Giardiasis outbreaks in the United States, 1971-2011.
Giardia intestinalis is the leading parasitic aetiology of human enteric infections in the United States, with an estimated 1·2 million cases occurring annually. To better understand transmission, we analysed data on all giardiasis outbreaks reported to the Centers for Disease Control and Prevention for 1971-2011. The 242 outbreaks, affecting ~41 000 persons, resulted from waterborne (74·8%), foodborne (15·7%), person-to-person (2·5%), and animal contact (1·2%) transmission. Most (74·6%) waterborne outbreaks were associated with drinking water, followed by recreational water (18·2%). Problems with water treatment, untreated groundwater, and distribution systems were identified most often during drinking water-associated outbreak investigations; problems with water treatment declined after the 1980s. Most recreational water-associated outbreaks were linked to treated swimming venues, with pools and wading pools implicated most often. Produce was implicated most often in foodborne outbreaks. Additionally, foods were most commonly prepared in a restaurant and contaminated by a food handler. Lessons learned from examining patterns in outbreaks over time can help prevent future disease. Groundwater and distribution system vulnerabilities, inadequate pool disinfection, fruit and vegetable contamination, and poor food handler hygiene are promising targets for giardiasis prevention measures.


N-nitroso compounds: detection in ambient air.
By use of a new, highly selective detection technique for N-nitroso compounds, which is sensitive to picogram quantities and which is based on the catalytic cleavage of the N-NO bond and the subsequent detection of the nitrosyl radical, dimethylnitrosamine has been found in concentrations of 0.02 to 0.96 part per billion in three out of five air samples from Baltimore, Maryland, and 0.014 to 0.051 part per billion in five out of six air samples from Belle, West Virginia. The sensitivity of the analytical procedures used was 1 part in 10(12). The presence of dimethylnitrosamine has been confirmed by using the new detector in conjunction with both a gas-liquid chromatograph and a high-performance liquid chromatograph. In addition, between one and three as yet unidentified N-nitroso compounds were detected in both cities. N-Nitroso compounds were not found in air samples from Philadelphia, Pennsylvania; Wilmington, Delaware; and Waltham, Massachusetts.


Building programmable jigsaw puzzles with RNA.
One challenge in supramolecular chemistry is the design of versatile, self-assembling building blocks to attain total control of arrangement of matter at a molecular level. We have achieved reliable prediction and design of the three-dimensional structure of artificial RNA building blocks to generate molecular jigsaw puzzle units called tectosquares. They can be programmed with control over their geometry, topology, directionality, and addressability to algorithmically self-assemble into a variety of complex nanoscopic fabrics with predefined periodic and aperiodic patterns and finite dimensions. This work emphasizes the modular and hierarchical characteristics of RNA by showing that small RNA structural motifs can code the precise topology of large molecular architectures. It demonstrates that fully addressable materials based on RNA can be synthesized and provides insights into self-assembly processes involving large populations of RNA molecules.


Structure, dynamics, and specificity of endoglucanase D from Clostridium cellulovorans.
The enzymatic degradation of cellulose is a critical step in the biological conversion of plant biomass into an abundant renewable energy source. An understanding of the structural and dynamic features that cellulases utilize to bind a single strand of crystalline cellulose and hydrolyze the β-1,4-glycosidic bonds of cellulose to produce fermentable sugars would greatly facilitate the engineering of improved cellulases for the large-scale conversion of plant biomass. Endoglucanase D (EngD) from Clostridium cellulovorans is a modular enzyme comprising an N-terminal catalytic domain and a C-terminal carbohydrate-binding module, which is attached via a flexible linker. Here, we present the 2.1-Å-resolution crystal structures of full-length EngD with and without cellotriose bound, solution small-angle X-ray scattering (SAXS) studies of the full-length enzyme, the characterization of the active cleft glucose binding subsites, and substrate specificity of EngD on soluble and insoluble polymeric carbohydrates. SAXS data support a model in which the linker is flexible, allowing EngD to adopt an extended conformation in solution. The cellotriose-bound EngD structure revealed an extended active-site cleft that contains seven glucose-binding subsites, but unlike the majority of structurally determined endocellulases, the active-site cleft of EngD is partially enclosed by Trp162 and Tyr232. EngD variants, which lack Trp162, showed a significant reduction in activity and an alteration in the distribution of cellohexaose degradation products, suggesting that Trp162 plays a direct role in substrate binding.


Wireless amplified nuclear MR detector (WAND) for high-spatial-resolution MR imaging of internal organs: preclinical demonstration in a rodent model.
To assess the feasibility of imaging deep-lying internal organs at high spatial resolution by imaging kidney glomeruli in a rodent model with use of a newly developed, wireless amplified nuclear magnetic resonance (MR) detector. This study was approved by the Animal Care and Use Committee at the National Institutes of Health/National Institute of Neurologic Disorder and Stroke. As a preclinical demonstration of this new detection technology, five different millimeter-scale wireless amplified nuclear MR detectors configured as double frequency resonators were chronically implanted on the medial surface of the kidney in five Sprague-Dawley rats for MR imaging at 11.7 T. Among these rats, two were administered gadopentetate dimeglumine to visualize renal tubules on T1-weighted gradient-refocused echo (GRE) images, two were administered cationized ferritin to visualize glomeruli on T2*-weighted GRE images, and the remaining rat was administered both gadopentetate dimeglumine and cationized ferritin to visualize the interleaved pattern of renal tubules and glomeruli. The image intensity in each pixel was compared with the local tissue signal intensity average to identify regions of hyper- or hypointensity. T1-weighted images with 70-μm in-plane resolution and 200-μm section thickness were obtained within 3.2 minutes to image renal tubules, and T2*-weighted images of the same resolution were obtained within 5.8 minutes to image the glomeruli. Hyperintensity from gadopentetate dimeglumine enabled visualization of renal tubules, and hypointensity from cationic ferritin enabled visualization of the glomeruli. High-spatial-resolution images have been obtained to observe kidney microstructures in vivo with a wireless amplified nuclear MR detector.


High-risk human papillomavirus infection in low risk women: incidence, patient characteristics, and clinical meaning for cervical cancer.
To investigate the incidence of high-risk human papillomavirus (HPV) infection and its clinical meaning. Total 28,339 women attending our hospital for routine gynecologic care underwent Papanicolaou test (PAP test) and high-risk HPV tests. Biopsies were taken from some women and their results were compared. The prevalence of high-risk HPV infection is 24.15%. And the women aged 20-29 years had the highest prevalence (32.3%) compared to 30-70 years (P<0.05). Of the 28,339 women, 1369 (4.83%) had positive PAP test (ASCUS, LSIL, HSIL). Of the 1369 PAP-positive patients, only 16 (1.17%) were negative for HPV test. Of the 1353 patients positive on both tests, 510 (37.7%) had lesions higher than CINII on histology. Of the 1,611 patients who underwent biopsies, 350 underwent the loop electrical excision procedure, with 339 (96%) being positive for HPV test, including 16 with CINI, 48 with CINII/III, 74 with CIS, and 16 with cervical cancer. HPV test had a positive predictive value of 40.7% and a negative predictive value of 100% for higher than CINII. Although HPV test has a burden of cost, considering its high negative predictive value, HPV test should be considered for more useful screening test.


Species-specific, P450- and sulfotransferase-mediated novel ring contraction of a naphthyridine-N-oxide compound in cynomolgus monkey.
BMS-A78277 (1) is a 5,10-dihydrobenzo[beta][1,8]naphthyridine-N-oxide compound that resides in a class of novel non-nucleoside reverse transcriptase inhibitors (NNRTIs), displaying improved activity against clinically relevant mutants of HIV-1 and possessing pharmacokinetic profiles amenable to once-daily dosing. In the course of investigating the nonclinical metabolism of 1, a circulating metabolite specific to the cynomolgus monkey was identified and subsequently characterized as the carboxyindole metabolite 2. The present investigation describes the biotransformation of this NNRTI in cynomolgus monkey, one which results in a net ring contraction of 1. The use of mass spectrometry and high field NMR analysis aided in the structural characterization of metabolite 2, the source of which originated from the urine and bile of cynomolgus monkeys receiving oral doses of 1. Preparation of a synthetic standard of 2 not only provided ultimate structural confirmation but also afforded ample material for biological testing. The metabolism of 1 was investigated in monkey hepatocytes and hepatic subcellular fractions. While microsomes were incapable of generating metabolite 2, incubation of 1 in monkey S9 fractions as well as hepatocytes resulted in measurable levels of the carboxyindole metabolite. Consequently, incubation of 1 in monkey hepatocytes, which were suspended in media containing (18)O-labeled water, resulted in the incorporation of (18)O into the carboxyindole metabolite, 2. These data implicate a mechanism involving the bioactivation of 1 to an electrophilic intermediate that upon hydrolysis undergoes a concerted ring contraction, resulting in the formation of 2. Previously confined to discussions regarding the metabolism of natural products and select aliphatic heterocycles, the present investigation extends the discussion of metabolism-mediated ring contraction to aromatics such as the present naphthyridine compound, 1.


