delay produced within the typical tolerances of volume manufacturing. This approach provides an alternative to the sizable delay lines and splitters that were previously employed for polarization compensation. We demonstrate adjustment of polarization-dependent parameters by the application of an external magnetic field to a ferromagnetic layer adjacent to the waveguide.


Estimating risk difference from relative association measures in meta-analysis can infrequently pose interpretational challenges.
Risk difference (RD) is often estimated from relative association measures generated by meta-analysis and a particular group's baseline risk. We describe a problematic situation in using this approach. We encountered a meta-analysis in which a confidence interval (CI) of relative risk (RR) overlapped 1.0; the point estimate favored treatment A, but when we used RR and median baseline risk to calculate a CI for RD, a greater portion of the CI favored treatment B (a result that some may find counterintuitive). We then calculated 10 different RDs from recently published meta-analyses in outcomes in which CIs of RR crossed 1.0 using three methods: estimation from RR, estimation from the odds ratio, and pooling RDs across trials. When RD is estimated from relative measures, the counterintuitive result occurred in 2 of 10 instances. This discordance of interpretation is because of the logarithmic transformation that makes CIs of relative measures asymmetric around their point estimates. When RD is estimated from relative association measures that are nonsignificant and this counterintuitive situation occurs, it may be more appropriate to pool RD across studies. Pooling is particularly valid when baseline risks across studies are homogeneous.


Infertility Causes and Pregnancy Outcome in Patients With Familial Mediterranean Fever and Controls.
Recurrent attacks of peritonitis due to familial Mediterranean fever (FMF) may lead to peritoneal adhesions and fallopian tube obstruction. Colchicine, which is the treatment of choice for FMF, may disturb cell division. Secondary amyloidosis, a complication of untreated FMF, may involve the testes and ovaries. Thus, FMF and colchicine may potentially affect fertility and pregnancy in patients with FMF. The aims of the study are to evaluate the causes of infertility and pregnancy outcome in FMF patients and to compare them with 2 groups: non-FMF patients with peritoneal female genital tuberculosis (FGTB) and normal healthy controls. This is a retrospective study in which FMF patients with reproductive disorders were recruited from the National Center of Medical Genetics and Primary Health Care in Yerevan, Armenia. The patients with FGTB and the healthy controls with reproductive problems were recruited successively from a large gynecology clinic in Yerevan. Genetic analyses for FMF were performed using ViennaLab StripAssay. The FMF group (211 patients) resembles the FGTB group (127 patients) regarding etiologies of infertility. However, in vitro fertilization (IVF) success rate and pregnancy outcome were comparable between the FMF patients and the control group (162 patients). Infertility in patients with FMF was clearly associated with a more severe disease and a lack of adequate colchicine treatment. Colchicine medication and controlled FMF disease do not adversely affect the reproductive system and pregnancy outcome. However, a lack of an appropriate colchicine treatment may cause infertility and poor pregnancy outcome.


Housing first for homeless persons with active addiction: are we overreaching?
More than 350 communities in the United States have committed to ending chronic homelessness. One nationally prominent approach, Housing First, offers early access to permanent housing without requiring completion of treatment or, for clients with addiction, proof of sobriety. This article reviews studies of Housing First and more traditional rehabilitative (e.g., "linear") recovery interventions, focusing on the outcomes obtained by both approaches for homeless individuals with addictive disorders. According to reviews of comparative trials and case series reports, Housing First reports document excellent housing retention, despite the limited amount of data pertaining to homeless clients with active and severe addiction. Several linear programs cite reductions in addiction severity but have shortcomings in long-term housing success and retention. This article suggests that the current research data are not sufficient to identify an optimal housing and rehabilitation approach for an important homeless subgroup. The research regarding Housing First and linear approaches can be strengthened in several ways, and policymakers should be cautious about generalizing the results of available Housing First studies to persons with active addiction when they enter housing programs.


