Trifluoperazine, a calmodulin antagonist, inhibits muscle cell fusion.
We investigated the effect of trifluoperazine (TFP), a calmodulin antagonist, on the fusion of chick skeletal myoblasts in culture. TFP was found to inhibit myoblast fusion. This effect occurs at concentrations that have been reported to inhibit Ca2+-calmodulin in vitro, and is reversed upon removal of TFP. In addition, other calmodulin antagonists, including chlorpromazine, N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W7), and N-(6-aminohexyl)-1-naphthalene-sulfonamide (W5), inhibit fusion at doses that correspond closely to the antagonistic effects of these drugs on calmodulin. The expression of surface acetylcholine receptor, a characteristic aspect of muscle differentiation, is not impaired in TFP-arrested myoblasts. Myoblasts inhibited from fusion by 10 microM TFP display impaired alignment. In the presence of the Ca2+ ionophore A23187, the fusion block by 10 microM TFP is partially reversed and myoblast alignment is restored. The presence and distribution of calmodulin in both prefusional myoblasts and fused muscle cells was established by immunofluorescence. We observed an apparent redistribution of calmodulin staining that is temporally correlated with the onset of myoblast fusion. Our findings suggest a possible role for calmodulin in the regulation of myoblast fusion.


Overexpression of ORMDL3 confers sexual dimorphism in diet-induced non-alcoholic steatohepatitis.
The bioactive sphingolipid metabolites ceramide and sphingosine-1-phosphate (S1P) accumulate with overnutrition and have been implicated in non-alcoholic steatohepatitis (NASH) development. ORMDL3, a negative regulator of the rate-limiting step in ceramide biosynthesis, has been identified as an obesity-related gene. Therefore, we assessed the role of ORMDL3 in diet-induced obesity and development of NASH. Globally overexpressing Ormdl3-Flag transgenic mice (ORMDL3TG) were fed a western high-fat, carbohydrate and cholesterol enriched diet, with high fructose-glucose drinking water. Physiological, biochemical and sphingolipidomic analyses were employed to measure the effect of ORMDL3 overexpression on NASH development. ORMDL3TG male but not female mice fed a western high-fat diet and sugar water had exacerbated adipocyte hypertrophy together with increased severity of white adipose inflammation and fibrosis. Hepatic steatosis, dyslipidemia, impaired glucose homeostasis, hyperinsulinemia, and insulin resistance were significantly more severe only in obese ORMDL3TG male mice that accompanied dramatic liver fibrosis, inflammation, and formation of hepatic crown-like structures, which are unique features of human and murine NASH. Obesogenic diet induces ORMDL expression in male mice but reduces it in females. Mechanistically, overexpression of Ormdl3 lowered the levels of S1P and ceramides only in obese female mice and antithetically increased them in tissues of obese males. ORMDL3TG male mice exhibited a much greater induction of the UPR, propagating ER stress that contributed to their early development of NASH. This study uncovered a previously unrecognized role for ORMDL3 in sexual dimorphism important for the development and progression of NASH.


Cow Milk and Intestinal Epithelial Cell-derived Extracellular Vesicles as Systems for Enhancing Oral Drug Delivery.
Ingestion is the preferred way for drug administration. However, many drugs have poor oral bioavailability, warranting the use of injections. Extracellular vesicles (EVs) from cow milk have shown potential utility in improving oral drug bioavailability. However, EVs produced by intestinal epithelial cells have not been investigated for this application. We compared the capacity of cow milk EVs and intestinal epithelial cell-derived counterparts to enhance oral drug bioavailability. EVs were isolated, fluorescently labelled, and loaded with curcumin (CUR) as a model poorly absorbable drug. These were then characterised before testing in an intestinal model (Caco-2). Epithelial cell-derived EVs showed notably higher cell uptake compared to cow milk EVs. Cell uptake was significantly higher in differentiated compared to undifferentiated cells for both types of EVs. While both milk- and cell-derived EVs improved the cell uptake and intestinal permeability of CUR (confirming oral drug bioavailability enhancement potential), epithelial cell EVs demonstrated a superior effect.


Two groups of diabetic KK-CAy mice specifically bred for high and low sensitivity to exogenous acetylcholine and beta 1-adrenergic stimulation: interaction of higenamine and aconitine on pulse rate.
Diabetic KK-CAy mice were specifically bred for high and low sensitivity to the addition of exogenous acetylcholine (ACh). The sensitivity to ACh was measured by the change in pulse rate 2 min after the administration of ACh (10 mg/kg, s.c.). The two groups of mice, with high and low sensitivity to ACh, were specially selected and mated sequentially until the 12th filial generation. Although higenamine (100 micrograms/kg, i.p.), a beta 1-adrenergic agonist (a compound derived from aconite), had no effect per se, it inhibited aconitine (another compound derived from aconite extract)-induced bradycardia within 30 s of administration in ACh-low sensitive mice but not in ACh-high sensitive mice. The effects of aconitine and higenamine alone did not differ between these two groups of mice. This demonstrates that the high muscarinic and high beta 1-adrenergic sensitive mice may be stratified into two groups based upon an antagonistic interaction between higenamine and aconitine.


Structured light scatteroscopy.
A new imaging approach, structured light scatteroscopy (SLS), is demonstrated, which offers rapid wide-field imaging of microscopic morphological variations in bulk tissue surfaces. Elastic scattering of light offers exquisite sensitivity to ultrastructural changes at multiple size scales ranging from nanometers to millimeters, but in bulk tissues the confounding effects of molecular absorption and strong multiple scattering of light often lead to a dramatic reduction in scatter contrast and specificity. It is demonstrated that the SLS using structured high spatial frequency illumination and detection to probe the tissue achieves direct, absorption-independent, high-resolution maps of the scattering response. The scattering response is observed to be dependent on both the wavelength and spatial frequency of choice, indicating a potential for multiscale probing of ultrastructural changes in superficial tissue layers. This methodology can be easily applied in most wide-field imaging systems.


Genetic and environmental influences on stages of alcohol use across adolescence and into young adulthood.
The progression to alcohol dependence unfolds across multiple stages, including the decision to initiate use, the development of regular patterns of use, and (for some individuals) the subsequent development of problems associated with alcohol use. Using data from two population-based, longitudinal twin studies, FinnTwin16 (FT16) and FinnTwin12 (FT12), we applied multiple stage genetic models (Heath et al., Twin Res. 5 (2002) 113) to better understand the extent to which genetic and environmental influences impact the initiation of alcohol use, frequency of use in adolescence and young adulthood, and alcohol problems in young adulthood. Shared environmental factors played a large role in initiation, and a more moderate role on frequency of use, and it was largely the same influences acting across these stages of use. However, there was no significant evidence of shared environmental influences on alcohol problems in early adulthood. Problems were largely influenced by genetic factors that overlapped with genetic influences on frequency of use. Unique environmental factors were largely specific to each stage, with some overlap between alcohol problems and frequency of use at age 25.


Understanding the information behaviours of carers of people with dementia: a critical review of models from information science.
The aim of this review is to discuss how existing models of information behaviour may help to improve provision of information to carers of people with dementia. The article analyses existing models of information behaviour derived from information science, describes studies that have examined these models in relation to health and discusses ways in which they help to understand the information behaviours of carers of people with dementia. A comprehensive review of the literature on studies of information needs and a critical examination of models of information behaviours in relation to health were undertaken. Two dominant paradigms in information science research were identified, involving system-centred and user-centred approaches. System-centred approaches and studies are limited in that they have made assumptions on the types of information that people may require, the way in which information should be provided, the timing of information provision and have homogenous treated groups, failing to take account of individual preferences. In contrast, user-centred approaches recognise the unique needs of individuals and that information needs are subjective and affected by a variety of factors. User-centred models of information behaviour, particularly Dervin's sense-making theory and Wilson's Information Seeking Behaviour Models, could be useful in developing a better understanding of the information behaviours of carers of people with dementia. Adopting a user-centred approach to study the information behaviours of carers of people with dementia will take account of individual needs. Testing existing models of information behaviour within this group may help to develop interventions to meet the needs of individual carers and people with dementia.


Puerarin ameliorated the behavioral deficits induced by chronic stress in rats.
The present study aimed to investigate the mechanisms underlying the antidepressant-like effects of puerarin via the chronic unpredictable stress (CUS) procedure in rats. Similar to Sertraline (Ser), Chronic treatment of puerarin (60 and 120 mg/kg, i.g) elicited the antidepressant-like effects by reversing the decreased sucrose preference in sucrose preference test (SPT), by blocking the increased latency to feed in novelty-suppressed feeding test (NSFT) and the increased immobility time in forced swimming test (FST) without affecting locomotor activity. However, acute puerarin treatment did not ameliorate the antidepressant- and anxiolytic- like effects in FST and NSFT, respectively. In addition, enzyme linked immunosorbent assay (ELISA) and high performance liquid chromatography-electrochemical detection (HPLC-ECD) showed that chronic treatment of puerarin (60 and 120 mg/kg, i.g) reversed the decreased levels of progesterone, allopregnanolone, serotonin (5-HT) and 5-Hydroxyindoleacetic acid (5-HIAA) in prefrontal cortex and hippocampus of post-CUS rats. Furthermore, puerarin (60 and 120 mg/kg, i.g) blocked the increased corticotropin releasing hormone (CRH), corticosterone (Cort) and adrenocorticotropic hormone (ACTH). Collectively, repeated administration of puerarin alleviated the behavioral deficits induced by chronic stress which was associated with the biosynthesis of neurosteroids, normalization of serotonergic system and preventing HPA axis dysfunction.