Overexpression of Cyclin E1 or Cdc25A leads to replication stress, mitotic aberrancies, and increased sensitivity to replication checkpoint inhibitors.
Oncogene-induced replication stress, for instance as a result of Cyclin E1 overexpression, causes genomic instability and has been linked to tumorigenesis. To survive high levels of replication stress, tumors depend on pathways to deal with these DNA lesions, which represent a therapeutically actionable vulnerability. We aimed to uncover the consequences of Cyclin E1 or Cdc25A overexpression on replication kinetics, mitotic progression, and the sensitivity to inhibitors of the WEE1 and ATR replication checkpoint kinases. We modeled oncogene-induced replication stress using inducible expression of Cyclin E1 or Cdc25A in non-transformed RPE-1 cells, either in a TP53 wild-type or TP53-mutant background. DNA fiber analysis showed Cyclin E1 or Cdc25A overexpression to slow replication speed. The resulting replication-derived DNA lesions were transmitted into mitosis causing chromosome segregation defects. Single cell sequencing revealed that replication stress and mitotic defects upon Cyclin E1 or Cdc25A overexpression resulted in genomic instability. ATR or WEE1 inhibition exacerbated the mitotic aberrancies induced by Cyclin E1 or Cdc25A overexpression, and caused cytotoxicity. Both these phenotypes were exacerbated upon p53 inactivation. Conversely, downregulation of Cyclin E1 rescued both replication kinetics, as well as sensitivity to ATR and WEE1 inhibitors. Taken together, Cyclin E1 or Cdc25A-induced replication stress leads to mitotic segregation defects and genomic instability. These mitotic defects are exacerbated by inhibition of ATR or WEE1 and therefore point to mitotic catastrophe as an underlying mechanism. Importantly, our data suggest that Cyclin E1 overexpression can be used to select patients for treatment with replication checkpoint inhibitors.


Tumor necrosis factor alpha in cerebrospinal fluid during bacterial, but not viral, meningitis. Evaluation in murine model infections and in patients.
To evaluate the potential role of cachectin/TNF-alpha in the pathogenesis of bacterial and viral meningitis, concentrations and kinetics of TNF-alpha were determined in cerebrospinal fluid (CSF). After intracerebral, but not systemic, infection with Listeria monocytogenes in mice, TNF-alpha was detected as early as 3 h after infection reaching maximum titers after 24 h. However, TNF-alpha was not found in serum during the course of Listeria infection. In contrast to bacterial meningitis, no TNF-alpha was detected at any time in CSF of mice suffering from severe lymphocytic choriomeningitis induced by intracerebral infection with lymphocytic choriomeningitis virus. This difference is striking since both model infections led to a massive infiltration of polymorphonuclear and mononuclear leukocytes into the meninges and CSF. The results found for the two model infections were paralleled by findings in humans; CSF from three out of three patients with bacterial meningitis examined during the first day of hospitalization showed significant levels of TNF-alpha; none of the CSF obtained later than 3 d after hospitalization was positive. In addition, similarly to what was found in mice with viral meningitis, zero out of seven patients with viral meningitis had detectable TNF-alpha in CSF.


Encephalitogenic T cell clones with variant receptor specificity.
We explored antigenic differences between guinea pig (GP)-basic protein (BP), rat (Rt)-BP, and respective peptides from the encephalitogenic region for Lewis rats by comparing the fine specificity of T lymphocyte lines and clones selected from animals primed with these Ag. Encephalitogenic T cell lines specific for GP-BP or Rt-BP predictably recognized the corresponding 72-89 and to a lesser degree the 72-84 (S55S) amino acid sequence. T cell lines selected from rats primed with GP-S55S responded preferentially to GP-S55S compared to other peptides. A T cell line raised to Rt-S55S, however, initially recognized the S55S and S72-89 peptides but were nearly unresponsive to the intact GP-BP or Rt-BP. T cell clones selected from the Rt-S55S line at that point had two distinct patterns of response: clones that recognized both of the BP and the S55S peptides adoptively transferred delayed-type hypersensitivity and experimental autoimmune encephalomyelitis. These clones also recognized residues 69-81 (S67) but not peptide S75-89. In contrast, T cell clones that responded only to synthetic peptides GP-S55S and Rt-S55S but not to the parent BP adoptively transferred delayed-type hypersensitivity but not disease in Lewis rats. The same clones failed to respond to either the S67 or the S75-89 sequences. These results demonstrate that the encephalitogenic Rt-S55S sequence houses a minimum of two T cell epitopes with differing specificities and functions. One epitope is immuno-dominant and resembles the encephalitogenic region of the intact BP molecule. The second non-encephalitogenic epitope is restricted to the S55S sequences and is not shared by the parent BP, the S67, or the S75-89 sequences. Both types of Rt-S55S-specific clones differ in fine specificity from encephalitogenic clones selected from GP-BP immunized rats, thus indicating that uniformity of T cell recognition of the encephalitogenic epitope is not an absolute condition for T cells to be encephalitogenic.


Double-blind randomized multicentre trial of acamprosate in maintaining abstinence from alcohol.
A prospective placebo-controlled, randomized double-blind study of Acamprosate at two dose levels in alcohol-dependent patients followed up for 12 months was performed. After detoxification, each of the 538 patients included was randomly assigned to one of three groups: 177 patients received placebo, 188 received Acamprosate at 1.3 g/day (low dose group) and 173 received 2.0 g/day (high dose group) for 12 months. This was followed by a single blind 6 month period on placebo. The patients' mean age was 43.2 +/- 8.6 years. Their mean daily alcohol intake was high (nearly 200 g/day) and of long duration (9.5 +/- 7.1 years). Abstinence figures followed the order high dose > low dose > placebo. The difference was significant at 6 months (P < or = 0.02) but not at 12 months (P = 0.096). The number of days of continuous abstinence after detoxification was 153 +/- 197 for the high-dose group versus 102 +/- 165 for the placebo group (P = 0.005), with the lose-dose group reporting 135 +/- 189 days. Clinic attendance was significantly better in the Acamprosate groups than in the placebo group at 6 months (P = 0.002) and 12 months (P = 0.005). During the 6-month post-treatment period, no increased relapse rate or residual drug effect was observed. The side effect profile for Acamprosate was good compared with controls with only diarrhoea being reported more frequently (P < 0.01). This study confirms the pharmacological efficacy of Acamprosate and its good acceptability. As an adjunct to psychotherapy, this study supports the inclusion of Acamprosate in a strategy for treating alcoholism.


Radical pancreaticoduodenectomy for benign disease.
Whipple's procedure is the treatment of choice for pancreatic and periampullary malignancies. Preoperative histological confirmation of malignancy is frequently unavailable and some patients will subsequently be found to have benign disease. Here, we review our experience with Whipple's procedure for patients ultimately proven to have benign disease. The medical records of all patients who underwent Whipple's procedure during a 15-year period (1987-2002) were reviewed; 112 patients underwent the procedure for suspected malignancy. In eight cases, the final histology was benign (7.1%). One additional patient was known to have benign disease at resection. The mean age was 50 years (range: 30-75). The major presenting features included jaundice (five), pain (two), gastric outlet obstruction (one), and recurrent gastrointestinal haemorrhage (one). Investigations included ultrasound (eight), computerised tomography (eight), endoscopic retrograde cholangiopancreatography (seven; of these, four patients had a stent inserted and three patients had sampling for cytology), and endoscopic ultrasound (two). The pathological diagnosis included benign biliary stricture (two), chronic pancreatitis (two), choledochal cyst (one), inflammatory pseudotumour (one), cystic duodenal wall dysplasia (one), duodenal angiodysplasia (one), and granular cell neoplasm (one). There was no operative mortality. Morbidity included intra-abdominal collection (one), anastomotic leak (one), liver abscess (one), and myocardial infarction (one). All patients remain alive and well at mean follow-up of 41 months. Despite recent advances in diagnostic imaging, 8% of the patients undergoing Whipple's procedure had benign disease. A range of unusual pathological entities can mimic malignancy. Accurate preoperative histological diagnosis may have allowed a less radical operation to be performed. Endoscopic ultrasound-guided fine needle aspirate (EUS-FNA) may reduce the need for Whipple's operation in benign pancreaticobiliary disease in the future.


