Transadaptation and Standardization of Tinnitus Primary Function Questionnaire in Hindi.
The term tinnitus is derived from the Latin word tinnire, meaning to ring. Although it is often referred to as "ringing in the ears, tinnitus can be perceived as many different sounds including hissing, clicking or whistling. India being a multilingual country needs to develop and standardize tinnitus questionnaire in Indian language. There are no significant test materials available with respect to Indian context based on Hindi, which can be routinely used by the professionals to assess the primary effect of tinnitus in patients. Thus, a need was felt to develop instrument in Hindi that can be used as per the ICF (International Classification of Functioning, Disability and Health) classification system in all over the country. The study aims to evaluate the impact of tinnitus on quality of life of clients having tinnitus with and without hearing loss by using primary function questionnaire (TPF-Hindi). The present study attempts to transadapt and standardize Tinnitus Primary function Questionnaire in Hindi. And administer both the questionnaires (TPFQ-H vs THQ-H) on the tinnitus clients with and without hearing loss and compare the impact of tinnitus in terms of quality of life in subjects with tinnitus with hearing loss and without hearing loss. A 12-item questionnaire was administered to 50 patients (tinnitus with hearing loss and without hearing loss) and compared between two questionnaires Tinnitus Primary Function Questionnaire and Tinnitus Handicapped Questionnaire (TPFQ-H & THQ-H). The results of the current study were analysed by using Statistical Package for the Social Sciences (SPSSv 20.0). Scores were positively correlated with the Tinnitus Handicap Questionnaire. There is significant correlation between both the scales and reliability between the scales is also good.


Epigallocatechin-3-gallate suppresses the global interleukin-1beta-induced inflammatory response in human chondrocytes.
Epigallocatechin-3-gallate (EGCG) is a bioactive polyphenol of green tea and exerts potent anti-inflammatory effects by inhibiting signaling events and gene expression. Interleukin-1beta (IL-1β) is the principal cytokine linked to cartilage degradation in osteoarthritis (OA). The objective of this study was to evaluate the global effect of EGCG on IL-1β-induced expression of proteins associated with OA pathogenesis in human chondrocytes. Primary OA chondrocytes were pretreated with EGCG (10 to 100 uM) and then stimulated with IL-1β (5 ng/ml) for 24 hours. Culture supernatants were incubated with cytokine antibody arrays and immunoreactive proteins (80 proteins) were visualized by enhanced chemiluminiscence. Effect of EGCG on IL-1β-induced expression of 18 selected genes was verified by Real time-PCR and effect on IL-6, IL-8 and tumor necrosis factor-alpha (TNF-α) production was determined using specific ELISAs. Western immunoblotting was used to analyze the effect of EGCG on the interleukin-1 receptor-associated kinase 1 (IRAK-1) and TNF receptor-associated factor 6 (TRAF-6) proteins in IL-1β-stimulated chondrocytes. The role of nuclear factor kappa-B (NF-κB) and mitogen activated protein kinases (MAPKs) in the regulation of selected genes and the mechanism involved in EGCG mediated modulation of these genes was determined by using specific inhibitors for NF- κB (MG132) and MAPKs (p38-MAPK, SB202190; JNK-MAPK, SP600125, ERK-MAPK, PD98059). Out of 80 proteins present on the array, constitutive expression of 14% proteins was altered by EGCG treatment. No significant stimulatory effect was observed on the proteins associated with cartilage anabolic response. Stimulation with IL-1β enhanced the expression of 29 proteins. Expression of all 29 proteins up-regulated by IL-1β was found to be suppressed by EGCG. EGCG also inhibited the expression of the signaling intermediate TRAF-6 at 50 and 100 uM concentrations (P < 0.05). Our results identified several new targets of EGCG, including epithelial neutrophil activating peptide-78 (ENA-78), granulocyte macrophage colony stimulation factor (GM-CSF), growth- related oncogene (GRO), GRO-α, IL-6, IL-8, monocyte chemotactic protein-1 (MCP-1), MCP-3, macrophage inflammatory protein-1beta (MIP-1β), granulocyte chemotactic protein-2 (GCP-2), MIP-3alpha, interferon-gamma-inducible protein-10 (IP-10), nucleosome assembly protein-2 (NAP-2) and leukemia inhibitory factor (LIF). The inhibitory effects of EGCG were mainly mediated by inhibiting the activation of NF-κB and c-Jun N-terminal Kinase (JNK)-MAPK in human chondrocytes. Our results suggest that the potential of EGCG in OA treatment/prevention may be related to its ability to globally suppress the inflammatory response in human chondrocytes. These results identify additional new targets of EGCG and advocate that EGCG may be a potent chondroprotective agent in OA.


Vibrational Stark spectroscopy for assessing ligand-binding strengths in a protein.
Nitrile groups are potentially useful spectroscopic probes in the infrared to characterize the binding and dynamics of ligands in proteins. This opens the possibility of locating and determining the binding mode of suitably labelled ligands in proteins based on optical spectroscopy, without the need for determining an X-ray structure. However, relating structure and spectroscopy requires means to accurately compute infrared spectra. This is investigated for benzonitrile (PhCN) in water, wild type (WT) and two lysozyme mutants in solution. The force field is validated by comparing with experimental data for benzonitrile in water which is the basis for computing the Stark shift and time scale for spectral diffusion of PhCN in WT and the L99A and L99G mutants of T4 lysozyme. The 1-d spectra for PhCN in WT and the two mutant proteins differ in their maximum absorption by up to 4 cm-1, which reflects the modified electrostatic environments in the three proteins. It is also tested whether extending from 1-d to 2-d infrared spectroscopy provides further discrimination in the ligand-binding modes. First, for PhCN in solution the frequency fluctuation correlation function (FFCF) decays to zero at short times whereas in the protein a pronounced static inhomogeneous component is found. Secondly, the decay time of the FFCF for the mutant to which PhCN binds most strongly has the longest decay time. It is demonstrated explicitly that the ligand-binding free energy with respect to the three protein variants correlates with the Stark shift. This makes 1-d infrared spectroscopy together with computations a valuable tool for characterizing binding modes and potentially binding locations in proteins.


Evaluating a Mental Health Program for Palestinian Refugees in Lebanon.
Medecins sans Frontière, an international non-governmental organization, initiated a mental health program for Palestinian refugees living in Lebanon. To evaluate the impact of the program after its completion, focus groups were conducted with three target groups: (1) patients, (2) staff, and (3) local community stakeholders. Participants voiced overall satisfaction with the program. The program provided easy access, good quality care, decreased stigma, as perceived by participants, and revealed a sense of community contentedness. In addition, several short-term outcomes were achieved, such as increasing the numbers of patients visiting the center/ receiving mental health treatment. However, lack of planning for sustainability and proper procedures for hand-over of the program constituted a major downfall. Program discontinuation posed ethical dilemmas, common in provisional interventions in underprivileged refugee communities.


Small intestinal neuroendocrine tumours and fibrosis: an entangled conundrum.
Small intestinal neuroendocrine tumours (SI-NETs) are neoplasms characterized by their ability to secrete biogenic amines and peptides. These cause distinct clinical pathology including carcinoid syndrome, marked by diarrhoea and flushing, as well as fibrosis, notably mesenteric fibrosis. Mesenteric fibrosis often results in significant morbidity by causing intestinal obstruction, oedema and ischaemia. Although advancements have been made to alleviate symptoms of carcinoid syndrome and prolong the survival of patients with SI-NETs, therapeutic options for patients with mesenteric fibrosis are still limited. As improved insight in the complex pathogenesis of mesenteric fibrosis is key to the development of new therapies, we evaluated the literature for known and putative mediators of fibrosis in SI-NETs. In this review, we discuss the tumour microenvironment, growth factors and signalling pathways involved in the complex process of fibrosis development and tumour progression in SI-NETs, in order to elucidate potential new avenues for scientific research and therapies to improve the management of patients suffering from the complications of mesenteric fibrosis.


SPREADING PROPERTIES OF LEECH EXTRACTS AND THE FORMATION OF LYMPH.
1. The injection of leech extracts into the skin increases its permeability, as shown both by the spread of fluid and of foreign particles through the dermis. The spread is followed some hours after the injection by more or less edema of the subcutaneous tissue. 2. A preliminary study of the chemical properties of the leech spreading factor indicates a similarity with the spreading factor prepared from testicle. 3. Attempts to separate the leech spreading and anticoagulating factors showed that the two have practically the same distribution in the leech body, extracts from the separated head being the most active. 4. It is undetermined whether two distinct factors are responsible for the spreading and anticoagulating properties of leech extracts. A chemical similarity is suggested by the fact that agents which affect the activity of one factor have a parallel effect on the other. 5. The mechanism of the spread produced by leech extracts and by other spreading agents is discussed.


Use of Antihypertensive Drugs and Ischemic Stroke Severity - Is There a Role for Angiotensin-II?
The increase in angiotensin II (Ang II) formation by selected antihypertensive drugs is said to exhibit neuroprotective properties, but this translation into improvement in clinical outcomes has been inconclusive. We undertook a study to investigate the relationship between types of antihypertensive drugs used prior to a stroke event and ischemic stroke severity. We hypothesized that use of antihypertensive drugs that increase Ang II formation (Ang II increasers) would reduce ischemic stroke severity when compared to antihypertensive drugs that suppress Ang II formation (Ang II suppressors). From the Malaysian National Neurology Registry, we included hypertensive patients with first ischemic stroke who presented within 48 hours from ictus. Antihypertensive drugs were divided into Ang II increasers (angiotensin-I receptor blockers (ARBs), calcium channel blockers (CCBs) and diuretics) and Ang II suppressors (angiotensin-converting-enzyme inhibitors (ACEIs) and beta blockers). We evaluated stroke severity during admission with the National Institute of Health Stroke Scale (NIHSS). We performed a multivariable logistic regression with the score being dichotomized at 15. Scores of less than 15 were categorized as less severe stroke. A total of 710 patients were included. ACEIs was the most commonly prescribed antihypertensive drug in patients using Ang II suppressors (74%) and CCBs, in patients prescribed with Ang II increasers at 77%. There was no significant difference in the severity of ischemic stroke between patients who were using Ang II increasers in comparison to patients with Ang II suppressors (OR: 1.32, 95%CI: 0.83-2.10, p = 0.24). In our study, we found that use of antihypertensive drugs that increase Ang II formation was not associated with less severe ischemic stroke as compared to use of antihypertensive drugs that suppress Ang II formation.