Consultations between nurse prescribers and patients with diabetes in primary care: A qualitative study of patient views.
There is a drive to improve the quality of service provision for patients with diabetes and to enable better self-management of this condition. The adoption of prescribing by nurses is increasing worldwide and can potentially enhance service provision. Evidence suggests that patients prefer services where their lifestyle factors and opinions are considered by healthcare professionals within a partnership approach. Few studies have explored patients' views about their consultations with a nurse prescriber. To explore the views patients with diabetes have about their consultations with nurse prescribers and any impact this may have on their medications management. A qualitative study involving semi-structured interviews and thematic analysis. Six primary care sites in which nurses prescribed medications for patients with diabetes in England. Data was collected in 2009. Interviews took place with 41 patients with diabetes from the case loads of 7 nurse prescribers. Findings are reported under three themes; the nurse consultation style, benefits of the nurse prescriber consultation and views on involvement and decision-making. Key aspects of the nurse consultation style were a non-hurried approach, care and rapport, approachability, continuity, and providing clear information based on specialist knowledge. Many benefits were described, including improved access to appropriate advice and medication, greater understanding and ability to self-manage, ability to address problems and improved confidence, trust and wellbeing. While patients were happy with the amount of information received and involvement they had decisions about their treatment, there was some controversy over the consistency of information provided on side-effects of treatment. The study provides new knowledge about what patients with diabetes value and benefit from in respect to care provided by nurse prescribers. Continuity of relationship, flexibility over consultation length, nurses' interpersonal skills and specialist diabetes knowledge were identified as crucial to good quality care. Patients require that nurse prescribers are skilled in providing a person-centred approach and have access to specialist training. The level of information and involvement offered to patients should reflect patients' requirements.


Cultural differences in end-of-life care.
The exact time of death for many intensive care unit patients is increasingly preceded by an end-of-life decision. Such decisions are fraught with ethical, religious, moral, cultural, and legal difficulties. Key questions surrounding this issue include the difference between withholding and withdrawing, when to withhold/withdraw, who should be involved in the decision-making process, what are the relevant legal precedents, etc. Cultural variations in attitude to such issues are perhaps expected between continents, but key differences also exist on a more local basis, for example, among the countries of Europe. Physicians need to be aware of the potential cultural differences in the attitudes not only of their colleagues, but also of their patients and families. Open discussion of these issues and some change in our attitude toward life and death are needed to enable such patients to have a pain-free, dignified death.


Clinical Outcomes and Predictors of Satisfaction in Patients with Improved Lithium Disilicate All-Ceramic Crowns.
The aim of this study was to determine the clinical outcomes and predictors of satisfaction in patients with lithium disilicate (LD) ceramic crowns. Clinical outcomes were assessed in 47 patients with 88 LD crowns using modified United States Public Health Service (USPHS) evaluation criteria and survival rates. The questionnaire for predictors included 3 aspects: (a) sociodemographic characteristics, (b) oral health habits (tooth brushing frequency, flossing frequency, and dental visits), and (c) satisfaction of the restorations (aesthetics, function, fit, cleansability, and chewing ability of the crowns, and overall satisfaction). Frequency distributions were computed using univariate and multivariate analysis. The Student t test and analysis of variance (ANOVA) were used to compare means across variables. Correlation analysis was done to assess the association between continuous variables. The age of crowns was 34.7 ± 9.7 months. The survival rate was 96.6% at 35.9 ± 9.2 months. There was a significant association between successful crown function and oral hygiene measures: tooth brushing (p˂ 0.001), dental visits (p = 0.006), and flossing (p = 0.009). A strong negative correlation was observed between aesthetic satisfaction (r = -0.717, p˂ 0.001) and chewing ability (r = -0.639, p˂ 0.001) with crown age. The linear regression model was significant for all predictors (p < 0.05) except overall satisfaction (p > 0.05). The LD crowns had long survival rates of 96.6% up to 35.9 ± 9.2 months and provided satisfactory clinical performance (low risk of failure). Oral hygiene habits such as brushing, flossing, and regular dental visits influenced patient satisfaction with LD crowns.


Functional assessment of component positioning in patients with groin pain after total hip arthroplasty as a tool to guide management.
Persisting groin pain post total hip arthroplasty (THA) is a common and complex issue that can be difficult to diagnose and manage. Acetabular component positioning is often implicated. We used a previously well described and validated functional positioning protocol to determine if functional acetabular malpositioning was a factor in groin pain post THA and hence to determine if acetabular revision would be indicated. We compared patient-specific functional acetabular positioning to traditional CT evaluation of cup position and assessment of anterior cup overhang. 39 patients with groin pain post-THA were investigated. Functional acetabular malpositioning was diagnosed in 31% (12/39). Revision THA was performed in those 12 patients, resulting in resolution of functional malpositioning (100%), with an overall accuracy of 5.6° (range 1-12), and resolution of groin pain in 67% (8/12). 33% (4/12) of the revised implants had functional positioning located outside the traditional "40/20 zone". Comparison with CT indicated that 40% (4/10) of implants with anterior overhang were well positioned, however only 50% (6/12) of functionally malpositioned implants had CT evidence of anterior cup prominence. Of the 8/12 revision patients who had resolution of their groin pain, only 1 had cup prominence. This study suggests that the utilisation of a patient specific functional positioning algorithm in the analysis of persistent groin pain following THA can assist in identifying the underlying cause of pain and help to guide treatment. For a functionally malpositioned acetabulum, revision surgery offers a potential resolution of groin pain.


Associations of vitamin d binding protein gene polymorphisms with the development of peripheral arthritis and uveitis in ankylosing spondylitis.
Genetic factors account for more than 90% of overall susceptibility to ankylosing spondylitis (AS), and recent studies have focused on non-major histocompatibility complex genes. Vitamin D binding protein (DBP) is a highly polymorphic protein that transports vitamin D and its metabolites. In addition to its sterol binding capacity, DBP has many other roles in the inflammatory and immune systems, and has been reported to be associated with autoimmune diseases. We investigated the association between DBP polymorphisms and susceptibility to AS. This case-control study was conducted in 223 patients with AS and 239 ethnically matched controls who were genotyped for 8 single-nucleotide polymorphisms (SNP) in the DBP and its promoter. Genomic DNA was isolated from peripheral blood leukocytes using the standard phenolchloroform method, and the GoldenGate assay was used for genotyping. No significant association was found between the susceptibility to AS and DBP polymorphisms. In a subgroup analysis of patients with AS, G alleles at rs222016 and rs222020 (OR 0.63, 95% CI 0.42-0.95, p = 0.03; OR 0.63, 95% CI 0.42-0.95, p = 0.03, respectively) and A allele at rs3733359 (OR 0.59, 95% CI 0.39-0.90, p = 0.01) showed the decreased risk of peripheral arthritis. G allele at rs4752 showed increased risk of uveitis (OR 2.04, 95% CI 1.12-3.72, p = 0.02). On the haplotype analyses, haplotype 2 (AGGA) protected against the development of peripheral arthritis (p = 0.01) and haplotype 3 (GAAG) was associated with an increased likelihood of uveitis (p = 0.02). DBP gene polymorphisms are associated with the development of peripheral arthritis and uveitis in Korean patients with AS. Given the influence of different DBP variants on the immune system, larger-scale studies are warranted to elucidate the role of DBP in the pathogenesis of AS.


Developing a successful self-rostering shift system.
Three wards of different sizes and with a mixture of full-time and part-time staff, qualified and unqualified, took part in a self-rostering trial lasting a total of six months. The results showed a high satisfaction rate among staff, with 82 per cent saying they were usually able to organise a work and home life balance compared to 38 per cent before the trial. In addition baseline sickness, and figures on the use of bank and agency staff, were taken for the preceding three months and repeated at six and 16 months after self-rostering was adopted. Two years later, staff on those wards are continuing with self-rostering and the concept has now been extended to other areas in the trust.


The diagnostic value of power Doppler measurements in the endometrium of women with postmenopausal bleeding.
The objective was to evaluate the accuracy of blood flow measurements by power Doppler in endometrial vessels in women with postmenopausal bleeding (PMB). Sixty consecutive patients with PMB participated in the study. Endometrial thickness and power Doppler measurements of small vessels in the endometrium and subendometrial tissue were performed prior to dilatation and curettage. Correlation between Doppler indices, endometrial thickness (by transvaginal sonography), and histopathologic examination was performed. A Student t test was used for statistical analysis with P < 0.05 as the level of significance. Fourteen positive cases were found: 11 endometrial cancers, 1 sarcoma, 1 simple hyperplasia, and 1 complex hyperplasia with atypia. Measurements of endometrial thickness using a cutoff point of 5 mm revealed a sensitivity of 78% and specificity of 45.6% (P > 0.05, NS) for detecting endometrial pathology. Power Doppler measurements (pulsatility index cutoff point = 1.0) revealed a sensitivity of 85. 7% and specificity of 89% (P = 0.001) for detecting endometrial pathology. The noninvasive methods for endometrial evaluation are not sensitive enough to exclude endometrial pathology. When invasive methods could not be performed, the combination of transvaginal sonography and power Doppler imaging provided the best results. When both modalities are negative, the probability of cancer is less than 5%.


Age and Living Situation as Key Factors in Understanding Changes in Alcohol Use during COVID-19 Confinement.
(1) The aim of the present study was to evaluate and characterize changes in alcohol use during the COVID-19 confinement in a sample of Spanish adults, analyzing their age and living situation as defining life cycle variables. (2) Method: Data from 3779 individuals were collected through a set of online surveys. AUDIT-C was used to measure the frequency of consumption, the average daily consumption, intensive consumption, risky consumption, and Standard Drink Units. (3) Results: Although alcohol consumption during confinement showed a significant general decline, age revealed important differences, with the decline being more pronounced in adults from 18 to 29 years old. The living situation also showed significant differences. The largest decreases in alcohol consumption were found in those who lived with their parents or other relatives, whereas those who lived alone or with a partner even increased their level of consumption. In addition, the data show a significant interaction between these two variables and gender. (4) Conclusions: Age and cohabitation processes are key factors in understanding the life situation of each individual during confinement and, consequently, in explaining consumption patterns. The results obtained provide interesting recommendations for designing prevention policies in both normal and crisis circumstances, emphasizing the need to understand alcohol use from a psychosocial perspective.