Radioiodine post-surgical remnant ablation in patients with differentiated thyroid cancer: news from the last 10 years.
Due to the growing incidence of differentiated thyroid carcinoma (DTC) and in particular of small papillary thyroid cancer observed in the last few decades, the indications, the activity of radioiodine (131I) to be administered, and the efficacy of post surgical thyroid 131I remnant ablation (RRA) have been widely discussed. In the last 10 years, the use of recombinant human TSH (rhTSH) or thyroid hormone withdrawal (THW) to stimulate the 131I remnant uptake has also interested many authors. The general agreement is that small (≤1 cm) intrathyroidal unifocal DTC with a favorable histology and no node metastases should not be submitted to RRA because of the low risk of relapse and cancer specific mortality. Conversely, RRA is indicated in patients with a higher risk level since it seems to reduce recurrence rates and mortality. The recent demonstration that the RRA preparation with rhTSH is as effective as THW using either high (100 mCi) or low (30 mCi) 131I activities suggests that rhTSH preparation and low activity of 131I should be considered as the standard of care for both low- and intermediate-risk DTC patients in the near future. Moreover, the use of low 131I activities and rhTSH reduces whole body radiation exposure and improves the quality of life which are very important advantages for DTC patients.


DAA treatment failures in a low-resource setting with a high burden of hepatitis C infections: a case series.
Globally, 58 million people are living with hepatitis C virus (HCV) infection and 1.5 million new patients are infected every year. The advent of direct acting antivirals (DAAs) has revolutionized the treatment of HCV, opening the door to the ambitious World Health Organization HCV infection elimination strategy by 2030. However, emerging resistance to DAAs could jeopardize any hope of achieving these targets. We discuss a series of 18 patients within a resource-limited setting, who after failing standard sofosbuvir-daclatasvir-based regimen also failed to respond to advanced pan-genotypic treatment regimens, i.e. sofosbuvir-velpatasvir, sofosbuvir-velpatasvir-ribavirin and sofosbuvir-velpatasvir-voxilaprevir. To avoid the spread of refractory HCV strains within the existing epidemic, we call for increased attention and research regarding patients failing treatment on standard pan-genotypic regimens and the spread of HCV-resistant strains within the communities.


Falstatin, a cysteine protease inhibitor of Plasmodium falciparum, facilitates erythrocyte invasion.
Erythrocytic malaria parasites utilize proteases for a number of cellular processes, including hydrolysis of hemoglobin, rupture of erythrocytes by mature schizonts, and subsequent invasion of erythrocytes by free merozoites. However, mechanisms used by malaria parasites to control protease activity have not been established. We report here the identification of an endogenous cysteine protease inhibitor of Plasmodium falciparum, falstatin, based on modest homology with the Trypanosoma cruzi cysteine protease inhibitor chagasin. Falstatin, expressed in Escherichia coli, was a potent reversible inhibitor of the P. falciparum cysteine proteases falcipain-2 and falcipain-3, as well as other parasite- and nonparasite-derived cysteine proteases, but it was a relatively weak inhibitor of the P. falciparum cysteine proteases falcipain-1 and dipeptidyl aminopeptidase 1. Falstatin is present in schizonts, merozoites, and rings, but not in trophozoites, the stage at which the cysteine protease activity of P. falciparum is maximal. Falstatin localizes to the periphery of rings and early schizonts, is diffusely expressed in late schizonts and merozoites, and is released upon the rupture of mature schizonts. Treatment of late schizionts with antibodies that blocked the inhibitory activity of falstatin against native and recombinant falcipain-2 and falcipain-3 dose-dependently decreased the subsequent invasion of erythrocytes by merozoites. These results suggest that P. falciparum requires expression of falstatin to limit proteolysis by certain host or parasite cysteine proteases during erythrocyte invasion. This mechanism of regulation of proteolysis suggests new strategies for the development of antimalarial agents that specifically disrupt erythrocyte invasion.


Maribavir use in practice for cytomegalovirus infection in French transplantation centers.
Maribavir (MBV), a UL97 inhibitor, shows good oral bioavailability, low host cell toxicity, and theoretical benefits to inhibit cross-resistant viruses. We herein examined clinical and virological outcomes of 12 patients, including 3 bone marrow recipients and 9 organ recipients infected with resistant cytomegalovirus (CMV) and treated with MBV during 2011-2012. All received at least 800-mg daily doses. They had developed clinical (12/12) and/or virological (11/12) resistance to CMV infection. Based on a decrease of viral load in blood >1.5 log copies/mL half of them responded to MBV treatment. The individual changes varied from a rapid decrease in viral load (n = 4) to no response (n = 3) with some late response slowly decreasing viremia (n = 3). In 2 cases MBV was used as secondary prophylaxis. No clear parameter emerged as a clinical surrogate for nonresponse to MBV. These results contrast with the lack of efficacy in phase III trials of MBV prophylaxis among stem cell recipients, which were possibly due to low doses or inadequate timing of drug initiation in the study. Additional clinical and surrogate laboratory markers are needed to determine antiviral responses to guide MBV use. Dosage ranging studies might benefit future MBV use.


Expression of potential resistance genes to the English grain aphid, Sitobion avenae, in wheat, Triticum aestivum.
The English grain aphid, Sitobion avenae (F.) (Homoptera: Aphididae), is a dominant and destructive pest in wheat, Triticum estivum L. (Poales: Poaceae), production regions in China and other grain-growing areas worldwide. Patterns of gene expression of the S. avenae-resistant synthetic wheat line 98-10-35, the S. avenae-susceptible line1376, and their hybrid population, and the differences in segments between 98-10-35/1376 F₃ resistant plants and resistant parents of 98-10-35, as well as those between the F3 resistant and susceptible populations, were examined with differential display reverse transcription PCR. The results showed that five patterns of differential expression were detected between the progeny and its resistant parents: 1) The gene was silenced in one of the parents; 2) Special expression showed in the progeny; 3) Expression was consistent with the resistant parents; 4) Up expression showed in the progeny but not in the parents; 5) Down expression showed in the progeny but not in the parents. Paired t-test results were not significant; however, the probability value (0.9158) indicated that gene expression on the RNA level were consistent with resistant bands found in F3 resistant individuals and resistant parents, as well as the F₃ resistant and susceptible populations. For both the F₃ of 98-10-35/1376 and the parents, the total number of amplified bands was 202, with an average of 25.3 per primer. The number of differential bands was 116, with an average of 14.5 per primer amplified and a polymorphism ratio of 56.3%. In the present study, differential expression genes in the resistant line 98-10-35 were all up-regulated. Among them, gene expression of resistant groups in the F₃ population was in agreement with patterns 2, 3, and 4. However, the susceptible line 1376 did not have this gene expression on the RNA level. This pattern is expected to be used to select and analyze target genes from the same F₃ population and the resistant parents. The results suggest that it can be employed as a new method for molecular assisted breeding.


How people interpret conditionals: shifts toward the conditional event.
We investigated how people interpret conditionals and how stable their interpretation is over a long series of trials. Participants were shown the colored patterns on each side of a 6-sided die and were asked how sure they were that a conditional holds of the side landing upward when the die is randomly thrown. Participants were presented with 71 trials consisting of all combinations of binary dimensions of shape (e.g., circles and squares) and color (e.g., blue and red) painted onto the sides of each die. In 2 experiments (N₁ = 66, N₂ = 65), the conditional event was the dominant interpretation, followed by conjunction, and material conditional responses were negligible. In both experiments, the percentage of participants giving a conditional event response increased from around 40% at the beginning of the task to nearly 80% at the end, with most participants shifting from a conjunction interpretation. The shift was moderated by the order of shape and color in each conditional's antecedent and consequent: Participants were more likely to shift if the antecedent referred to a color. In Experiment 2 we collected response times: Conditional event interpretations took longer to process than conjunction interpretations (mean difference = 500 ms). We discuss implications of our results for mental models theory and probabilistic theories of reasoning.