Sleep apnoea severity independently predicts glycaemic health in nondiabetic subjects: the ESADA study.
Obstructive sleep apnoea (OSA) is associated with increased risk of dysglycaemia but the intimate link of these conditions with obesity makes discerning an independent relationship between them challenging. Glycosylated haemoglobin (HbA1c) levels predict adverse cardiovascular outcomes in nondiabetics but there is a lack of population-level data exploring the relationship of HbA1c with OSA. A cross-sectional analysis of 5294 participants in the multinational European Sleep Apnoea Cohort (European Sleep Apnoea Database) study was performed, assessing the relationship of OSA severity with HbA1c levels in nondiabetic subjects, with adjustment for confounding factors. HbA1c levels correlated significantly with OSA severity in univariate analysis. Following adjustment for confounding factors, apnoea-hypopnoea index (AHI) (standardised β 0.158; p<0.001), along with nocturnal hypoxaemia, predicted HbA1c. Adjusted mean HbA1c levels were lower in the lowest AHI quartile (5.24%, 95% CI 5.21-5.27%) than in the second (5.37%, 95% CI 5.34-5.40%), third (5.44%, 95% CI 5.41-5.47%) or highest (5.50%, 95% CI 5.46-5.53%) quartiles. Subjects in the higher quartiles had significantly greater adjusted odds ratios of HbA1c level ≥6.0% than those in the first quartile. In stratified analyses, OSA severity predicted glycaemic health irrespective of sleep study modality, sex, obesity or daytime sleepiness. OSA severity independently predicts glycaemic health in nondiabetic subjects. Further studies should assess the impact of OSA treatment on glycaemic health and elucidate underlying mechanisms.


Effect of Chronotherapy of Antihypertensives in Chronic Kidney Disease: A Randomized Control Trial.
There is a higher prevalence of non-dipping pattern in hypertensive chronic kidney disease (CKD) patients. Nocturnal hypertension has been shown to predict cardiovascular mortality and morbidity and is often superior to daytime blood pressure. We studied the effect of shifting or adding antihypertensive to night time on blood pressure profile of CKD III-IV patients. In this single-center, prospective, randomized controlled trial, eligible participants were adults from eastern India aged 18-65 years with CKD stages 3 and 4, with a non-dipping pattern on ambulatory blood pressure monitor (ABPM). The intervention group received all the antihypertensives in the night time whereas the standard care group continued to take the medication in the morning. Both groups were followed up for 1 year. The primary outcome was the number of patients changed from non-dippers to dippers in the standard care group and intervention group. Secondary outcomes included a change in estimated glomerular filtration rate (eGFR) and change in the cardiac structure. 39 patients in the intervention group and 36 patients in the standard care group were analyzed. 10 patients (26%) reverted to dipping pattern in the intervention group as compared to none in the standard care group. Mean changes in eGFR were -2.55 and -0.18 mL/min/1.73 m2 in the standard care and intervention group at the end of the study, respectively. Between-group difference in eGFR was significant at 1 year (5.22 [95% CI, 4.3-6.1] ml/min/1.73 m2); (P = 0.03). The cardiac structure showed no significant changes in either group. Bedtime administration of antihypertensives reverted non-dippers to dippers and slowed the decline in eGFR in CKD stages 3 and 4 compared to morning administration of antihypertensives.


Multiple P2Y receptor subtypes in the apical membranes of polarized epithelial cells.
Apical ATP, ATP, UTP and UDP evoked transient increases in short circuit current (I(SC), a direct measure of transepithelial ion transport) in confluent Caco-2 cells grown on permeable supports. These responses were mediated by a population of at least three pharmacologically distinct receptors. Experiments using cells grown on glass coverslips showed that ATP and UTP consistently increased intracellular free calcium ([Ca(2+)](i)) whilst sensitivity to UDP was variable. Cross desensitization experiments suggested that the responses to UTP and ATP were mediated by a common receptor population. Messenger RNA transcripts corresponding to the P2Y(2), P2Y(4) and P2Y(6) receptors genes were detected in cells grown on Transwell membranes by the reverse transcriptase - polymerase chain reaction. Identical results were obtained for cells grown on glass. Experiments in which I(SC) and [Ca(2+)](i) were monitored simultaneously in cells on Transwell membranes, confirmed that apical ATP and UTP increased both parameters and showed that the UDP-evoked increase in I(SC) was accompanied by a [Ca(2+)](i)-signal. Ionomycin consistently increased [Ca(2+)](i) in such polarized cells but caused no discernible change in I(SC). However, subsequent application of apical ATP or UTP evoked a small rise in I(SC) but no rise in [Ca(2+)](i). UDP evoked no such response. As well as evoking increases in [Ca(2+)](i), the ATP/UTP-sensitive receptors present in Caco-2 cells thus allow direct control over ion channels in the apical membrane. The UDP-sensitive receptors, however, appear to simply evoke a rise in [Ca(2+)](i).