Dopamine depletion alters phosphorylation of striatal proteins in a model of Parkinsonism.
Nigrostriatal dopamine depletion disrupts striatal medium spiny neuron morphology in Parkinson's disease and modulates striatal synaptic plasticity in animal models of parkinsonism. We demonstrate that long-term nigrostriatal dopamine depletion in the rat induces evolving changes in the phosphorylation of striatal proteins critical for synaptic plasticity. Dopamine depletion increased the phosphorylation of the alpha isoform of calcium-calmodulin-dependent protein kinase II (CaMKIIalpha) at Thr286, a site associated with enhanced autonomous kinase activity, but did not alter total levels of CaMKIIalpha or other synaptic proteins. Dopamine depletion decreased CaMKIIalpha levels in postsynaptic density-enriched fractions without significant changes in other proteins. The activity of protein phosphatase 1 (PP1), a postsynaptic phosphatase that dephosphorylates CaMKII, is regulated by DARPP-32 (dopamine- and cAMP-regulated phosphoprotein of 32 kDa). Dopamine depletion had no effect on DARPP-32 phosphorylation at Thr34, but increased DARPP-32 phosphorylation at Thr75. Levodopa administration reversed the increased phosphorylation of both CaMKIIalpha and DARPP-32. Normal ageing increased the levels of PP1(gamma1 isoform) but decreased levels of the PP1gamma1-targeting proteins spinophilin and neurabin. Elevated phosphorylations of CaMKIIalpha and DARPP-32 were maintained for up to 20 months after dopamine depletion. However, phosphorylation of the CaMKII-PP1 substrate, Ser831 in the glutamate receptor GluR1 subunit, was increased only after sustained (9-20 months) dopamine depletion. Interaction of ageing-related changes in PP1 with the dopamine depletion-induced changes in CaMKIIalpha may account for enhanced GluR1 phosphorylation only after long-term dopamine depletion. These evolving changes may impact striatal synaptic plasticity, Parkinson's disease progression and the changing efficacy and side-effects associated with dopamine replacement therapy.


A Prospective Hospital-based Surveillance to Estimate Rotavirus Disease Burden in Bhutanese Children under 5 Years of Age.
As part of efforts to develop an informed policy for rotavirus vaccination, this prospective study was conducted to estimate the burden of rotavirus diarrhea among children less than 5 years old attended to the Department of Pediatrics, Jigme Dorji Wangchuk National Referral Hospital (JDWNRH), Thimphu, Bhutan. The duration of the study was three years, extending from February 2010 through December 2012. We estimated the frequency of hospitalization in the pediatric ward and dehydration treatment unit (DTU) for diarrhea and the number of events attributable to rotavirus infection among children under 5 years of age. During the study period, a total of 284 children (1 in 45) were hospitalized in the pediatric ward, and 2,220 (1 in 6) in the DTU with diarrhea among children residing in the Thimphu district. Group A rotavirus was detected in 32.5% and 18.8% of the stool samples from children hospitalized in the pediatric ward, respectively. Overall, 22.3% of the stool samples were rotavirus-positive, and the majority (90.8%) of them was detected in children under 2 years of age. From this study, we estimated that the annual incidence of hospitalization in the pediatric ward and DTU due to rotavirus diarrhea was 2.4/1000 (95% CI 1.7-3.4) and 10.8/1000 (95% CI 9.1-12.7) children, respectively. This study revealed that rotavirus is a major cause of diarrhea in Bhutanese children in Thimphu district and since no study has been performed previously, represents an important finding for policy discussions regarding the adoption of a rotavirus vaccine in Bhutan.


Foot and ankle surgery in Australia: a descriptive analysis of the Medicare Benefits Schedule database, 1997-2006.
Foot and ankle problems are highly prevalent in the general community and a substantial proportion of people seek surgical treatment to alleviate foot pain and deformity. However, the epidemiology of foot and ankle surgery has not been examined in detail. Therefore, the aim of this study was to examine patterns and costs of private sector foot surgery provision in Australia. Data pertaining to all foot and ankle surgical procedures for the calendar years 1997-2006 were extracted from the Australian Medicare Benefits Schedule (MBS) database and were cross-tabulated by sex and age. Descriptive analyses were undertaken to assess sex and age differences in the number and type of procedures performed and to assess for temporal trends over the ten year assessment period. The total cost to Medicare of subsiding surgeons' fees in 2006 was also determined. During the 1997-2006 period, 996,477 surgical procedures were performed on the foot and ankle by private surgeons in Australia. Approximately equal numbers of procedures were performed on males (52%) and females (48%). However, males were more likely to undergo toenail, ankle, clubfoot, tarsal coalition and congenital vertical talus surgery, whereas females were more likely to undergo lesser toe, first metatarsophalangeal joint (MPJ), neuroma, heel, rearfoot and lesser MPJ surgery. The total number of procedures was stable over the assessment period, however there was a relative increase in the number of procedures performed on people aged over 55 years. The total contribution of Medicare to subsiding surgeons' fees for procedures performed in 2006 was $14 M. Foot and ankle surgery accounts for a considerable degree of healthcare expenditure in Australia, and the number of procedures in those aged over 55 years is increasing. Given the ageing demographics of the Australian population, the future public health and economic impact of foot morbidity is likely to be substantial. Strategies need to be implemented to ensure that the surgical labour force is adequate to address this increasing demand.


Cell-type-specific genome editing with a microRNA-responsive CRISPR-Cas9 switch.
The CRISPR-Cas9 system is a powerful genome-editing tool useful in a variety of biotechnology and biomedical applications. Here we developed a synthetic RNA-based, microRNA (miRNA)-responsive CRISPR-Cas9 system (miR-Cas9 switch) in which the genome editing activity of Cas9 can be modulated through endogenous miRNA signatures in mammalian cells. We created miR-Cas9 switches by using a miRNA-complementary sequence in the 5΄-UTR of mRNA encoding Streptococcus pyogenes Cas9. The miR-21-Cas9 or miR-302-Cas9 switches selectively and efficiently responded to miR-21-5p in HeLa cells or miR-302a-5p in human induced pluripotent stem cells, and post-transcriptionally attenuated the Cas9 activity only in the target cells. Moreover, the miR-Cas9 switches could differentially control the genome editing by sensing endogenous miRNA activities within a heterogeneous cell population. Our miR-Cas9 switch system provides a promising framework for cell-type selective genome editing and cell engineering based on intracellular miRNA information.


Quality of life and mortality assessment in patients with major cardiac events in the postoperative period.
Cardiovascular complications in the postoperative period are associated with high mortality and morbidity. Few studies have assessed the degree of dependence in these patients and their perception of health. The objective of this study was to assess the mortality and the quality of life in patients who developed major cardiac events (MCE) in the postoperative period. Retrospective study carried out in a Surgical Intensive Care Unit (SICU), between March 2006 and March 2008. The patients were assessed regarding the occurrence of CE. Six months after the hospital discharge, the Short-Form-36 (SF-36) questionnaire was filled out and dependence was assessed in relation to activities of daily living (ADL). The comparisons between independent groups of patients were carried out using Student's t test. The comparison between each variable and the occurrence of CE was carried out by logistic regression and included all patients. Of the 1,280 patients that met the inclusion criteria, 26 (2%) developed MCE. The univariate analysis identified as independent determinants for the development of major cardiac events: ASA physical status, hypertension, ischemic heart disease, congestive heart disease and score of the Revised Cardiac Risk Index (RCRI). The six-month mortality after the SICU discharge was 35%. Of the 17 surviving patients, 13 completed the questionnaires. Thirty-one percent of them reported that their general health was better on the day they answered the questionnaire, when compared to 12 months before. Sixty-nine percent of the patients were dependent in instrumental ADL e 15% in personal ADL. The development of MCE has a significant impact on the duration of hospital stay and mortality rates. Six months after the discharge from the SICU, more than 50% of the patients were dependent in at least one instrumental ADL.


[Evolution in the treatment of juvenile nasopharyngeal angiofibroma].
Juvenile nasopharyngeal angiofibroma (JNA) is a rare benign tumour in adolescent males. It may be associated with a significant morbidity because of its anatomical location and its locally destructive growth pattern. Severe haemorrhage constitutes a high risk in JNA and its surgical management could be complex. We retrospectively analysed the clinical data from 43 patients with JNA surgically treated in our Department from 1993 until 2010. Mean postoperative follow-up time was 85 months. Analysis was performed on 42 males and one female. Mean patient age was 16 years old. The most common presenting symptoms were unilateral epistaxis (56%) and nasal obstruction (56%). Using the Fisch staging scale, tumours were classified as stage I in 2 patients, stage II in 9, stage III-a in 13, stage III-b in 13 and stage IV-a in 6. Preoperative selective embolisation was performed on 32 patients (74%). Thirty-three patients (77%) underwent an open surgical approach and 10 (23%) were treated by endoscopic approach. Complete resection of the lesion was achieved in 35 patients (81%) and tumour recurrence was observed in 2 (5%). All lesions treated via transnasal endoscopic approach were stage I and stage II lesions. Surgery is the treatment of choice for JNA. An endoscopic approach is feasible for early-stage lesions (Fisch I and II) and conservative external approaches are still useful in advanced stages (Fisch III and IV). The open approaches proved helpful with respect to exposure, safety, cosmetic outcome and low morbidity. Preoperative embolisation, if possible, is mandatory.


In vivo delivery of plasmid DNA by lipid nanoparticles: the influence of ionizable cationic lipids on organ-selective gene expression.
Ionizable cationic lipids play a critical role in developing new gene therapies for various biomedical applications, including COVID-19 vaccines. However, it remains unclear whether the formulation of lipid nanoparticles (LNPs) using DLin-MC3-DMA, an optimized ionizable lipid clinically used for small interfering RNA (siRNA) therapy, also facilitates high liver-selective transfection of other gene therapies such as plasmid DNA (pDNA). Here we report the first investigation into pDNA transfection efficiency in different mouse organs after intramuscular and intravenous administration of lipid nanoparticles (LNPs) where DLin-MC3-DMA, DLin-KC2-DMA or DODAP are used as the ionizable cationic lipid component of the LNP. We discovered that these three benchmark lipids previously developed for siRNA delivery followed an unexpected characteristic rank order in gene expression efficiency when utilized for pDNA. In particular, DLin-KC2-DMA facilitated higher in vivo pDNA transfection than DLin-MC3-DMA and DODAP, possibly due to its head group pKa and lipid tail structure. Interestingly, LNPs formulated with either DLin-KC2-DMA or DLin-MC3-DMA exhibited significantly higher in vivo protein production in the spleen than in the liver. This work sheds light on the importance of the choice of ionizable cationic lipid and nucleic acid cargo for organ-selective gene expression. The study also provides a new design principle towards the formulation of more effective LNPs for biomedical applications of pDNA, such as gene editing, vaccines and immunotherapies.