Prospective study of postoperative lumbar epidural hematoma: incidence and risk factors.
Prospective clinical series. To determine the incidence, volume, and extent of postoperative epidural hematoma resulting in thecal sac compression, and to identify risk factors correlated with measured hematoma volumes. Risk factors for postoperative hematoma development have been retrospectively determined in small populations of symptomatic patients. A prospective study of hematoma characteristics and associated risk factors in a consecutive series of patients could significantly enhance our understanding of postoperative hematoma. Preoperative magnetic resonance imaging and clinical data on 13 pre- and intraoperative risk factors were prospectively collected on 50 consecutive patients undergoing lumbar decompression surgery with or without fusion. Postoperative magnetic resonance imagings were performed within 2 to 5 days of surgery. Thecal sac cross-sectional area was calculated at each disc space. Relative thecal sac compression due to hematoma was calculated at all levels where postoperative cross-sectional area was smaller than preoperative. Hematoma volumes were calculated. Multivariate analysis identified risk factors associated with postoperative hematoma volume. After decompression, 58% of patients developed epidural hematoma of sufficient magnitude to compress the thecal sac beyond its preoperative state at one or more levels. None developed new postoperative neurologic deficits. A mean of 1.4 levels were decompressed. Hematoma extended over a mean of 1.9 levels. Maximal thecal sac compression due to hematoma occurred at an adjacent, nondecompressed level in 28% of patients. Multivariate analysis found age greater than 60, multilevel procedures, and preoperative international normalized ratio to be associated with larger hematoma volumes. Lumbar decompression surgery results in a 58% incidence of asymptomatic compressive postoperative epidural hematoma. Adjacent level compression by hematoma occurs in 28% of patients. Advanced age, multilevel procedures, and international normalized ratio are independently associated with postoperative hematoma volume.


Applying a One Health Approach in Global Health and Medicine: Enhancing Involvement of Medical Schools and Global Health Centers.
Multidisciplinary and multisectoral approaches such as One Health and related concepts (e.g., Planetary Health, EcoHealth) offer opportunities for synergistic expertise to address complex health threats. The connections between humans, animals, and the environment necessitate collaboration among sectors to comprehensively understand and reduce risks and consequences on health and wellbeing. One Health approaches are increasingly emphasized for national and international plans and strategies related to zoonotic diseases, food safety, antimicrobial resistance, and climate change, but to date, the possible applications in clinical practice and benefits impacting human health are largely missing. In 2018 the "Application of the One Health Approach to Global Health Centers" conference held at the Albert Einstein College of Medicine convened experts involved in One Health policy and practice. The conference examined issues relevant to One Health approaches, sharing examples of challenges and successes to guide application to medical school curricula and clinical practice for human health. This paper presents a synthesis of conference proceedings, framed around objectives identified from presentations and audience feedback. The following objectives provide opportunities for One Health involvement and benefits for medical schools and global health centers by: 1) Improving One Health resource sharing in global health and medical education; 2) Creating pathways for information flow in clinical medicine and global health practice; 3) Developing innovative partnerships for improved health sector outcomes; and 4) Informing and empowering health through public outreach. These objectives can leverage existing resources to deliver value to additional settings and stakeholders through resource efficiency, more holistic and effective service delivery, and greater ability to manage determinants of poor health status. We encourage medical and global health educators, practitioners, and students to explore entry points where One Health can add value to their work from local to global scale.


Mechanically Induced Chromatin Condensation Requires Cellular Contractility in Mesenchymal Stem Cells.
Mechanical cues play important roles in directing the lineage commitment of mesenchymal stem cells (MSCs). In this study, we explored the molecular mechanisms by which dynamic tensile loading (DL) regulates chromatin organization in this cell type. Our previous findings indicated that the application of DL elicited a rapid increase in chromatin condensation through purinergic signaling mediated by ATP. Here, we show that the rate and degree of condensation depends on the frequency and duration of mechanical loading, and that ATP release requires actomyosin-based cellular contractility. Increases in baseline cellular contractility via the addition of an activator of G-protein coupled receptors (lysophosphatidic acid) induced rapid ATP release, resulting in chromatin condensation independent of loading. Conversely, inhibition of contractility through pretreatment with either a RhoA/Rock inhibitor (Y27632) or MLCK inhibitor (ML7) abrogated ATP release in response to DL, blocking load-induced chromatin condensation. With loading, ATP release occurred very rapidly (within the first 10-20 s), whereas changes in chromatin occurred at a later time point (∼10 min), suggesting a downstream biochemical pathway mediating this process. When cells were pretreated with blockers of the transforming growth factor (TGF) superfamily, purinergic signaling in response to DL was also eliminated. Further analysis showed that this pretreatment decreased contractility, implicating activity in the TGF pathway in the establishment of the baseline contractile state of MSCs (in the absence of exogenous ligands). These data indicate that chromatin condensation in response to DL is regulated through the interplay between purinergic and RhoA/Rock signaling, and that ligandless activity in the TGF/bone morphogenetic proteins signaling pathway contributes to the establishment of baseline contractility in MSCs.


Quantitative maps of protein phosphorylation sites across 14 different rat organs and tissues.
Deregulated cellular signalling is a common hallmark of disease, and delineating tissue phosphoproteomes is key to unravelling the underlying mechanisms. Here we present the broadest tissue catalogue of phosphoproteins to date, covering 31,480 phosphorylation sites on 7,280 proteins quantified across 14 rat organs and tissues. We provide the data set as an easily accessible resource via a web-based database, the CPR PTM Resource. A major fraction of the presented phosphorylation sites are tissue-specific and modulate protein interaction networks that are essential for the function of individual organs. For skeletal muscle, we find that phosphotyrosines are over-represented, which is mainly due to proteins involved in glycogenolysis and muscle contraction, a finding we validate in human skeletal muscle biopsies. Tyrosine phosphorylation is involved in both skeletal and cardiac muscle contraction, whereas glycogenolytic enzymes are tyrosine phosphorylated in skeletal muscle but not in the liver. The presented phosphoproteomic method is simple and rapid, making it applicable for screening of diseased tissue samples.


Rapamycin increases oxidative stress response gene expression in adult stem cells.
Balancing quiescence with proliferation is of paramount importance for adult stem cells in order to avoid hyperproliferation and cell depletion. In some models, stem cell exhaustion may be reversed with the drug rapamycin, which was shown can suppress cellular senescencein vitro and extend lifespan in animals. We hypothesized that rapamycin increases the expression of oxidative stress response genes in adult stem cells, and that these gene activities diminish with age. To test our hypothesis, we exposed mice to rapamycin and then examined the transcriptome of their spermatogonial stem cells (SSCs). Gene expression microarray analysis revealed that numerous oxidative stress response genes were upregulated upon rapamycin treatment, including superoxide dismutase 1, glutathione reductase, and delta-aminolevulinate dehydratase. When we examined the expression of these genes in 55-week-old wild type SSCs, their levels were significantly reduced compared to 3-week-old SSCs, suggesting that their downregulation is coincident with the aging process in adult stem cells. We conclude that rapamycin-induced stimulation of oxidative stress response genes may promote cellular longevity in SSCs, while a decline in gene expression in aged stem cells could reflect the SSCs' diminished potential to alleviate oxidative stress, a hallmark of aging.


Accuracy of radiographer reporting of paediatric brain CT.
Radiographer reporting has been studied for plain films and for ultrasonography, but not in paediatric brain CT in the emergency setting. To study the accuracy of radiographer reporting in paediatric brain CT. We prospectively collected 100 paediatric brain CT examinations. Films were read from hard copies using a prescribed tick sheet. Radiographers with 12 years' and 3 years' experience, respectively, were blinded to the history and were not trained in diagnostic film interpretation. The radiographers' results were compared with those of a consultant radiologist. Three categories were defined: abnormal scans, significant abnormalities and insignificant abnormalities. Both radiographers had an accuracy of 89.5% in reading a scan correctly as abnormal, and radiographer 1 had a sensitivity of 87.8% and radiographer 2 a sensitivity of 96%. Radiographer 1 had an accuracy in detecting a significant abnormality of 75% and radiographer 2 an accuracy of 48.6%, and the sensitivities for this category were 61.6% and 52.9%, respectively. Results for detecting the insignificant abnormalities were poorer. Selected radiographers could play an effective screening role, but lacking the sensitivity required for detecting significant abnormality, they could not be the final diagnostician. We recommend that the study be repeated after both radiographers have received formal training in interpretation of paediatric brain CT.


