Small interfering RNA: Discovery, pharmacology and clinical development-An introductory review.
Post-transcriptional gene silencing targets and degrades mRNA transcripts, silencing the expression of specific genes. RNA interference technology, using synthetic structurally well-defined short double-stranded RNA (small interfering RNA [siRNA]), has advanced rapidly in recent years. This introductory review describes the utility of siRNA, by exploring the underpinning biology, pharmacology, recent advances and clinical developments, alongside potential limitations and ongoing challenges. Mediated by the RNA-induced silencing complex, siRNAs bind to specific complementary mRNAs, which are subsequently degraded. siRNA therapy offers advantages over other therapeutic approaches, including ability of specifically designed siRNAs to potentially target any mRNA and improved patient adherence through infrequent administration associated with a very long duration of action. Key pharmacokinetic and pharmacodynamic challenges include targeted administration, poor tissue penetration, nuclease inactivation, rapid renal elimination, immune activation and off-target effects. These have been overcome by chemical modification of siRNA and/or by utilising a range of delivery systems, increasing bioavailability and stability to allow successful clinical translation. Patisiran (hereditary transthyretin-mediated amyloidosis) was the first licensed siRNA, followed by givosiran (acute hepatic porphyria), lumasiran (primary hyperoxaluria type 1) and inclisiran (familial hypercholesterolaemia), which all use N-acetylgalactosamine (GalNAc) linkage for effective liver-directed delivery. Others are currently under development for indications varying from rare genetic diseases to common chronic non-communicable diseases (hypertension, cancer). Technological advances are paving the way for broader clinical use. Ongoing challenges remain in targeting organs beyond the liver and reaching special sites (e.g., brain). By overcoming these barriers, siRNA therapy has the potential to substantially widen its therapeutic impact.


Identification of miRNAs differentially expressed in human epilepsy with or without granule cell pathology.
The microRNAs (miRNAs) are small size non-coding RNAs that regulate expression of target mRNAs at post-transcriptional level. miRNAs differentially expressed under pathological conditions may help identifying mechanisms underlying the disease and may represent biomarkers with prognostic value. However, this kind of studies are difficult in the brain because of the cellular heterogeneity of the tissue and of the limited access to fresh tissue. Here, we focused on a pathology affecting specific cells in a subpopulation of epileptic brains (hippocampal granule cells), an approach that bypasses the above problems. All patients underwent surgery for intractable temporal lobe epilepsy and had hippocampal sclerosis associated with no granule cell pathology in half of the cases and with type-2 granule cell pathology (granule cell layer dispersion or bilamination) in the other half. The expression of more than 1000 miRNAs was examined in the laser-microdissected dentate granule cell layer. Twelve miRNAs were differentially expressed in the two groups. One of these, miR487a, was confirmed to be expressed at highly differential levels in an extended cohort of patients, using RT-qPCR. Bioinformatics searches and RT-qPCR verification identified ANTXR1 as a possible target of miR487a. ANTXR1 may be directly implicated in granule cell dispersion because it is an adhesion molecule that favors cell spreading. Thus, miR487a could be the first identified element of a miRNA signature that may be useful for prognostic evaluation of post-surgical epilepsy and may drive mechanistic studies leading to the identification of therapeutic targets.


[Conformational aspects of beta-trypsin interactions with substrates and pancreatic trypsin inhibitor. II. Structure of tetrahedral adducts and acylenzyme].
Theoretical conformational analysis of the tetrahedral complexes of trypsin with the N-acetyl-L-lysine methyl amide, which are formed at the acylation and the deacylation stages of the catalytical act has been carried out. The lowest energy conformations are shown to be productive ones. All favorable structures of N-acetyl-L-lysyl-trypsin and N-acetyl-L-arginyl-trypsin acylenzymes have been analysed. The global conformations of both complexes are found to be very similar with the structures providing for a transition to the second tetrahedral state. Conformations of the nonbonded, tetrahedral and acyl complexes with N-acetyl-L-lysine methyl amide are compared and the differences in orientation of atomic groups participating in the catalysis are revealed. All changes of optimal structures of the complexes indispensable for the catalytical process are shown to proceed in a spontaneous way without introduction of any intramolecular strain.


Comparison of 0.25% Ropivacaine for Intraperitoneal Instillation v/s Rectus Sheath Block for Postoperative Pain Relief Following Laparoscopic Cholecystectomy: A Prospective Study.
As Laparoscopic Cholecystectomy (LC) is not a totally pain free procedure, with the pain being most intense on the day of surgery and on the following day. Various techniques are available for postoperative pain relief like intraperitoneal instillation of local anaesthetics and rectus sheath block (RSB)which may provide effective pain relief. To compare the efficacy of preemptive administration (initiated before the surgical procedure) of intraperitoneal instillation and rectus sheath block using ropivacaine for postoperative analgesia after laparoscopic cholecystectomy. A total of 75 selected patients were randomly assigned to three equal groups as Group R, who received bilateral RSB with 0.25 % ropivacaine 15 ml on either side; Group I, who received intraperitoneal instillation of 0.25% ropivacaine 50 ml and Group C (Control group), who received only rescue analgesic on pain. These were compared regarding postoperative analgesia in terms of Visual Analog Scale (0-10 cm), Prince Henry Hospital Pain Score (0-3), time to first dose of rescue analgesic (tramadol), total rescue analgesic consumption in 48 hours, patient satisfaction scores (1-7) and adverse effects. The time to first rescue analgesic was significantly longer in Group R (16.16±4.73h) and Group I (7.84±1.34h) as compared to Group C (1.72±0.67h), p<0.001. Mean tramadol consumption in 48h for each patient was significantly less in Group R (148±54.92mg) and Group I (202±33.78mg) as compared to Group C (298±22.73mg) p<0.001. Postoperative pain scores were also significantly less in Group R and Group I as compared to Group C during first 6 hours, p<0.05. The difference in above parameters was also significant between Group R and Group I, p<0.05. Thus order of postoperative analgesia effect was: Group R > Group I > Group C. Rescue analgesic requirement showed a 32.21% reduction in Group I and 50.33% reduction in Group R as compared to Group C. Patient Satisfaction Scores (PSS) showed a significant difference among groups with acceptable PSS scores as: Group R (92%) v/s Group I (40%) v/s Group C (20%) p<0.001. Pre-emptive administration of rectus sheath block or intraperitoneal instillation of 0.25% ropivacaine was found effective in providing better postoperative analgesia as compared to control group after laparoscopic cholecystectomy. Among these two techniques, rectus sheath block was found to be superior over intraperitoneal instillation.


Effects and side-effects of surgery for snoring and obstructive sleep apnea--a systematic review.
Many patients undergo surgery for snoring and sleep apnea, although the efficacy and safety of such procedures have not been clearly established. Our aim was systematically to review studies of the efficacy and adverse effects of surgery for snoring and obstructive sleep apnea. Systematic review. PubMed and Cochrane databases were searched in September 2007. Randomized controlled trials of surgery vs. sham surgery or conservative treatment in adults, with daytime sleepiness, quality of life, apnea-hypopnea index, and snoring as outcomes were included. Observational studies were also reviewed to assess adverse effects. Evidence of effect required at least two studies of medium and high quality reporting the same result. Four studies of benefits and 45 studies of adverse effects were included. There was no significant effect on daytime sleepiness and quality of life after laser-assisted uvulopalatoplasty and radiofrequency ablation. The apnea-hypopnea index and snoring was reduced in one trial after laser-assisted uvulopalatoplasty but not in another trial. Subjective snoring was reduced in one trial after radiofrequency ablation. No trial investigating the effect of any other surgical modality met the inclusion criteria. Persistent side-effects occurred after uvulopalatopharyngoplasty and uvulopalatoplasty in about half the patients and difficulty in swallowing, globus sensation and voice changes were especially common. Only a small number of randomized controlled trials with a limited number of patients assessing some surgical modalities for snoring or sleep apnea are available. These studies do not provide any evidence of effect from laser-assisted uvulopalatoplasty or radiofrequency ablation on daytime sleepiness, apnea reduction, quality of life or snoring. We call for research of randomized, controlled trials of surgery other than uvulopalatopharyngoplasty and uvulopalatoplasty, as they are related to a high risk of long-term side-effects, especially difficulty swallowing.


Cdk1-mediated phosphorylation of Cdc7 suppresses DNA re-replication.
To maintain genetic stability, the entire mammalian genome must replicate only once per cell cycle. This is largely achieved by strictly regulating the stepwise formation of the pre-replication complex (pre-RC), followed by the activation of individual origins of DNA replication by Cdc7/Dbf4 kinase. However, the mechanism how Cdc7 itself is regulated in the context of cell cycle progression is poorly understood. Here we report that Cdc7 is phosphorylated by a Cdk1-dependent manner during prometaphase on multiple sites, resulting in its dissociation from origins. In contrast, Dbf4 is not removed from origins in prometaphase, nor is it degraded as cells exit mitosis. Our data thus demonstrates that constitutive phosphorylation of Cdc7 at Cdk1 recognition sites, but not the regulation of Dbf4, prevents the initiation of DNA replication in normally cycling cells and under conditions that promote re-replication in G2/M. As cells exit mitosis, PP1α associates with and dephosphorylates Cdc7. Together, our data support a model where Cdc7 (de)phosphorylation is the molecular switch for the activation and inactivation of DNA replication in mitosis, directly connecting Cdc7 and PP1α/Cdk1 to the regulation of once-per-cell cycle DNA replication in mammalian cells.


Role of CHD5 in human cancers: 10 years later.
CHD5 was first identified because of its location on 1p36 in a region of frequent deletion in neuroblastomas. CHD5 (chromodomain-helicase-DNA-binding-5) is the fifth member of a family of chromatin remodeling proteins, and it probably functions by forming a nucleosome remodeling and deacetylation (NuRD) complex that regulates transcription of particular genes. CHD5 is preferentially expressed in the nervous system and testis. On the basis of its position, pattern of expression, and function in neuroblastoma cells and xenografts, CHD5 was identified as a tumor suppressor gene (TSG). Evidence soon emerged that CHD5 also functioned as a TSG in gliomas and a variety of other tumor types, including breast, colon, lung, ovary, and prostate cancers. Although one copy of CHD5 is deleted frequently, inactivating mutations of the remaining allele are rare. However, DNA methylation of the CHD5 promoter is found frequently, and this epigenetic mechanism leads to biallelic inactivation. Furthermore, low CHD5 expression is strongly associated with unfavorable clinical and biologic features as well as outcome in neuroblastomas and many other tumor types. Thus, based on its likely involvement as a TSG in neuroblastomas, gliomas, and many common adult tumors, CHD5 may play an important developmental role in many other tissues besides the nervous system and testis.