Coupled tomography and distinct-element-method approach to exploring the granular media microstructure in a jamming hourglass.
We describe an approach for exploring microscopic properties of granular media that couples x-ray microtomography and distinct-element-method (DEM) simulations through image analysis. We illustrate it via the study of the intriguing phenomenon of instant arching in an hourglass (in our case a cylinder filled with a polydisperse mixture of glass beads that has a small circular shutter in the bottom). X-ray tomography provides three-dimensional snapshots of the microscopic conditions of the system both prior to opening the shutter, and thereafter, once jamming is completed. The process time in between is bridged using DEM simulation, which settles to positions in remarkably good agreement with the x-ray images. Specifically designed image analysis procedures accurately extract the geometrical information, i.e., the positions and sizes of the beads, from the raw x-ray tomographs, and compress the data representation from initially 5 gigabytes to a few tens of kilobytes per tomograph. The scope of the approach is explored through a sensitivity analysis to input data perturbations in both bead sizes and positions. We establish that accuracy of size--much more than position--estimates is critical, thus explaining the difficulty in considering a mixture of beads of different sizes. We further point to limits in the replication ability of granular flows away from equilibrium; i.e., the difficulty of numerically reproducing chaotic motion.


Building a Social Network One Choice at a Time.
Newcomers to a social network show preferential attachment, a tendency to befriend those with many friends. Here, we show that preferential attachment is equivalent to a form of 'probability matching' commonly found in studies of decision-making. This equivalence, whereby newcomers probability match to a social signal akin to popularity, marries network science to the study of decision-making and raises new questions about how individual psychology impacts the social structure of groups. We asked people to view a visualization of a social network and to select group members whom they would like to meet and befriend. People varied in how strongly they weighed popularity and this was mildly correlated with aspects of their personality. Individual differences in preferential attachment affect the structure and connectivity of the network that emerges.


Complete genome sequence of bacteriochlorophyll-synthesizing bacterium Porphyrobacter neustonensis DSM 9434.
The genus Porphyrobacter belongs to aerobic anoxygenic phototrophic bacteria cluster. Porphyrobacter neustonensis DSM 9434 was isolated from a eutrophic freshwater pond in Australia, and is able to synthesize Bacteriochlorophyll a as well as grow under aerobic conditions. It is the type species of the genus Porphyrobacter. Here we describe the characteristics of the strain DSM 9434, including the genome sequence and annotation, synthesis of BChl a, and metabolic pathways of the organism. The genome of strain DSM 9434 comprises 3,090,363 bp and contains 2,902 protein-coding genes, 47 tRNA genes and 6 rRNA genes. Strain DSM 9434 encodes 46 genes which participate in BChl a synthesis and this investigation shed light on the evolution and functional implications regarding bacteriochlorophyll synthesis.


Goniospectrophotometric space curves of diffraction gratings and their applicability as appearance fingerprints.
The bidirectional reflectance distribution functions of diffraction gratings were calculated by applying diffraction theory and transformed into goniospectrophotometric space curves. Gratings with parallel sinusoidal grooves having periods of 1-3.5 μm and amplitudes below 0.2 μm were analyzed. The obtained goniospectrophotometric space curves consist of lines with different slopes and possible interconnections. The slope of the lines is directly connected to the grating period and the length to the period and the amplitude. Such curves could be regarded as a simple appearance fingerprint of a diffraction grating. The ability of portable multiangle spectrophotometers to provide them for diffraction gratings is analyzed.


Noise, clutter, and determinism in infrared spectral signatures under varying meteorological conditions.
Infrared remote detection of chemical and biological agents in a complex environment depends on the ability to recognize threat signatures and differentiate them from the signatures of innocuous materials. In this paper, we addressed the methods of producing the constraint spectra needed to ensure reliable operation in a meteorologically changing environment. We collected arrays of background spectra of ground, woods, and low-angle sky on an irregular basis over a period of a year. Based on the hypothesis that the concentration fluctuations of species in the sensor's field of view can be exploited to form signatures, the standard deviations of the array (the result is characteristic of all fluctuations) and the difference array (the result is characteristic of sensor fluctuations) were computed. Subtracting these two spectra and filtering the result produced a spectrum, which is a measure of the IR fluctuations in the scene. The resulting set of scene spectra were processed into aberrant noise, and deterministic groups by numerical filtering and statistical methods.