Changes over time in risk factors for cardiovascular disease and use of lipid-lowering drugs in HIV-infected individuals and impact on myocardial infarction.
Because of the known relationship between exposure to combination antiretroviral therapy and cardiovascular disease (CVD), it has become increasingly important to intervene against risk of CVD in human immunodeficiency virus (HIV)-infected patients. We evaluated changes in risk factors for CVD and the use of lipid-lowering therapy in HIV-infected individuals and assessed the impact of any changes on the incidence of myocardial infarction. The Data Collection on Adverse Events of Anti-HIV Drugs Study is a collaboration of 11 cohorts of HIV-infected patients that included follow-up for 33,389 HIV-infected patients from December 1999 through February 2006. The proportion of patients at high risk of CVD increased from 35.3% during 1999-2000 to 41.3% during 2005-2006. Of 28,985 patients, 2801 (9.7%) initiated lipid-lowering therapy; initiation of lipid-lowering therapy was more common for those with abnormal lipid values and those with traditional risk factors for CVD (male sex, older age, higher body mass index [calculated as the weight in kilograms divided by the square of the height in meters], family and personal history of CVD, and diabetes mellitus). After controlling for these, use of lipid-lowering drugs became relatively less common over time. The incidence of myocardial infarction (0.32 cases per 100 person-years [PY]; 95% confidence interval [CI], 0.29-0.35 cases per 100 PY) appeared to remain stable. However, after controlling for changes in risk factors for CVD, the rate decreased over time (relative rate in 2003 [compared with 1999-2000], 0.73 cases per 100 PY [95% CI, 0.50-1.05 cases per 100 PY]; in 2004, 0.64 cases per 100 PY [95% CI, 0.44-0.94 cases per 100 PY]; in 2005-2006, 0.36 cases per 100 PY [95% CI, 0.24-0.56 cases per 100 PY]). Further adjustment for lipid levels attenuated the relative rates towards unity (relative rate in 2003 [compared with 1999-2000], 1.06 cases per 100 PY [95% CI, 0.63-1.77 cases per 100 PY]; in 2004, 1.02 cases per 100 PY [95% CI, 0.61-1.71 cases per 100 PY]; in 2005-2006, 0.63 cases per 100 PY [95% CI, 0.36-1.09 cases per 100 PY]). Although the CVD risk profile among patients in the Data Collection on Adverse Events of Anti-HIV Drugs Study has decreased since 1999, rates have remained relatively stable, possibly as a result of a more aggressive approach towards managing the risk of CVD.


A Saccharomyces cerevisiae mitochondrial DNA fragment activates Reg1p-dependent glucose-repressible transcription in the nucleus.
As part of an effort to identify random carbon-source-regulated promoters in the Saccharomyces cerevisiae genome, we discovered that a mitochondrial DNA fragment is capable of directing glucose-repressible expression of a reporter gene. This fragment (CR24) originated from the mitochondrial genome adjacent to a transcription initiation site. Mutational analyses identified a GC cluster within the fragment that is required for transcriptional induction. Repression of nuclear CR24-driven transcription required Reg1p, indicating that this mitochondrially derived promoter is a member of a large group of glucose-repressible nuclear promoters that are similarly regulated by Reg1p. In vivo and in vitro binding assays indicated the presence of factors, located within the nucleus and the mitochondria, that bind to the GC cluster. One or more of these factors may provide a regulatory link between the nucleus and mitochondria.


Optical dipole model for photodetection in the near field.
Near-field photodetection optical microscopy (NPOM) is a scanning probe technique that has been developed to perform nanometer-scale optical intensity mapping and spectroscopy. In NPOM a nanometer-scale photodiode detector absorbs power directly as it is scanned in the near field of an illuminated sample surface. A model of photodetection in the near and intermediate fields is presented. A brief review of far-field absorption is given for comparison. Far-field absorption measurements measure the imaginary part of the polarizability to first order. In contrast, photodetection in the near field measures the real part of the polarizability. Other aspects of near-field photodetection are also examined, including contrast mechanisms and lateral resolution. NPOM measurements performed on isolated 300-nm spheres show good agreement with the theory.


Langerhans cell histiocytosis: promises and caveats of targeted therapies in high-risk and CNS disease.
Langerhans cell histiocytosis (LCH) is a rare myeloid neoplasm driven by activating mutations in the MAPK pathway, most commonly BRAF-V600E and MAP2K1. It affects children and adults, with a wide spectrum of clinical presentations ranging from self-limited to multisystem (MS) life-threatening forms. LCH is defined by the accumulation of CD1a+/CD207+ cells in different organs, and patients with liver, spleen, or hematopoietic system involvement have a higher risk of mortality. Patients with neurodegeneration (ND) have devastating outcomes and are resistant to systemic therapies. MS-LCH is treated with risk-adapted therapy, but many patients require multiple salvage regimens that are myelosuppressive and expensive. MAPK inhibitors are increasingly being used, but most patients relapse upon discontinuation of therapy. Here, we review the management of central nervous system disease and how novel cerebrospinal fluid biomarkers might predict patients at high risk of ND who could benefit from early MAPK inhibition. Further, we discuss treatment strategies for refractory/relapsed (R/R) LCH, with a focus on MAPK inhibitors' efficacy and challenges (ie, the unknown): long-term toxicity in children, optimal duration, if they are curative, whether it is safe to combine them with chemotherapy, and their high price tag. Lastly, emerging strategies, such as the new panRAF inhibitor (Day 101) in patients with R/R LCH, ERK1/2 or CSF1R inhibition in patients with MEK1/2 inhibitor resistance, and targeting the microenvironment (checkpoint plus MEK inhibition) or senescent cells (mTOR or BCL-XL inhibitors) in R/R patients, are also examined.


Fibroblast growth factor homologous factors are intracellular signaling proteins.
Fibroblast growth factors (FGFs) mediate cell growth, differentiation, migration, and morphogenesis by binding to the extracellular domain of cell surface receptors, triggering receptor tyrosine phosphorylation and signal transduction [1-5]. FGF homologous factors (FHFs) were discovered within vertebrate DNA sequence databases by virtue of their sequence similarity to FGFs [3, 6, 7], but the mechanism of FHF action has not been reported. We show here that FHF-1 is associated with the MAP kinase (MAPK) scaffold protein Islet-Brain-2 (IB2) [8] in the brain and in specific cell lines. FHF/IB2 interaction is highly specific, as FHFs do not bind to the related scaffold protein IB1(JIP-1b) [9, 10], nor can FGF-1 bind to IB2. We further show that FHFs enable IB2 to recruit a specific MAPK in transfected cells, and our data suggest that the scaffolds IB1 and IB2 have different MAPK specificities. Hence, FHFs are intracellular components of a tissue-specific protein kinase signaling module.


Clinical outcomes after open locked intramedullary nailing of closed femoral shaft fractures for adult patients in a Nigerian Hospital.
Femoral shaft fractures are common injuries in adults. Closed locked intramedullary nailing is the recommended treatment for femoral shaft fractures due to its high union rate. The objective of this study is to determine the outcome of management of closed femoral shaft fractures in adult patients, using open locked intramedullary nailing. This is a prospective study which was carried out on all adult patients aged 16 years and above who presented within 2 weeks of sustaining closed femoral shaft fractures to the accident and emergency unit of a University Teaching Hospital in Nigeria from January 2013 to December 2013. Pathological fractures were excluded from the study. The procedure was carried out using standard techniques, and each patient was followed up for a minimum of 1 year. Forty-three patients were recruited into the study. They had a mean age of 36.9 ± 11.7 years, with a male to female ratio of 2.9:1. The most common cause of closed femoral shaft fractures was road traffic crashes (95.3%), with motorcycle-related injuries found to be the highest type (56.1%). The rate of union in the study was 95.3%. The average time to radiological union was 14.0 ± 1.2 weeks while the mean time to painless full weight bearing was 14.2 ± 1.2 weeks. Among the complications encountered were broken nails (4.7%), infection, loosening of the distal screw, and limb length discrepancy (2.3% each). Using Thoresen's criteria, excellent results were obtained in 93% and poor results in 4.7% of patients. Open locked intramedullary nailing gives excellent clinical outcomes with high union and low complication rates in the management of closed femoral shaft fractures in adult patients.