The effective mitigation of greenhouse gas emissions from rice paddies without compromising yield by early-season drainage.
Global rice production systems face two opposing challenges: the need to increase production to accommodate the world's growing population while simultaneously reducing greenhouse gas (GHG) emissions. Adaptations to drainage regimes are one of the most promising options for methane mitigation in rice production. Whereas several studies have focused on mid-season drainage (MD) to mitigate GHG emissions, early-season drainage (ED) varying in timing and duration has not been extensively studied. However, such ED periods could potentially be very effective since initial available C levels (and thereby the potential for methanogenesis) can be very high in paddy systems with rice straw incorporation. This study tested the effectiveness of seven drainage regimes varying in their timing and duration (combinations of ED and MD) to mitigate CH4 and N2O emissions in a 101-day growth chamber experiment. Emissions were considerably reduced by early-season drainage compared to both conventional continuous flooding (CF) and the MD drainage regime. The results suggest that ED+MD drainage may have the potential to reduce CH4 emissions and yield-scaled GWP by 85-90% compared to CF and by 75-77% compared to MD only. A combination of (short or long) ED drainage and one MD drainage episode was found to be the most effective in mitigating CH4 emissions without negatively affecting yield. In particular, compared with CF, the long early-season drainage treatments LE+SM and LE+LM significantly (p<0.01) decreased yield-scaled GWP by 85% and 87% respectively. This was associated with carbon being stabilised early in the season, thereby reducing available C for methanogenesis. Overall N2O emissions were small and not significantly affected by ED. It is concluded that ED+MD drainage might be an effective low-tech option for small-scale farmers to reduce GHG emissions and save water while maintaining yield.


Admission Glasgow Coma Scale Score as a Predictor of Outcome in Patients Without Traumatic Brain Injury.
The Glasgow Coma Scale was developed in 1974 as an injury severity score to assess and predict outcome after traumatic brain injury. The tool is now used to score depth of impaired consciousness in patients with and without traumatic brain injury. However, evidence supporting the use of the Glasgow Coma Scale in the latter group is limited. To assess Glasgow Coma Scale score on hospital admission as a predictor of outcome in patients without traumatic brain injury. This was a secondary analysis of prospectively collected data from 3507 patients admitted to 4 hospitals between October 2015 and October 2019. Patients with a primary diagnosis of traumatic brain injury were excluded from this study. The mean age of the 3507 participants in the study was 57 years. Participants were primarily female (52%), White (77%), and non-Hispanic (89%). On admission, 90% of patients had a modified Rankin Scale score of 0 to 3 and 72% had a Glasgow Coma Scale score of 13 to 15 (mild injury). Generalized estimating equation modeling indicated that admission Glasgow Coma Scale score did not predict modified Rankin Scale score at discharge in patients not diagnosed with traumatic brain injury (Glasgow Coma Scale score <8: z = -7.89, P < .001; Glasgow Coma Scale score 8-12: z = -4.17, P < .001). The Glasgow Coma Scale is not recommended for use in patients without traumatic brain injury; clinicians should use a more appropriate and validated clinical assessment instrument for this patient population.


Multivariable analysis of total cholesterol levels in male Swiss Armed Forces conscripts 2006-2012 (N = 174,872).
Cholesterol is an important contributor to morbidity and mortality risks due to its association with obesity, cardiovascular disease, and cancer. A system of mandatory military conscription is a useful tool for disease-risk monitoring in a given male population. Swiss military conscription data are representative for more than 90% of a given male birth cohort (with Swiss citizenship). The medical examination also includes voluntary laboratory testing, for which approximately 65% of the young men present at conscription give consent. Here we present the temporal and subgroup analyses of total serum cholesterol levels (TCL) among Swiss conscripts from 2006 to 2012 (N = 174,872; mean age = 19.75 years). The voluntary blood samples were tested by a central laboratory (Viollier AG) with identical measurement standards and strict quality control. To test differences in TCL by socioeconomic occupational status, sports test performance, Body Mass Index (BMI), age, and place of residence of the conscripts we used a multivariable regression model with TCL as dependent variable. Mean TCL decreased significantly, by 0.125 mmol/l (95% CI 0.108-0.142, p < 0.001) from 4.225 mmol/l (95% CI 4.210-4.240) in 2006 to 4.100 mmol/l (95% CI 4.091-4.109) in 2012. Similarly, the prevalence of conscripts with an elevated TCL ≥ 5.17 mmol/l decreased from ≥ 10.2% prior to 2011 to 6.9% in 2011 and 8.2% in 2012. Multivariate regression showed an association between elevated TCL and lower socioeconomic occupational status, lower sports test performance, higher BMI, higher age, and area of residence. There was no longer a significant increase in mean TCL among the three grades of obesity (BMI ≥ 30.0 kg/m2) as defined by the WHO. Within the BMI categories of normal weight and overweight, TCL was stratified by sports performance (better sports performance = lower TCL). Decreasing TCL in 2011 and 2012 fits the known pattern of conscripted persons' stabilizing BMI and sports test performance of the conscripts in recent years. However, small temporal drifts within the laboratory analyses cannot be ruled out as confounding factors. In conclusion, identifying subgroups with unfavorable lipid profiles will contribute to the continuing success of intensified public health programs.


The genetics of isoflurane-induced developmental neurotoxicity.
Neurotoxicity induced by early developmental exposure to volatile anesthetics is a characteristic of organisms across a wide range of species, extending from the nematode C. elegans to mammals. Prevention of anesthetic-induced neurotoxicity (AIN) will rely upon an understanding of its underlying mechanisms. However, no forward genetic screens have been undertaken to identify the critical pathways affected in AIN. By characterizing such pathways, we may identify mechanisms to eliminate isoflurane induced AIN in mammals. Chemotaxis in adult C. elegans after larval exposure to isoflurane was used to measure AIN. We initially compared changes in chemotaxis indices between classical mutants known to affect nervous system development adding mutants in response to data. Activation of specific genes was visualized using fluorescent markers. Animals were then treated with rapamycin or preconditioned with isoflurane to test effects on AIN. Forty-four mutations, as well as pharmacologic manipulations, identified two pathways, highly conserved from invertebrates to humans, that regulate AIN in C. elegans. Activation of one stress-protective pathway (DAF-2 dependent) eliminates AIN, while activation of a second stress-responsive pathway (endoplasmic reticulum (ER) associated stress) causes AIN. Pharmacologic inhibition of the mechanistic Target of Rapamycin (mTOR) blocks ER-stress and AIN. Preconditioning with isoflurane prior to larval exposure also inhibited AIN. Our data are best explained by a model in which isoflurane acutely inhibits mitochondrial function causing activation of responses that ultimately lead to ER-stress. The neurotoxic effect of isoflurane can be completely prevented by manipulations at multiple points in the pathways that control this response. Endogenous signaling pathways can be recruited to protect organisms from the neurotoxic effects of isoflurane.


Estimation of RF energy absorbed in the brain from mobile phones in the Interphone Study.
The objective of this study was to develop an estimate of a radio frequency (RF) dose as the amount of mobile phone RF energy absorbed at the location of a brain tumour, for use in the Interphone Epidemiological Study. We systematically evaluated and quantified all the main parameters thought to influence the amount of specific RF energy absorbed in the brain from mobile telephone use. For this, we identified the likely important determinants of RF specific energy absorption rate during protocol and questionnaire design, we collected information from study subjects, network operators and laboratories involved in specific energy absorption rate measurements and we studied potential modifiers of phone output through the use of software-modified phones. Data collected were analysed to assess the relative importance of the different factors, leading to the development of an algorithm to evaluate the total cumulative specific RF energy (in joules per kilogram), or dose, absorbed at a particular location in the brain. This algorithm was applied to Interphone Study subjects in five countries. The main determinants of total cumulative specific RF energy from mobile phones were communication system and frequency band, location in the brain and amount and duration of mobile phone use. Though there was substantial agreement between categorisation of subjects by cumulative specific RF energy and cumulative call time, misclassification was non-negligible, particularly at higher frequency bands. Factors such as adaptive power control (except in Code Division Multiple Access networks), discontinuous transmission and conditions of phone use were found to have a relatively minor influence on total cumulative specific RF energy. While amount and duration of use are important determinants of RF dose in the brain, their impact can be substantially modified by communication system, frequency band and location in the brain. It is important to take these into account in analyses of risk of brain tumours from RF exposure from mobile phones.


Immunohistochemical distribution of enkephalin, substance P, and somatostatin in the brainstem of the leopard frog, Rana pipiens.
The brainstems of frogs contain many of the neurochemicals that are found in mammals. However, the clustering of nuclei near the ventricles makes it difficult to distinguish individual cell groups. We addressed this problem by combining immunohistochemistry with tract tracing and an analysis of cell morphology to localize neuropeptides within the brainstem of Rana pipiens. We injected a retrograde tracer, Fluoro-Gold, into the spinal cord, and, in the same frog, processed adjacent sections for immunohistochemical location of antibodies to the neuropeptides enkephalin (ENK), substance P (SP), and somatostatin (SOM). SOM+ cells were more widespread than cells containing immunoreactivity (ir) to the other substances. Most reticular nuclei in frog brainstem contained ir to at least one of these chemicals. Cells with SOM ir were found in nucleus (n.) reticularis pontis oralis, n. reticularis magnocellularis, n. reticularis paragigantocellularis, n. reticularis dorsalis, the optic tectum, n. interpeduncularis, and n. solitarius. ENK-containing cell bodies were found in n. reticularis pontis oralis, n. reticularis dorsalis, the nucleus of the solitary tract, and the tectum. The midbrain contained most of the SP+ cells. Six nonreticular nuclei (griseum centrale rhombencephali, n. isthmi, n. profundus mesencephali, n. interpeduncularis, torus semicircularis laminaris, and the tectum) contained ir to one or more of the substances but did not project to the spinal cord. The descending tract of V, and the rubrospinal, reticulospinal, and solitary tracts contained all three peptides as did the n. profundus mesencephali, n. isthmi, and specific tectal layers. Because the distribution of neurochemicals within the frog brainstem is similar to that of amniotes, our results emphasize the large amount of conservation of structure, biochemistry, and possibly function that has occurred in the brainstem, and especially in the phylogenetically old reticular formation.