Regional glucose utilization and blood flow following graded forebrain ischemia in the rat: correlation with neuropathology.
Regional patterns of cerebral glucose utilization (rCMRglc) and blood flow (rCBF) were examined in the early recovery period following transient forebrain ischemia in order to correlate early postischemic physiological events with regionally selective patterns of ischemic neuropathology. Wistar rats were subjected to 30 or 60 minutes of graded forebrain ischemia by a method combining unilateral occlusion of the common carotid artery with moderate elevation of intracranial pressure and mild hypotension; this procedure results in a high-grade ischemic deficit affecting chiefly the lateral neocortex, striatum, and hippocampus ipsilateral to the carotid occlusion. Simultaneous measurements of rCMRglc and rCBF made in regional tissue samples after 2 and 4 hours of postischemic recirculation using a double-tracer radioisotopic strategy revealed a disproportionately high level of glucose metabolism relative to blood flow in the early postischemic striatum, owing to the resumption of nearly normal rCMRglc in the face of depressed flow. In contrast, the neocortex, which had been equally ischemic, showed parallel depressions of both metabolism and blood flow during early recovery. Light microscopy at 4 and 8 hours after recovery revealed the striatum to be the predominant locus of ischemic neuronal alterations, whereas neocortical lesions were much less prominent in extent and severity at this time. The resumption of normal levels of metabolism in the setting of a disproportionate depression of rCBF in the early postischemic period may accentuate the process of neuronal injury initiated by ischemia and may contribute to the genesis of neuronal necrosis in "selectively vulnerable" areas of the forebrain.


Schistosome egg antigen(s) presentation and regulatory activity by macrophages isolated from vigorous or immunomodulated liver granulomas of Schistosoma mansoni-infected mice.
Previously, it was shown that macrophages isolated from liver granulomas of Schistosoma mansoni-infected mice constituted about 30% of the granuloma cell population, were highly activated, and more than 50% displayed H-2 I region-associated antigens. Moreover, the degree of macrophage maturation/activation correlated with the intensity of the granulomatous response. Thus, inflammatory macrophages from the vigorous granulomas of acutely infected mice (VigGrM phi) displayed greater phagocytic and tumoricidal activity than did M phi from the T suppressor cell-modulated granulomas (ModGrM phi) of chronically infected animals. No difference was seen, however, in the percentage of Ia-bearing M phi isolated from vigorous or modulated liver granulomas. Presently, accessory activity of GrM phi was assayed by reconstitution of the proliferative responses of M phi-depleted lymphocytes. Both VigGrM phi and ModGrM phi were able to reconstitute the Con-A induced proliferation of normal thymoctyes. Moreover, both types of GrM phi could reconstitute parasite egg antigen-specific proliferation of the mesenteric lymph node cells from acutely infected mice. Reconstitution was dependent on M phi that displayed I-A and I-E subregion-encoded determinants. In higher doses the activated VigGrM phi were more suppressive toward lymphocytic proliferation than were the less activated ModGrM phi. Apparently, the degree of M phi suppressor activity was in direct relationship to the state of M phi maturation/activation. These data indicate that the inflammatory M phi of the schistosome granuloma may be involved in the focal immune accessory/regulatory events that occur within these T cell-mediated lesions.


Esophageal capsule endoscopy in patients refusing conventional endoscopy for the study of suspected esophageal pathology.
Esophageal capsule endoscopy is a well tolerated procedure that does not require sedation and has proved its value for the study of the esophagus. The aim of our study was to assess the feasibility, accuracy, safety and acceptability of esophageal capsule endoscopy for the study of gastroesophageal reflux disease and esophageal varices in patients who refused conventional upper gastrointestinal endoscopy. Thirty consecutive examinations performed in 28 patients (15 men/13 women; mean age: 58.5+/-12.4 years; range: 23-87 years) were reviewed. Twenty-five examinations were performed in 23 patients presenting with chronic gastroesophageal reflux disease symptoms and the remaining five were carried out in patients with cirrhosis for screening of esophageal varices. The procedures were done with the new PillCam ESO that harbors two viewing cupules and takes a total of 14 frames/s. Technical data (total recording time, esophageal transit time and Z-line visualization), clinical findings (Savary-Miller grade; presence and characteristics of esophageal varices or portal hypertension gastropathy), and patient's opinion (quality and comfort questionnaire) were analyzed. All the patients ingested and excreted the capsule without complications. Two examinations in the gastroesophageal reflux disease group were repeated; one was issued as a follow-up and the other because no images were recorded in the first capsule endoscopy. From the former, only the satisfaction questionnaire was analyzed. Mean total recording time and esophageal transit time were 1224 and 243.79 s, respectively (range: 2-1192 s). Complete study of the Z-line was possible in 23 examinations (23/29; 79.3%). Esophageal erosions were seen in 58.33% (14/24) of the examinations carried out in patients with gastroesophageal reflux disease. Nine out of 14 patients (64.29%) presented with grade I esophagitis and the other five (35.71%) with grade II esophagitis. Among the five patients with cirrhosis, small varices were visualized in one (1/5; 20%), while large varices with red spots were evidenced in the remaining three (3/5; 60%). All four patients showing esophageal varices were found to have portal hypertension gastropathy. In most examinations, patients found the capsule easy to swallow (28/30; 93.33%), asymptomatic (29/30; 96.66%), evaluated the procedure as comfortable (29/30; 96.66%), and would repeat it if necessary (30/30; 100%). Esophageal capsule endoscopy is an adequate alternative diagnostic method for the study of gastroesophageal reflux disease and for the screening of esophageal varices in patients refusing to undergo conventional upper gastrointestinal endoscopy.


Control of haemodynamic response to tracheal intubation in cigarette smokers compared with non-smokers.
Some authors have demonstrated that a bolus dose of 1 microg/kg followed by an infusion rate of 0.5 microg/kg/min is adequate to attenuate the haemodynamic response to laringoscopy and tracheal intubation. In this study we have evaluated the efficacy of Remifentanil in controlling haemodynamic and some neuroendocrine responses to tracheal intubation in smokers compared with non-smokers. We studied 126 patients, ASA I-II, aged 20-49 yr, submitted laparoscopic cholecystectomy (66 male, 60 female); sixty-three patients were non-smokers and 63 patients smoked 10 or more cigarettes per day. Anaesthesia was induced with thiopental 3-5 mg/kg and remifentanil 1 microg/kg. Vecuronium 0.1 mg/kg was administrated to facilitate tracheal intubation. Immediately after intubation heart rate of smokers (mean 101.2 +/- 17 beat/min) was significantly higher (p < 0.001) than non-smokers (mean 90.2 +/- 14 beat/min) and also the neuroendocrine responses of smokers (epinephrine value 155 +/- 173 pcg/ml; norepinephrine value 276 +/- 164 pcg/ml) was significantly higher (p < 0.01) than non-smokers (epinephrine 95 +/- 75, norepinephrine 154 +/- 76). These findings may be clinically important to evaluate the risk of ischaemic heart diseases.


Hyperspectral Analysis for Discriminating Herbicide Site of Action: A Novel Approach for Accelerating Herbicide Research.
In agricultural weed management, herbicides are indispensable, yet innovation in their modes of action (MOA)-the general mechanisms affecting plant processes-has slowed. A finer classification within MOA is the site of action (SOA), the specific biochemical pathway in plants targeted by herbicides. The primary objectives of this study were to evaluate the efficacy of hyperspectral imaging in the early detection of herbicide stress and to assess its potential in accelerating the herbicide development process by identifying unique herbicide sites of action (SOA). Employing a novel SOA classification method, eight herbicides with unique SOAs were examined via an automated, high-throughput imaging system equipped with a conveyor-based plant transportation at Purdue University. This is one of the earliest trials to test hyperspectral imaging on a large number of herbicides, and the study aimed to explore the earliest herbicide stress detection/classification date and accelerate the speed of herbicide development. The final models, trained on a dataset with nine treatments with 320 samples in two rounds, achieved an overall accuracy of 81.5% 1 day after treatment. With the high-precision models and rapid screening of numerous compounds in only 7 days, the study results suggest that hyperspectral technology combined with machine learning can contribute to the discovery of new herbicide MOA and help address the challenges associated with herbicide resistance. Although no public research to date has used hyperspectral technology to classify herbicide SOA, the successful evaluation of herbicide damage to crops provides hope to accelerate the progress of herbicide development.


Brominated and organophosphorus flame retardants in body wipes and house dust, and an estimation of house dust hand-loadings in Dutch toddlers.
Children generally have higher Flame Retardant (FR) concentrations in serum compared to other age groups. Toddler behavior enhances direct contact with house dust since their frequent presence proximate to the floor, and their mouthing behavior. This study aimed to thoroughly investigate FR levels in body wipes of toddlers 8-18 months old) and in indoor dust using a noninvasive sampling technique. In this cross-sectional study, body wipes from hands, mouth and back, and indoor household dust samples were collected in twenty-one families and analyzed for one brominated- and seven organophosphorus FRs (polybrominated diphenyl ether 209 (BDE209), tris(2-chloroisopropyl) phosphate (TCIPP), tris(chloroethyl) phosphate (TCEP), tris(1,3-dichloroisopropyl) phosphate (TDCIPP), tris(phenyl) phosphate (TPHP), tris(methylphenyl) phosphate (TMPP), resorcinol bis(diphenyl phosphate) and bisphenol A bis(diphenyl phosphate)). Accelerated solvent extraction was used for extraction and the extract was measured with liquid chromatography combined with mass spectrometry. Non-parametric correlation analyses were performed to assess associations. All FRs were detected in body- and indoor dust samples (median range: 1.0ng/hand wipe (BDE209) to 65ng/hand wipe (TCIPP)) and were mostly correlated with each other. We estimated that approximately 260mg dust (range 50-880mg) accumulated on toddler's hands per day. Hand-to-mouth frequency was negatively associated with FR levels in wipes (τ = -0.38, p = 0.04). With increasing age FR concentrations (BDE209, TCEP, TDCIPP, TPHP and TMPP) on hands decreased significantly (p = 0.01-0.03). Girls had significantly less FRs (TCEP, TCIPP, TPHP and TMPP) on the hands (p = 0.01-0.03) than boys. This is to the best of the authors' knowledge the first study in Europe that measured brominated- as well as organophosphorus FRs in several types of body wipes from toddlers and that estimated the amount of house dust that accumulates on toddler's hands.