Perceptions of vascular access for intravenous systemic therapy and risk factors for lymphedema in early-stage breast cancer-a patient survey.
The choice of vascular access for systemic therapy administration in breast cancer remains an area of clinical equipoise, and patient preference is not consistently acknowledged. Using a patient survey, we evaluated the patient experience with vascular access during treatment for early-stage breast cancer and explored perceived risk factors for lymphedema. Patients who had received systemic therapy for early-stage breast cancer were surveyed at 2 Canadian cancer centres. Responses were received from 187 patients (94%). The route of vascular access was peripheral intravenous line (IV) in 24%, a peripherally inserted central catheter (picc) in 42%, and a surgically inserted central catheter (port) in 34%. Anthracycline-based regimens were associated with a greater use of central vascular access devices (cvads- that is, a picc or port; 86/97, 89%). Trastuzumab use was associated with greater use of ports (49/64, 77%). Although few patients (7%) reported being involved in the decisions about vascular access, most were satisfied or very satisfied (88%) with their access type. Patient preference centred mainly on avoiding delays in the initiation of chemotherapy. Self-reported rates of complications (183 evaluable responses) were infiltration with peripheral IVs (9/44, 20%), local skin infections with piccs (7/77, 9%), and thrombosis with ports (4/62, 6%). Perceived risk factors for lymphedema included use of the surgical arm for blood draws (117/156, 75%) and blood pressure measurement (115/156, 74%). Most patients reported being satisfied with the vascular access used for their treatment. Improved education and understanding about the evidence-based requirements for vascular access are needed. Perceived risk factors for lymphedema remain variable and are not evidence-based.


Life expectancy, economic inequality, homicide, and reproductive timing in Chicago neighbourhoods.
In comparisons among Chicago neighbourhoods, homicide rates in 1988-93 varied more than 100-fold, while male life expectancy at birth ranged from 54 to 77 years, even with effects of homicide mortality removed. This "cause deleted" life expectancy was highly correlated with homicide rates; a measure of economic inequality added significant additional prediction, whereas median household income did not. Deaths from internal causes (diseases) show similar age patterns, despite different absolute levels, in the best and worst neighbourhoods, whereas deaths from external causes (homicide, accident, suicide) do not. As life expectancy declines across neighbourhoods, women reproduce earlier; by age 30, however, neighbourhood no longer affects age specific fertility. These results support the hypothesis that life expectancy itself may be a psychologically salient determinant of risk taking and the timing of life transitions.


Glucocorticoid receptor gene-based SNP analysis in patients with recurrent major depression.
Dysregulation of the hypothalamic-pituitary-adrenal axis, one of the stress-response systems, is one of the key neurobiological features of major depression (MDD). Data supporting the notion that glucocorticoid-mediated feedback inhibition is impaired in MDD come from a multitude of studies demonstrating nonsuppression of cortisol secretion following administration of the synthetic glucocorticoid dexamethasone. We examined whether genetic variations in the glucocorticoid receptor gene (Nuclear Receptor Subfamily 3, Group C, Member 1; NR3C1) could be associated with increased susceptibility for MDD using a whole gene-based association analysis of single nucleotide polymorphisms (SNPs). Four SNPs were identified in NR3C1 and genotyped in two well-diagnosed samples of patients with MDD ascertained in Belgium and northern Sweden, and matched control samples. In total, 314 MDD patients and 354 control individuals were included in the study. In the Belgian sample, we observed significant allele (p=0.02) and genotype (p=0.02) association with an SNP in the promoter region (NR3C1-1); in the Swedish sample, we observed significant allele (p=0.02) and genotype (p=0.02) association with the R23K SNP. The haplotype association studies showed modest evidence for an involvement of the 5' region of the NR3C1 gene in the genetic vulnerability for MDD. This study suggests that polymorphisms in the 5' region of the NR3C1 gene may play a role in the genetic vulnerability for MDD.


Direct binding of cytosolic NDP kinases to membrane lipids is regulated by nucleotides.
In spite of their complete lack of any structural features that characterize membrane proteins, cytosolic nucleoside-diphosphate kinases (NDPKs) have been found repeatedly to associate with membranes. In some instances the recruitment of cytosolic NDPKs to membranes was attributed to interactions with peripheral or integral membrane proteins, but in many cases the mechanism underlying the association of NDPKs with membranes remained unknown. We show here that cytosolic NDPKs bind directly to membrane lipids in a dynamic process that is controlled by its substrates, nucleoside tri- and diphosphates, and can be fully reconstituted with chemically defined, protein-free phospholipids and recombinant NDPK, or with purified NDPK. Our results uncover a novel mechanism for the reversible targeting of soluble NDPKs to membranes, where they may act as a reservoir of high energy phosphate, supporting the operation of membrane-based processes that utilize nucleotides other than ATP, such as intracellular traffic and phospholipid biosynthesis.