How Diverse Is Your Universe? An Activity for Students to Reflect on Ethnoracial Diversity During Orientation.
Physicians' unconscious biases have been linked to health disparities within ethnic and racially diverse communities. Addressing these unconscious biases is difficult but may be ameliorated by raising individuals' awareness of the ethnoracial makeup of their personal and professional networks and reflecting on whether it needs to be expanded while in medical school. First- and second-year students were provided with an overview of the ethnoracial makeup of individuals within the state, community, and medical school as a means to reflect on the ethnoracial makeup of their future patient population. Following this overview, students engaged in an activity adapted from the University of Houston, which allowed them to visually represent the ethnoracial diversity within their networks. Written reflections on the adapted activity were collected, analyzed using manifest content analysis, and reported according to themes. The results indicated that the activity was valuable in helping students visualize their current exposure to ethnoracially diverse individuals (143 of 357 responses [40%]) and reflect on their need to expand the level of ethnoracial diversity in their lives (47 of 357 responses [13%]). Additionally, students provided comments to help improve the activity when used in another institution. Assisting students in raising their awareness of the ethnoracial diversity in their personal and professional networks is a step toward addressing the unconscious biases that emerge in physicians while in clinical practice. This activity, designed to raise students' awareness of ethnoracial diversity, originated in Augusta, Georgia, but can be adapted to any state.


Online popularity of JUUL and Puff Bars in the USA: 2019-2020.
The US Food and Drug Administration (FDA) announced a flavour ban on cartridge-based e-cigarettes in January 2020. It is unclear whether e-cigarette users will switch to disposable vaping products with a variety of kids-appealing flavours available. We performed piece-wise regression and autoregressive integrated moving average (ARIMA) algorithms to compare the relative search volume (RSV) of JUUL and Puff Bar (a disposable vaping product) using the 1-year Google Trends data from 24 February 2019 to 20 February 2020, separated by three events that may have spurred changes in RSV for each product. The RSV for JUUL was relatively stable before Trump Administration announced plans to ban flavoured e-cigarettes. After that, the RSV for JUUL dropped sharply (rate of change=-8.8 per week) from 11 September 2019 to 17 October 2019 when JUUL Labs announced to halt online sales of some flavoured products, and the RSV resumed the decreasing trend after FDA announced enforcement policy of cartridge-based e-cigarettes on 2 January 2020. In comparison, the RSV for Puff Bar started to increase after 11 September 2019 with a low rate of change (0.6) until 17 October 2019. After that, the increase in RSV for Puff Bar accelerated. The RSV of puff bars surpassed that of JUUL during the week of 2 February 2020. The popularity of Puff Bar on Google Search suggests that users may replace cartridge-based vaping products with disposable e-cigarettes in the circumvention of the partial flavour ban. Continuous surveillance and further assessment are needed to prevent potential loopholes in tobacco regulation.


Two-photon fluorescence correlation spectroscopy through a dual-clad optical fiber.
We report on the utilization of a dual-clad optical fiber for two-photon excited fluorescence correlation spectroscopy (FCS). High excitation efficiency is obtained by sending prechirped ultrafast pulses through the single-mode core of the fiber, while the fluorescence collection efficiency is enhanced because of the larger numerical aperture of the inner cladding. We show that the utilization of a dual-clad fiber is critical for ultrasensitive two-photon fluorescence detection. Our system has the ability to detect fluorescent nanospheres as small as 12 nm in radius. Quantum dots of radius 7 nm are also measured and show excellent signal to noise ratio. The particle sizes obtained from the fiber FCS system were confirmed by measurements using a commercial dynamic light scattering (DLS) system.


The BUD4 protein of yeast, required for axial budding, is localized to the mother/BUD neck in a cell cycle-dependent manner.
A and alpha cells of the yeast Saccharomyces cerevisiae exhibit an axial budding pattern, whereas a/alpha diploid cells exhibit a bipolar pattern. Mutations in BUD3, BUD4, and AXL1 cause a and alpha cells to exhibit the bipolar pattern, indicating that these genes are necessary to specify the axial budding pattern (Chant, J., and I. Herskowitz. 1991. Cell. 65:1203-1212; Fujita, A., C. Oka, Y. Arikawa, T. Katagi, A. Tonouchi, S. Kuhara, and Y. Misumi. 1994. Nature (Lond.). 372:567-570). We cloned and sequenced BUD4, which codes for a large, novel protein (Bud4p) with a potential GTP-binding motif. Bud4p is expressed and localized to the mother/bud neck in all cell types. Most mitotic cells contain two apparent rings of Bud4 immunoreactive staining, as observed for Bud3p (Chant, J., M. Mischke, E. Mitchell, I. Herskowitz, and J.R. Pringle. 1995. J. Cell Biol. 129: 767-778). Early G1 cells contain a single ring of Bud4p immunoreactive staining, whereas cells at START and in S phase lack these rings. The level of Bud4p is also regulated in a cell cycle-dependent manner. Bud4p is inefficiently localized in bud3 mutants and after a temperature shift of a temperature-sensitive mutant, cdc12, defective in the neck filaments. These observations suggest that Bud4p and Bud3p cooperate to recognize a spatial landmark (the neck filaments) during mitosis and support the hypothesis that they subsequently become a landmark for establishing the axial budding pattern in G1.


Role of chromosomes in assembly of meiotic and mitotic spindles.
The assembly of a mitotic spindle requires the interaction of microtubules with chromosomes. As a cell enters mitosis, long microtubules are converted to short ones, as microtubules become unstable. Dynamic microtubules are then stabilised by chromosomes, forming a bipolar spindle. In this review, we discuss the different roles of kinetochores and chromosome arms during spindle assembly. Kinetochores, required for proper chromosomes segregation, capture microtubules and maintain attachment. Chromosome arms greatly enhance microtubule stability, and alone can be sufficient for spindle assembly.


In-ear vital signs monitoring using a novel microoptic reflective sensor.
Cardiovascular diseases are among the most common causes of death in industrial countries. In order to take preventive actions, it is of great interest, to both physicians and patients, to determine cardiovascular risk factors early. To address this problem, a wearable in-ear measuring system (IN-MONIT) for 24/7 monitoring of vital parameters has been developed. The central component is a microoptic reflective sensor located inside the auditory canal. From the measured photoplethysmographic curves, heart activity and heart rate can be derived. In this paper, we describe the optoelectronic sensor concept and the autonomous design of the IN-MONIT measurement system. For the assessment of heart rate, different algorithms are introduced and the performance of the developed sensor system is evaluated in relation to conventional systems. In addition, the robustness to external artifacts is evaluated and artifact reduction strategies are considered.


Cholera toxin can catalyze ADP-ribosylation of cytoskeletal proteins.
Cholera toxin catalyzes transfer of radiolabel from [32P]NAD+ to several peptides in particulate preparations of human foreskin fibroblasts. Resolution of these peptides by two-dimensional gel electrophoresis allowed identification of two peptides of Mr = 42,000 and 52,000 as peptide subunits of a regulatory component of adenylate cyclase. The radiolabeling of another group of peptides (Mr = 50,000 to 65,000) suggested that cholera toxin could catalyze ADP-ribosylation of cytoskeletal proteins. This suggestion was confirmed by showing that incubation with cholera toxin and [32P]NAD+ caused radiolabeling of purified microtubule and intermediate filament proteins.


Managing incidental findings and research results in genomic research involving biobanks and archived data sets.
Biobanks and archived data sets collecting samples and data have become crucial engines of genetic and genomic research. Unresolved, however, is what responsibilities biobanks should shoulder to manage incidental findings and individual research results of potential health, reproductive, or personal importance to individual contributors (using "biobank" here to refer both to collections of samples and collections of data). This article reports recommendations from a 2-year project funded by the National Institutes of Health. We analyze the responsibilities involved in managing the return of incidental findings and individual research results in a biobank research system (primary research or collection sites, the biobank itself, and secondary research sites). We suggest that biobanks shoulder significant responsibility for seeing that the biobank research system addresses the return question explicitly. When reidentification of individual contributors is possible, the biobank should work to enable the biobank research system to discharge four core responsibilities to (1) clarify the criteria for evaluating findings and the roster of returnable findings, (2) analyze a particular finding in relation to this, (3) reidentify the individual contributor, and (4) recontact the contributor to offer the finding. We suggest that findings that are analytically valid, reveal an established and substantial risk of a serious health condition, and are clinically actionable should generally be offered to consenting contributors. This article specifies 10 concrete recommendations, addressing new biobanks as well as those already in existence.


Evidence for a role of sphingosine-1 phosphate in cardiovascular remodelling in Fabry disease.
A hallmark of Fabry disease is the concomitant development of left-ventricular hypertrophy and arterial intima-media thickening, the pathogenesis of which is thought to be related to the presence of a plasmatic circulating growth-promoting factor. We therefore characterized the plasma of patients with Fabry disease in order to identify this factor. Using a classical biochemical strategy, we isolated and identified sphingosine-1 phosphate (S1P) as a proliferative factor present in the plasma of patients with Fabry disease. Plasma S1P levels were significantly higher in 17 patients with Fabry disease compared with 17 healthy controls (225 +/- 40 vs. 164 +/- 17 ng/mL; P = 0.005). There was a positive correlation between plasma S1P levels and both common carotid artery intima-media thickness and left-ventricular mass index (r(2) = 0.47; P = 0.006 and r(2) = 0.53; P = 0.0007, respectively). In an experimental model, mice treated with S1P developed cardiovascular remodelling similar to that observed in patients with Fabry disease. Sphingosine-1 phosphate participates in cardiovascular remodelling in Fabry disease. Our findings have implications for the treatment of cardiovascular involvement in Fabry disease.