The history of the British Dental Association Benevolent Fund: wartimes and between - Part 3.
By the end of the 1910s, the British Dental Association Benevolent Fund was well-established and financially sound. Subscriptions and donations continued to grow but were never quite enough for the treasurer or indeed the committee, who were unable to fulfil all requests for help. Wartime restrictions on paper use meant that publicity was entirely in the hands for the editor of the British Dental Journal, as no separate annual reports were produced. Call-ups to military service decreased the numbers of volunteers willing or able to serve on the committee but applications for assistance in these years continued to grow. The introduction of the 1921 Dentist Act raised issues of resourcing and the future of the Charity for the committee. Novel fundraising suggestions were rife. Volunteer almoners were needed to monitor the increasing number of grant recipients.


Do junior doctors take sick leave?
Nosocomial infections place a heavy burden on overstretched health services. An audit of junior doctors' sick leave behaviour was undertaken in 1993 and again in 2001. The object was to ascertain the level of common infectious illness and to investigate whether junior doctors were remaining at work inappropriately. The doctors were asked if any factors had influenced their decision to take sick leave or not. Between the two audits several initiatives have been introduced to improve the working conditions of junior doctors, including the New Deal to reduce hours of work. Eighty one junior doctors in a large teaching hospital participated in 1993 and 110 in 2001. The number reporting an infectious illness in the previous six months was similar (61.7% in 1993, 68.2% in 2001). There had been a significant increase in the percentage of infectious illness episodes for which the doctors took sick leave (15.1% in 1993, 36.8% in 2001, p < 0.001). The most common reason for taking less sick leave than was felt necessary was concern about colleagues having to do extra work (72% in 1993, 68% in 2001). Consultant pressure was cited by 26% (1993) and 20% (2001). Use of the staff occupational health unit was minimal, with none of the ill doctors contacting the department in 1993 and only three in 2001. Overall, despite the reduction in the number of infectious doctors not taking sick leave, the majority remained at work. Fundamental changes are needed if potentially infected doctors are not to present a risk of iatrogenic infection.


Evaluation of a statewide non-name-based HIV surveillance system.
Recent advances in AIDS-related therapies have delayed the onset of AIDS-defining illnesses and reduced the usefulness of AIDS surveillance in assessing the incidence of early HIV disease and estimating future needs of the HIV-infected population. These changes have prompted renewed interest in expanding surveillance to include HIV and have engendered national debate on whether an HIV surveillance system should be based on reports of the names of infected individuals or employ non-name-based data codes. In 1994, the state of Maryland implemented a program to require HIV surveillance by unique identifier (UI) patient code. This evaluation of Maryland's program found that when complete, the 12-digit UI number provided a virtually unduplicated count 99.8% unique, was 99.9% unique with only the last four digits of the U.S. government Social Security Number (SSN), date of birth (DOB), and race, and 77.7% unique if the last four digits of the SSN were missing. Health care providers were willing to create the UI, with DOB and gender present 98.3% and 98.8% of the time, race was complete 84.1% and last four digits of SSN were complete 72.4%. The overall completeness of reporting for HIV tests was 87.8%.and 84.8%, respectively, using different methodologies. Evidence from the Maryland UI evaluation demonstrates that a non-name-based system can provide accurate, timely and valid data concerning the scope of the HIV epidemic, without the creation of state-wide name-based registry.


Sexual and bladder dysfunction after total mesorectal excision for benign diseases.
An evaluation was made of sexual and bladder dysfunction combined with quality of life (QoL) score after a proctocolectomy with total mesorectal excision (TME) for benign diseases. An in-depth questionnaire was mailed to all patients (n = 94) who had undergone a proctocolectomy with TME followed by an ileal pouch-anal anastomosis (IPAA) in the period from 1989 through 1994. Seventy-six patients responded (81%) with a mean age of 34 +/- 10.6 years and a mean follow-up of 33 +/- 18.3 months. Sexual activity, satisfaction and libido were preserved better in males than in females. Severe sexual dysfunction was found in two males (permanent retrograde ejaculation) and in one female (complete vaginal dryness). No severe bladder dysfunction was found, although minor dysfunction, such as stress and urge incontinence, occurred frequently. QoL was significantly increased postsurgery. Moreover, no evident correlation was demonstrated between QoL and sexual and bladder dysfunction. A relatively low incidence of severe sexual and bladder disorders was found following proctocolectomy with TME and IPAA, whereas minor dysfunction was a relatively common finding. Nevertheless, a significant increase was found in postoperative QoL compared to preoperative QoL.


Multi-country health surveys: are the analyses misleading?
The aim of this paper was to review the types of approaches currently utilized in the analysis of multi-country survey data, specifically focusing on design and modeling issues with a focus on analyses of significant multi-country surveys published in 2010. A systematic search strategy was used to identify the 10 multi-country surveys and the articles published from them in 2010. The surveys were selected to reflect diverse topics and foci; and provide an insight into analytic approaches across research themes. The search identified 159 articles appropriate for full text review and data extraction. The analyses adopted in the multi-country surveys can be broadly classified as: univariate/bivariate analyses, and multivariate/multivariable analyses. Multivariate/multivariable analyses may be further divided into design- and model-based analyses. Of the 159 articles reviewed, 129 articles used model-based analysis, 30 articles used design-based analyses. Similar patterns could be seen in all the individual surveys. While there is general agreement among survey statisticians that complex surveys are most appropriately analyzed using design-based analyses, most researchers continued to use the more common model-based approaches. Recent developments in design-based multi-level analysis may be one approach to include all the survey design characteristics. This is a relatively new area, however, and there remains statistical, as well as applied analytic research required. An important limitation of this study relates to the selection of the surveys used and the choice of year for the analysis, i.e., year 2010 only. There is, however, no strong reason to believe that analytic strategies have changed radically in the past few years, and 2010 provides a credible snapshot of current practice.


Blood transfusion practices and blood-banking services in a Kenyan hospital.
To identify ways to improve the operation of blood-screening programs and to decrease the inappropriate use of blood by evaluating blood-transfusion practices and blood-banking services in a Kenyan hospital. Prospective cohort. The study was conducted in a rural district hospital in western Kenya between September 1990 and July 1991. We collected data on all transfusion requests (blood donation, grouping, HIV screening) and blood recipients (age, sex, diagnosis, and for a 3-month period on the pediatric, maternity, and female wards, admission hemoglobin and outcome). During the 11-month study period, 799 patients received 927 transfusions: 67% were children < 15 years of age, 27% were adult women and 6% were adult men. Transfusions were often delayed due to reliance on patient-recruited donors. Patients who received blood donated on or after the date of request waited longer for transfusion (median, 3 days) than patients who received blood that had been banked and screened before the request (median, 1 day). Patient-recruited donors had a higher HIV-seropositivity rate than volunteer donors (13.4 and 4.6%, respectively; chi 2 test, P < 0.001). Overall, 47% of pediatric transfusions were classified as inappropriate: 23% did not meet the criteria of having hemoglobin < 5.0 g/dl and clinical evidence of respiratory distress, and 27% were transfused 2 or more days after requested. Among adults, 68% received one unit of blood or less. Improved laboratory services, reduction of unnecessary transfusions, and increased recruitment of volunteer donors are critical for improving the appropriate and timely use of blood and reducing transfusion-associated HIV transmission.


The Drosophila cell cycle kinase PAN GU forms an active complex with PLUTONIUM and GNU to regulate embryonic divisions.
Early embryonic cell cycles in Drosophila consist of rapidly alternating S and M phases. Three genes, pan gu (png), plutonium (plu), and giant nuclei (gnu) coordinate these early S-M cycles by ensuring adequate Cyclin B protein levels. Mutations in any of these genes result in unregulated DNA replication and a lack of mitosis ("giant nuclei" phenotype). png encodes a serine/threonine protein kinase, and plu and gnu encode small, novel proteins. We show that PNG, PLU, and GNU constitute a novel protein kinase complex that specifically regulates S-M cell cycles. All three proteins are required for PNG kinase activity and are phosphorylated by PNG in vitro. Yeast two-hybrid screening revealed a direct interaction between PNG and PLU, and their co-expression is required for physical association and activation of PNG kinase. Artificial dimerization of PLU via fusion to either GST or FK506 binding protein (in the presence of dimerizing agent) abrogates the requirement for GNU to activate PNG kinase. We propose a model in which GNU normally regulates embryonic cell cycles by promoting transient dimerization of a core PNG/PLU complex, thereby stimulating PNG kinase activity.


Cloning and expression of a viral phosphoprotein: structure suggests vesicular stomatitis virus NS may function by mimicking an RNA template.
The phosphoprotein (NS) gene from the Indiana serotype of vesicular stomatitis virus (VSV; Mudd-Summers strain) was cloned and sequenced. The NS gene encodes a protein of 265 amino acids which was expressed from a simian virus 40 vector in COS cells. The post-translational modification characteristic of viral NS, the extensive phosphorylation of a cluster of serine and threonine residues, was also evident in recombinant NS protein. The NS gene displays a property common to the phosphoprotein genes of negative-strand RNA viruses: the phosphoprotein mRNA has a second open reading frame (ORF) which could encode a small (7500 mol. wt.) protein. Both measles virus and Sendai virus employ the second ORF of their phosphoprotein gene, and the resultant proteins have an amino acid composition similar to that predicted for the VSV ORF. Comparison of phosphoproteins from different VSV strains revealed two conserved domains that we propose are critical for the function of NS in transcription and replication.


Pattern of methylation of two genes coding for housekeeping functions.
The distribution of sites that can be methylated was analyzed in the Chinese hamster adenine phosphoribosyl-transferase (aprt) gene and the patterns of methylation of this gene and the mouse dihydrofolate reductase (dhfr) gene were studied by using CpG restriction enzymes. Both genes were found to be unmethylated completely at their 5'-end region and methylated heavily throughout the rest of the gene. Because the hamster aprt gene can be inhibited by DNA methylation in vivo, the results suggest that 5' undermethylation of this gene may be a necessary condition for its expression. The pattern of methylation of each of these two genes was similar in sperm and all other somatic tissue DNAs. This is in contrast to many tissue-specific genes that were found to be highly methylated in sperm DNA and undermethylated in the tissue in which they are expressed. This result is consistent with the fact that both aprt and dhfr are key enzymes in the biosynthesis of nucleotides and therefore expected to be synthesized in all cells.