Impulse control disorders in Parkinson's disease: clinical characteristics and implications.
Impulse control disorders (ICDs), specifically those related to excessive gambling, eating, sex and shopping, have been observed in a subset of people with Parkinson's disease (PD). Although some initial case reports claimed that dopamine replacement therapies, particularly dopamine agonists, cause ICDs, more recent, larger and better controlled studies indicate a more complicated picture. While dopamine replacement therapy use is related to ICDs, other vulnerabilities, some related to PD and/or its treatment directly and others seemingly unrelated to PD, have also been associated with ICDs in PD. This suggests a complex etiology with multiple contributing factors. As ICDs occur in a sizable minority of PD patients and can be associated with significant distress and impairment, further investigation is needed to identify factors that can predict who may be more likely to develop ICDs. Clinical implications are discussed and topics for future research are offered.


Antioxidant role of oleuropein on midbrain and dopaminergic neurons of substantia nigra in aged rats.
Oleuropein is a phenolic compound which is present in the olive leaf extract. The purpose of the present study was to investigate the neuroprotective effect of oleuropein as an antioxidant agent on the substantia nigra in aged rats. Twenty 18-month-old Wistar rats (450-550 g) were randomly divided into control and experimental groups. The experimental group received a daily single dose of 50 mg/kg of oleuropein by oral gavage for 6 months. The control group received only distilled water. All rats were sacrificed two hours after the last gavage and the brains were removed and midbrains were cut. One part of the midbrains were homogenized and centrifuged. The tissue supernatant was assayed for lipid peroxidation (LPO) and antioxidant enzyme activities. The other part of midbrains fixed and embedded in paraffin, then processed for Nissl and immunohistochemistry (IHC) staining. Data was analyzed using SPSS by t-test. Differences were considered significant for P<0.05. The level of LPO in midbrain of the rats was decreased significantly in the experimental group, but superoxide dismutase, catalase and glutathione peroxidase activities were increased in experimental group compared to control group (P<0.05). Morphometric analyses showed significantly that the experimental group had more neurons in substantia nigra pars compacta (SNc) either in Nissl or IHC staining when compared to control (P<0.05). The results of the present study indicate that treatment of the old rats with oleuropein reduces the oxidative damage in SNc by increasing the antioxidant enzyme activities.


Application of the five stages of grief to diabetic limb loss and amputation.
A potentially underappreciated member of the multidisciplinary approach to diabetic limb salvage is that of psychiatry. Diabetic patients are more likely to experience depression, and diabetic patients with depression are more likely to undergo an amputation. Also, both diabetes and depression independently increase the healthcare costs in the United States. The objective of the present investigation was to increase knowledge among diabetic foot practitioners with respect to psychiatric and other mental health patient-care issues, specifically the potential application of the 5 stages of grief to diabetic limb loss and amputation. We enlisted the assistance of a clinical professor from the psychiatry department at our institution to review the 5 stages of grief, provide context specific for application to diabetic limb loss, and offer clinically relevant guidelines for surgeons to better understand and communicate with their patients at each stage. The 5 stages reviewed were denial, anger, bargaining, depression, and acceptance. We hope that the present review will increase the body of knowledge with respect to relevant psychiatric issues and the diabetic foot and provide a starting point for increased awareness with respect to this important, yet underappreciated, aspect of patient care.


Maternal and child health in the occupied Palestinian territory.
The Countdown to 2015 intervention coverage indicators in the occupied Palestinian territory are similar to those of other Arab countries, although there are gaps in continuity and quality of services across the continuum of the perinatal period. Since the mid 1990s, however, access to maternity facilities has become increasingly unpredictable. Mortality rates for infants (age </=1 year) and children younger than 5 years have changed little, and the prevalence of stunting in children has increased. Living conditions have worsened since 2006, when the elected Palestinian administration became politically and economically boycotted, resulting in unprecedented levels of Palestinian unemployment, poverty, and internal conflict, and increased restrictions to health-care access. Although a political solution is imperative for poverty alleviation, sustainable development, and the universal right to health care, women and children should not have to wait. Urgent action from international and local decision makers is needed for sustainable access to high-quality care and basic health entitlements.