Smokeless tobacco cessation support in dental hospitals in Pakistan: Dentists and dental patients' perspectives on current practices, support needed and opportunities available.
Despite evidence on the effectiveness of tobacco cessation interventions in dental settings, the implementation remains low, especially for smokeless tobacco (ST). The purpose of this study was to develop an understanding of the influences governing the implementation of ST cessation support in dental hospitals. A multi-centre qualitative study was conducted at two tertiary-care dental hospitals, in Pakistan. Semi-structured interview guide, guided by the Capability-Opportunity-Motivation-Behaviour (COM-B) model, were used to capture the views of dentists (n=12) and dental patients (n=12), regarding ST cessation support in dental hospitals. Framework approach was used to thematically analyse the data. Screening of ST users in routine dental practice was seldom practiced and the cessation support offered was brief advice. Barriers identified by dentists included: fear of offending and stereotyping patients; lack of knowledge and skills; lack of privacy; lack of belief in the effectiveness of behavioural support; lack of time and workload pressure; ST use amongst dentists; lack of referral systems and; the absence of a mandatory requirement of offering ST cessation support. Facilitators included: delivering support through junior dentists and the length of interaction between the dentist and the patient. Naswar was the most common ST product used by dental patients. Patients reported receiving negligible cessation support from any healthcare provider. A range of influences governing the implementation of ST cessation support in dental hospitals were identified. These findings can inform the implementation of behavioural interventions for ST cessation in dental and other clinical settings, in low and middle-income countries. Smokeless Tobacco (ST) control considerably lags, in comparison to the control of combustible tobacco. This is the first study that qualitatively explores the implementation of ST cessation support in dental settings in Pakistan. Utilizing the 'Capability-Opportunity-Motivation-Behaviour' model, it provides an in-depth understanding of the inability of dentists in implementing effective behavioural interventions for ST cessation support in routine dental practice. Highlighting the striking discrepancy between the patients' need for and receptivity towards cessation support and the dentists' concerns over their patients' receptivity towards cessation support, it calls for the need for effective implementation strategies to optimize dentist-led tobacco cessation interventions in low-resource settings.


A poisonous surprise under the coat of the African crested rat.
Plant toxins are sequestered by many animals and the toxicity is frequently advertised by aposematic displays to deter potential predators. Such 'unpalatability by appropriation' is common in many invertebrate groups and also found in a few vertebrate groups. However, potentially lethal toxicity by acquisition has so far never been reported for a placental mammal. Here, we describe complex morphological structures and behaviours whereby the African crested rat, Lophiomys imhausi, acquires, dispenses and advertises deterrent toxin. Roots and bark of Acokanthera schimperi (Apocynaceae) trees are gnawed, masticated and slavered onto highly specialized hairs that wick up the compound, to be delivered whenever the animal is bitten or mouthed by a predator. The poison is a cardenolide, closely resembling ouabain, one of the active components in a traditional African arrow poison long celebrated for its power to kill elephants.


Risk factors for hypertension in a national cohort study.
Hypertension continues to be a major public health problem in the United States. We used data from the National Health and Nutrition Examination Survey Epidemiologic Followup Study (1971-1984) to examine predictors of hypertension for the 7,073 participants free from hypertension at the baseline examination. The follow-up period averaged 10 years. Body mass index was positively related to the probability of hypertension developing among white men (n = 2,370), white women (n = 3,949), black men (n = 231), and black women (n = 523). Education was inversely associated with the probability of hypertension developing among white women and was of borderline significance among white men and black women. In a subanalysis of white men (n = 1,790) and white women (n = 3,063) who completed the 24-hour recall dietary questionnaire, dietary consumption of sodium, calcium, and potassium did not predict the development of hypertension. The failure of our study to support findings relating intake of dietary cations to the development of hypertension may be attributable to imprecision in the measurement of dietary data and misclassification of hypertension status. These data reinforce the importance of weight control in the primary prevention of hypertension.


The new scope of virus taxonomy: partitioning the virosphere into 15 hierarchical ranks.
Virus taxonomy emerged as a discipline in the middle of the twentieth century. Traditionally, classification by virus taxonomists has been focussed on the grouping of relatively closely related viruses. However, during the past few years, the International Committee on Taxonomy of Viruses (ICTV) has recognized that the taxonomy it develops can be usefully extended to include the basal evolutionary relationships among distantly related viruses. Consequently, the ICTV has changed its Code to allow a 15-rank classification hierarchy that closely aligns with the Linnaean taxonomic system and may accommodate the entire spectrum of genetic divergence in the virosphere. The current taxonomies of three human pathogens, Ebola virus, severe acute respiratory syndrome coronavirus and herpes simplex virus 1 are used to illustrate the impact of the expanded rank structure. This new rank hierarchy of virus taxonomy will stimulate further research on virus origins and evolution, and vice versa, and could promote crosstalk with the taxonomies of cellular organisms.


Retrieving the optical transmission matrix of a multimode fiber using the extended Kalman filter.
Characterizing the transmission matrix (TM) of a multimode fiber (MMF) benefits many fiber-based applications and allows in-depth studies on the physical properties. For example, by modulating the incident field, the knowledge of the TM allows one to synthesize any optical field at the distill end of the MMF. However, the extraction of optical fields usually requires holographic measurements with interferometry, which complicates the system design and introduces additional noise. In this work, we developed an efficient method to retrieve the TM of the MMF in a referenceless optical system. With pure intensity measurements, this method uses the extended Kalman filter (EKF) to recursively search for the optimum solution. To facilitate the computational process, a modified speckle-correlation scatter matrix (MSSM) is constructed as a low-fidelity initial estimation. This method, termed EKF-MSSM, only requires 4N intensity measurements to precisely solve for N unknown complex variables in the TM. Experimentally, we successfully retrieved the TM of the MMF with high precision, which allows optical focusing with the enhancement (>70%) close to the theoretical value. We anticipate that this method will serve as a useful tool for studying physical properties of the MMFs and potentially open new possibilities in a variety of applications in fiber optics.


The Prevalence of Stimulant and Antidepressant Use by Australian Children and Adolescents with Attention-Deficit/Hyperactivity Disorder and Major Depressive Disorder: A National Survey.
To identify the prevalence of stimulant and antidepressant medication use by children and adolescents with symptoms meeting the criteria for attention-deficit/hyperactivity disorder (ADHD) and major depressive disorder (MDD) in Australia. To identify factors associated with stimulant and antidepressant use by children and adolescents in Australia. Data are from a nationally representative sample of 4- to 17-year-olds (n = 6310). Parents completed the Diagnostic Interview Schedule for Children-Version IV (DISC-IV) and the Strengths and Difficulties Questionnaire. Eleven- to 17-year-olds completed a self-report version of the DISC-IV MDD module. Interviewers recorded prescribed medications used by participants in the previous 2 weeks. During a 2-week period, 1.3% of all 4- to 17-year-olds and 13.7% of those with symptoms meeting the criteria for ADHD had used stimulant medication, while 0.9% of all 4- to 17-year-olds and 13.4% with MDD had used antidepressants. In total, 22.6% of those using stimulant medications and 57.7% using antidepressant medications did not have symptoms meeting criteria for ADHD or MDD, respectively. Among 11- to 17-year-olds, 5.6% of those with adolescent-only-reported MDD, 10.9% of those with parent/carer-only-reported MDD, and 25.7% of those with MDD reported by both parents/carers and adolescents were using antidepressant medications. Only a minority of 4- to 17-year-olds with ADHD and MDD were being treated with stimulant or antidepressant medication. The percentage of adolescents with MDD using antidepressant medications varied depending on whether adolescents, parents/carers, or both identified the presence of MDD. This highlights the importance of using information from both these informants when assessing and treating adolescent depressive disorder.


The influence of educational level and oral hygiene behaviours on DMFT index and CPITN index in an adult Italian population: an epidemiological study.
To estimate the influence of educational level and oral hygiene behaviours on the prevalence and severity of dental caries and periodontal disease in an adult Italian population attending the Oral Hygiene department of a public Dental Clinic. Dental caries was diagnosed according to the World Health Organization criteria. The DMFT index (decayed, missing, filled tooth) was used to record the dental caries' experience. The periodontal status was assessed using the community periodontal index of treatment needs (CPITN). Questionnaires on educational level and oral hygiene behaviours were also collected. A total of 350 patients were enrolled. The mean DMFT value reported was 4.37 ± 3.06, and higher values were observed for male patients (P < 0.05). Increased CPITN scores and DMFT values were significantly correlated with lower level of education (P < 0.05). Subjects of high educational status showed significantly better oral hygiene habits (P < 0.05). The oral health status, in terms of periodontal disease and dental caries, appears correlated with patients' educational level.


Analytical Platforms 3: Processing Samples via the RPPA Pipeline to Generate Large-Scale Data for Clinical Studies.
Reverse phase protein array (RPPA) is a functional proteomics technology amenable to moderately high throughputs of samples and antibodies. The University of Texas MD Anderson Cancer Center RPPA Core Facility has implemented various processes and techniques to maximize RPPA throughput; key among them are maximizing array configuration and relying on database management and automation. One major tool used by the RPPA Core is a semi-automated RPPA process management system referred to as the RPPA Pipeline. The RPPA Pipeline, developed with the aid of MD Avnderson's Department of Bioinformatics and Computational Biology and InSilico Solutions, has streamlined sample and antibody tracking as well as advanced quality control measures of various RPPA processes. This chapter covers RPPA Core processes associated with the RPPA Pipeline workflow from sample receipt to sample printing to slide staining and RPPA report generation that enables the RPPA Core to process at least 13,000 samples per year with approximately 450 individual RPPA-quality antibodies. Additionally, this chapter will cover results of large-scale clinical sample processing, including The Cancer Genome Atlas Project and The Cancer Proteome Atlas.


Reassessing Graphene Absorption and Emission Spectroscopy.
We present a new paradigm for understanding optical absorption and hot electron dynamics experiments in graphene. Our analysis pivots on assigning proper importance to phonon-assisted indirect processes and bleaching of direct processes. We show indirect processes figure in the excess absorption in the UV region. Experiments which were thought to indicate ultrafast relaxation of electrons and holes, reaching a thermal distribution from an extremely nonthermal one in under 5-10 fs, instead are explained by the nascent electron and hole distributions produced by indirect transitions. These need no relaxation or ad-hoc energy removal to agree with the observed emission spectra and fast pulsed absorption spectra. The fast emission following pulsed absorption is dominated by phonon-assisted processes, which vastly outnumber direct ones and are always available, connecting any electron with any hole any time. Calculations are given, including explicitly calculating the magnitude of indirect processes, supporting these views.