Substrate conformation modulates aggrecanase (ADAMTS-4) affinity and sequence specificity. Suggestion of a common topological specificity for functionally diverse proteases.
Protease-substrate interactions are governed by a variety of structural features. Although the substrate sequence specificities of numerous proteases have been established, "topological specificities," whereby proteases may be classified based on recognition of distinct three-dimensional structural motifs, have not. The aggrecanase members of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family cleave a variety of proteins but do not seem to possess distinct sequence specificities. In the present study, the topological substrate specificity of ADAMTS-4 (aggrecanase-1) was examined using triple-helical or single-stranded poly(Pro) II helical peptides. Substrate topology modulated the affinity and sequence specificity of ADAMTS-4 with K(m) values indicating a preference for triple-helical structure. In turn, non-catalytic ADAMTS-4 domains were critical for hydrolysis of triple-helical and poly(Pro) II helical substrates. Comparison of ADAMTS-4 with MMP-1 (collagenase 1), MMP-13 (collagenase 3), trypsin, and thermolysin using triple-helical peptide (THP) and single-stranded peptide (SSP) substrates demonstrated that all five proteases possessed efficient "triple-helical peptidase" activity and fell into one of two categories: (k(cat)/K(m))(SSP) > (k(cat)/K(m))(THP) (thermolysin, trypsin, and MMP-13) or (k(cat)/K(m))(THP) > or = (k(cat)/K(m))(SSP) and (K(m))(SSP) > (K(m))(THP) (MMP-1 and ADAMTS-4). Overall these results suggest that topological specificity may be a guiding principle for protease behavior and can be utilized to design specific substrates and inhibitors. The triple-helical and single-stranded poly(Pro) II helical peptides represent the first synthetic substrates successfully designed for aggrecanases.


KDM2B Recruitment of the Polycomb Group Complex, PRC1.1, Requires Cooperation between PCGF1 and BCORL1.
KDM2B recruits H2A-ubiquitinating activity of a non-canonical Polycomb Repression Complex 1 (PRC1.1) to CpG islands, facilitating gene repression. We investigated the molecular basis of recruitment using in vitro assembly assays to identify minimal components, subcomplexes, and domains required for recruitment. A minimal four-component PRC1.1 complex can be assembled by combining two separately isolated subcomplexes: the DNA-binding KDM2B/SKP1 heterodimer and the heterodimer of BCORL1 and PCGF1, a core component of PRC1.1. The crystal structure of the KDM2B/SKP1/BCORL1/PCGF1 complex illustrates the crucial role played by the PCGF1/BCORL1 heterodimer. The BCORL1 PUFD domain positions residues preceding the RAWUL domain of PCGF1 to create an extended interface for interaction with KDM2B, which is unique to the PCGF1-containing PRC1.1 complex. The structure also suggests how KDM2B might simultaneously function in PRC1.1 and an SCF ubiquitin ligase complex and the possible molecular consequences of BCOR PUFD internal tandem duplications found in pediatric kidney and brain tumors.


Implementation of electronic checklists in an oncology medical record: initial clinical experience.
The quality of any medical treatment depends on the accurate processing of multiple complex components of information, with proper delivery to the patient. This is true for radiation oncology, in which treatment delivery is as complex as a surgical procedure but more dependent on hardware and software technology. Uncorrected errors, even if small or infrequent, can result in catastrophic consequences for the patient. We developed electronic checklists (ECLs) within the oncology electronic medical record (EMR) and evaluated their use and report on our initial clinical experience. Using the Mosaiq EMR, we developed checklists within the clinical assessment section. These checklists are based on the process flow of information from one group to another within the clinic and enable the processing, confirmation, and documentation of relevant patient information before the delivery of radiation therapy. The clinical use of the ECL was documented by means of a customized report. Use of ECL has reduced the number of times that physicians were called to the treatment unit. In particular, the ECL has ensured that therapists have a better understanding of the treatment plan before the initiation of treatment. An evaluation of ECL compliance showed that, with additional staff training, > 94% of the records were completed. The ECL can be used to ensure standardization of procedures and documentation that the pretreatment checks have been performed before patient treatment. We believe that the implementation of ECLs will improve patient safety and reduce the likelihood of treatment errors.


Electroconvulsive therapy for self-injurious behaviour in autism spectrum disorders: recognizing catatonia is key.
Self-injurious behaviour (SIB) is a devastating condition frequently encountered in autism spectrum disorders (ASDs) that can lead to dangerous tissue injury and profound psychosocial difficulty. An increasing number of reports over the past decade have demonstrated the swift and well tolerated resolution of intractable SIB with electroconvulsive therapy (ECT) when psychopharmacological and behavioural interventions are ineffective. The current article provides a review of the salient literature, including the conceptualization of repetitive self-injury along the catatonia spectrum, and further clarifies the critical distinction between ECT and contingent electric shock. We searched electronically for literature regarding ECT for self-injurious behaviour from 1982 to present, as the first known report was published in 1982. Eleven reports were identified that presented ECT in the resolution of self-injury in autistic or intellectually disabled patients, and another five reports discussed such in typically developing individuals. These reports and related literature present such self-injury along the spectrum of agitated catatonia, with subsequent implications for ECT. Intractable self-injury remains a significant challenge in ASDs, especially when patients do not respond adequately to behavioural and psychopharmacological interventions. ECT is well tolerated and efficacious treatment for catatonia, and can confer marked reduction in SIB along the agitated catatonia spectrum.


How do medical specialists value their own intercultural communication behaviour? A reflective practice study.
Intercultural communication behaviour of doctors with patients requires specific intercultural communication skills, which do not seem structurally implemented in medical education. It is unclear what motivates doctors to apply intercultural communication skills. We investigated how purposefully medical specialists think they practise intercultural communication and how they reflect on their own communication behaviour. Using reflective practice, 17 medical specialists independently watched two fragments of videotapes of their own outpatient consultations: one with a native patient and one with a non-native patient. They were asked to reflect on their own communication and on challenges they experience in intercultural communication. The interviews were open coded and analysed using thematic network analysis. The participants experienced only little differences in their communication with native and non-native patients. They mainly mentioned generic communication skills, such as listening and checking if the patient understood. Many participants experienced their communication with non-native patients positively. The participants mentioned critical incidences of intercultural communication: language barriers, cultural differences, the presence of an interpreter, the role of the family and the atmosphere. Despite extensive experience in intercultural communication, the participants of this study noticed hardly any differences between their own communication behaviour with native and non-native patients. This could mean that they are unaware that consultations with non-native patients might cause them to communicate differently than with native patients. The reason for this could be that medical specialists lack the skills to reflect on the process of the communication. The participants focused on their generic communication skills rather than on specific intercultural communication skills, which could either indicate their lack of awareness, or demonstrate that practicing generic communication is more important than applying specific intercultural communication. They mentioned well-known critical incidences of ICC: language barriers, cultural differences, the presence of an interpreter, the role of the family and the atmosphere. Nevertheless, they showed a remarkably enthusiastic attitude overall was noteworthy. A strategy to make doctors more aware of their intercultural communication behaviour could be a combination of experiential learning and ICC training, for example a module with reflective practice.


Tumor protein 53-induced nuclear protein 1 is a major mediator of p53 antioxidant function.
p53 exerts its tumor suppressor function mainly through transcriptional induction of target genes involved in several processes, including cell cycle checkpoints, apoptosis, and regulation of cell redox status. p53 antioxidant function is dependent on its transcriptional activity and proceeds by sequential induction of antioxidant and proapoptotic targets. However, none of the thus far renowned p53 targets have proved able to abolish on their own the intracellular reactive oxygen species (ROS) accumulation caused by p53 deficiency, therefore pointing to the existence of other prominent and yet unknown p53 antioxidant targets. Here, we show that TP53INP1 represents such a target. Indeed, TP53INP1 transcript induction on oxidative stress is strictly dependent on p53. Mouse embryonic fibroblasts (MEF) and splenocytes derived from TP53INP1-deficient (inp1(-/-)) mice accumulate intracellular ROS, whereas overexpression of TP53INP1 in p53-deficient MEFs rescues ROS levels to those of p53-proficient cells, indicating that TP53INP1 antioxidant function is p53 independent. Furthermore, accumulation of ROS in inp1(-/-) cells on oxidant challenge is associated with decreased expression of p53 targets p21/Cdkn1a, Sesn2, TAp73, Puma, and Bax. Mutation of p53 Ser(58) (equivalent to human p53 Ser(46)) abrogates transcription of these genes, indicating that TP53INP1-mediated p53 Ser(58) phosphorylation is implicated in this process. In addition, TP53INP1 deficiency results in an antioxidant (N-acetylcysteine)-sensitive acceleration of cell proliferation. Finally, TP53INP1 deficiency increases oxidative stress-related lymphoma incidence and decreases survival of p53(+/-) mice. In conclusion, our data show that TP53INP1 is a major actor of p53-driven oxidative stress response that possesses both a p53-independent intracellular ROS regulatory function and a p53-dependent transcription regulatory function.