Work disability after diagnosis of hand-arm vibration syndrome.
Our aim was to study the course of vasospastic and sensorineural symptoms after the clinical diagnosis of hand-arm vibration syndrome (HAVS), and the association of current HAVS symptoms with occupational status, self-evaluation of health, quality of life, and work ability. We gathered all HAVS cases diagnosed at the Finnish Institute of Occupational Health in Helsinki and Tampere during 1990-2008. A questionnaire was sent to all these patients (n = 241). Altogether 149 of them (62 %) returned the questionnaire. Cumulative lifelong vibration exposure was evaluated on the basis of the data in the patient files. On average, 8.5 years after the diagnosis of HAVS, approximately one-third of the patients reported improvement in symptoms of vibration-induced white finger (VWF) and the sensorineural symptoms. Young age and shorter exposure time were associated with improvement in VWF symptoms (p = 0.033 and p < 0.001, respectively). Persistent or deteriorated symptoms of both VWF and sensorineural symptoms were associated with lowered work ability, quality of life (EQ-5D), and general health, also after adjusting for age, smoking, and diseases other than HAVS. The patients' own prediction of work ability in 2 years was more negative if the VWF symptoms or sensorineural symptoms had continued after diagnosis of HAVS (p = 0.065 and p = 0.001, respectively). Our results suggest that in about two-thirds of the patients, the HAVS symptoms may stabilize or deteriorate in the follow-up. Considering the effects on work ability, timely prevention measures should be taken more actively to help patients continue their working careers.


A genome-wide association study of nonsynonymous SNPs identifies a type 1 diabetes locus in the interferon-induced helicase (IFIH1) region.
In this study we report convincing statistical support for a sixth type 1 diabetes (T1D) locus in the innate immunity viral RNA receptor gene region IFIH1 (also known as mda-5 or Helicard) on chromosome 2q24.3. We found the association in an interim analysis of a genome-wide nonsynonymous SNP (nsSNP) scan, and we validated it in a case-control collection and replicated it in an independent family collection. In 4,253 cases, 5,842 controls and 2,134 parent-child trio genotypes, the risk ratio for the minor allele of the nsSNP rs1990760 A --> G (A946T) was 0.86 (95% confidence interval = 0.82-0.90) at P = 1.42 x 10(-10).


Perceptually bistable three-dimensional figures evoke high choice probabilities in cortical area MT.
The role of the primate middle temporal area (MT) in depth perception was examined by considering the trial-to-trial correlations between neuronal activity and reported depth sensations. A set of moving random dots portrayed a cylinder rotating about its principal axis. In this structure-from-motion stimulus, the direction of rotation is ambiguous and the resulting percept undergoes spontaneous fluctuations. The stimulus can be rendered unambiguous by the addition of binocular disparities. We trained monkeys to report the direction of rotation in a set of these stimuli, one of which had zero disparity. Many disparity-selective neurons in area MT are selective for the direction of rotation defined by disparity. Across repeated presentations of the ambiguous (zero-disparity) stimulus, there was a correlation between neuronal firing and the reported direction of rotation, as found by Bradley et al. (1998). Quantification of this effect using choice probabilities (Britten et al., 1996) allowed us to demonstrate that the correlation cannot be explained by eye movements, behavioral biases, or attention to spatial location. MT neurons therefore appear to be involved in the perceptual decision process. The mean choice probability (0.67) was substantially larger than that reported for MT neurons in a direction discrimination task (Britten et al., 1996). This implies that MT neurons make a different contribution to the two tasks. For the depth task, either the pool of neurons used is smaller or the correlation between neurons in the pool is larger.


Ineffectiveness of the Doppler Ophthalmic Test (DOT) in post-endarterectomy evaluation.
In an effort to evaluate the effectiveness of the Doppler Ophthalmic Test (DOT) following carotid endarterectomy, a large group of patients was examined both pre- and postoperatively with non-invasive techniques. The DOT, which has proven to be a useful non-invasive diagnostic test for the determination of significant carotid artery stenosis, was found to be persistently abnormal in 46% of patients with a preoperative positive test. This occurred in spite of the fact that operative arteriography, direct ultrasonic auscultation, and Doppler imaging studies were all within normal limits. It is suggested from this study that the Doppler Ophthalmic Test alone is not adequate to follow patients postoperatively, especially if an abnormal study persists following a satisfactory endarterectomy. Other non-invasive techniques, which employ direct ultrasonic imaging of the carotid flow, may be more accurate in determining vessel patency.