Hepatic Metabolic, Inflammatory, and Stress-Related Gene Expression in Growing Mice Consuming a Low Dose of Trans-10, cis-12-Conjugated Linoleic Acid.
Dietary trans-10, cis-12-conjugated linoleic acid (trans-10, cis-12-CLA) fed to obese and nonobese rodents reduces body fat but leads to greater liver mass due to steatosis. The molecular mechanisms accompanying such responses remain largely unknown. Our study investigated the effects of chronic low trans-10, cis-12-CLA supplementation on hepatic expression of 39 genes related to metabolism, inflammation, and stress in growing mice. Feeding a diet supplemented with 0.3% trans-10, cis-12-CLA (wt/wt basis) for 6 weeks increased liver mass and concentration of long-chain fatty acids (LCFAs) in liver, while adipose tissue mass decreased markedly. These changes were accompanied by greater expression of genes involved in LCFA uptake (Cd36), lipogenesis, and triacylglycerol synthesis (Acaca, Gpam, Scd, Pck1, Plin2). Expression of these genes was in line with upregulation of the lipogenic transcription factor Srebf1. Unlike previous studies where higher >0.50% of the diet) doses of trans-10, cis-12-CLA were fed, we found greater expression of genes associated with VLDL assembly/secretion (Mttp, Cideb), ketogenesis (Hmgcs2, Bdh1), and LCFA oxidation (Acox1, Pdk4) in response to trans-10, cis-12-CLA. Dietary CLA, however, did not affect inflammation- and stress-related genes. Results suggested that a chronic low dose of dietary CLA increases liver mass and lipid accumulation due to activation of lipogenesis and insufficient induction of LCFA oxidation and VLDL assembly/secretion.


Reproducing the molecular subclassification of peripheral T-cell lymphoma-NOS by immunohistochemistry.
Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of mature T-cell malignancies; approximately one-third of cases are designated as PTCL-not otherwise specified (PTCL-NOS). Using gene-expression profiling (GEP), we have previously defined 2 major molecular subtypes of PTCL-NOS, PTCL-GATA3 and PTCL-TBX21, which have distinct biological differences in oncogenic pathways and prognosis. In the current study, we generated an immunohistochemistry (IHC) algorithm to identify the 2 subtypes in paraffin tissue using antibodies to key transcriptional factors (GATA3 and TBX21) and their target proteins (CCR4 and CXCR3). In a training cohort of 49 cases of PTCL-NOS with corresponding GEP data, the 2 subtypes identified by the IHC algorithm matched the GEP results with high sensitivity (85%) and showed a significant difference in overall survival (OS) (P = .03). The IHC algorithm classification showed high interobserver reproducibility among pathologists and was validated in a second PTCL-NOS cohort (n = 124), where a significant difference in OS between the PTCL-GATA3 and PTCL-TBX21 subtypes was confirmed (P = .003). In multivariate analysis, a high International Prognostic Index score (3-5) and the PTCL-GATA3 subtype identified by IHC were independent adverse predictors of OS (P = .0015). Additionally, the 2 IHC-defined subtypes were significantly associated with distinct morphological features (P < .001), and there was a significant enrichment of an activated CD8+ cytotoxic phenotype in the PTCL-TBX21 subtype (P = .03). The IHC algorithm will aid in identifying the 2 subtypes in clinical practice, which will aid the future clinical management of patients and facilitate risk stratification in clinical trials.


EMDR for children with medically related subthreshold PTSD: short-term effects on PTSD, blood-injection-injury phobia, depression and sleep.
Background: Paediatric illness, injury and medical procedures are potentially traumatic experiences with a range of possible negative psychosocial consequences. To prevent psychosocial impairment and improve medical adherence, evidence-based psychotherapy should be offered if indicated. Eye movement desensitization and reprocessing (EMDR) has been found to reduce symptoms of posttraumatic stress disorder (PTSD) in adults. The evidence for the use with children is promising. Furthermore, recent studies indicate its effectiveness for the treatment of other psychological symptomatology. However, the effectiveness of EMDR in children with subthreshold PTSD after medically related trauma has not yet been investigated. Objective: Investigating the short-term effectiveness of EMDR on posttraumatic stress, anxiety, depression and sleep problems in children with subthreshold PTSD after hospitalization through a randomized controlled trial (RCT). Method: Following baseline screening of 420 children from various Dutch hospitals, 74 children (4-15 years old) with medically related subthreshold PTSD were randomized to EMDR (n = 37) or care-as-usual (CAU; n = 37). Follow-up assessment took place after M = 9.7 weeks. Generalized Estimating Equation (GEE) analyses were performed to examine the effectiveness of EMDR compared to CAU. Results: Children in both groups improved significantly over time on all outcomes. However, the EMDR group improved significantly more as to child-reported symptoms of blood-injection-injury (BII) phobia and depression, and child-, and parent-reported sleep problems of the child. There was no superior effect of EMDR compared to CAU on subthreshold PTSD symptom reduction. Conclusions: EMDR did not perform better than CAU in reducing PTSD symptoms in a paediatric sample of children with subthreshold PTSD after hospitalization. However, the study results indicate that EMDR might be superior in reducing symptoms of blood-injection-injury phobia, depression and sleep problems.


Preventing neuronal edema increases network excitability after traumatic brain injury.
Edema is an important target for clinical intervention after traumatic brain injury (TBI). We used in vivo cellular resolution imaging and electrophysiological recording to examine the ionic mechanisms underlying neuronal edema and their effects on neuronal and network excitability after controlled cortical impact (CCI) in mice. Unexpectedly, we found that neuronal edema 48 hours after CCI was associated with reduced cellular and network excitability, concurrent with an increase in the expression ratio of the cation-chloride cotransporters (CCCs) NKCC1 and KCC2. Treatment with the CCC blocker bumetanide prevented neuronal swelling via a reversal in the NKCC1/KCC2 expression ratio, identifying altered chloride flux as the mechanism of neuronal edema. Importantly, bumetanide treatment was associated with increased neuronal and network excitability after injury, including increased susceptibility to spreading depolarizations (SDs) and seizures, known agents of clinical worsening after TBI. Treatment with mannitol, a first-line edema treatment in clinical practice, was also associated with increased susceptibility to SDs and seizures after CCI, showing that neuronal volume reduction, regardless of mechanism, was associated with an excitability increase. Finally, we observed an increase in excitability when neuronal edema normalized by 1 week after CCI. We conclude that neuronal swelling may exert protective effects against damaging excitability in the aftermath of TBI and that treatment of edema has the potential to reverse these effects.


Opportunities for earlier diagnosis of type 1 diabetes in children: A case-control study using routinely collected primary care records.
The epidemiology of type 1 diabetes mellitus (T1DM) suggests diagnostic delays may contribute to children developing diabetic ketoacidosis at diagnosis. We sought to quantify opportunities for earlier diagnosis of T1DM in primary care. A matched case-control study of children (0-16 years) presenting to UK primary care, examining routinely collected primary care consultation types and National Institute for Health and Care Excellence (NICE) warning signs in the 13 weeks before diagnosis. Our primary analysis included 1920 new T1DM cases and 7680 controls. In the week prior to diagnosis more cases than controls had medical record entries (663, 34.5% vs 1014, 13.6%, odds ratio 3.46, 95% CI 3.07-3.89; p<0.0001) and the incidence rate of face-to-face consultations was higher in cases (mean 0.32 vs 0.11, incidence rate ratio 2.90, 2.61-3.21; p<0.0001). The preceding week entries were found in 330 cases and 943 controls (17.2% vs 12.3%, OR 1.49, 1.3-1.7, p<0.0001), but face-to-face consultations were no different (IRR 1.08 (0.9-1.29, p=0.42)). There may be opportunities to reduce time to diagnosis for up to one third of cases, by up to two weeks. Diagnostic opportunities might be maximised by measures that improve access to primary care, and public awareness of T1DM.


Inhibition of swarming motility of Pseudomonas aeruginosa by branched-chain fatty acids.
Pseudomonas aeruginosa is capable of moving by swimming, swarming, and twitching motilities. In this study, we investigated the effects of fatty acids on Pseudomonas aeruginosa PAO1 motilities. A branched-chain fatty acid (BCFA)--12-methyltetradecanoic acid (anteiso-C15:0)--has slightly repressed flagella-driven swimming motility and completely inhibited a more complex type of surface motility, i.e. swarming, at a concentration of 10 microg mL(-1). In contrast, anteiso-C15:0 exhibited no effect on pili-mediated twitching motility. Other BCFAs and unsaturated fatty acids tested in this study showed similar inhibitory effects on swarming motility, although the level of inhibition differed between these fatty acids. These fatty acids caused no significant growth inhibition in liquid cultures. Straight-chain saturated fatty acids such as palmitic acid were less effective in swarming inhibition. The wetness of the PAO1 colony was significantly reduced by the addition of anteiso-C15:0; however, the production of rhamnolipids as a surface-active agent was not affected by the fatty acid. In addition to motility repression, anteiso-C15:0 caused 31% repression of biofilm formation by PAO1, suggesting that BCFA could affect the multiple cellular activities of Pseudomonas aeruginosa.


Heading in the right direction: thermodynamics-based network analysis and pathway engineering.
Thermodynamics-based network analysis through the introduction of thermodynamic constraints in metabolic models allows a deeper analysis of metabolism and guides pathway engineering. The number and the areas of applications of thermodynamics-based network analysis methods have been increasing in the last ten years. We review recent applications of these methods and we identify the areas that such analysis can contribute significantly, and the needs for future developments. We find that organisms with multiple compartments and extremophiles present challenges for modeling and thermodynamics-based flux analysis. The evolution of current and new methods must also address the issues of the multiple alternatives in flux directionalities and the uncertainties and partial information from analytical methods.


How Heart Valves Evolve to Adapt to an Extreme-Pressure System: Morphologic and Biomechanical Properties of Giraffe Heart Valves.
Heart valves which exist naturally in an extreme-pressure system must have evolved in a way to resist the stresses of high pressure. Giraffes are interesting as they naturally have a blood pressure twice that of humans. Thus, knowledge regarding giraffe heart valves may aid in developing techniques to design improved pressure-resistant biological heart valves. Heart valves from 12 giraffes and 10 calves were explanted and subjected to either biomechanical or morphological examinations. Strips from the heart valves were subjected to cyclic loading tests, followed by failure tests. Thickness measurements and analyses of elastin and collagen content were also made. Valve specimens were stained with hematoxylin and eosin, elastic van Gieson stain, Masson's trichrome and Fraser-Lendrum stain, as well as immunohistochemical reactions for morphological examinations. The aortic valve was shown to be 70% (95% CI 42-103%) stronger in the giraffe than in its bovine counterpart (p <0.001). No significant difference was found between mitral or pulmonary valves. After normalization for collagen, no significant differences were found in strength between species. The giraffe aortic valve was found to be significantly stiffer than the bovine aortic valve (p <0.001), with no significant difference between mitral and pulmonary valves. On a dry weight basis, the aortic (10.9%), pulmonary (4.3%), and mitral valves (9.6%) of giraffes contained significantly more collagen than those of calves. The elastin contents of the pulmonary valves (2.5%) and aortic valves (1.5%) were also higher in giraffes. The greater strength of the giraffe aortic valve is most likely due to a compact collagen construction. Both, collagen and elastin contents were higher in giraffes than in calves, which would make giraffe valves more resistant to the high-pressure forces. However, collagen also stiffens and thickens the valves. The mitral leaflets showed similar (but mostly insignificant) trends in strength, stiffness, and collagen content.