A gradient of Wnt activity positions the neurosensory domains of the inner ear.
The auditory and vestibular organs of the inner ear and the neurons that innervate them originate from Sox2-positive and Notch-active neurosensory domains specified at early stages of otic development. Sox2 is initially present throughout the otic placode and otocyst, and then it becomes progressively restricted to a ventro-medial domain. Using gain- and loss-of-function approaches in the chicken otocyst, we show that these early changes in Sox2 expression are regulated in a dose-dependent manner by Wnt/beta-catenin signalling. Both high and very low levels of Wnt activity repress Sox2 and neurosensory competence. However, intermediate levels allow the maintenance of Sox2 expression and sensory organ formation. We propose that a dorso-ventral (high-to-low) gradient and wave of Wnt activity initiated at the dorsal rim of the otic placode progressively restricts Sox2 and Notch activity to the ventral half of the otocyst, thereby positioning the neurosensory competent domains in the inner ear.


Pathologic response and surgical outcomes in patients undergoing nephrectomy following receipt of immune checkpoint inhibitors for renal cell carcinoma.
To evaluate the pathologic response, safety, and feasibility of nephrectomy following receipt of immune checkpoint inhibition (ICI) for renal cell carcinoma (RCC). Patients who underwent nephrectomy for RCC after exposure to nivolumab monotherapy or combination ipilimumab/nivolumab were reviewed. Primary surgical outcomes included operative time (OT), estimated blood loss (EBL), length of stay (LOS), readmission rates, and complication rates. Pathologic response in the primary and metastatic sites constituted secondary outcomes. Eleven nephrectomies (10 radical, 1 partial) were performed in 10 patients after ICI with median postoperative follow-up 180 days. Six patients received 1 to 4 cycles of ipilimumab/nivolumab, while 5 received 2 to 12 infusions of nivolumab preoperatively. Five surgeries were performed laparoscopically, and 4 patients underwent concomitant thrombectomy. One patient exhibited complete response (pT0) to ICI, and 3/4 patients who underwent metastasectomy for hepatic, pulmonary, or adrenal lesions exhibited no detectable malignancy in any of the metastases resected. No patients experienced any major intraoperative complications, and all surgical margins were negative. Median OT, EBL, and LOS were 180 minutes, 100 ml, and 4 days, respectively. Four patients experienced a complication, including 3 that were addressed with interventional radiology procedures. One patient died of progressive disease >3 months after surgery, and 1 patient succumbed to pulmonary embolism complicated by sepsis. No complications or readmissions were noted in 6 patients. Nephrectomy following ICI for RCC is safe and technically feasible with favorable surgical outcomes and pathologic response. Timing of the nephrectomy relative to checkpoint dosing did not seem to impact outcome. Biopsies of lesions responding radiographically to ICI may warrant attention prior to surgical excision.


The burden of endometriosis: costs and quality of life of women with endometriosis and treated in referral centres.
This study aimed to calculate costs and health-related quality of life of women with endometriosis-associated symptoms treated in referral centres. A prospective, multi-centre, questionnaire-based survey measured costs and quality of life in ambulatory care and in 12 tertiary care centres in 10 countries. The study enrolled women with a diagnosis of endometriosis and with at least one centre-specific contact related to endometriosis-associated symptoms in 2008. The main outcome measures were health care costs, costs of productivity loss, total costs and quality-adjusted life years. Predictors of costs were identified using regression analysis. Data analysis of 909 women demonstrated that the average annual total cost per woman was €9579 (95% confidence interval €8559-€10 599). Costs of productivity loss of €6298 per woman were double the health care costs of €3113 per woman. Health care costs were mainly due to surgery (29%), monitoring tests (19%) and hospitalization (18%) and physician visits (16%). Endometriosis-associated symptoms generated 0.809 quality-adjusted life years per woman. Decreased quality of life was the most important predictor of direct health care and total costs. Costs were greater with increasing severity of endometriosis, presence of pelvic pain, presence of infertility and a higher number of years since diagnosis. Our study invited women to report resource use based on endometriosis-associated symptoms only, rather than drawing on a control population of women without endometriosis. Our study showed that the economic burden associated with endometriosis treated in referral centres is high and is similar to other chronic diseases (diabetes, Crohn's disease, rheumatoid arthritis). It arises predominantly from productivity loss, and is predicted by decreased quality of life.


Chronic illness and wellbeing: using nursing practice to foster resillence as resistance.
Chronic non-communicable diseases such as heart disease, cancer and diabetes are the biggest killers worldwide. Chronic conditions include heart disease, stroke, diabetes, cancer, chronic respiratory disease and are often comorbid with mental illness. Over 60 years ago, the British Medical Journal reported an association between mental illness and poor physical health (Philips, 1934). Comorbid mental illness and physical illness incrementally worsens health compared with mental illness alone or any of the chronic non-communicable diseases such as cardiovascular diseases, cancers, chronic respiratory diseases and diabetes alone. This paper aims to open the dialogue related to optimising, through nursing intervention, a patient's self-righting and self-management factors in the context of comorbid chronic conditions such as heart disease, stroke, diabetes, cancer, chronic respiratory disease, with mental illness. Self-management and self-righting capabilities are now being considered integral to reducing the negative impact of chronic conditions such as mental illness. Personal characteristics associated with resilience comprise optimism, an active or adaptable coping style and the ability to elicit social support. Existing resilience factors can be assessed for by nurses and optimised through interventions when patients with chronic conditions are in care. Representing over 70% of the global health workforce, nurses are well positioned to enact such practice enhancements to facilitate better outcomes for patients.


Action biases perceptual decisions toward expected outcomes.
We predict how our actions will influence the world around us. Prevailing models in the action control literature propose that we use these predictions to suppress or "cancel" perception of expected action outcomes, to highlight more informative surprising events. However, contrasting normative Bayesian models in sensory cognition suggest that we are more, not less, likely to perceive what we expect-given that what we expect is more likely to occur. Here we adjudicated between these models by investigating how expectations influence perceptual decisions about action outcomes in a signal detection paradigm. Across three experiments, participants performed one of two manual actions that were sometimes accompanied by brief presentation of expected or unexpected visual outcomes. Contrary to dominant cancellation models but consistent with Bayesian accounts, we found that observers were biased to report the presence of expected action outcomes. There were no effects of expectation on sensitivity. Computational modeling revealed that the action-induced bias reflected a sensory bias in how evidence was accumulated rather than a baseline shift in decision circuits. Expectation effects remained in Experiments 2 and 3 when orthogonal cues indicated which finger was more likely to be probed (i.e. task-relevant). These biases toward perceiving expected action outcomes are suggestive of a mechanism that would enable generation of largely veridical representations of our actions and their consequences in an inherently uncertain sensory world. (PsycInfo Database Record (c) 2021 APA, all rights reserved).


Long-Term Access to Sit-Stand Workstations in a Large Office Population: User Profiles Reveal Differences in Sitting Time and Perceptions.
Background: To decrease the detrimental health effects of prolonged sitting, the implementation of sit-stand workstations is a commonly used intervention for office workers. Most studies on this topic evaluated the effects of newly introduced sit-stand workstations. The objective of this study was to determine how often and how long the standing option is used and how the use of sit-stand workstations is perceived in office workers with long-term access to these workstations. Methods: Using an online survey, 1098 office employees responded to questions about frequency of usage of the sit-stand workstation, sitting time, physical activity, and positive and negative perceptions of the use of the sit-stand workstations. Results: Based on the frequency of use, three user groups were identified: non-users (32.1%), monthly/weekly users (37.5%) and daily users (30.4%). Non-users reported to sit more, stand less and have longer bouts of sitting, compared to monthly/weekly users, and these differences were even larger compared to daily users. A higher proportion of daily users perceived the use of the sit-stand workstation as being more healthy and appealing and making them more productive and energetic compared to the non-users. A higher proportion of the non-users perceived it as being uncomfortable, distracting, and unpractical, compared to the other user groups. Conclusions: The differences between the three identified user groups with respect to sitting, standing and perceptions of sit-stand workstations, might be helpful in tailoring future interventions to reduce occupational sitting time, to increase the reach, effectiveness and sustainability.


Evaluation of the adequacy of levothyroxine replacement therapy in patients with central hypothyroidism.
As there are few data on the evaluation of the adequacy of levothyroxine (L-T4) therapy in patients with central hypothyroidism (CH), a prospective study was performed to assess the accuracy of various parameters in the follow-up of 37 CH patients. Total and free thyroid hormones, TSH, and a series of clinical and biochemical indexes of peripheral thyroid hormone action have been evaluated off and on L-T4 therapy. Samples were taken before the daily administration of L-T4. In all patients off therapy, clinical hypothyroidism and low levels of free T4 (FT4) were observed, whereas values of FT3, total T4, and total T3 were below the normal range in 73%, 57%, and 19% of cases, respectively. Most of the indexes of thyroid hormone action were significantly modified after L-T4 withdrawal and exhibited significant correlation with free thyroid hormone levels. During L-T4 replacement therapy, 32 patients had circulating levels of FT4 and FT3 and indexes within the normal range with a mean L-T4 daily dose of 1.5 +/- 0.3 microg/kg BW. Despite normal serum FT4, 3 patients had borderline high values of FT3 and a clear elevation of serum-soluble interleukin-2 receptor concentrations, suggesting overtreatment. Low or borderline low FT4/FT3 levels indicated undertreatment in 2 patients. The clinical parameters lack the required specificity for the diagnosis or follow-up of CH patients. The L-T4 daily dose should be established, taking into account the weight, the age, and the presence of other hormone deficiencies or pharmacological treatment of CH patients. In conclusion, our results indicate that the diagnosis of CH is reached at best by measuring TSH and FT4 concentrations. In the evaluation of the adequacy of L-T4 replacement therapy, both FT4 and FT3 serum levels together with some biochemical indexes of thyroid hormone action are all necessary to a more accurate disclosure of over- or undertreated patients.