Upregulation of forces and morphogenic asymmetries in dorsal closure during Drosophila development.
Tissue dynamics during dorsal closure, a stage of Drosophila development, provide a model system for cell sheet morphogenesis and wound healing. Dorsal closure is characterized by complex cell sheet movements, driven by multiple tissue specific forces, which are coordinated in space, synchronized in time, and resilient to UV-laser perturbations. The mechanisms responsible for these attributes are not fully understood. We measured spatial, kinematic, and dynamic antero-posterior asymmetries to biophysically characterize both resiliency to laser perturbations and failure of closure in mutant embryos and compared them to natural asymmetries in unperturbed, wild-type closure. We quantified and mathematically modeled two processes that are upregulated to provide resiliency--contractility of the amnioserosa and formation of a seam between advancing epidermal sheets, i.e., zipping. Both processes are spatially removed from the laser-targeted site, indicating they are not a local response to laser-induced wounding and suggesting mechanosensitive and/or chemosensitive mechanisms for upregulation. In mutant embryos, tissue junctions initially fail at the anterior end indicating inhomogeneous mechanical stresses attributable to head involution, another developmental process that occurs concomitant with the end stages of closure. Asymmetries in these mutants are reversed compared to wild-type, and inhomogeneous stresses may cause asymmetries in wild-type closure.


Cortactin-Src kinase signaling pathway is involved in N-syndecan-dependent neurite outgrowth.
N-syndecan (syndecan-3) was previously isolated as a cell surface receptor for heparin-binding growth-associated molecule (HB-GAM) and suggested to mediate the neurite growth-promoting signal from cell matrix-bound HB-GAM to the cytoskeleton of neurites. However, it is unclear whether N-syndecan would possess independent signaling capacity in neurite growth or in related cell differentiation phenomena. In the present study, we have transfected N18 neuroblastoma cells with a rat N-syndecan cDNA and show that N-syndecan transfection clearly enhances HB-GAM-dependent neurite growth and that the transfected N-syndecan distributes to the growth cones and the filopodia of the neurites. The N-syndecan-dependent neurite outgrowth is inhibited by the tyrosine kinase inhibitors herbimycin A and PP1. Biochemical studies show that a kinase activity, together with its substrate(s), binds specifically to the cytosolic moiety of N-syndecan immobilized to an affinity column. Western blotting reveals both c-Src and Fyn in the active fractions. In addition, cortactin, tubulin, and a 30-kDa protein are identified in the kinase-active fractions that bind to the cytosolic moiety of N-syndecan. Ligation of N-syndecan in the transfected cells by HB-GAM increases phosphorylation of c-Src and cortactin. We suggest that N-syndecan binds a protein complex containing Src family tyrosine kinases and their substrates and that N-syndecan acts as a neurite outgrowth receptor via the Src kinase-cortactin pathway.


Detection of gene fusions in rhabdomyosarcoma by reverse transcriptase-polymerase chain reaction assay of archival samples.
Alveolar rhabdomyosarcoma is a pediatric soft-tissue tumor that is often difficult to distinguish from other small round-cell tumors. The PAX3-FKHR and PAX7-FKHR gene fusions that result from chromosomal translocations in this tumor provide potential molecular diagnostic markers. To apply these molecular markers to commonly available archival material, we used reverse transcriptase-polymerase chain reaction and oligonucleotide hybridization methodology to develop an assay capable of identifying PAX3-FKHR and PAX7-FKHR fusion transcripts in formalin-fixed, paraffin-embedded tissue. Use of a control assay for wild-type FKHR mRNA indicated that RNA was successfully isolated, reverse-transcribed, and amplified in 15 of 16 archival cases. Comparison of assay results for the PAX3-FKHR and PAX7-FKHR fusions with standard molecular assays of paired frozen material revealed that all eight cases of known fusion-positive rhabdomyosarcoma were correctly identified and distinguished as PAX3-FKHR or PAX7-FKHR. The seven cases of known fusion-negative rhabdomyosarcoma showed no evidence of either product. These results indicate that we have developed a molecular assay that accurately identifies the fusion transcripts characteristic of alveolar rhabdomyosarcoma in archival samples. This assay will be useful for diagnosis and for retrospective clinicopathologic correlative studies.