Increased flow rate and papaverine increase K+ exchange in perfused rat hind-limb skeletal muscle.
This study tested the hypothesis that increases in perfusate flow rate result in increased rates of unidirectional and net K+ transport in rat hind-limb skeletal muscle at rest. Ten neurally and vascularly isolated hind limbs, with arterial and venous catheters placed proximal to the popliteal region, were perfused for 10-min periods at flow rates (presented in a random order) of 0.27, 0.42, 0.63, 0.84, or 1.05 mL x min(-1) x g(-1). Potassium extraction and unidirectional K+ influx were determined using 42K, and arterial perfusion pressure was measured continuously. Increases in flow rate resulted in decreases in K+ extraction and increases in unidirectional K+ influx, unidirectional K+ efflux, and net K+ efflux. The increases in K+ flux were associated with increases in oxygen uptake, glucose uptake, and lactate release. In separate experiments (n = 5), the vasodilator papaverine (10(-4) M) did not further vasodilate the vasculature of resting hind limbs, suggesting that the hind limbs in this preparation were fully vasodilated. Papaverine, at constant flow, resulted in a nearly 1.5-fold increase in K+ extraction, a doubling of unidirectional K+ influx, and increases in unidirectional K+ efflux and net K+ efflux. It is concluded that physiological increases in flow rate result in increases in K+ transport in isolated, perfused rat hind-limb skeletal muscle. Furthermore, papaverine appeared to induce an increase in skeletal muscle membrane permeability to K+.


Rapid Construction of ZnO@ZIF-8 Heterostructures with Size-Selective Photocatalysis Properties.
To selectively remove heavy metal from dye solution, inspired by the unique pore structure of ZIF-8, we developed a synthetic strategy for rapid construction of ZnO@ZIF-8 heterostructure photocatalyst for selective reduction of Cr(VI) between Cr(VI) and methylene blue (MB). In particular, ZnO@ZIF-8 core-shell heterostructures were prepared by in situ ZIF-8 crystal growth using ZnO colloidal spheres as template and zinc source within 8-60 min. The shell of the resulting ZnO@ZIF-8 core-shell heterostructure with a uniform thickness of around 30 nm is composed of ZIF-8 crystal polyhedrons. The concentration of organic ligand 2-methylimidazole (Hmim) was found to be crucial for the formation of ZnO@ZIF-8 core-shell heterostructures. Different structures, ZnO@ZIF-8 core-shell spheres and separate ZIF-8 polyhedrons could be formed by altering Hmim concentration, which significantly influences the balance between rate of Zn(2+) release from ZnO and coordinate rate. Importantly, such ZnO@ZIF-8 core-shell heterostructures exhibit size-selective photocatalysis properties due to selective adsorption and permeation effect of ZIF-8 shell. The as-synthesized ZnO@ZIF-8 heterostructures exhibited enhanced selective reduction of Cr(VI) between Cr(VI) and MB, which may find application in the dye industry. This work not only provides a general route for rapid fabrication of such core-shell heterostructures but also illustrates a strategy for selectively enhanced photocatalysis performance by utilizing adsorption and size selectivity of ZIF-8 shell.


Dorsal induction from dorsal vegetal cells in Xenopus occurs after mid-blastula transition.
We performed some experiments to investigate the temporal and spatial details of the dorsal induction exerted by dorsal vegetal cells in Xenopus embryo. Two dorsal vegetal cells (D1 cells) were transplanted into the ventral vegetal region of a recipient at the 32-cell stage. At various times after transplantation, the ventral animal-equatorial part was explanted and cultured. The explants isolated 5.5 h after transplantation (time 5.5) elongated and formed somites. In RT-PCR analysis, the expression of dorsal gene, chordin was activated in the explants isolated after time 4.0 (about the 4000-cell stage which corresponds to the mid blastula transition (MBT)) at control stage 10. In another series of experiments, ventral animal-equatorial and dorsal vegetal parts were isolated from the 4000-cell stage embryos and they were combined for 2.0-2.5 h. These ventral animal-equatorial explants elongated and formed somites. The chordin expression was also observed in the explants. But the 32- and 256-cell stage dorsal vegetal cells failed to exert the dorsalizing activity within the 2.0-2.5 h of the conjugation. These results suggest that 2 h contact after MBT is necessary and sufficient for the dorsal induction from the dorsal vegetal cells and it occurs as a result of the zygotic gene expression. Consistent with this idea, the zygotic dorsal genes, siamois and chordin were expressed on the upper regions of the transplanted D1 descendants at stage 10. Furthermore, this region began to gastrulate when the D1 cell was transplanted with upside-down orientation. Our data indicate that the upper region of the D1 descendants by itself act as the Spemann organizer rather than the Nieuwkoop center.


Characterization of retroviral and lentiviral vectors pseudotyped with xenotropic murine leukemia virus-related virus envelope glycoprotein.
Retroviral and lentiviral vectors are effective gene delivery vehicles that are being evaluated in clinical trials. Variations in the viral envelope (Env) glycoproteins, which are used to pseudotype retroviral or lentiviral vectors, can alter vector performance, including stability, titers, host range, and tissue tropism. Xenotropic murine leukemia virus (MLV)-related virus (XMRV) is a novel human retrovirus identified in patients with prostate cancer. XMRV targets XPR1 cell surface receptor, which is expressed in a broad range of human tissues including hematopoietic stem cells. Pseudotyping with XMRV Env would allow targeting of XPR1-expressing tissues. Here, we characterized XMRV Env-pseudotyped retroviral and lentiviral vectors. Although HIV and MLV vectors were poorly pseudotyped with wild-type XMRV Env, replacement of the C-terminal 11 amino acid residues in the transmembrane domain of XMRV Env with the corresponding 6 amino acid residues of amphotropic MLV Env (XMRV/R(ampho)) significantly increased XMRV Env-pseudotyped HIV and MLV vector titers. The transduction efficiency in human CD34(+) cells when using the XMRV/R(ampho)-pseudotyped HIV vector (10-20%) was comparable to that achieved when using the same infectious units of vesicular stomatitis virus G glycoprotein-pseudotyped vector (25%); thus the modified XMRV Env offers an alternative pseudotyping strategy for XPR1-mediated gene delivery.


Single-cell epigenomics: Recording the past and predicting the future.
Single-cell multi-omics has recently emerged as a powerful technology by which different layers of genomic output-and hence cell identity and function-can be recorded simultaneously. Integrating various components of the epigenome into multi-omics measurements allows for studying cellular heterogeneity at different time scales and for discovering new layers of molecular connectivity between the genome and its functional output. Measurements that are increasingly available range from those that identify transcription factor occupancy and initiation of transcription to long-lasting and heritable epigenetic marks such as DNA methylation. Together with techniques in which cell lineage is recorded, this multilayered information will provide insights into a cell's past history and its future potential. This will allow new levels of understanding of cell fate decisions, identity, and function in normal development, physiology, and disease.


Effect of Ramadan fasting on salivary IgA, serum IgA, IL-17, and IL-22 levels.
Background: nutritional factors might affect the number and function of immune cells for instance the production of cytokines and immunoglobulins. Ramadan fasting is intermittent abstinence from eating and drinking for almost four weeks. Aim: The present study aimed to investigate the influence of intermittent fasting on serum IgA, salivary IgA (sIgA), interleukin (IL)-17, and IL-22 levels. Methods: 40 healthy men aged 19-29 years were evaluated before and during the fourth week of Ramadan fasting for IgA levels by the nephelometric method as well as salivary IgA (sIgA), IL-17, and IL-22 amounts using enzyme-linked immunosorbent assay (ELISA). Results: serum IgA levels reduced significantly at the end of Ramadan fasting (225.8 ± 87 vs. 196 ± 70 mg/dl) (p-value<0.001); however, sIgA amounts did not differ between before and the last week of Ramadan. Serum IL-17 reduced significantly (2.93 ± 1.51 vs. 2.17 ± 1.33 pg/ml) (p-value = 0.006) whereas IL-22 levels remained approximately unchanged. Summary: four weeks of intermittent fasting during Ramadan reduced the serum levels of IgA and IL-17 but did not affect the production of sIgA and IL-22. These findings indicate a limited impact of intermittent fasting on mucosal immunity.


[Effects of chelation therapy with succimer in young rabbits of moderate lead poisoning].
To explore the effect of chelation therapy with succimer (DMSA) in male rabbits of moderate lead poisoning during juvenile stage. Twenty-four 45-day-old male New Zealand rabbits were randomly divided into three groups (therapy group, TG; positive control group, PG and negative control group, NG, n=8). The TG and PG were orally exposed to lead acetate (5 mg x kg(-1) x d(-1)) for 6 weeks. Rabbits in TG were orally supplied DMSA 1050 mg/m2 in the first week and 700 mg/m2 in the next two weeks, while the other two groups wren't blood and urinary samples of all rabbits were collected per week. The tissues and organs of all rabbits were collected after 12 weeks. The blood lead levels (BLLs) were determined by atomic absorption spectrometer. The urine lead levels and the lead contents of tissue and organ were determined by inductively coupled plasma-mass spectrometry. Histopathology of tissue and organ was observed by light microscope. Compared with PG, the lead level in the morning urine of TG with DMSA chelating was increased significantly. The level was peaked at (1246.96 +/- 157.91) microg/L on the first day after chelating. While the base line was (40.97 +/- 1.77) microg/L before chelating. Meanwhile, the BLLs were sharply declined from (429.63 +/- 10.82) microg/L to (238.50 +/- 11.82) microg/L. The urine lead levels of TG decreased through the 3-week chelating and 3-week discontinuation. The urine lead levels of these two groups were significantly different (F=2934.35, P<0.01). Compared to each two groups in these three groups, there were significant difference (P<0.01). The authors found the reversion of BLLs in first week after stop chelating. The BLLs of PG presented the slow course of declining in the same time, were (135.50 +/- 7.09) microg/L, very close to the level of TG for (149.88 +/- 11.39) microg/L. Compared with treatment discontinuation for 3 weeks, the urine lead levels and the body weight gain of the therapy group increased more than that of PG, and the BLLs and the lead concentrations in tissues and organs decreased more than that of PG, and histopathology in the liver tissues and testicle tissues were improved. DMSA chelating for the rodent models of moderate lead poisoning might reduce the BLLs and soft tissue lead contents quickly and effectively, decrease toxic effects of lead in a short period of time, thus alleviate the impairment of lead poisoning on tissues and organs by decreasing lead burden, and bring out improvement on the growth retardation caused by lead poisoning.


Autofluorescence in samples obtained from chronic biofilm infections--"all that glitters is not gold".
When looking at tissue sections of ex vivo samples, autofluorescence can be a major cause of artifacts and misinterpretations. We here reiterate evidence that autofluorescing granules, often hemosiderin but also ceroid or mucinogen granules, are severe obstacles when imaging and diagnosing biofilm infections through fluorescent imaging techniques. We used confocal laser scanning microscopy with spectral analysis for autofluorescence detection as well as standard histological stains in order to identify the culprit and show that these granules might very well be mistaken for bacterial biofilms. Furthermore, we hypothesize that the increased amount of autofluorescing granules may be a consequence of prolonged inflammation as a consequence of chronic biofilm infections.