An Integrated Linkage Map of Three Recombinant Inbred Populations of Pea (Pisum sativum L.).
Biparental recombinant inbred line (RIL) populations are sets of genetically stable lines and have a simple population structure that facilitates the dissection of the genetics of interesting traits. On the other hand, populations derived from multiparent intercrosses combine both greater diversity and higher numbers of recombination events than RILs. Here, we describe a simple population structure: a three-way recombinant inbred population combination. This structure was easy to produce and was a compromise between biparental and multiparent populations. We show that this structure had advantages when analyzing cultivar crosses, and could achieve a mapping resolution of a few genes.


Two-year observation of pulmonary function in workers exposed to low concentrations of toluene diisocyanate.
Pulmonary effects were investigated in 106 toluene diisocyanate (TDI)-exposed workers and 39 referents in 64 exposed workers and 21 referents on the 2-year follow-up study in 1982. Means of individual, time-weighted average (TWA) exposure concentrations measured by personal monitors were approximately 0.001 ppm in each year. Short-term exposure at 0.02 ppm or over was observed in 9.3% of the collected samples in 1980 and 1.9% in 1982. Pulmonary function was assessed by measurements of maximum expiratory flow-volume curve and respiratory impedance. No differences were observed in the means of the pulmonary functions and their individual daily changes between the TDI workers and the referents. Significant intra-individual, two-year decreases were observed in some pulmonary function parameters in both groups, but when the effect of aging was adjusted, the decreases disappeared. Prevalences of respiratory symptoms in the TDI workers were not significantly higher than those in the referents, except for irritating complaints of the eyes and throat probably due to peak exposure to TDI and/or co-existing irritants. Eight of the TDI workers had episodes of acute asthmatic reactions shortly after having begun their TDI jobs and most parameters in their maximum expiratory flow were significantly less than the predicted values, though exposure concentrations when the episodes occurred could not be defined. From these results, it was suggested that TDI exposure at the levels of 0.001 ppm may not induce adverse pulmonary effects when the workers are not hypersusceptive to TDI.


Probing Mobility-Selected Saccharide Isomers: Selective Ion-Molecule Reactions and Wavelength-Specific IR Activation.
Differential Ion Mobility Spectrometry (DIMS) provides orthogonal separation to mass spectrometry, and DIMS combined with the high sensitivity of a quadrupole ion-trap is shown to be useful for the separation and identification of saccharides. A comprehensive analysis of the separation of anomers (α- and β-methylated glucose) and epimers (α-methylated glucose and mannose) ionized with Li(+), Na(+), and K(+) is performed. DIMS separation is found to be better for saccharides cationized with the two latter species. The corresponding resolving power for the two glucose anomers with Na(+) is found to be very close to the corresponding drift-tube IMS value. The lithiated complexes are investigated further using a combination of infrared spectroscopy integrated to ion-trap mass spectrometry and quantum chemical calculations. Together with DIMS, consistent results are obtained. It is found that two competing structural motifs might be at play, depending on the subtle balance between the maximization of the coordination of the metal cation and the intrinsic conformational energetics of the saccharide, which is for a large part driven by hydrogen bonding. The comparison of simulated and observed spectra clearly shows that a band at ∼3400 cm(-1) is specific to a structural motif found in the lithiated glucose complexes, which could explain the trends observed in the DIMS spectra of the saccharide complexes. It is shown that DIMS-MS/MS using wavelength specific IR activation would provide a new orthogonal dimension to mass spectrometry.


Living history of physiology: Bodil Schmidt-Nielsen.
In 2005, The American Physiological Society initiated The Living History of Physiology Project to recognize senior members who have made extraordinary contributions during their career to the advancement of the discipline and profession of physiology. Each physiologist will be interviewed for archival purposes, and the video tape will be available from the American Physiological Society Headquarters. In addition, a biographical profile of the recipient will be published in Advances in Physiology Education.


Principles of a brain-computer interface (BCI) based on real-time functional magnetic resonance imaging (fMRI).
A brain-computer interface (BCI) based on functional magnetic resonance imaging (fMRI) records noninvasively activity of the entire brain with a high spatial resolution. We present a fMRI-based BCI which performs data processing and feedback of the hemodynamic brain activity within 1.3 s. Using this technique, differential feedback and self-regulation is feasible as exemplified by the supplementary motor area (SMA) and parahippocampal place area (PPA). Technical and experimental aspects are discussed with respect to neurofeedback. The methodology now allows for studying behavioral effects and strategies of local self-regulation in healthy and diseased subjects.


Childhood prediction models for hypertension later in life: a systematic review.
Hypertension, even during childhood, increases the risk of developing atherosclerosis and cardiovascular disease. Therefore, starting prevention of hypertension early in the life course could be beneficial. Prediction models might be useful for identifying children at increased risk of developing hypertension, which may enable targeted primordial prevention of cardiovascular disease. To provide an overview of childhood prediction models for future hypertension. Embase and Medline were systematically searched. Studies were included that were performed in the general population, and that reported on development or validation of a multivariable model for children to predict future high blood pressure, prehypertension or hypertension. Data were extracted using the CHARMS checklist for prediction modelling studies. Out of 12 780 reviewed records, six studies were included in which 18 models were presented. Five studies predicted adulthood hypertension, and one predicted adolescent prehypertension/hypertension. BMI and current blood pressure were most commonly included as predictors in the final models. Considerable heterogeneity existed in timing of prediction (from early childhood to late adolescence) and outcome measurement. Important methodological information was often missing, and in four studies information to apply the model in new individuals was insufficient. Reported area under the ROC curves ranged from 0.51 to 0.74. As none of the models were validated, generalizability could not be confirmed. Several childhood prediction models for future hypertension were identified, but their value for practice remains unclear because of suboptimal methods, limited information on performance, or the lack of external validation. Further validation studies are indicated.


Public Health Resilience Checklist for High-Consequence Infectious Diseases-Informed by the Domestic Ebola Response in the United States.
The experiences of communities that responded to confirmed cases of Ebola virus disease in the United States provide a rare opportunity for collective learning to improve resilience to future high-consequence infectious disease events. Key informant interviews (n = 73) were conducted between February and November 2016 with individuals who participated in Ebola virus disease planning or response in Atlanta, Georgia; Dallas, Texas; New York, New York; or Omaha, Nebraska; or had direct knowledge of response activities. Participants represented health care; local, state, and federal public health; law; local and state emergency management; academia; local and national media; individuals affected by the response; and local and state governments. Two focus groups were then conducted in New York and Dallas, and study results were vetted with an expert advisory group. Participants focused on a number of important areas to improve public health resilience to high-consequence infectious disease events, including governance and leadership, communication and public trust, quarantine and the law, monitoring programs, environmental decontamination, and waste management. Findings provided the basis for an evidence-informed checklist outlining specific actions for public health authorities to take to strengthen public health resilience to future high-consequence infectious disease events.


Highly efficient silver particle layers on glass substrate synthesized by the sonochemical method for surface enhanced Raman spectroscopy purposes.
A fast method for preparing of silver particle layers on glass substrates with high application potential for using in surface enhanced Raman spectroscopy (SERS) is introduced. Silver particle layers deposited on glass cover slips were generated in one-step process by reduction of silver nitrate using several reducing agents (ethylene glycol, glycerol, maltose, lactose and glucose) under ultrasonic irradiation. This technique allows the formation of homogeneous layers of silver particles with sizes from 80nm up to several hundred nanometers depending on the nature of the used reducing agent. Additionally, the presented method is not susceptible to impurities on the substrate surface and it does not need any additives to capture or stabilize the silver particles on the glass surface. The characteristics of prepared silver layers on glass substrate by the above mentioned sonochemical approach was compared with chemically prepared ones. The prepared layers were tested as substrates for SERS using adenine as a model analyte. The factor of Raman signal enhancement reached up to 5·10(5). On the contrary, the chemically prepared silver layers does not exhibit almost any pronounced Raman signal enhancement. Presented sonochemical approach for preparation of silver particle layers is fast, simple, robust, and is better suited for reproducible fabrication functional SERS substrates than chemical one.


Dentin sialophosphoprotein (DSPP) gene-silencing inhibits key tumorigenic activities in human oral cancer cell line, OSC2.
We determined recently that dentin sialophosphoprotein (DSPP), a member of the SIBLING (Small integrin-binding ligand N-linked glycoproteins) family of phosphoglycoproteins, is highly upregulated in human oral squamous cell carcinomas (OSCCs) where upregulation is associated with tumor aggressiveness. To investigate the effects of DSPP-silencing on the tumorigenic profiles of the oral cancer cell line, OSC2, short-hairpin RNA (shRNA) interference was employed to silence DSPP in OSC2 cells. Multiple regions of DSPP transcript were targeted for shRNA interference using hDSP-shRNA lentiviral particles designed to silence DSPP gene expression. Control shRNA plasmid encoding a scrambled sequence incapable of degrading any known cellular mRNA was used for negative control. Following puromycin selection of stable lines of DSSP-silenced OSC2 cells, phenotypic hallmarks of oral carcinogenesis were assayed by western blot and RT-PCR analyses, MTT (cell-viability), colony-formation, modified Boyden-Chamber (migration and invasion), and flow cytometry (cell-cycle and apoptosis) analyses. DSPP-silenced OSC2 cells showed altered cell morphology, reduced viability, decreased colony-formation ability, decreased migration and invasion, G0/G1 cell-cycle arrest, and increased tumor cell sensitivity to cisplatin-induced apoptosis. Furthermore, MMP-2, MMP-3, MMP-9, VEGF, Ki-67, p53, and EGFR were down-regulated. There was a direct correlation between the degree of DSPP-silencing and MMP suppression, as indicated by least squares regression: MMP-2 {(y = 0.850x, p<0.001) (y = 1.156x, p<0.001)}, MMP-3 {(y = 0.994x, p<0.001) (y = 1.324x, p = 0.004)}, and MMP-9 {(y = 1.248x, p = 0.005, y = 0.809, p = 0.013)}. DSPP-silencing in OSC2 cell decreased salient hallmarks of oral tumorigenesis and provides the first functional evidence of a potential key role for DSPP in oral cancer biology. The down-regulation of MMP-2, MMP-3, MMP-9, p53 and VEGF in DSPP-silenced OSC2 cells provides a significant functional/molecular framework for deciphering the mechanisms of DSPP activities in oral cancer biology.