Evolutionary Timeline of Genetic Delivery and Gene Therapy.
There are more than 3,500 genes that are being linked to hereditary diseases or correlated with an elevated risk of certain illnesses. As an alternative to conventional treatments with small molecule drugs, gene therapy has arisen as an effective treatment with the potential to not just alleviate disease conditions but also cure them completely. In order for these treatment regimens to work, genes or editing tools intended to correct diseased genetic material must be efficiently delivered to target sites. There have been many techniques developed to achieve such a goal. In this article, we systematically review a variety of gene delivery and therapy methods that include physical methods, chemical and biochemical methods, viral methods, and genome editing. We discuss their historical discovery, mechanisms, advantages, limitations, safety, and perspectives.


Stage IV rectal cancer with liver metastases: is there a benefit to resection of the primary tumor?
Resection of primary and liver lesions is the optimal management of Stage IV rectal cancer with liver metastases. For patients with extensive liver metastases, FOLFOX and FOLFIRI have improved resection rates and survival. We compared survival outcomes in patients with Stage IV rectal cancer with liver metastases undergoing staged or synchronous resection with those undergoing primary rectal resection only or no resection at all. Patients with metastatic rectal cancer to liver were identified from a colorectal cancer database from 2002 to 2008. Patients received neoadjuvant chemoradiation and adjuvant FOLFOX or FOLFIRI therapy. The outcomes for patients who underwent synchronous resection, staged resection, resection of rectal tumor only, and no resection with chemotherapy only were compared. Statistical analysis was determined by ANOVA. Survival was determined using the Kaplan-Meier method. Seventy-four patients were identified: 30 synchronous resections, 13 staged resections, 22 primary resection only, and 9 no resection. Median follow-up was 23 months (range = 4-58 months). Sixty-five percent of patients underwent liver resection with 26% rendered eligible for resection after adjuvant therapy. Those who underwent primary resection only had shorter median survival than those who underwent either staged or synchronous liver resection (31 vs. 47 vs. 46 months, respectively; P = 0.17). Survival was no different for synchronous versus staged resection (P = 0.6). Volume of liver disease predicted resectability (P = 0.001). Without liver resection, 2-year survival was approximately 60%. Palliative surgery was required in six of nine patients who did not undergo resection of their primary tumor. Current chemotherapeutic regimens lead to improved survival in patients with unresectable liver metastases. Upfront chemotherapy in the asymptomatic patient compared with resection of the primary tumor does not appear to significantly affect survival. However, given that 60% of patients were alive after 2 years, resection of the primary lesion for palliative reasons and local control must be considered.


Coupled plasma filtration adsorption: rationale, technical development and early clinical experience.
The adjuvant treatment of sepsis remains a major therapeutic challenge. Blood purification is theoretically appealing if the humoral theory of sepsis is accepted as the basis for intervention. In this setting, blood purification would provide a broad-based restoration of humoral homeostasis thereby avoiding both excessive inflammation and counterinflammation. Several techniques of blood purification have been tried or are under active investigation. One of these is the so-called coupled plasma filtration adsorption (CPFA). CPFA is a novel extracorporeal blood purification therapy aimed at nonselectively reducing the circulating levels and activities of both pro- and anti-inflammatory mediators during sepsis and multiorgan failure. In vitro studies have shown CPFA to be effective in binding a broad range of such mediators proving its technical efficacy. Subsequent animal models have shown a beneficial effect on survival in endotoxemia. These studies have provided the necessary technical developments and biologic rationale for initial human studies. Two phase I/IIa clinical studies have now been performed. Both studies have shown that CPFA improves blood pressure and restores immune function in patients with severe sepsis and multiorgan dysfunction. In this article, we will discuss some of the basic principles involved in sorbent technology, and how these may contribute to treatment efficacy, review animal experiments with CPFA and finally discuss the results of recent human studies and their implications.


Safety Net Care and Midlevel Dental Practitioners: A Case Study of the Portion of Care That Might Be Performed Under Various Setting and Scope-of-Practice Assumptions.
We sought to determine the proportion of dental care provided at safety net-type clinics that might be performed by midlevel practitioners. Data were obtained on 157,328 procedures performed in 2012 at the clinics associated with a Midwestern dental school. Based on procedure codes, we determined the overall proportion, as well as the proportion of visits and patients' care, that could have been performed by 3 types of practitioners. Overall, 48% to 66% of all procedures could have been performed by a midlevel dental practitioner. Nearly half of all visits, and roughly a third of all patients, could have been entirely cared for by a practitioner trained in prophylaxis and with evaluation capabilities. Such practitioners could handle roughly 80% of the visits at the community-based clinic and more than half of the visits at the hospital-based clinic. A midlevel practitioner with training in prophylaxis has the potential to alleviate much of the burden on the dental safety net because much of the need among vulnerable populations falls well within their scope of practice.


Percutaneous Closure Device for the Carotid artery: An integrated review and design analysis.
Endovascular thrombectomies (EVTs) are the current standard of care therapy for treating acute ischemic strokes. While access through the femoral or radial arteries is routine, up to 20% of EVTs through these sites are unable to access the cerebral vasculature on the first pass. These shortcomings are commonly due to tortuous vasculature, atherosclerotic arteries, and type III aortic arch, seen especially in the elderly population. Recent studies have shown the benefits of accessing the cerebral vasculature through a percutaneous direct carotid puncture (DCP), which can reduce the time of the procedure by half. However, current vascular closure devices (VCDs) designed for the femoral artery are not suited to close the carotid artery due to the anatomical differences. This unmet clinical need further limits a DCP approach. Thus, to foster safe adoption of this potential approach, a VCD designed specifically for the carotid artery is needed. In this review, we outline the major biomechanical properties and shortcomings of current VCDs and propose the requirements necessary to effectively design and develop a carotid closure device.


Reporting normal endometrial cells in Pap smears: an outcome appraisal.
The purpose of this study was to determine the clinical relevance of reporting the presence of normal endometrial cells in the Pap smears of women over the age of 35 years and the significance of this practice as it relates to patient management. From January 1992 to December 1995, normal endometrial cells were reported in 206 consecutive Pap smears of women over the age of 35 years. Clinical follow-up was available for all patients, including the results of diagnostic procedures whenever performed. Of the 206 women with normal endometrial cells in their Pap smears, 162 presented with the chief complaint of abnormal vaginal bleeding. They were all evaluated by direct endometrial sampling, resulting in detection of 10 endometrial hyperplasias and 7 endometrial carcinomas. The remaining 44 women who were clinically asymptomatic were followed up with only routine annual gynecologic examinations for a minimum of 3 years. All had negative clinical courses. Reporting the presence of normal endometrial cells in Pap smears has little, if any, impact on subsequent patient management. Women who present with abnormal uterine bleeding are worked up for endometrial disease regardless of their Pap smear findings. In clinically asymptomatic patients, practitioners may, and in our experience often do, choose to disregard normal endometrial cells in Pap smear reports. The negative follow-up for the asymptomatic women in our study supports this practice. Therefore, reporting the presence of normal endometrial cells in Pap smears is of no clinical relevance and may, in fact, create a management dilemma for clinicians.


Interferon-inducible ubiquitin E2, Ubc8, is a conjugating enzyme for protein ISGylation.
Protein ISGylation is unique among ubiquitin-like conjugation systems in that the expression and conjugation processes are induced by specific stimuli, mainly via the alpha/beta interferon signaling pathway. It has been suggested that protein ISGylation plays a special role in the immune response, because of its interferon-signal dependency and its appearance only in higher eukaryotic organisms. Here, we report the identification of an ISG15-conjugating enzyme, Ubc8. Like other components of the protein ISGylation system (ISG15, UBE1L, and UBP43), Ubc8 is an interferon-inducible protein. Ubc8 clearly mediates protein ISGylation in transfection assays. The reduction of Ubc8 expression by small interfering RNA causes a decrease in protein ISGylation in HeLa cells upon interferon treatment. Neither UbcH7/UbcM4, the closest homologue of Ubc8 among known ubiquitin E2s, nor the small ubiquitin-like modifier E2 Ubc9 supports protein ISGylation. These findings strongly suggest that Ubc8 is a major ISG15-conjugating enzyme responsible for protein ISGylation upon interferon stimulation. Furthermore, we established an assay system to detect ISGylated target proteins by cotransfection of ISG15, UBE1L, and Ubc8 together with a target protein to be analyzed. This method provides an easy and effective way to identify new targets for the ISGylation system and will facilitate related studies.


[Effects of the nutritional status of patients submitted to elective surgery].
Between march 5, 1987 and september 10, 1987, 30 patients were operated in elective way of the alimentary tract. They were evaluated to determine the nutritional status and their relation to the postoperative morbidity-mortality. The variables evaluated were weight, height, arm circumference, triceps skinfold thicknesses and serum albumin, lymphocytes and transferrin. 60% of those patients had a normal nutrition, and didn't have morbidity-mortality. 30% had light malnutrition; in this group there was 1 demise and 33% morbidity. 33% had mild malnutrition and 6.7% (2 patients) severe malnutrition. Both patients died because the postoperative complications. The most useful predictive variables were albumin and lymphocytes.


Burst kinetics of single calcium-activated potassium channels in cultured rat muscle.
Burst kinetics of single Ca-activated K channels in excised patches of surface membrane from cultured rat muscle were studied using the patch-clamp technique. Channel activity was separated into bursts using a calculated gap derived from the distribution of shut intervals. Shut intervals greater than the calculated gap were taken as gaps between bursts. The distribution of burst duration was described as the sum of two exponentials with mean durations of about 0.8 and 24 msec (1 microM-Cai, + 20 mV), suggesting two classes of bursts (short and long). The composition of short and long bursts was determined from comparisons of the distributions of open intervals, unit bursts (bursts of single openings), and openings/burst. Short bursts consisted mainly of single openings to the open channel state of short mean lifetime. Long bursts consisted of one or more openings to the (compound) open-channel state of long mean lifetime, plus, in fewer than 70% of the long bursts, one or more openings to the short open-channel state. The frequency of occurrence of bursts from each class first increased and then decreased with increasing [Ca]i, with the number of long bursts increasing at a greater rate than the number of short bursts. The number of openings/short burst was relatively independent of [Ca]i, while the number of openings/long burst increased, often more than linearly, with increasing [Ca]i. This increase arose almost entirely from an increase in openings to the long open state. These results suggest that openings to the long open state typically require the binding of three or more Ca ions, and openings to the short open state typically require the binding of at least one Ca ion. This is the case whether the openings occur in isolation as bursts of single openings or in bursts composed of both types of openings. An obvious burst of channel activity would occur when the channel opens and closes several times without losing all its bound Ca. The power relationship between [Ca]i and the percentage of time spent in the open state is accounted for in terms of the effects of [Ca]i upon mean channel open time, openings/burst, and burst rate. A model is presented that describes quantitatively many features of the burst kinetics of the Ca-activated K channel for constant [Ca]i.