Decreased hepatic insulin-like growth factor (IGF)-I and increased IGF binding protein-1 and -2 gene expression in experimental uremia.
The imbalance between normal insulin-like growth factor-I (IGF-I) and markedly increased IGF binding protein (IGFBP) plasma levels plays a pathogenic role for growth retardation and catabolism in children with chronic renal failure. To investigate the mechanism of these alterations, experiments were performed in an experimental model of uremia in rats (5/6 nephrectomy) and in pair-fed and ad libitum-fed sham-operated controls Using a specific solution hybridization/RNase protection assay, we observed a marked reduction of hepatic IGF-I messenger RNA (mRNA) abundance at steady state in uremic animals (37 +/- 5% of control) compared both with pair-fed (65 +/- 10%) and ad libitum-fed controls (100 +/- 11%) (P < 0.001). Reduced IGF-I gene expression was clearly organ-specific; it was most pronounced in liver (significant vs., pair-fed controls) and lung and muscle tissue (significant vs., ad libitum-fed controls); no change was observed in kidney and heart tissue. To determine a potential mechanism of reduced hepatic IGF-I gene expression in uremia, the hepatic GH receptor gene expression in the same experimental animals was analyzed by specific solution hybridization/RNase protection assay. Uremic animals had a 20-30% reduction of hepatic GH receptor mRNA abundance compared with controls. Hepatic GHBP expression in uremia was decreased in parallel. Despite the reduction of hepatic IGF-I mRNA abundance, plasma IGF-I levels in uremia were not different from ad libitum-fed controls. This discrepancy is explained by an increased concentration of IGFBPs in uremic plasma. By RIA, plasma IGFBP-1 levels in uremia were increased 4-fold; by Western immunoblot, plasma IGFBP-2 levels were increased 7-fold and plasma IGFBP-4 levels were increased 2-fold compared with both control groups. Intact IGFBP-3 (M(r), approximately 48 kDa) and low molecular IGFBP-3 fragments were not significantly different among the three groups. By Northern blot analysis, hepatic IGFBP-1 mRNA levels in uremia were 2-fold higher than in controls. IGFBP-2 mRNA abundance in liver tissue was increased 4-fold, whereas in kidney there was a significant reduction of IGFBP-2 mRNA (30% of control). IGFBP-4 mRNA was increased by 50% in kidney but not in liver. Plasma insulin and corticosterone levels were not different among the groups. Our study shows that hepatic IGF-I gene expression was specifically reduced in uremia, partially as the consequence of a reduced hepatic GH receptor gene expression. One of the mechanisms contributing to increased IGFBP levels in uremia is increased hepatic gene expression of IGFBP-1 and IGFBP-2. The imbalance between reduced hepatic IGF-I production and increased hepatic IGFBP-1 and 2 production is likely to play a pathogenic role for catabolism and growth failure in CRF.


Physician-prompting statin therapy intervention improves outcomes in patients with coronary heart disease.
To evaluate the effectiveness of a posthospital discharge intervention that prompted physicians to increase the use and effectiveness of statins (3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors) in patients with coronary heart disease (CHD). Participants were 612 patients with CHD who were admitted to a coronary care unit. The control group (303 patients admitted from October 1-December 31, 1998) received no follow-up intervention. The intervention group (309 patients admitted fromJanuary 1-March 31, 1999) had follow-up letters sent or phone calls made to their primary care physicians with patient-specific recommendations concerning assessment of lipid profiles and statin therapy. Over a 2-year follow-up period, assessment of lipid profiles, use of therapy, and adverse clinical outcomes were compared between the control and intervention groups. At hospital discharge, there was no significant difference in the use of statins between the groups. At each reported follow-up interval, the percentages of patients having lipid profiles measured, being treated with a statin, receiving titrated dosages of a statin, and achieving low-density lipid (LDL) cholesterol goals set by the National Cholesterol Education Program (NCEP) were significantly greater in the intervention group compared with the control group (all p<0.05). At the end of the 2-year follow-up period, nearly three-fourths (72%) of the intervention group were receiving a statin, compared with 43% of the control group. In addition, 55% of the intervention group achieved their NCEP LDL goal, compared with only 10% of the control group. Recurrent myocardial infarction, hospitalization for myocardial ischemia, coronary revascularization, and cardiovascular mortality were significantly reduced in the intervention group compared with the control group (all p<0.05). A relatively simple physician-prompting intervention significantly increased assessment of lipid status, frequency of statin use, achievement of LDL treatment goals, and titration of lipid drug dosages. In addition, the improved use of statins significantly reduced adverse cardiovascular outcomes. This intervention tool should be more broadly applied in patient populations eligible to receive these agents.


Objective measurement of dyskinesia in Parkinson's disease using a force plate.
Clinical investigation of levodopa-induced dyskinesia (LID) in Parkinson's disease (PD) is limited because of lack of objective measurements and no consensus on use of a standard measuring tool. Currently, clinical trials use subject-completed diaries of dyskinesia throughout the day or investigator-administered clinical rating scales. An objective and valid method of measuring LID would reduce bias, variability, and decrease the time and number needed in trials of potential anti-dyskinetic agents. We have investigated using a force plate under standing subjects, which records movement of the center of pressure (CoP) to quantify LID over a levodopa (L-dopa) cycle. Twenty-two PD subjects (15 with LID, 7 without LID) admitted to an inpatient research facility had their PD meds withheld overnight, followed by a 2 hours intravenous L-dopa infusion the next day. The root mean squared of the velocity in the anterior-posterior direction (RMSV) derived from an analysis of the CoP, and, the modified Abnormal Involuntary Movement Scale (mAIMS) were performed repeatedly for 6 hours, initially as subjects were OFF before the infusion, through infusion until OFF again. There was a high correlation between the area under the curve (AUC) of the mAIMS and the RMSV within and between subjects. As a measure of LID, RMSV had excellent validity and reliability between subjects, and using a force plate was feasible. Sensitivity to changes in LID was initially demonstrated but should be repeated. Thus, CoP recordings on a force plate can objectively quantify LID in PD and may be very useful in clinical trials or other investigations of dyskinesia.


An optimized procedure for fluorescence-activated cell sorting (FACS) isolation of autonomic neural progenitors from visceral organs of fetal mice.
During development neural crest (NC)-derived neuronal progenitors migrate away from the neural tube to form autonomic ganglia in visceral organs like the intestine and lower urinary tract. Both during development and in mature tissues these cells are often widely dispersed throughout tissues so that isolation of discrete populations using methods like laser capture micro-dissection is difficult. They can however be directly visualized by expression of fluorescent reporters driven from regulatory regions of neuron-specific genes like Tyrosine hydroxylase (TH). We describe a method optimized for high yields of viable TH+ neuronal progenitors from fetal mouse visceral tissues, including intestine and lower urogenital tract (LUT), based on dissociation and fluorescence-activated cell sorting (FACS). The Th gene encodes the rate-limiting enzyme for production of catecholamines. Enteric neuronal progenitors begin to express TH during their migration in the fetal intestine and TH is also present in a subset of adult pelvic ganglia neurons . The first appearance of this lineage and the distribution of these neurons in other aspects of the LUT, and their isolation has not been described. Neuronal progenitors expressing TH can be readily visualized by expression of EGFP in mice carrying the transgene construct Tg(Th-EGFP)DJ76Gsat/Mmnc. We imaged expression of this transgene in fetal mice to document the distribution of TH+ cells in the developing LUT at 15.5 days post coitus (dpc), designating the morning of plug detection as 0.5 dpc, and observed that a subset of neuronal progenitors in the coalescing pelvic ganglia express EGFP. To isolate LUT TH+ neuronal progenitors, we optimized methods that were initially used to purify neural crest stem cells from fetal mouse intestine. Prior efforts to isolate NC-derived populations relied upon digestion with a cocktail of collagenase and trypsin to obtain cell suspensions for flow cytometry. In our hands these methods produced cell suspensions from the LUT with relatively low viability. Given the already low incidence of neuronal progenitors in fetal LUT tissues, we set out to optimize dissociation methods such that cell survival in the final dissociates would be increased. We determined that gentle dissociation in Accumax (Innovative Cell Technologies, Inc), manual filtering, and flow sorting at low pressures allowed us to achieve consistently greater survival (>70% of total cells) with subsequent yields of neuronal progenitors sufficient for downstream analysis. The method we describe can be broadly applied to isolate a variety of neuronal populations from either fetal or adult murine tissues.