Impact of Diagnosis and Therapy on Cognitive Function in Urea Cycle Disorders.
Individuals with urea cycle disorders (UCDs) often present with intellectual and developmental disabilities. The major aim of this study was to evaluate the impact of diagnostic and therapeutic interventions on cognitive outcomes in UCDs. This prospective, observational, multicenter study includes data from 503 individuals with UCDs who had comprehensive neurocognitive testing with a cumulative follow-up of 702 patient-years. The mean cognitive standard deviation score (cSDS) was lower in symptomatic than in asymptomatic (p < 0.001, t test) individuals with UCDs. Intellectual disability (intellectual quotient < 70, cSDS < -2.0) was associated with the respective subtype of UCD and early disease onset, whereas height of the initial peak plasma ammonium concentration was inversely associated with neurocognitive outcomes in mitochondrial (proximal) rather than cytosolic (distal) UCDs. In ornithine transcarbamylase and argininosuccinate synthetase 1 deficiencies, we did not find evidence that monoscavenger therapy with sodium or glycerol phenylbutyrate was superior to sodium benzoate in providing cognitive protection. Early liver transplantation appears to be beneficial for UCDs. It is noteworthy that individuals with argininosuccinate synthetase 1 and argininosuccinate lyase deficiencies identified by newborn screening had better neurocognitive outcomes than those diagnosed after the manifestation of first symptoms. Cognitive function is related to interventional and non-interventional variables. Early detection by newborn screening and early liver transplantation appear to offer greater cognitive protection, but none of the currently used nitrogen scavengers was superior with regard to long-term neurocognitive outcome. Further confirmation could determine these variables as important clinical indicators of neuroprotection for individuals with UCDs. ANN NEUROL 2019.


Geographic variability in potentially discretionary red blood cell transfusions after coronary artery bypass graft surgery.
A number of established regional quality improvement collaboratives have partnered to assess and improve care across their regions under the umbrella of the Cardiac Surgery Quality Improvement (IMPROVE) Network. The first effort of the IMPROVE Network has been to assess regional differences in potentially discretionary transfusions (<3 units red blood cells [RBCs]). We examined 11,200 patients undergoing isolated nonemergent coronary artery bypass graft surgery across 56 medical centers in 4 IMPROVE Network regions between January 2008 and June 2012. Each center submitted the most recent 200 patients who received 0, 1, or 2 units of RBC transfusion during the index admission. Patient and disease characteristics, intraoperative practices, and percentage of patients receiving RBC transfusions were collected. Region-specific transfusion rates were calculated after adjusting for pre- and intraoperative factors among region-specific centers. There were small but significant differences in patient case mix across regions. RBC transfusions of 1 or 2 units occurred among 25.2% of coronary artery bypass graft procedures (2826 out of 11,200). Significant variation in the number of RBC units used existed across regions (no units, 74.8% [min-max, 70.0%-84.1%], 1 unit, 9.7% [min-max, 5.1%-11.8%], 2 units, 15.5% [min-max, 9.1%-18.2%]; P < .001). Variation in overall transfusion rates remained after adjustment (9.1%-31.7%; P < .001). Delivery of small volumes of RBC transfusions was common, yet varied across geographic regions. These data suggest that differences in regional practice environments, including transfusion triggers and anemia management, may contribute to variability in RBC transfusion rates.


Effect of root death and decay on dissipation of polycyclic aromatic hydrocarbons in the rhizosphere of yellow sweet clover and tall fescue.
A 12-mo greenhouse study was conducted to evaluate the contribution of root death and decay on the dissipation of polycyclic aromatic hydrocarbons (PAHs) in rhizosphere soil. The contaminated soil was previously treated by land-farming, but residual PAHs remained after treatment. Tall fescue (Festuca arundinacea Schreb.) and yellow sweet clover (Melilotus officinalis Lam.) were the target plants. To specifically evaluate the effect of root decay on contaminant dissipation, plants were treated with glyphosate, a broad spectrum herbicide, to induce root decay. Although tall fescue treatments had the highest root and shoot biomass and root surface area, this plant did not result in the highest contaminant degradation rates. Significant differences were noted between treatments for seven PAHs, with the active yellow sweet clover resulting in 60 to 75% degradation of these compounds. Induced root death and decay did not produce a significant enhancement of PAH degradation. The PAH microbial degrader populations in the vegetated treatments were more than 100 times greater than those in the unvegetated control. The phospholipid fatty acid (PLFA) structural group profile shifted over the growing period, indicating a change in the community structure. In conclusion, phytoremediation was shown to be an effective polishing tool for PAH-affected soil previously subjected to biological treatment.


Gene diversity and biochemical markers among some Muslim populations of Uttar Pradesh.
A sample of 312 individuals belonging to Sheikh (80), Pathan (54), Ansari (82), Syed (33), Saifi (33) and Hindu Bania (30) populations were surveyed for four protein and three enzyme loci comprising 12 alleles. The markers used were protein (transferrin, haptoglobin, haemoglobin) and enzyme (lactate dehydrogenase, phosphogluconate dehydrogenase, adenylate kinase). Except caeruloplasmin, variants were found among all loci, though no rare variant appears. The populations show genetic equilibrium for all of the loci. Our gene frequencies show some difference from earlier studies for Muslims in general, there being no biradari wise study among Sunni Muslims earlier done anywhere in India or abroad. Heterozygosity ranged from 0.0123 to 0.1994 (Sheikh), 0.0182 to 0.2046 (Pathan), 0.0239 to 0.1844 (Ansari), 0.0587 to 0.3966 (Syed), 0 to 0.3909 (Bania); the average DST and GST values for the seven marker loci were 0.001032 and 0.00879, respectively. The results are discussed. The gene frequency study shows closer relationship of Ansaris and Saifis with native Hindu Banias, and may reflect on their probable conversion in the not remote past.


Advancing the application of systems thinking in health: managing rural China health system development in complex and dynamic contexts.
This paper explores the evolution of schemes for rural finance in China as a case study of the long and complex process of health system development. It argues that the evolution of these schemes has been the outcome of the response of a large number of agents to a rapidly changing context and of efforts by the government to influence this adaptation process and achieve public health goals. The study draws on several sources of data including a review of official policy documents and academic papers and in-depth interviews with key policy actors at national level and at a sample of localities. The study identifies three major transition points associated with changes in broad development strategy and demonstrates how the adaptation of large numbers of actors to these contextual changes had a major impact on the performance of the health system. Further, it documents how the Ministry of Health viewed its role as both an advocate for the interests of health facilities and health workers and as the agency responsible for ensuring that government health system objectives were met. It is argued that a major reason for the resilience of the health system and its ability to adapt to rapid economic and institutional change was the ability of the Ministry to provide overall strategy leadership. Additionally, it postulates that a number of interest groups have emerged, which now also seek to influence the pathway of health system development. This history illustrates the complex and political nature of the management of health system development and reform. The paper concludes that governments will need to increase their capacity to analyze the health sector as a complex system and to manage change processes.


Evidence of a sex-dependent association between the MSX1 locus and nonsyndromic cleft lip with or without cleft palate in the Chilean population.
Prior studies have implicated an involvement of the Msx1 homeobox gene in cleft palate in mice and its homolog in humans (called MSX1 in the HOX7 gene, located on chromosome 4). In this study we present evidence of a sex-dependent association between MSX1 and non-syndromic cleft lip/palate (NSCLP) in the Chilean population. The sample included 73 NSCLP cases, 37 from multiplex families (Mx), 36 from simplex families (Sx), and 87 controls. Polymerase chain reaction amplification of the MSX1 intragenic microsatellite (CA)n-sequence shows significant (p = 0.035) differences in the allele frequencies between NSCLP-Mx males and control males. These differences are mainly due to frequency differences in allele *2 (173 base pairs) among cases (21.9%) and controls (13.2%). When the NSCLP cases are subdivided by sex and positive family history (Mx versus Sx), the Mx males (27.8%) as well as the total NSCLP-Mx cases (25.7%) showed significantly higher frequencies of allele *2, compared to controls (11.4% and 13.2%, respectively). Analysis of the genotype data indicates that the relative risk for NSCLP is greater for persons carrying allele *2 (i.e., odds ratio [OR] larger than 1), reaching significance for all Mx cases (OR = 2.67; 95% confidence interval [CI], 1.10 to 6.52) and even more pronounced for Mx males (OR = 3.33; 95% CI, 1.08 to 10.32). Taken together, these findings support the hypothesis that the genetic variation at the MSX1 locus is a predisposing gene involved in sex-dependent susceptibility to clefting and that it also differentiates simplex from multiplex families.


Potential clinical benefits of cell therapy in coronary heart disease: an update.
Cell therapy is a central issue of regenerative medicine and is raising a growing interest in the scientific community, but its full therapeutic potential in coronary heart disease (CHD) has not been reached yet. Several different methods, cell types, delivery routes, and supporting techniques have been attempted and improved to elicit cardiac regeneration in CHD, but only some of them showed a really convincing potential for the use in clinical practice. Here we provide an update on approaches and clinical trials of cell therapy applied to CHD, which are ongoing or that have been realized in the last 5 years. Moreover, we discuss the evidence collected so far in favor or against the validity of stem cell therapy for CHD. In particular, we review and comment the recent advances in cell therapy applied to CHD, the most promising cell types, delivery strategies, biochemical and engineering techniques that have been adopted in this context.