Randomized phase II trial of sunitinib on an intermittent versus continuous dosing schedule as first-line therapy for advanced renal cell carcinoma.
Sunitinib has shown antitumor activity with a manageable safety profile as metastatic renal cell carcinoma (RCC) treatment, when given by the standard intermittent schedule as well as a continuous daily dosing (CDD) schedule. A trial was conducted to compare the schedules. Patients with treatment-naive, clear cell advanced RCC were randomly assigned 1:1 to receive sunitinib 50 mg/d for 4 weeks followed by 2 weeks off treatment (schedule 4/2; n = 146) or 37.5 mg/d on the CDD schedule (n = 146) for up to 2 years. The primary end point was time to tumor progression. Median time to tumor progression was 9.9 months for schedule 4/2 and 7.1 months for the CDD schedule (hazard ratio, 0.77; 95% CI, 0.57 to 1.04; P = .090). No significant difference was observed in overall survival (23.1 v 23.5 months; P = .615), commonly reported adverse events, or patient-reported kidney cancer symptoms. Schedule 4/2 was statistically superior to CDD in time to deterioration, a composite end point of death, progression, and disease-related symptoms (P = .034). CONCLUSION; There was no benefit in efficacy or safety for continuous dosing of sunitinib compared with the approved 50 mg/d dose on schedule 4/2. Given the numerically longer time to tumor progression with the approved 50 mg/d dose on schedule 4/2, adherence to this dose and schedule remains the treatment goal for patients with advanced RCC.


Resorbable Plates Prevent Regression in Pediatric Mandibular Distraction Osteogenesis.
Mandibular distraction osteogenesis is an important technique to correct the pediatric hypoplastic mandible. Regression of the mandible after distraction continues to be a challenge, with some studies reporting up to 22% to 56% decrease toward predistraction length in the first year. We hypothesize that a resorbable plate placed after removal of the distractor will provide additional stability to newly formed bone, allowing further time for consolidation and minimizing regression. This is a retrospective review of neonatal and pediatric patients between 2010 and 2015 who had mandibular distraction osteogenesis performed by the senior author (D.J.). Five patients, including 1 control and 4 intervention subjects, who had preoperative, postdistraction, and follow-up imaging were evaluated. Mandibular distraction was performed via internal devices (KLS Martin, Jacksonville, Fla) and followed by a consolidation period of 6 to 8 weeks. At the second operation when the distractors were removed, the intervention group had a SonicWeld Resorbx (KLS Martin) resorbable plate placed across the bone regenerate. A digital imaging and communications in medicine viewer was used to create a 3-dimensional reconstruction of each of the computed tomography scans and measurements of the mandible were made in the lateral cephalometric view. We then determined distraction distance, regression distance and percent regression. The average mandible distraction distance was 8.2 mm for the control group, and 9.9 mm for the intervention group (range, 6.7-12 mm). The average distance that the mandible regressed after distraction (regression distance) was 2.7 mm for the control group, and 0.5 mm for the intervention group (range, 0.2-0.9 mm). The average percent regression was 32.9% for the control group, and 5.1% for the intervention group (range 1.8 to 7.5%). The results of this study suggest that resorbable plate placement after removal of distractors helps prevent regression in pediatric mandibular distraction osteogenesis. The improvement was readily apparent both radiographically and clinically.


[Use of combined lidocaine and bupivacaine spinal-epidural anesthesia in vascular surgery of lower limbs].
The efficacy of combined spinal epidural anesthesia (CSEA) for femoral to distal artery bypass surgery is assessed. Thirty-eight patients were divided into 3 groups. CSE block was performed at L3-L4 (26G pensil-point spinal needle and 18G catheter). In group I (n = 14) blocking was induced with 20 mg of 1% lidocaine and maintained with 2% lidocaine through an epidural catheter. Group II (n = 15) received spinal plain 20 mg of 0.5% bupivacaine. In group III (n = 9) the initial dose of plain 0.5% bupivacaine was 3 ml (15 mg); 5 min after the first bolus the incremental dose of plain bupivacaine 3 ml (15 mg) was injected and spinal needle was withdrawn. Epidural 0.5% bupivacaine was injected as needed. Sensory blockade was assessed by the pinprick test. Two patients in group I (18.2%) were in need of general anesthesia (GA) (inadequate dissemination of solution in 1 case and catheter kinking in the other). In group II GA had to be performed in 1 case because surgery was longer than planned. In group III anesthesia was effective in all cases. The mean level of sensory block in group I was T11.2 +/- 0.4, in group II T9.4 +/- 0.5, and in group III T8.6 +/- 0.55 (p > 0.05). The mean dose of bupivacaine per segment in groups II and III was 1.47 +/- 0.08 and 2.08 +/- 0.01, respectively (p < 0.05), use of epidural catheter during surgery 40 and 11%, respectively (p < 0.05). Arterial pressure drop was 10.8, 14.1, and 11.6% in groups I, II, and III, respectively. The mean total dose was 735 +/- 89 mg (172 +/- 14 mg/h) in the lidocaine group, in group II 38.5 +/- 6.4 mg (10.25 +/- 1.15 mg/h), and in group III 32.8 +/- 2.8 mg (7.46 +/- 1.67 mg/h) (p < 0.05). No neurological problems or PDPH were observed in any of the patients and no vasoactive drugs were needed. CSEA is fit for anesthesia for peripheral vascular surgery. CSEA with double spinal injection is preferable, for it provides a longer anesthesia and hemodynamic stability.


Understanding the influences on successful quality improvement in emergency general surgery: learning from the RCS Chole-QuIC project.
Acute gallstone disease is the highest volume Emergency General Surgical presentation in the UK. Recent data indicate wide variations in the quality of care provided across the country, with national guidance for care delivery not implemented in most UK hospitals. Against this backdrop, the Royal College of Surgeons of England set up a 13-hospital quality improvement collaborative (Chole-QuIC) to support clinical teams to reduce time to surgery for patients with acute gallstone disease requiring emergency cholecystectomy. Prospective, mixed-methods process evaluation to answer the following: (1) how was the collaborative delivered by the faculty and received, understood and enacted by the participants; (2) what influenced teams' ability to improve care for patients requiring emergency cholecystectomy? We collected and analysed a range of data including field notes, ethnographic observations of meetings, and project documentation. Analysis was based on the framework approach, informed by Normalisation Process Theory, and involved the creation of comparative case studies based on hospital performance during the project. Chole-QuIC was delivered as planned and was well received and understood by participants. Four hospitals were identified as highly successful, based upon a substantial increase in the number of patients having surgery in line with national guidance. Conversely, four hospitals were identified as challenged, achieving no significant improvement. The comparative analysis indicate that six inter-related influences appeared most associated with improvement: (1) achieving clarity of purpose amongst site leads and key stakeholders; (2) capacity to lead and effective project support; (3) ideas to action; (4) learning from own and others' experience; (5) creating additional capacity to do emergency cholecystectomies; and (6) coordinating/managing the patient pathway. Collaborative-based quality improvement is a viable strategy for emergency surgery but success requires the deployment of effective clinical strategies in conjunction with improvement strategies. In particular, achieving clarity of purpose about proposed changes amongst key stakeholders was a vital precursor to improvement, enabling the creation of additional surgical capacity and new pathways to be implemented effectively. Protected time, testing ideas, and the ability to learn quickly from data and experience were associated with greater impact within this cohort.


Cell cycle accumulation of the proliferating cell nuclear antigen PCN-1 transitions from continuous in the adult germline to intermittent in the early embryo of C. elegans.
The proliferating cell nuclear antigen (PCNA or PCN-1 in C. elegans), an essential processivity factor for DNA polymerase δ, has been widely used as a marker of S-phase. In C. elegans early embryos, PCN-1 accumulation is cyclic, localizing to the nucleus during S-phase and the cytoplasm during the rest of the cell cycle. The C. elegans larval and adult germline is an important model systems for studying cell cycle regulation, and it was observed that the cell cycle regulator cyclin E (CYE-1 in C. elegans) displays a non-cyclic, continuous accumulation pattern in this tissue. The accumulation pattern of PCN-1 has not been well defined in the larval and adult germline, and the objective of this study was to determine if the accumulation pattern is cyclic, as in other cells and organisms, or continuous, similar to cyclin E. To study the larval and adult germline accumulation of PCN-1 expressed from its native locus, we used CRISPR/Cas9 technology to engineer a novel allele of pcn-1 that encodes an epitope-tagged protein. S-phase nuclei were labeled using EdU nucleotide incorporation, and FLAG::PCN-1 was detected by antibody staining. All progenitor zone nuclei, including those that were not in S-phase (as they were negative for EdU staining) showed PCN-1 accumulation, indicating that PCN-1 accumulated during all cell cycle phases in the germline progenitor zone. The same result was observed with a GFP::PCN-1 fusion protein expressed from a transgene. pcn-1 loss-of-function mutations were analyzed, and pcn-1 was necessary for robust fertility and embryonic development. In the C. elegans early embryo as well as other organisms, PCN-1 accumulates in nuclei only during S-phase. By contrast, in the progenitor zone of the germline of C. elegans, PCN-1 accumulated in nuclei during all cell cycle stages. This pattern is similar to accumulation pattern of cyclin E. These observations support the model that mitotic cell cycle regulation in the germline stem and progenitor cells is distinct from somatic cells, as it does not heavily rely on cyclic accumulation of classic cell cycle proteins.