Chlorido[2,3,5,6-tetra-kis-(tert-butyl-sulfanylmeth-yl)phenyl-κ(3) S (2),C (1),S (6)]palladium(II) dichloro-methane monosolvate.
The title compound, [Pd(C26H45S4)Cl]·CH2Cl2, crystallizes with a disordered dichloro-methane solvent mol-ecule [occupancy ratio = 0.67 (4):0.33 (4)]. Two of the tert-butyl groups are also disordered [occupancy ratios = 0.70 (5):0.30 (5) and 0.63 (4):0.37 (4)]. Although the pincer ligand offers the possibility for coordination of two different metal atoms, the present structure shows only the coordination of a single Pd(II) atom in a typical S-C-S tridentate pincer manner. The Pd(II) atom is in a slightly distorted square-planar environment with the two tert-butyl-sulfanyl groups arranged in a trans con-formation and with a chloride ligand trans to the σ-bonded aromatic C atom. The structure exhibits a durene-like ligand frame, forming a dihedral angle of 13.6 (4)° with the metal coordination (Pd/S/S/Cl/C) environment. It is noteworthy that the tert-butyl groups are found in a syn arrangement, this being different to that found previously by Loeb, Shimizu & Wisner [(1998). Organometallics, 17, 2324-2327].


Hepatitis C virus infection: an update.
Hepatitis C virus (HCV) infection is widely prevalent world-wide particularly in high-risk individuals including patients on hemodialysis (HD). Parenteral route seems to be the main route of transmission of this infection though other routes also exist. The high prevalence of HCV infection in patients on HD who have never received blood transfusions points to the dialysis machine as one of the likely sources of transmission of infection. The reported prevalence of anti-HCV in patients on HD in Saudi Arabia is about 68%. The natural history of HCV shows a tendency to progress to chronic liver disease including chronic active hepatitis, cirrhosis and hepatocellular carcinoma. The prevalence of anti-HCV in the general population varies from 0.5 to 3% and thus vigorous screening should be applied to all potential blood donors. A policy of public health education should be designed and implemented on the general public and health workers to reduce the non-parenteral spread of HCV. Separate HD machines should be considered for patients who are anti-HCV positive. Such preventive measures are vital because treatment of HCV with interferon has met with mixed success so far.


Teleost kidney in evaluation of xenobiotic toxicity and elimination.
The unique anatomy of the marine teleost kidney and the development of techniques to take advantage of that anatomy are discussed and several examples of their use to study xenobiotic excretion and toxicity are documented. These include: 1) characterization of renal organic anion transport, the system that mediates excretion of many such compounds and their metabolites; 2) assessment of the relative roles of transport, metabolism, and plasma binding in xenobiotic excretion; 3) examination of the morphological aspects of cellular injury; and 4) analysis of subcellular sites of heavy metal toxicity. Also presented are applications of new techniques in membrane physiology to the study of the effects of xenobiotics. It is suggested that continued application of the comparative approach, as exemplified by the use of the marine teleost kidney, will provide powerful and versatile tools for investigation of fundamental biological processes and the interactions of foreign compounds with those processes.


AKAP79/150 interacts with AC8 and regulates Ca2+-dependent cAMP synthesis in pancreatic and neuronal systems.
Protein kinase A anchoring proteins (AKAPs) provide the backbone for targeted multimolecular signaling complexes that serve to localize the activities of cAMP. Evidence is accumulating of direct associations between AKAPs and specific adenylyl cyclase (AC) isoforms to facilitate the actions of protein kinase A on cAMP production. It happens that some of the AC isoforms (AC1 and AC5/6) that bind specific AKAPs are regulated by submicromolar shifts in intracellular Ca(2+). However, whether AKAPs play a role in the control of AC activity by Ca(2+) is unknown. Using a combination of co-immunoprecipitation and high resolution live cell imaging techniques, we reveal an association of the Ca(2+)-stimulable AC8 with AKAP79/150 that limits the sensitivity of AC8 to intracellular Ca(2+) events. This functional interaction between AKAP79/150 and AC8 was observed in HEK293 cells overexpressing the two signaling molecules. Similar findings were made in pancreatic insulin-secreting cells and cultured hippocampal neurons that endogenously express AKAP79/150 and AC8, which suggests important physiological implications for this protein-protein interaction with respect to Ca(2+)-stimulated cAMP production.


(1S,3R,8R)-2,2-Di-chloro-3,7,7,10-tetra-methyl-tri-cyclo-[18.104.22.168(1,3)]dodec-9-en-11-one.
The title compound, C16H22Cl2O, was synthesized from β-himachalene (3,5,5,9-tetra-methyl-2,4a,5,6,7,8-hexa-hydro-1H-benzo-cyclo-heptene), which was isolated from the essential oil of the Atlas cedar (Cedrus Atlantica). The mol-ecule is built up from fused six- and seven-membered rings and an additional three-membered ring arising from the reaction of himachalene with di-chloro-carbene. The six-membered ring has an envelope conformation, with the C atom belonging to the three-membered ring forming the flap, whereas the seven-membered ring displays a screw-boat conformation; the dihedral angle between the rings (all atoms) is 59.65 (14)°.


Second-sphere contributions to substrate-analogue binding in iron(III) superoxide dismutase.
A combination of spectroscopic and computational methods has been employed to explore the nature of the yellow and pink low-temperature azide adducts of iron(III) superoxide dismutase (N(3)-FeSOD), which have been known for more than two decades. Variable-temperature variable-field magnetic circular dichroism (MCD) data suggest that both species possess similar ferric centers with a single azide ligand bound, contradicting previous proposals invoking two azide ligands in the pink form. Complementary data obtained on the azide complex of the Q69E FeSOD mutant reveal that relatively minor perturbations in the metal-center environment are sufficient to produce significant spectral changes; the Q69E N(3)-FeSOD species is red in color at all temperatures. Resonance Raman (RR) spectra of the wild-type and Q69E mutant N(3)-FeSOD complexes are consistent with similar Fe-N(3) units in all three species; however, variations in energies and relative intensities of the RR features associated with this unit reveal subtle differences in (N(3)(-))-Fe(3+) bonding. To understand these differences on a quantitative level, density functional theory and semiempirical INDO/S-CI calculations have been performed on N(3)-FeSOD models. These computations support our model that a single azide ligand is present in all three N(3)-FeSOD adducts and suggest that their different appearances reflect differences in the Fe-N-N bond angle. A 10 degrees increase in the Fe-N-N bond angle is sufficient to account for the spectral differences between the yellow and pink wild-type N(3)-FeSOD species. We show that this bond angle is strongly affected by the second coordination sphere, which therefore might also play an important role in orienting incoming substrate for reaction with the FeSOD active site.


Transient N-6-Methyladenosine Transcriptome Sequencing Reveals a Regulatory Role of m6A in Splicing Efficiency.
Splicing efficiency varies among transcripts, and tight control of splicing kinetics is crucial for coordinated gene expression. N-6-methyladenosine (m6A) is the most abundant RNA modification and is involved in regulation of RNA biogenesis and function. The impact of m6A on regulation of RNA splicing kinetics is unknown. Here, we provide a time-resolved high-resolution assessment of m6A on nascent RNA transcripts and unveil its importance for the control of RNA splicing kinetics. We find that early co-transcriptional m6A deposition near splice junctions promotes fast splicing, while m6A modifications in introns are associated with long, slowly processed introns and alternative splicing events. In conclusion, we show that early m6A deposition specifies the fate of transcripts regarding splicing kinetics and alternative splicing.


Does in situ replacement of a staphylococcal infected vascular graft with a rifampicin impregnated gelatin sealed Dacron graft reduce the incidence of subsequent infection?
The aim of this study was to treat methicillin-resistant Staphylococcus aureus (MRSA) or S. epidermidis prosthetic vascular graft infections by in situ replacement with a rifampicin bonded Gelsoft graft. Interposition grafts were placed in the carotid artery of 56 sheep and the graft surface directly inoculated with 10(8) colony forming units of MRSA or S. epidermidis. At three weeks, grafts were harvested and sheep allocated to three groups. In the MRSA group, sheep received grafts soaked in 1.2 mg/ml (12), 10 mg/ml (10) and no rifampicin (7). For S. epidermidis, sheep received grafts soaked in 1.2 mg/ml (10), 10 mg/ml (9) and no rifampicin (8). There were two deaths, in the MRSA study group. Remaining sheep were euthanased and grafts harvested three weeks following regrafting. Swabs were taken to assess bacterial growth in the perigraft tissues, and external and internal graft surfaces. A 3-5 mm segment of graft was incubated in broth medium. For MRSA, no statistical difference between the groups was reached for any of the measured parameters. For S. epidermidis, a significant reduction was reached for total infected specimens in the 10 mg/ml group compared to both control (p<0.001) and 1.2 mg/ml (p<0.005) groups. Graft re-infection was also less likely to occur with S. epidermidis than MRSA. Replacement of S. epidermidis infected vascular grafts with 10 mg/ml rifampicin soaked Gelsoft graft is effective in reducing subsequent S. epidermidis infection. This conclusion cannot be extended to MRSA infected vascular grafts.


Tubular cells produce FGF2 via autophagy after acute kidney injury leading to fibroblast activation and renal fibrosis.
Following acute kidney injury (AKI), renal tubular cells may stimulate fibroblasts in a paracrine fashion leading to interstitial fibrosis, but the paracrine factors and their regulation under this condition remain elusive. Here we identify a macroautophagy/autophagy-dependent FGF2 (fibroblast growth factor 2) production in tubular cells. Upon induction, FGF2 acts as a key paracrine factor to activate fibroblasts for renal fibrosis. After ischemic AKI in mice, autophagy activation persisted for weeks in renal tubular cells. In inducible, renal tubule-specific atg7 (autophagy related 7) knockout (iRT-atg7-KO) mice, autophagy deficiency induced after AKI suppressed the pro-fibrotic phenotype in tubular cells and reduced fibrosis. Among the major cytokines, tubular autophagy deficiency in iRT-atg7-KO mice specifically diminished FGF2. Autophagy inhibition also attenuated FGF2 expression in TGFB1/TGF-β1 (transforming growth factor, beta 1)-treated renal tubular cells. Consistent with a paracrine action, the culture medium of TGFB1-treated tubular cells stimulated renal fibroblasts, and this effect was suppressed by FGF2 neutralizing antibody and also by fgf2- or atg7-deletion in tubular cells. In human, compared with non-AKI, the renal biopsies from post-AKI patients had higher levels of autophagy and FGF2 in tubular cells, which showed significant correlations with renal fibrosis. These results indicate that persistent autophagy after AKI induces pro-fibrotic phenotype transformation in tubular cells leading to the expression and secretion of FGF2, which activates fibroblasts for renal fibrosis during maladaptive kidney repair.Abbreviations: 3-MA: 3-methyladnine; ACTA2/α-SMA: actin alpha 2, smooth muscle, aorta; ACTB/β-actin: actin, beta; AKI: acute kidney injury; ATG/Atg: autophagy related; BUN: blood urea nitrogen; CCN2/CTGF: cellular communication network factor 2; CDKN2A/p16: cyclin dependent kinase inhibitor 2A; CKD: chronic kidney disease; CM: conditioned medium; COL1A1: collagen, type I, alpha 1; COL4A1: collagen, type IV, alpha 1; CQ: chloroquine; ECM: extracellular matrix; eGFR: estimated glomerular filtration rate; ELISA: enzyme-linked immunosorbent assay; FGF2: fibroblast growth factor 2; FN1: fibronectin 1; FOXO3: forkhead box O3; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HAVCR1/KIM-1: hepatitis A virus cellular receptor 1; IHC: immunohistochemistry; IRI: ischemia-reperfusion injury; ISH: in situ hybridization; LTL: lotus tetragonolobus lectin; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PDGFB: platelet derived growth factor, B polypeptide; PPIB/cyclophilin B: peptidylprolyl isomerase B; RT-qPCR: real time-quantitative PCR; SA-GLB1/β-gal: senescence-associated galactosidase, beta 1; SASP: senescence-associated secretory phenotype; sCr: serum creatinine; SQSTM1/p62: sequestosome 1; TASCC: TOR-autophagy spatial coupling compartment; TGFB1/TGF-β1: transforming growth factor, beta 1; VIM: vimentin.