A functional polymorphism in the 5HTR2C gene associated with stress responses also predicts incident cardiovascular events.
Previously we have shown that a functional nonsynonymous single nucleotide polymorphism (rs6318) of the 5HTR2C gene located on the X-chromosome is associated with hypothalamic-pituitary-adrenal axis response to a stress recall task, and with endophenotypes associated with cardiovascular disease (CVD). These findings suggest that individuals carrying the rs6318 Ser23 C allele will be at higher risk for CVD compared to Cys23 G allele carriers. The present study examined allelic variation in rs6318 as a predictor of coronary artery disease (CAD) severity and a composite endpoint of all-cause mortality or myocardial infarction (MI) among Caucasian participants consecutively recruited through the cardiac catheterization laboratory at Duke University Hospital (Durham, NC) as part of the CATHGEN biorepository. Study population consisted of 6,126 Caucasian participants (4,036 [65.9%] males and 2,090 [34.1%] females). A total of 1,769 events occurred (1,544 deaths and 225 MIs; median follow-up time = 5.3 years, interquartile range = 3.3-8.2). Unadjusted Cox time-to-event regression models showed, compared to Cys23 G carriers, males hemizygous for Ser23 C and females homozygous for Ser23C were at increased risk for the composite endpoint of all-cause death or MI: Hazard Ratio (HR) = 1.47, 95% confidence interval (CI) = 1.17, 1.84, p = .0008. Adjusting for age, rs6318 genotype was not related to body mass index, diabetes, hypertension, dyslipidemia, smoking history, number of diseased coronary arteries, or left ventricular ejection fraction in either males or females. After adjustment for these covariates the estimate for the two Ser23 C groups was modestly attenuated, but remained statistically significant: HR = 1.38, 95% CI = 1.10, 1.73, p = .005. These findings suggest that this functional polymorphism of the 5HTR2C gene is associated with increased risk for CVD mortality and morbidity, but this association is apparently not explained by the association of rs6318 with traditional risk factors or conventional markers of atherosclerotic disease.


Topography in the Drosophila visual system.
The Drosophila visual system offers an excellent opportunity for studying the development of proper retinotopic connections at the level of individual identifiable cell types. Recent work suggests that, despite obvious anatomical and developmental differences, at least some of the general developmental strategies operating in the Drosophila visual system parallel observations made previously for vertebrates. The extensive repertoire of powerful genetic and molecular techniques available in Drosophila can now be directed towards determining whether these parallels also reflect similarities in the underlying molecular mechanisms.


Regulation of AMPA receptor function by the human memory-associated gene KIBRA.
KIBRA has recently been identified as a gene associated with human memory performance. Despite the elucidation of the role of KIBRA in several diverse processes in nonneuronal cells, the molecular function of KIBRA in neurons is unknown. We found that KIBRA directly binds to the protein interacting with C-kinase 1 (PICK1) and forms a complex with α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPARs), the major excitatory neurotransmitter receptors in the brain. KIBRA knockdown accelerates the rate of AMPAR recycling following N-methyl-D-aspartate receptor-induced internalization. Genetic deletion of KIBRA in mice impairs both long-term depression and long-term potentiation at hippocampal Schaffer collateral-CA1 synapses. Moreover, KIBRA knockout mice have severe deficits in contextual fear learning and memory. These results indicate that KIBRA regulates higher brain function by regulating AMPAR trafficking and synaptic plasticity.


Real-World Budget Impact of the Adoption of Insulin Glargine Biosimilars in Primary Care in England (2015-2018).
Lantus, the reference insulin glargine used for the treatment of diabetes, lost its patent protection in 2014, opening the market to biosimilar competitors. First, to analyze the adoption rates of insulin glargine biosimilars in primary care in England and estimate the savings realized and missed, since an insulin glargine biosimilar was first used, and second, to assess potential variations in adoption rates across Clinical Commissioning Groups (CCGs). Data sets capturing information on all insulin glargine items prescribed by all general practitioners up to December 2018 were used. Total costs of insulin glargine and uptake rates of biosimilars were calculated. The real-world budget impact was estimated assuming the cost of reference insulin glargine for all items and comparing the total costs in this scenario with the total costs in the real world. The missed savings were estimated assuming the cost of biosimilars for all insulin glargine items. Choropleth maps were generated to assess potential variations in uptake across CCGs. Insulin glargine biosimilars generated savings of £900,000 between October 2015 (time of first prescription) and December 2018. The missed savings amounted to £25.6 million in this period, indicating that only 3.42% of the potential savings were achieved. The analyses demonstrated a large level of variation in the uptake of insulin glargine biosimilars across CCGs, with market shares ranging from 0 to 53.3% (December 2018). These results may encourage decision makers in England to promote the use of best-value treatments in primary care and to reevaluate variation across CCGs.