Protein instability and immunogenicity: roadblocks to clinical application of injectable protein delivery systems for sustained release.
Protein instability and immunogenicity are two main roadblocks to the clinical success of novel protein drug delivery systems. In this commentary, we discuss the need for more extensive analytical characterization in relation to concerns about protein instability in injectable drug delivery systems for sustained release. We then will briefly address immunogenicity concerns and outline current best practices for using state-of-the-art analytical assays to monitor protein stability for both conventional and novel therapeutic protein dosage forms. Next, we provide a summary of the stresses on proteins arising during preparation of drug delivery systems and subsequent in vivo release. We note the challenges and difficulties in achieving the absolute requirement of quantitatively assessing the degradation of protein molecules in a drug delivery system. We describe the potential roles for academic research in further improving protein stability and developing new analytical technologies to detect protein degradation byproducts in novel drug delivery systems. Finally, we provide recommendations for the appropriate approaches to formulation design and assay development to ensure that stable, minimally immunogenic formulations of therapeutic proteins are created. These approaches should help to increase the probability that novel drug delivery systems for sustained protein release will become more readily available as effective therapeutic agents to treat and benefit patients.


DNA repair: relationship to drug and radiation resistance, metastasis and growth factors.
DNA repair mechanisms are important for the recovery of both normal and malignant tissues from radiation and chemotherapy. Drug 'resistance' may merely reflect the similarity of cancer to normal tissues. Investigating the normal repair mechanisms by cloning human DNA repair genes will permit a much better comparison. Therapeutic inhibition of DNA repair may be possible with poly-ADP-ribose polymerase inhibitors. A differential effect may be obtained since less-differentiated cells have a higher poly-ADP-ribose polymerase activity. Clinical application of repair inhibitors can be achieved by using antimetabolites such as high-dose hydroxyurea which produces levels of 1-3 mmol litre -1/24 hours. The whole cell and tissue response to DNA damage is more complex than removal of adducts and joining strand breaks. DNA damage can result in an increase in growth-factor receptors, the release of soluble mediators that affect undamaged cells and stimulation of plasminogen activator. These changes may enhance growth and recovery as well as bypass or repair the damage. The generation of heterogeneity in a tumour population may be mediated by DNA rearrangements. Genetic instability is much higher in metastatic clones and a comparison of DNA strand-break repair in a metastatic and a non-metastatic line showed more rapid repair in the former. Aberrant use of DNA repair stimulated by growth factors may mediate tumour progression and heterogeneity as well as drug resistance.


Chronic epi-inositol has an anxiolytic-like effect in the plus-maze model in rats.
Behavioural effects of myo-inositol have been demonstrated in animal models of psychiatric disorders and in clinical trials in psychiatric patients. These effects of myo-inositol were thought to be consequences of its being a substrate for the enzyme phosphatidylinositol (PI) synthase. Recently, epi-inositol, an unnatural stereoisomer of myo-inositol that is not a substrate for PI synthase, was found to also have effects similar to those of myoinositol to reverse lithium-pilocarpine seizures. The present study measured the behaviour of rats in an elevated plus-maze model of anxiety after chronic treatment of 11 daily i.p. injections of epi-inositol, myo-inositol, or control solution. Epi-inositol reduced levels of anxiety-like behaviour in rats compared with controls, and its effect was stronger than that of myo-inositol. The increased relative effect of epi-inositol could be due to slower metabolism or to a different mechanism of action.


Immunochemical properties of the lipopolysaccharide O-antigen of Vibrio cholerae O1 in relation to its chemical structure.
D-Glucuronic acid and D-glucosamine have an immunodominant role in the lipopolysaccharide (LPS) O-antigen of both the Ogawa and the Inaba subtypes of Vibrio cholerae O1. This was evident from the pronounced inhibitory effect on the LPS precipitin reaction demonstrated by these monosaccharides and by oligosaccharides containing either of them which were isolated from LPS hydrolysate. There was a considerable decrease in the antibody-combining capacity of chemically modified LPS in which the carboxyl group of the glucuronic acid had been reduced. Similarly, on deamination, the O-specific polysaccharide fraction of the LPS molecule from both subtypes completely lost the ability to precipitate the LPS antibody.


Autophagy inhibition induces atrophy and myopathy in adult skeletal muscles.
Autophagy is required for cellular survival and for the clearance of damaged proteins and altered organelles. Excessive autophagy activation contributes to muscle loss in different catabolic conditions. However, the function of basal autophagy for homeostasis of skeletal muscle was unknown. To clarify this issue we have generated conditional and inducible knockout mice for the critical gene Atg7, to block autophagy specifically in skeletal muscle. Atg7 null muscles reveal an unexpected phenotype which is characterized by muscle atrophy, weakness and features of myofiber degeneration. Morphological, biochemical and molecular analyses of our autophagy knockout mice show the presence of protein aggregates, abnormal mitochondria, accumulation of membrane bodies, sarcoplasmic reticulum distension, vacuolization, oxidative stress and apoptosis. Moreover, autophagy inhibition does not protect skeletal muscles from atrophy during denervation and fasting, but instead promotes greater muscle loss. In conclusion, autophagy plays a critical role for myofiber maintenance and its activation is crucial to avoid accumulation of toxic proteins and dysfunctional organelles that, in the end, would lead to atrophy and weakness.


Identification of a 80 kDa calmodulin-binding protein as a new Ca2+/calmodulin-dependent kinase by renaturation blotting assay (RBA).
We surveyed rabbit brain cytosol for a new Ca2+/calmodulin (CaM)-dependent kinase. The renaturation blotting assay (RBA) exploits the ability of blotted SDS-denatured proteins to regain enzymic activity after guanidine treatment. Using RBA, we found that the eluate of rabbit brain cytosol from a CaM affinity column contains at least four electrophoretically distinct protein kinase bands which were autophosphorylated in a Ca2+/CaM-dependent manner. The 49 kDa band and the 60 kDa band were alpha and beta subunit of CaM kinase II, and the 42 kDa band was presumed to be CaM kinase I, but the 80 kDa band could not be attributed to any reported Ca2+/CaM-dependent protein kinases. The 80 kDa protein kinase was isolated by three-step chromatography. We examined the phosphorylation of exogenous substrates by 80 kDa protein kinase, and histone IIIs and myosin light chain were phosphorylated in a Ca2+/CaM-dependent manner. W-7, a specific inhibitor for calmodulin, inhibited this kinase activity, but KN-62, a specific inhibitor for CaM kinase II, had no effect on this protein kinase activity. Autoradiography using boiled rabbit brain homogenate as substrate showed three intrinsic substrates (80 kDa, 60 kDa and 42 kDa), which were phosphorylated in a Ca2+/CaM-dependent manner. These findings suggest that a new Ca2+/CaM-dependent protein kinase could be identified by the RBA.


The antisense RNA of the par locus of pAD1 regulates the expression of a 33-amino-acid toxic peptide by an unusual mechanism.
The par stability determinant of the Enterococcus faecalis plasmid pAD1 is the first antisense RNA-regulated post-segregational killing system (PSK) identified in a Gram-positive organism. Par encodes two small, convergently transcribed RNAs, designated RNA I and RNA II, which are the toxin and antidote of the par PSK system respectively. RNA I encodes an open reading frame of 33 codons designated fst. The results presented here demonstrate that the peptide encoded by fst is the par toxin. The fst sequence was shown to be sufficient for cell killing, and removal of the final codon inactivated the toxin. In vitro translation reactions of purified RNA I transcript produced a product of the expected size for the fst-encoded peptide. This product was not produced when purified RNA II transcript was added to the translation reaction. Toeprint analysis demonstrated that purified RNA II was able to inhibit ribosome binding to RNA I. These data suggest that fst expression is regulated by RNA II via an antisense RNA mechanism. In vitro translation studies and toeprint analyses also indicated that fst expression is internally regulated by a stem-loop structure at the 5' end of RNA I. Removal of this structure resulted in better ribosome binding to RNA I and a 300-fold increase in production of the fst-encoded peptide. Finally, RNA II was shown to be less stable than RNA I in vivo, providing a basis for the selective expression of fst in plasmid-free cells.


Evaluation of the S100 protein A12 as a biomarker of neonatal sepsis.
Introduction: Sepsis has a grave impact on neonatal morbidity and mortality. Proper timely diagnosis and a subsequently tailored management are crucial to improving neonatal outcome and survival. New diagnostic methods are needed and much effort is directed to this objective. In this work, we aimed to evaluate S100A12 protein as a biomarker of neonatal sepsis.Materials and methods: In this prospective single-center study, 118 preterm and term neonates were enrolled and assigned to four groups: controls, infants with no infection, infants with probable infection and infants with proven infection. Clinical and routine laboratory data, the serum levels of S100A12 and additional cytokines (interleukin (IL)-1β, IL-2, IL-6, IL-17A, IL-18, IL-22, IL-10, and interferon (IFN)-γ) were assessed. Using stepwise multivariate logistic regression analysis, S100A12 protein was evaluated as a biomarker of neonatal infection.Results: Significant differences of the parameters of complete blood count and level of C-reactive protein were documented between the study/the four groups. The studied marker S100A12, as well as IL-6 and IL-10, were highly significant (p < .001) between infected and control groups. S100A12 had a sensitivity of 96.8% and a specificity of 93.3%. Even after adjusting for the confounding factors sex, body weight, gestational age, mode of delivery, number of pregnancies, premature rupture of membranes, and preeclampsia S100A12 remained significant between the infected and control groups.Conclusions: S100A12 may be considered as a new biomarker of neonatal sepsis.


Viral chemokine-binding proteins.
The chemokines are a large family of small signaling proteins that bind to G-protein-coupled receptors (GPCRs) on target cells and mediate the directional migration of immune cells into sites of infection or inflammation. The large DNA viruses, particularly the poxviruses and herpesviruses, have evolved several mechanisms to corrupt the normal functioning of the chemokine network. Two strategies rely on mimicking chemokines or chemokine receptors. A third strategy involves the production of secreted chemokine-binding proteins (CKBPs) that exhibit no sequence similarity to any known host proteins, yet function to competitively bind and inhibit the interactions of chemokines with cognate receptors. Each strategy has provided unique insights into the elusively complex world of the chemokines. Here, we focus on recent advances made in the understanding of secreted CKBPs encoded by poxviruses and herpesviruses. A better understanding of how viral CKBPs function to manipulate the immune response may provide further clues as to how to develop specific therapeutic agents to abrogate chemokine-mediated disease conditions.