Synthetic Flavonoids, Aminoisoflavones: Interaction and Reactivity with Metal-Free and Metal-Associated Amyloid-β Species.
Metal ion homeostasis in conjunction with amyloid-β (Aβ) aggregation in the brain has been implicated in Alzheimer's disease (AD) pathogenesis. To uncover the interplay between metal ions and Aβ peptides, synthetic, multifunctional small molecules have been employed to modulate Aβ aggregation in vitro. Naturally occurring flavonoids have emerged as a valuable class of compounds for this purpose due to their ability to modulate both metal-free and metal-induced Aβ aggregation. Although, flavonoids have shown anti-amyloidogenic effects, the structural moieties of flavonoids responsible for such reactivity have not been fully identified. In order to understand the structure-interaction-reactivity relationship within the flavonoid family for metal-free and metal-associated Aβ, we designed, synthesized, and characterized a set of isoflavone derivatives, aminoisoflavones (1-4), that displayed reactivity (i.e., modulation of Aβ aggregation) in vitro. NMR studies revealed a potential binding site for aminoisoflavones between the N-terminal loop and central helix on prefibrillar Aβ different from the non-specific binding observed for other flavonoids. The absence or presence of the catechol group differentiated the binding affinities and enthalpy/entropy balance between aminoisoflavones and Aβ. Furthermore, having a catechol group influenced the binding mode with fibrillar Aβ. Inclusion of additional substituents moderately tuned the impact of aminoisoflavones on Aβ aggregation. Overall, through these studies, we obtained valuable insights on the requirements for parity among metal chelation, intermolecular interactions, and substituent variation for Aβ interaction.


Cardiomyocyte ATP release through pannexin 1 aids in early fibroblast activation.
Fibrosis following myocardial infarction is associated with increases in arrhythmias and sudden cardiac death. Initial steps in the development of fibrosis are not clear; however, it is likely that cardiac fibroblasts play an important role. In immune cells, ATP release from pannexin 1 (Panx1) channels acts as a paracrine signal initiating activation of innate immunity. ATP has been shown in noncardiac systems to initiate fibroblast activation. Therefore, we propose that ATP release through Panx1 channels and subsequent fibroblast activation in the heart drives the development of fibrosis in the heart following myocardial infarction. We identified for the first time that Panx1 is localized within sarcolemmal membranes of canine cardiac myocytes where it directly interacts with the postsynaptic density 95/Drosophila disk large/zonula occludens-1-containing scaffolding protein synapse-associated protein 97 via its carboxyl terminal domain (amino acids 300-357). Induced ischemia rapidly increased glycosylation of Panx1, resulting in increased trafficking to the plasma membrane as well as increased interaction with synapse-associated protein 97. Cellular stress enhanced ATP release from myocyte Panx1 channels, which, in turn, causes fibroblast transformation to the activated myofibroblast phenotype via activation of the MAPK and p53 pathways, both of which are involved in the development of cardiac fibrosis. ATP release through Panx1 channels in cardiac myocytes during ischemia may be an early paracrine event leading to profibrotic responses to ischemic cardiac injury.


Natural killer (NK) cell stimulatory factor increases the cytotoxic activity of NK cells from both healthy donors and human immunodeficiency virus-infected patients.
Natural killer cell stimulatory factor (NKSF), or interleukin 12 (IL-12), is a heterodimeric lymphokine produced by B cells that has multiple effects on T and NK cell functions. NKSF at concentrations as low as 0.4 pM enhances the spontaneous cytotoxic activity of peripheral blood lymphocytes (PBL) against a variety of tumor-derived target cell lines and virus-infected target cells. The combined treatment of PBL with NKSF and IL-2 results in a less than additive enhancement of cytotoxicity. NKSF enhances the cytotoxic activity of spontaneously cytotoxic CD16+CD5- NK cells and does not confer cytotoxic activity to CD16-CD5+ T cells. PBL from patients infected with human immunodeficiency virus (HIV) have significantly lower cytotoxic activity against tumor-derived target cells and virus-infected target cells than PBL from control healthy donors. Treatment of PBL from HIV-infected patients with NKSF and/or IL-2 results in an increase of NK cell cytotoxicity against both types of target cells to levels similar to or higher than those of untreated PBL from healthy donors. PBL from HIV-infected patients produce interferon gamma in response to NKSF and/or IL-2, although at levels 5- or 10-fold lower than those produced by PBL from healthy donors. The multiple biological effects of NKSF, its activity at very low molar concentrations, and its ability to synergize with other physiological stimuli suggest that NKSF/IL-12 is a lymphokine likely to have physiological importance and considerable therapeutic potential.


Candida albicans response regulator gene SSK1 regulates a subset of genes whose functions are associated with cell wall biosynthesis and adaptation to oxidative stress.
Ssk1p of Candida albicans is a putative response regulator protein of the Hog1 two-component signal transduction system. In Saccharomyces cerevisiae, the phosphorylation state of Ssk1p determines whether genes that promote the adaptation of cells to osmotic stress are activated. We have previously shown that C. albicans SSK1 does not complement the ssk1 mutant of S. cerevisiae and that the ssk1 mutant of C. albicans is not sensitive to sorbitol. In this study, we show that the C. albicans ssk1 mutant is sensitive to several oxidants, including hydrogen peroxide, t-butyl hydroperoxide, menadione, and potassium superoxide when each is incorporated in yeast extract-peptone-dextrose (YPD) agar medium. We used DNA microarrays to identify genes whose regulation is affected by the ssk1 mutation. RNA from mutant cells (strain CSSK21) grown in YPD medium for 3 h at 30 degrees C was reverse transcribed and then compared with similarly prepared RNA from wild-type cells (CAF2). We observed seven genes from mutant cells that were consistently up regulated (three-fold or greater compared to CAF2). In S. cerevisiae, three (AHP1, HSP12, and PYC2) of the seven genes that were up regulated provide cells with an adaptation function in response to oxidative stress; another gene (GPH1) is regulated under stress conditions by Hog1p. Three other genes that are up regulated encode a cell surface protein (FLO1), a mannosyl transferase (MNN4-4), and a putative two-component histidine kinase (CHK1) that regulates cell wall biosynthesis in C. albicans. Of the down-regulated genes, ALS1 is a known cell adhesin in C. albicans. Verification of the microarray data was obtained by reverse transcription-PCR for HSP12, AHP1, CHK1, PYC2, GPH1, ALS1, MNN4-4, and FLO1. To further determine the function of Ssk1p in the Hog1p signal transduction pathway in C. albicans, we used Western blot analysis to measure phosphorylation of Hog1p in the ssk1 mutant of C. albicans when grown under either osmotic or oxidative stress. We observed that Hog1p was phosphorylated in the ssk1 mutant of C. albicans when grown in a hyperosmotic medium but was not phosphorylated in the ssk1 mutant when the latter was grown in the presence of hydrogen peroxide. These data indicate that C. albicans utilizes the Ssk1p response regulator protein to adapt cells to oxidative stress, while its role in the adaptation to osmotic stress is less certain. Further, SSK1 appears to have a regulatory function in some aspects of cell wall biosynthesis. Thus, the functions of C. albicans SSK1 differ from those of S. cerevisiae SSK1.


In situ measurement and statistical modelling of Escherichia coli decay in small rivers.
Numerous studies have been carried out on the environmental factors associated with the decay of faecal bacteria in open (fresh or marine) waters. The present study aimed at understanding the fate of bacteria in small streams (flow <20 m3 s(-1)) for which there is a lack of knowledge. An original in situ protocol was developed for measuring the die-off of Escherichia coli (E. coli) from wastewater treatment plants. Based upon 80 values of the decay first-order parameter (K or its inverse T90), collected from five rivers in Normandy (France), a median T90 of 10 h and a minimal T90 of 1.3 h were obtained. K was then modelled as a linear function of variables made up from flow, water temperature and suspended particulate matter (SPM). The set of significant co-variables did not include light indicators. E. coli decay is inversely related to the river flow and it becomes highly significant below 0.3 m3 s(-1). The positive effect of small flows on die-off is increased by water temperature over 15 degrees C, whereas it could be reduced by SPM. The major co-variable of the model (p < 10(-9)) is an empiric composite variable integrating the effect of flow and temperature that explains more than 40% of the variance of K. We interpreted this as an expression of predation by benthic micro-grazers which could be the main cause of E. coli die-off in small streams in temperate countries.


Tandem mass spectrometric analysis of a mixture of isobars using the survival yield technique.
Collision induced dissociation tandem mass spectrometry experiments were performed to unequivocally separate compounds from an isobaric mixture of two products. The Survival Yield curve was obtained and is shown to consist in a linear combination of the curves corresponding to the two components separately. For such a mixture, a plateau appears on the diagram in lieu of the continuous decrease expected allowing for the structural study of the two components separately. The width of the plateau critically relates to the fragmentation parameters of the two molecular ions, which need to be sufficiently different structurally for the plateau to be observed. However, at constant fragmentation parameters, we have observed the width significantly increases at large m/z. This makes the separation more and more efficient as isobars have larger m/z and the technique complementary to those applicable at low m/z only. We have observed that the vertical position of the plateau relates linearly to the relative concentration of the two compounds that may be useful for quantification. Repeatability was estimated at 2% on a quadrupole ion trap. An advantage of using survival yield curves only, is that a priori knowledge of the respective fragmentation patterns of the two isobars becomes unnecessary. Consequently, similar performances are obtained if fragments are isobaric, which is also demonstrated in our study. The critical case of reverse peptides, at low m/z and similar fragmentation parameters, is also presented as a limitation of the method.


Plasma membrane receptors for ecotropic murine retroviruses require a limiting accessory factor.
A retroviral vector was used to express various amounts of the receptor (ecoR) for ecotropic host range murine retroviruses on naturally barren hamster, mink, and human cells. These cells and murine cells were then incubated for 2 h with dilutions of a helper-free ecotropic retrovirus that encodes human growth hormone, and the number of infected cells was later determined by growth hormone-specific immunofluorescence. For all cells under the conditions of these studies, virus adsorption was the limiting step of infection and the cellular capacities for infection were unsaturated either at cell surfaces or at intracellular sites. Thus, infections occurred at low multiplicities of infection per cell and were directly proportional to virus and cell concentrations, and only a small percentage (ca. 5%) of the infectious virions became adsorbed from the medium during the 2-h incubations. Although increasing the adsorption by raising virus or cell concentrations results in more infections in the cultures, increasing adsorption by raising the number of ecoR above a low threshold had no effect on infections. Thus, cells with a low number of ecoR were infected as efficiently as highly adsorbing cells that contained many times more ecoR. To reconcile these results, we conclude that only a small, set number of cell surface ecoR can be functional for infection and that all excess ecoR can only bind virus into an unsalvageable pool. Therefore, retroviral receptors on single cells are functionally diverse. Our results suggest that activity of ecoR in infection requires a limiting second cellular component.