Relaxation effect of CGP-48506, EMD-57033, and dobutamine in ejecting and isovolumically beating rabbit hearts.
Because it is not known whether ejection influences the negative effect of the Ca(2+)-sensitizing drugs on ventricular relaxation, we extended our previous analysis of stress-dependent relaxation in isovolumic beats to encompass ejecting beats and evaluated the relationships between both the time of onset of relaxation and the rate of relaxation and wall stress in a broader analysis framework. Furthermore, because the sites of action of the Ca(2+)-sensitizing drugs CGP-48506 and EMD-57033 may be different, and thus CGP-48506 may have fewer adverse effects on resting muscle length or force, we compared these two drugs to test the hypothesis that CGP-48506 would have less effect than EMD-57033 on relaxation in the isolated buffer-perfused rabbit heart. This analysis of stress-dependent relaxation in both ejecting and isovolumic beats readily differentiates between the negative lusitropic effect of 2 x 10(-6) M EMD-57033, the negligible lusitropic effect of 6 x 10(-6) M CGP-48506, and the positive lusitropic effect of 1.25 x 10(-6) M dobutamine. Furthermore, comparison of the effect of the two Ca(2+)-sensitizing drugs in ejecting versus isovolumic contractions shows that CGP-48506 affects relaxation differently in ejecting contractions than it does in isovolumic contractions, whereas EMD-57033 affects relaxation similarly in both ejecting and isovolumic contractions.


Atogepant for migraine.
Atogepant is a selective oral, small-molecule calcitonin gene-related peptide (CGRP) receptor antagonist that has been approved for preventive treatment of migraine. CGRP is a neuropeptide involved in vasodilation and cerebrovascular regulation. CGRP is the most potent vasoactive constituent of the cerebrovascular trigeminal neuronal system and has a key role in migraine. Medications targeting CGRP are being used as migraine preventive and abortive treatments.


Extracellular vesicles from in vivo liver tissue accelerate recovery of liver necrosis induced by carbon tetrachloride.
Extracellular vesicles (EVs) are nano-sized vesicles composed of proteolipid bilayers carrying various molecular signatures of the cells. As mediators of intercellular communications, EVs have gained great attention as new therapeutic agents in the field of nanomedicine. Therefore, many studies have explored the roles of cell-derived EVs isolated from cultured hepatocytes or stem cells as inducer of liver proliferation and regeneration under various pathological circumstances. However, study investigating the role of EVs directly isolated from liver tissue has not been performed. Herein, to understand the pathophysiological role and to investigate the therapeutic potential of in vivo liver EVs, we isolated EVs from both normal and carbon tetrachloride (CCl4)-induced damaged in vivo liver tissues. The in vivo EVs purified from liver tissues display typical features of EVs including spherical morphology, nano-size, and enrichment of tetraspanins. Interestingly, administration of both normal and damaged liver EVs significantly accelerated the recovery of liver tissue from CCl4-induced hepatic necrosis. This restorative action was through the induction of hepatocyte growth factor at the site of the injury. These results suggest that not only normal liver EVs but also damaged liver EVs play important pathophysiological roles of maintaining homeostasis after tissue damage. Our study, therefore, provides new insight into potentially developing in vivo EV-based therapeutics for preventing and treating liver diseases.


Cardiovascular risk factors and the metabolic syndrome in pediatric nonalcoholic fatty liver disease.
Nonalcoholic fatty liver disease (NAFLD), the most common cause of liver disease in children, is associated with obesity and insulin resistance. However, the relationship between NAFLD and cardiovascular risk factors in children is not fully understood. The objective of this study was to determine the association between NAFLD and the presence of metabolic syndrome in overweight and obese children. This case-control study of 150 overweight children with biopsy-proven NAFLD and 150 overweight children without NAFLD compared rates of metabolic syndrome using Adult Treatment Panel III criteria. Cases and controls were well matched in age, sex, and severity of obesity. Children with NAFLD had significantly higher fasting glucose, insulin, total cholesterol, low-density lipoprotein cholesterol, triglycerides, systolic blood pressure, and diastolic blood pressure than overweight and obese children without NAFLD. Subjects with NAFLD also had significantly lower high-density lipoprotein cholesterol than controls. After adjustment for age, sex, race, ethnicity, body mass index, and hyperinsulinemia, children with metabolic syndrome had 5.0 (95% confidence interval, 2.6 to 9.7) times the odds of having NAFLD as overweight and obese children without metabolic syndrome. NAFLD in overweight and obese children is strongly associated with multiple cardiovascular risk factors. The identification of NAFLD in a child should prompt global counseling to address nutrition, physical activity, and avoidance of smoking to prevent the development of cardiovascular disease and type 2 diabetes.


The osteoprotective effect of psoralen in ovariectomy-induced osteoporotic rats via stimulating the osteoblastic differentiation from bone mesenchymal stem cells.
Psoralea corylifolia extract has been reported to promote bone formation in osteoporotic animals. Psoralen (PSO), a flavonoid glycoside, as the active component of P corylifolia L, is effective in increasing new bone-forming osteoblasts in parietal bone defects. However, the effect and molecular mechanisms of PSO on bone mesenchymal stem cells (bMSCs) in the osteoporotic state are widely unknown. This study was designed to evaluate the osteoprotective effect of PSO in ovariectomy (OVX)-induced rats and to seek possible molecular mechanisms of PSO in bMSCs. We observed the osteogenic effect of PSO (3-month treatment) on osteoporotic rat models induced by OVX via testing bone densitometry, histomorphometries, and immunohistochemistry in vivo. Alkaline phosphatase staining and colony-forming unit-fibroblast and colony-forming unit-adipocyte assays were performed to evaluate the differentiation potential of bMSCs ex vivo. In addition, the molecular targets of PSO in bMSCs were detected by stem cell microarray analysis of 256 genes and confirmed by real-time reverse transcription-polymerase chain reaction. Micro-CT morphometry analysis showed that PSO significantly improved bone mass indicators including increased trabecular thickness and decreased trabecular space. Meanwhile, PSO elevated the well-known osteogenic marker osteocalcin level in OVX-induced osteoporotic rats. Next, in ex vivo studies, we revealed that PSO facilitated alkaline phosphatase staining and increased the colony-forming unit-fibroblasts. Based on gene expression profile analysis, we screened a set of genes dysregulated in OVX but reversed by PSO treatment. These genes were highly enriched in the Notch signaling pathway, which was documented to play a role in bMSC differentiation. Our findings show that PSO promotes bone mass in OVX-induced osteoporotic rats. This effect of PSO is highly related to the stimulation of differentiation of bMSCs to osteoblasts.


GASOLINE: detecting germline and somatic structural variants from long-reads data.
Long-read sequencing allows analyses of single nucleic-acid molecules and produces sequences in the order of tens to hundreds kilobases. Its application to whole-genome analyses allows identification of complex genomic structural-variants (SVs) with unprecedented resolution. SV identification, however, requires complex computational methods, based on either read-depth or intra- and inter-alignment signatures approaches, which are limited by size or type of SVs. Moreover, most currently available tools only detect germline variants, thus requiring separate computation of sample pairs for comparative analyses. To overcome these limits, we developed a novel tool (Germline And SOmatic structuraL varIants detectioN and gEnotyping; GASOLINE) that groups SV signatures using a sophisticated clustering procedure based on a modified reciprocal overlap criterion, and is designed to identify germline SVs, from single samples, and somatic SVs from paired test and control samples. GASOLINE is a collection of Perl, R and Fortran codes, it analyzes aligned data in BAM format and produces VCF files with statistically significant somatic SVs. Germline or somatic analysis of 30[Formula: see text] sequencing coverage experiments requires 4-5 h with 20 threads. GASOLINE outperformed currently available methods in the detection of both germline and somatic SVs in synthetic and real long-reads datasets. Notably, when applied on a pair of metastatic melanoma and matched-normal sample, GASOLINE identified five genuine somatic SVs that were missed using five different sequencing technologies and state-of-the art SV calling approaches. Thus, GASOLINE identifies germline and somatic SVs with unprecedented accuracy and resolution, outperforming currently available state-of-the-art WGS long-reads computational methods.


Self-supervised learning in medicine and healthcare.
The development of medical applications of machine learning has required manual annotation of data, often by medical experts. Yet, the availability of large-scale unannotated data provides opportunities for the development of better machine-learning models. In this Review, we highlight self-supervised methods and models for use in medicine and healthcare, and discuss the advantages and limitations of their application to tasks involving electronic health records and datasets of medical images, bioelectrical signals, and sequences and structures of genes and proteins. We also discuss promising applications of self-supervised learning for the development of models leveraging multimodal datasets, and the challenges in collecting unbiased data for their training. Self-supervised learning may accelerate the development of medical artificial intelligence.