A pigmented mycelial antibiotic in Streptomyces coelicolor: control by a chromosomal gene cluster.
Streptomyces coelicolor was found to produce a third secondary metabolite, in addition to the antibiotics methylenomycin A and actinorhodin previously described. This is a red pigmented, highly non-polar compound with antibiotic activity against certain Gram-positive bacteria. Mutants lacking the red compound fell into five cosynthetic classes. Representatives of each of the five classes were mapped to the chromosome of the producing organism, in a closely linked cluster. Genetic studies provided evidence that this new metabolite is distinct from actinorhodin and indicated that the two pigments do not share parts of the same biosynthetic pathway.


Impact of coeliac disease on dietary habits and quality of life.
Coeliac disease (CD) affects approximately 1% of the population in the UK and is managed by the life-long adherence to a strict gluten-free diet (GFD). Adhering to a GFD is practically difficult and not only affects dietary patterns, but also can affect many other aspects of daily life. The present study aimed to investigate the effect of CD and a GFD on dietary habits and quality of life of a cohort of adult biopsy diagnosed coeliac patients who reside in England. The cohort was composed of 146 adult biopsy-diagnosed CD patients, who were all members of the Coeliac UK charity. Participants responded to a self-administered questionnaire considering dietary habits and quality of life. A food frequency questionnaire (FFQ) was used to assess dietary compliance. Generally, English CD patients reported to be in good physical and emotional health, although there were reports of anxiety and depression as a result of CD, most likely as a result of exclusion from social and leisure activities. The cohort reported high levels of dietary compliance (96%) which was supported by FFQ responses. However, there were reports of intentional gluten intake during social situations and when eating take-away foods. The FFQ revealed further examples of gluten ingestion, presumably unintentional, particularly through the consumption of breakfast cereals and starch-based sauces such as cheese sauce, custard and ketchup. The present study revealed that CD affects a wide range of daily activities and that gluten consumption may be more common than anticipated with possible consequences on health.


VATS anatomic lung resections-the European experience.
Video-assisted thoracoscopic surgery (VATS) has undergone significant evolution over several decades. Although endoscopic instruments continued to improve, it was not until 1992 that the first VATS lobectomy for lung cancer was performed. Despite significant seeding of such procedure in several thoracic units globally, the uptake was slow and frustrating. Many surgeons considered it complex and unsafe being skeptic about its oncological validity. The last decade has witnessed significant change of practice in many thoracic units with a new generation of VATS thoracic surgeons. Additionally the technique has been refined, standardized and proved its validity and superiority in lung cancer treatment.


Corticotropin-releasing hormone during pregnancy is associated with infant temperament.
During pregnancy corticotropin-releasing hormone (CRH) is released into maternal and fetal circulation from the placenta. Elevated concentrations of placental CRH are associated with spontaneous preterm birth, but the consequences for infant development, independent of birth outcome, are unknown. In this study, the effects of placental CRH on infant temperament were examined in a sample of 248 full-term infants. Maternal blood samples were collected at 19, 25 and 31 weeks of gestation for CRH analysis. Infant temperament was assessed with measures of fear and distress at 2 months of age. Infants of mothers with low CRH at 25 weeks of gestation scored lower in fear and distress at 2 months. CRH at 19 and 31 weeks' gestation was not significantly associated with measures of infant temperament, suggesting the possibility that there is a sensitive period for its effects. These data suggest that prenatal exposure to CRH may exert influences that persist into the postnatal period.


Eosinophilic pneumonia and atypical mycobacterial infection.
Chronic eosinophilic pneumonia is a relatively uncommon condition associated with fever, malaise, weight loss, dyspnea, and hypoxia. The chest radiograph displays peripheral air-space consolidation, often referred to as the negative of pulmonary edema. The characteristic pathologic findings in the lung include marked accumulation of eosinophils in the alveolar spaces, often with the formation of eosinophilic abscesses. We report the history and pathologic findings in a patient who fit the clinical, radiologic, and pathologic features of chronic eosinophilic pneumonia, but who went on to develop cavitary atypical mycobacterial infection and died. We believe that this case represents an unusual response to an atypical mycobacterial infection.


[Bronchopulmonary dysplasia in newborns]].
Published data on the modern state-of-art of bronchopulmonary dysplasia in newborns were analyzed as a complication of prolonged artificial pulmonary ventilation (APV). Agents that can affect an underformed lung in an immature newborn and which can trigger a chronic pathology yet on the 1st month life are under discussion. Modern concepts related with the development of bronchopulmonary dysphasia in newborns after a prolonged APV as well as the key issues of treatment and prevention of the mentioned disease are in the focus of attention.


Macrophage activation syndrome in patients with adult-onset Still's disease under tocilizumab treatment: A single-center observational study.
Macrophage activation syndrome (MAS) developed under tocilizumab treatment poses a diagnostic challenge. This study aims to demonstrate the frequency and the clinical features of MAS developed in patients with adult-onset Still's disease (AOSD) receiving tocilizumab. The consecutive AOSD patients treated with tocilizumab in our institution from April 2008 to March 2020 were studied. The frequency of clinically diagnosed MAS during tocilizumab treatment, their conformity to the several criteria relevant for MAS, and laboratory characteristics compared to AOSD flare were investigated. Of the 20 AOSD patients treated with tocilizumab, six developed clinically diagnosed MAS, four immediately after starting tocilizumab and two after long-term treatment. Some of them had already met the MAS criteria before starting tocilizumab. At MAS diagnosis, although some did not meet the MAS criteria due to lack of fever and/or the lower ferritin levels, all consistently showed sharp increases in ferritin along with marked abnormal changes in two or more different markers of organ damage, unlike the AOSD flares. MAS is not a rare complication in AOSD patients receiving tocilizumab. The clinical similarities between systemic AOSD and MAS, and substantial alterations in MAS features by inhibition of interleukin-6 signaling may limit the utility of the existing diagnostic/classification criteria in diagnosing MAS under tocilizumab treatment. The emergence of abnormalities in MAS-related organ damage markers with a rapid elevation of ferritin should be considered as MAS development in AOSD patients receiving tocilizumab even if the patients are afebrile or have relatively low ferritin levels.


The Evolutionary Arms Race between Virus and NK Cells: Diversity Enables Population-Level Virus Control.
Viruses and natural killer (NK) cells have a long co-evolutionary history, evidenced by patterns of specific NK gene frequencies in those susceptible or resistant to infections. The killer immunoglobulin-like receptors (KIR) and their human leukocyte antigen (HLA) ligands together form the most polymorphic receptor-ligand partnership in the human genome and govern the process of NK cell education. The KIR and HLA genes segregate independently, thus creating an array of reactive potentials within and between the NK cell repertoires of individuals. In this review, we discuss the interplay between NK cell education and adaptation with virus infection, with a special focus on three viruses for which the NK cell response is often studied: human immunodeficiency virus (HIV), hepatitis C virus (HCV) and human cytomegalovirus (HCMV). Through this lens, we highlight the complex co-evolution of viruses and NK cells, and their impact on viral control.


Machine learning techniques and drug design.
The interest in the application of machine learning techniques (MLT) as drug design tools is growing in the last decades. The reason for this is related to the fact that the drug design is very complex and requires the use of hybrid techniques. A brief review of some MLT such as self-organizing maps, multilayer perceptron, bayesian neural networks, counter-propagation neural network and support vector machines is described in this paper. A comparison between the performance of the described methods and some classical statistical methods (such as partial least squares and multiple linear regression) shows that MLT have significant advantages. Nowadays, the number of studies in medicinal chemistry that employ these techniques has considerably increased, in particular the use of support vector machines. The state of the art and the future trends of MLT applications encompass the use of these techniques to construct more reliable QSAR models. The models obtained from MLT can be used in virtual screening studies as well as filters to develop/discovery new chemicals. An important challenge in the drug design field is the prediction of pharmacokinetic and toxicity properties, which can avoid failures in the clinical phases. Therefore, this review provides a critical point of view on the main MLT and shows their potential ability as a valuable tool in drug design.


Isolation and characterization of exopolysaccharide with immunomodulatory activity from fermentation broth of Morchella conica.
BACKGROUND AND THE PURPOSE OF THIS STUDY: Mushroom polysaccharides have traditionally been used for the prevention and treatment of a multitude of disorders like infectious illnesses, cancers and various autoimmune diseases. In vitro and in vivo studies suggest that certain polysaccharides affect immune system function. Morchella conica (M. conica) is a species of rare edible mushroom whose multiple medicinal functions have been proven. Thus, the objective of this study is to isolate and characterize of exopolysaccharide from submerged mycelial culture of M. conica, and to evaluate its immunomodulatory activity. A water-soluble Morchella conica Polysaccharides (MCP) were extracted and isolated from the fermentation broth of M. conica through a combination of DEAE-cellulose and Sephacryl S-300 HR chromatograph. NMR and IR spectroscopy has played a developing role in identification of polysaccharide with different structure and composition from fungal and plant sources, as well as complex glycosaminoglycans of animal origin. Thus, NMR and IR spectroscopy were used to analyze the chemical structure and composition of the isolated polysaccharide. Moreover, the polysaccharide was tested for its immunomodulatory activity at different concentrations using in vitro model. The results showed that MCP may significantly modulate nitric oxide production in macrophages, and promote splenocytes proliferation. Analysis from HPLC, infrared spectra and nuclear magnetic resonance spectroscopy showed that MCP was a homogeneous mannan with an average molecular weight of approximately 81.2 kDa. The glycosidic bond links is →6)-α-D-Man p-(1→. The results suggested that the extracted MCP may modulate nitric oxide production in macrophages and promote splenocytes proliferation, and it may act as a potent immunomodulatory agent.


Indomethacin suppression of the estradiol-induced proliferative response of the seminal vesicles.
Effects of indomethacin (1.25 mg/kg of body weight and 1.25 mg/rat) on estradiol-induced (250 micrograms/rat and 25 micrograms/rat) mitotic activity of the seminal vesicles and the ventral prostate in rats were examined. The obtained results show clearly that indomethacin restrains the seminal vesicles proliferative reaction induced by estradiol, whereas the ventral prostate is not sensitive to estradiol, and indomethacin does not affect its mitotic activity. These results suggest that prostaglandins are involved in the mitogenic effect of estradiol on the seminal vesicles.