A polytope DNA vaccine elicits multiple effector and memory CTL responses and protects against human papillomavirus 16 E7-expressing tumour.
Vaccine-induced CD8 T cells directed to tumour-specific antigens are recognised as important components of protective and therapeutic immunity against tumours. Where tumour antigens have pathogenic potential or where immunogenic epitopes are lost from tumours, development of subunit vaccines consisting of multiple individual epitopes is an attractive alternative to immunising with whole tumour antigen. In the present study we investigate the efficacy of two DNA-based multiepitope ('polytope') vaccines containing murine (H-2b) and human (HLA-A*0201)-restricted epitopes of the E7 oncoprotein of human papillomavirus type 16, in eliciting tumour-protective cytotoxic T-lymphocyte (CTL) responses. We show that the first of these polytopes elicited powerful effector CTL responses (measured by IFN-gamma ELISpot) and long-lived memory CTL responses (measured by functional CTL assay and tetramers) in immunised mice. The responses could be boosted by immunisation with a recombinant vaccinia virus expressing the polytope. Responses induced by immunisation with polytope DNA alone partially protected against infection with recombinant vaccinia virus expressing the polytope. Complete protection was afforded against challenge with an E7-expressing tumour, and reduced growth of nascent tumours was observed. A second polytope differing in the exact composition and order of CTL epitopes, and lacking an inserted endoplasmic reticulum targeting sequence and T-helper epitope, induced much poorer CTL responses and failed to protect against tumour challenge. These observations indicate the validity of a DNA polytope vaccine approach to human papillomavirus E7-associated carcinoma, and underscore the importance of design in polytope vaccine construction.


Saliva as a biomarker for head and neck squamous cell carcinoma: in vitro detection of cytotoxic effects by using the plating efficiency index.
Worldwide, the incidence of head and neck squamous cell carcinoma (HNSCC) is increasing. The development of an effective prevention program would provide a promising opportunity to control this disease. We applied the 'plating efficiency index' of Chinese hamster lung fibroblasts to demonstrate the presence of cytotoxic effects in saliva samples from cancer patients as well as from healthy probands. Correlations between individual risk factors and the cytotoxic effects of saliva specimens were analysed and evaluated. Saliva samples were obtained from male patients (n=43) with carcinomas of the upper aerodigestive tract, and from a healthy age-matched male control group (n=131) with different tobacco and alcohol consumption habits. The extraction of non-stimulated sober saliva was chosen for this investigation. In vitro cytotoxicity of the saliva was detected using the permanent mammalian cell line V79 (lung fibroblasts) of Chinese hamsters. The determination of the plating efficiency index was made in the logarithmic growth phase of the initial cell culture. A reduction in cell colonies (plating efficiency) of at least 50% was defined as strongly cytotoxic. A significant direct correlation was found between daily tobacco consumption and the reduction of plating efficiency (p<0.0001). We found only a moderate increase in the cytotoxicity of tested saliva samples in correlation with daily alcohol uptake. The difference between tumor patients and healthy probands was highly significant (chi(2) test; p=0.001). Using the method of logistic regression analysis, we found a 3.6-fold increased cancer risk in probands with cytotoxic saliva (p<0.001). According to our results, the plating efficiency index seems to be a suitable method for the detection of increased cancer risk. In combination with several effective biomarkers on cytogenetic end-points, it may help to establish biomonitoring programs for secondary cancer prevention.


Probing the catalytic sites of triosephosphate isomerase by 31P-NMR with reversibly and irreversibly binding substrate analogues.
We have explored the degree of independence of the two catalytic centers, interactions between the catalytic centers and the subunit-subunit contact sites, and different conformations of triosephosphate isomerase (TPI), by simultaneously employing irreversibly (covalent) and reversibly binding substrate analogues and monitoring their 31P-NMR resonances. 3-Chloroacetol phosphate (CAP) was bound to the active site by reaction with Glu165. The resulting, inactive (CAP-TPI)2 complex exhibited two distinct 31P-NMR resonances which were independent of pH and represent two conformational forms of the enzyme. Dissociation in guanidine hydrochloride followed by redimerization resulted in a single conformation. This was observed with the enzyme from chicken, rabbit and yeast. The inactive (CAP-TPI)2 dimer was mixed with native TPI, and dissociated/reassociated to form heterodimers (CAP-TPI)(TPI) in which one subunit contained the CAP label and the other subunit was unmodified. This hybrid migrated intermediate between the native and CAP-modified enzyme on nondenaturing PAGE. The heterodimer exhibited 50% the activity of the native dimer, but kinetic properties were otherwise indistinguishable. The reversibly binding transition state analogue, 2-phosphoglycolate (PGA), was used to probe the remaining vacant active site of the heterodimer. Bound PGA exhibited a pH-independent 31P-NMR resonance which was readily distinguishable from resonances of CAP-TPI and free PGA. No differences were observed in the binding of PGA to the vacant subunit of the heterodimer or the native dimer, further pointing to the independent nature of the two catalytic centers. However, the (CAP-TPI)(TPI) heterodimer was more susceptible to subunit dissociation in guanidine hydrochloride than the native dimer. Thus, it appears that the two active sites function completely independently of each other, but that the binding of CAP at the active center loosens the subunit-subunit contact. In addition, the two forms of the enzyme-inhibitor complex trapped by reaction with CAP may represent conformations with the hinged lid or flexible loop (residues 166-176) in the open and closed positions.


Intrajoint comparisons of gene expression patterns in human osteoarthritis suggest a change in chondrocyte phenotype.
Osteoarthritis (OA) is a degenerative cartilage disease with varying degrees of severity within a given joint. The purpose of this study was to define a sampling procedure for comparing human minimal and advanced OA cartilage in the same patient and to determine basic patterns of gene expression in these regions. A specific hypothesis under study was that the expression level of Bcl-2 would correlate with Sox9 and aggrecan mRNA expression in vivo as has been demonstrated in vitro. Femoral condylar advanced OA cartilage was located within 1cm of overt lesions, and minimal cartilage was taken from areas with no obvious surface defects. Histological sections were scored for disease severity and chondroitin sulfate and hydroxyproline content was determined. The expression level of nine specific genes (aggrecan, collagen type II, Bcl-2, Sox9, Link protein, osteopontin, and MMP-13, -3, and -9) was determined by quantitative real time PCR. The scores for fibrillation, chondrocyte cloning, and proteoglycan depletion were significantly different between advanced and minimal OA cartilage. The advanced OA cartilage had significantly less chondroitin sulfate than the minimal OA cartilage. Osteopontin mRNA expression showed a 3.6-fold increase in advanced compared to minimal OA cartilage. In contrast, the level of mRNA coding for aggrecan, link protein, Bcl-2, Sox9 and MMP-3, -9, -13 were all decreased in advanced compared to minimal cartilage in the majority of the patients studied. Collagen type II mRNA expression displayed a wide-range of variation. A statistically significant correlation was observed both between Bcl-2 and Sox9 mRNA level, and between Bcl-2 and aggrecan mRNA expression. The patient matched comparison of minimal and advanced OA cartilage revealed differences in cellular and tissue characteristics, and changes in gene expression that may be involved in OA progression. In addition, Bcl-2 may also play a role in regulating the expression of aggrecan through Sox9 in vivo as well as in vitro.


Diagnostic efficacy of non-breath-hold magnetic resonance cholangiopancreatography.
Magnetic resonance cholangiopancreatography (MRCP) is an emerging imaging technique for direct visualization of biliary and pancreatic ducts without the need for an invasive procedure, ionizing radiation, or iodine contrast media administration. The purpose of this study was to evaluate the efficacy of non-breath-hold MRCP in depicting normal and diseased biliary and pancreatic ducts. A retrospective analysis of 162 patients who underwent MRCP was performed, and a comparison between MRCP and direct cholangiography was made. The overall accuracy of MRCP in diagnosing malignant and benign pancreaticobiliary diseases was also calculated. MRCP depicted more than three hepatic segments in 99% of patients with dilated intrahepatic ducts and in 63% of patients with nondilated intrahepatic ducts. MRCP demonstrated the main hepatic duct, gallbladder, and cystic duct in 100%, 89%, and 72% of patients, respectively. The dilated extrahepatic duct, nondilated extrahepatic duct, dilated pancreatic duct, and nondilated pancreatic duct were visualized in 100%, 98%, 95%, and 77% of patients, respectively. The accuracy of MRCP in diagnosing hepatolithiasis, cholecystolithiasis, and choledocholithiasis was 96%, 97%, and 96%, respectively. The obstruction levels and characteristics determined by MRCP were in agreement with those determined from direct cholangiography in 98% of malignant obstructions and 89% of benign obstructions. The overall accuracy of a combination of MRCP and conventional magnetic resonance imaging in diagnosing pancreaticobiliary diseases was 81% for malignant diseases, 86% for benign diseases, and 82% for stone diseases. We conclude that non-breath-hold MRCP can reliably depict normal and diseased pancreaticobiliary ducts except for cystic ducts and nondilated pancreatic ducts.


[Eyelid tick bite].
We describe a patient who was bitten on his right upper eyelid by a common canine tick. The tick was impregnated with gasoline prior to being removed. Tick bites are uncommon in ophthalmologic practice. They occur more frequently in rural areas, especially in spring and summer. Correct identification of the problem and treatment is essential. The tick needs to be completely removed in order to avoid the embedding of different parts of the insect. Zoonoses, such as rickettsiosis, arbovirus and paralysis due to ticks must be ruled out.


beta-Endorphin-induced stimulation of central sympathetic outflow: inhibitory modulation by central noradrenergic neurons.
Endorphins, a class of endogenous opioid peptides, produce a diversity of physiological effects attributable to their actions in the central nervous system. We have reported previously the effect of intracisternal (i.c.) administration of synthetic human beta-endorphin to increase plasma catecholamine concentrations, presumably by acting at opioid receptors in the brain to stimulate the central sympathetic outflow to adrenal medulla and peripheral sympathetic nerve terminals. This beta-endorphin-induced increase in plasma catecholamine concentration is dose-dependent, inhibited by naloxone and blocked by bilateral adrenal gland denervation or ganglionic blockade with chlorisondamine. We also provided evidence supporting involvement of central cholinergic, somatostatin and angiotensin II neuronal systems in modulating this beta-endorphin-induced effect. In this study we examined the possibility of central noradrenergic regulation of beta-endorphin-induced catecholamine secretion. Simultaneous i.c. administration of norepinephrine with beta-endorphin blunted the plasma epinephrine response to i.c. beta-endorphin in conscious, unrestrained rats bearing chronic i.a. cannulas. On the other hand, depletion of brain norepinephrine by prior i.c. 6-hydroxydopamine (6-OHDA) treatment potentiated the plasma epinephrine response to i.c. beta-endorphin. In addition, rats pretreated with 6-OHDA appeared supersensitive to the blunting effect of i.c. norepinephrine on beta-endorphin-induced catecholamine secretion. Specifically, i.c. norepinephrine at a dose insufficient to reduce beta-endorphin-induced catecholamine secretion in vehicle-treated rats prevented beta-endorphin-induced epinephrine and norepinephrine secretion in rats whose brains had been depleted of norepinephrine by prior 6-OHDA treatment.(ABSTRACT TRUNCATED AT 250 WORDS)


B-cell-derived lymphokines: regulatory effects on the immune system.
While both regulatory and effector functions within the T cell lineage have been extensively studied, B lymphocytes have been considered merely a source of humoral antibodies until recently. With the exception of antibody feedback, only a limited number of investigations have been designed to explore the possibility that B cells may exert regulatory activity. The purpose of this review is to analyze how B cells can modulate themselves by generating signals (lymphokines), which under certain circumstances may initiate a chain of regulatory events. Only factors exclusively produced by B cells are considered. While, for the most part, this review is dedicated to recent studies on nonimmunoglobulin B cell factors, the role played by Ig molecules as immunoregulatory linker molecules, independently of their antigenic specificity, will be briefly examined. Furthermore, some basic questions regarding the biological significance of these regulatory mechanisms will be raised.


Synchrotron radiation in radiology: radiology techniques based on synchrotron sources.
The characteristics of synchrotron X-ray sources--quite different from those of conventional sources--are exploited by several new imaging techniques. These techniques expand the capabilities of conventional radiology and find interesting application in special cases. We briefly review the basic principle, applications and limitations of the most important of them: monochromatic mammography, two-wavelength digital subtraction angiography, phase-contrast/edge-enhancement imaging, diffraction-enhanced imaging and microtomography.


Ultrafast transient absorption spectroscopy of doped P3HT films: distinguishing free and trapped polarons.
It is generally presumed that the vast majority of carriers created by chemical doping of semiconducting polymer films are coulombically trapped by the counteranion, with only a small fraction that are free and responsible for the increased conductivity essential for organic electronic applications. At higher doping levels, it is also possible for bipolarons to form, which are expected to be less conductive than single polarons. Unfortunately, there is no simple way to distinguish free polarons, trapped polarons and bipolarons using steady-state spectroscopy. Thus, in this work, we use ultrafast transient absorption spectroscopy to study the dynamics of polarons in 2,3,5,6-tetrafluoro-7,7,8,8-tetracyanoquinodimethane (F4TNCQ)-doped films of poly(3-hexylthiophene-2,5-diyl) (P3HT) as a function of dopant concentration and excitation wavelength. When exciting on the red side of the polaron P1 transition, our transient absorption spectra and kinetics match well with what is expected for free 2-D-delocalized polarons; the measurements are not consistent with a recent theory of doped conjugated polymer electronic structure that suggests that the half-filled state lies deeper in the conduction band rather than in the bandgap. As we tune the excitation wavelength to the blue, our measurements reveal an increasing amount of slower transient kinetics that are consistent with the presence of coulombically-trapped polarons rather than bipolarons. Taking advantage of their distinct ultrafast relaxation kinetics as a type of action spectroscopy, we are able to extract the steady-state absorption spectra of free and trapped polarons as a function of dopant concentration. By comparing the results to theoretical models, we determine that in F4TCNQ-doped P3HT films, trapped polarons sit ∼0.4 nm away from the anion while free polarons reside between 0.7 and 0.9 nm from the counteranion. Perhaps counterintuitively, the ratio of trapped to free polarons increases at higher doping levels, an observation that is consistent with a plateau in the concentration-dependent conductivity of F4TCNQ-doped P3HT films.


Quantitative visualization of photosynthetic pigments in tea leaves based on Raman spectroscopy and calibration model transfer.
Photosynthetic pigments participating in the absorption, transformation and transfer of light energy play a very important role in plant growth. While, the spatial distribution of foliar pigments is an important indicator of environmental stress, such as pests, diseases and heavy metal stress. In this paper, in situ quantitative visualization of chlorophyll and carotenoid was realized by combining the Raman spectroscopy with calibration model transfer, and a laboratory Raman spectral model was successfully extended to a portable field spectral measurement. Firstly, a nondestructive and fast model for determination of chlorophyll and carotenoid in tea leaf was established based on confocal micro-Raman spectrometer in the laboratory. Then the spectral model was extended to a real-time foliar map scanning spectra of a field portable Raman spectrometer through calibration model transfer, and the spectral variation between the confocal micro-Raman spectrometer in the laboratory and the portable Raman spectrometer were effectively corrected by the direct standardization (DS) algorithm. The portable map scanning Raman spectra of the tea leaves after the model transfer were got into the established quantitative determination model to predict the concentration of photosynthetic pigments at each pixel of the tea leaves. The predicted photosynthetic pigments concentration of each pixel was imaged to illustrate the distribution map of foliar pigments. Statistical analysis showed that the predicted pigment contents were highly correlated with the real contents. It can be concluded that the Raman spectroscopy was applicable for in situ, non-destructive and rapid quantitative detecting and imaging of photosynthetic pigment concentration in tea leaves, and the spectral detection model established based on the laboratory Raman spectrometer can be applied to a portable field spectrometer for quantitatively imaging of the foliar pigments.


Accuracy of automated identification of delayed diagnosis of pediatric appendicitis and sepsis in the ED.
Delayed diagnoses of serious emergency conditions can lead to morbidity in children, but are challenging to identify and measure. We developed and piloted an automated tool for identifying delayed diagnosis of two serious conditions commonly seen in the ED using administrative data. We identified cases with a final diagnosis of appendicitis or sepsis in a freestanding children's hospital from 2008 to 2018, with any hospital ED encounter within the preceding 7 days. Two investigators reviewed a subset of these cases using the electronic health records (EHR) to determine if there was a delayed diagnosis and interrater reliability was assessed using the intraclass correlation coefficient (ICC). An automated tool was applied to the same cases to assess its positive predictive value (PPV) to identify those with a delayed diagnosis, using the manual chart review as the gold standard. The tool used number of days since visit, presence of a related diagnosis on the initial visit, and whether or not the patient was discharged. Previous ED encounters preceded 91/3703 (2.5%) appendicitis cases and 159/1754 (9.1%) sepsis cases; 78 cases of each were sampled for review. In manual review, 73.4% and 22.8% were thought to have delayed diagnoses; reviewer agreement was excellent (appendicitis ICC 0.77, 95% CI 0.62 to 0.86 and sepsis ICC 0.77, 95% CI 0.43 to 0.89). The PPVs of the automated tool for determination of delayed diagnosis for appendicitis within 1, 3 or 7 days were 96.2%, 95.1% and 93.6%, respectively. For sepsis, the PPVs were 71.4%, 63.6% and 41.2% within 1, 3 or 7 days, respectively. This automated tool performed well compared with expert EHR review. Performance was stronger for appendicitis. Further tool refinement could improve performance.


Use of ferrous fumarate to fortify foods for infants and young children.
Ferrous fumarate is currently recommended for use in the fortification of foods for infants and young children. This recommendation is based on the compound's good sensory properties and on results from isotope studies in adults that reported similar iron absorption values for ferrous fumarate and ferrous sulphate (relative bioavailability [RBV] of ferrous fumarate, 100). However, later isotope studies conducted on both iron-replete and iron-deficient young children found that iron absorption from ferrous fumarate was only about 30% of that achieved from ferrous sulphate (RBV, 30). The reasons for the differences observed in adults compared with children are unclear but could be related to the following factors: lower iron status in children resulting in greater iron absorption via upregulation from ferrous sulphate but not from ferrous fumarate; reduced gastric acid secretion in children leading to retarded dissolution of ferrous fumarate; or an influence of added ascorbic acid on RBV. Ferrous fumarate-fortified complementary foods have been demonstrated to improve iron status in iron-deficient infants and, more recently, to prevent iron deficiency equally as well as ferrous sulphate in iron-replete infants. However, current evidence indicates that iron-deficient infants and young children may absorb iron from ferrous fumarate less well than iron from ferrous sulfate and that, for equivalent efficacy, complementary foods targeted at such infants and young children should contain more iron in the form of fumarate.


Smartphone Monitoring of Mood Instability in Young Depressed Patients: A Latent-class Analyses.
This study captured daily and weekly mood ratings using a smartphone from bipolar disorder (BD) and unipolar major depression disorder (MDD) subjects at high (HRMDD) and low risk (LRMDD) for developing Bipolar Disorder (BD) and healthy controls (HC). 40 subjects (18 - 30 yr) (6 BD, 13 HRMDD, 16 LRMDD and 5 HC) were studied and a total of 2401 daily and 744 weekly ratings were collected. HRMDD and LRMDD subjects were naturalistically treated with antidepressants. We investigate if latent-class analyses of ratings can detect mood instability among MDD and BD groups. Our analyses revealed four underlying mood states correlating with clinical mood states. There was a trend for greater number of state changes in BD and HRMDD subjects compared to LRMDD and HC groups. Smartphone ratings may adequately capture mood instability in BD subjects and at risk HRMDD subjects and offers a prudent way for monitoring development of serious manic symptoms.


Ultrasensitive coincidence fluorescence detection of single DNA molecules.
We have detected individual DNA molecules labeled with two different fluorophores in solution by using two-color excitation and detection of coincidence fluorescence bursts. The confocal volumes of the two excitation lasers were carefully matched so that the volume overlap was 30% of the total confocal volume illuminated. This method greatly reduces the level of background fluorescence and, hence, extends the sensitivity of single molecule detection down to 50 fM. At these concentrations, the dual-labeled DNA is detectable in the presence of a 1000-fold excess of single-fluorophore-labeled DNA. We demonstrate that we can detect 100 fM dual-labeled DNA diluted in 1 microM unlabeled DNA, which was not possible with single color detection. This method can be used to detect rare molecules in complex mixtures.


Localization of the Fanconi anemia complementation group D gene to a 200-kb region on chromosome 3p25.3.
Fanconi anemia (FA) is a rare autosomal recessive disease manifested by bone-marrow failure and an elevated incidence of cancer. Cells taken from patients exhibit spontaneous chromosomal breaks and rearrangements. These breaks and rearrangements are greatly elevated by treatment of FA cells with the use of DNA cross-linking agents. The FA complementation group D gene (FANCD) has previously been localized to chromosome 3p22-26, by use of microcell-mediated chromosome transfer. Here we describe the use of noncomplemented microcell hybrids to identify small overlapping deletions that narrow the FANCD critical region. A 1.2-Mb bacterial-artificial-chromosome (BAC)/P1 contig was constructed, bounded by the marker D3S3691 distally and by the gene ATP2B2 proximally. The contig contains at least 36 genes, including the oxytocin receptor (OXTR), hOGG1, the von Hippel-Lindau tumor-suppressor gene (VHL), and IRAK-2. Both hOGG1 and IRAK-2 were excluded as candidates for FANCD. BACs were then used as probes for FISH analyses, to map the extent of the deletions in four of the noncomplemented microcell hybrid cell lines. A narrow region of common overlapping deletions limits the FANCD critical region to approximately 200 kb. The three candidate genes in this region are TIGR-A004X28, SGC34603, and AA609512.