Plasma adiponectin complexes have distinct biochemical characteristics.
Adipocytes release the secretory protein adiponectin in a number of different higher-order complexes. Once synthesized and assembled in the secretory pathway of the adipocyte, these complexes circulate as biochemically distinct and stable entities with little evidence of interchange between the different forms that include a high-molecular-weight (HMW) species, a hexamer (low-molecular-weight form), and a trimeric form of the complexes. Here, we validate a high-resolution gel filtration method that reproducibly separates the three complexes in recombinant adiponectin and adiponectin from human and murine samples. We demonstrate that the HMW form is prominently reduced in male vs. female subjects and in obese, insulin-resistant vs. lean, insulin-sensitive individuals. A direct comparison of human and mouse adiponectin demonstrates that the trimer is generally more abundant in human serum. Furthermore, when the production of adiponectin is reduced, either by obesity or in mice carrying only a single functional allele of the adiponectin locus, then the amount of the HMW form is selectively reduced in circulation. The complex distribution of adiponectin can be regulated in several ways. Both mouse and human HMW adiponectin are very stable under basic conditions but are exquisitely labile under acidic conditions below pH 7. Murine and human adiponectin HMW forms also display differential susceptibility to the presence of calcium in the buffer. A mutant form of adiponectin unable to bind calcium is less susceptible to changes in calcium concentrations. However, the lack of calcium binding results in a destabilization of the structure. Disulfide bond formation (at position C39) is also important for complex formation. A mutant form of adiponectin lacking C39 prominently forms HMW and trimer but not the low-molecular-weight form. Injection of adiponectin with a fluorescent label reveals that over time, the various complexes do not interconvert in vivo. The stability of adiponectin complexes highlights that the production and secretion of these forms from fat cells has a major influence on the circulating levels of each complex.


Identification of rCop-1, a new member of the CCN protein family, as a negative regulator for cell transformation.
By using a model system for cell transformation mediated by the cooperation of the activated H-ras oncogene and the inactivated p53 tumor suppressor gene, rCop-1 was identified by mRNA differential display as a gene whose expression became lost after cell transformation. Homology analysis indicates that rCop-1 belongs to an emerging cysteine-rich growth regulator family called CCN, which includes connective-tissue growth factor, CYR61, CEF10 (v-src inducible), and the product of the nov proto-oncogene. Unlike the other members of the CCN gene family, rCop-1 is not an immediate-early gene, it lacks the conserved C-terminal domain which was shown to confer both growth-stimulating and heparin-binding activities, and its expression is lost in cells transformed by a variety of mechanisms. Ectopic expression of rCop-1 by retroviral gene transfers led to cell death in a transformation-specific manner. These results suggest that rCop-1 represents a new class of CCN family proteins that have functions opposing those of the previously identified members.


Cranberry components inhibit interleukin-6, interleukin-8, and prostaglandin E production by lipopolysaccharide-activated gingival fibroblasts.
Periodontitis is a chronic inflammatory disease that affects the tooth supporting tissues. Gingival fibroblasts are the most abundant cells in periodontal tissues and participate actively in the host inflammatory response to periodontopathogens, which is known to mediate local tissue destruction in periodontitis. The aim of this study was to investigate the effect of a proanthocyanidin-enriched cranberry fraction, prepared from cranberry juice concentrate, on inflammatory mediator production by gingival fibroblasts stimulated by the lipopolysaccharide (LPS) of Aggregatibacter actinomycetemcomitans. Interleukin (IL)-6, IL-8, and prostaglandin E(2) (PGE(2)) production by fibroblasts treated with the cranberry fraction and stimulated by A. actinomycetemcomitans LPS was evaluated by enzyme-linked immunosorbent assay. Changes induced by A. actinomycetemcomitans LPS and the cranberry fraction in the expression and phosphorylation state of fibroblast intracellular signaling proteins were characterized by antibody microarrays. The LPS-induced IL-6, IL-8, and PGE(2) responses of gingival fibroblasts were inhibited by treatment with the cranberry fraction. This fraction was found to inhibit fibroblast intracellular signaling proteins, a phenomenon that may lead to a down-regulation of activating protein-1 activity. Cranberry components also reduced cyclooxygenase 2 expression. This study suggests that cranberry juice contains molecules with interesting properties for the development of new host-modulating therapeutic strategies in the adjunctive treatment of periodontitis.


Screening of international immigrants, refugees, and adoptees.
Frequently clinicians are faced with screening and providing preventive care to immigrants, refugees, and international adoptees. Evidence-based medicine on which to base screening protocols for these populations is lacking. It is important to review all health and vaccination records of the patient. In addition to acute symptoms, one should inquire about the symptoms of diseases prevalent in the country of origin or transit (e.g., hematuria). Many unexpected pathologic conditions may be detected by a thorough physical examination. If a reliable immunization record is presented, one need not repeat the vaccines or check titers. Remaining vaccines should be administered according to ACIP guidelines, except for certain populations (e.g., adoptees). Routine laboratory screening tests should include CBC with differential, stool for ova and parasites, urinalysis, general chemistry profile, serology for hepatitis B, and tests for HIV and syphilis. A tuberculin skin test should be performed on all immigrants, and a chest radiograph should be obtained for any patient with symptoms or a positive PPD. Lead level, hepatitis C, and TSH should be obtained for all children and most adoptees. In addition, special screening tests (e.g., for malaria, hepatitis C, and STIs) may be indicated in high-risk populations. A more organized screening system that emphasizes evidence-based and population-specific screening protocols and better communication between international, federal, state, and local levels is needed in the United States.


Sexual preference, gender, and blame attributions in adolescent sexual assault.
The study investigated the impact of victim sexual orientation, perpetrator gender, and participant gender on judgements toward a 15-year-old male victim of a depicted sexual assault. One hundred and eight-eight participants (97 male, 91 female) read a hypothetical scenario depicting the sexual assault of a 15-year-old male victim where the victim's sexual orientation and the perpetrator's gender were varied between subjects. Participants then completed a questionnaire assessing their attributions toward both the victim and the perpetrator. Results revealed that male participants blamed the victim more than female participants when the victim was both gay and attacked by a male perpetrator. All participants, regardless of gender, made more positive judgements toward the female as opposed to male perpetrator. Results are discussed in relation to gender role stereotypes and homophobia.


Regulatory flexibility in the Nrf2-mediated stress response is conferred by conformational cycling of the Keap1-Nrf2 protein complex.
The transcription factor NF-E2 p45-related factor 2 (Nrf2), a master regulator of cytoprotective genes, is controlled by dimeric Kelch-like ECH associated protein 1 (Keap1), a substrate adaptor protein for Cullin3/RING-box protein 1 ubiquitin ligase, which normally targets Nrf2 for ubiquitination and degradation but loses this ability in response to electrophiles and oxidants (inducers). By using recombinant proteins and populations of cells, some of the general features of the regulation of Nrf2 by Keap1 have been outlined. However, how the two proteins interact at a single-cell level is presently unknown. We now report the development of a quantitative Förster resonance energy transfer-based system using multiphoton fluorescence lifetime imaging microscopy and its application for investigating the interaction between Nrf2 and Keap1 in single live cells. By using this approach, we found that under homeostatic conditions, the interaction between Keap1 and Nrf2 follows a cycle in which the complex sequentially adopts two distinct conformations: "open," in which Nrf2 interacts with a single molecule of Keap1, followed by "closed," in which Nrf2 binds to both members of the Keap1 dimer. Inducers disrupt this cycle by causing accumulation of the complex in the closed conformation without release of Nrf2. As a consequence, free Keap1 is not regenerated, and newly synthesized Nrf2 is stabilized. On the basis of these findings, we propose a model we have named the "cyclic sequential attachment and regeneration model of Keap1-mediated degradation of Nrf2." This previously unanticipated dynamism allows rapid transcriptional responses to environmental changes and can accommodate multiple modes of regulation.


Despite Buffers, Experimental Forest Clearcuts Impact Amphibian Body Size and Biomass.
Forest buffers are a primary tool used to protect wetland-dependent wildlife. Though implemented widely, buffer efficacy is untested for most amphibian species. Consequently, it remains unclear whether buffers are sufficient for maintaining amphibian populations and if so, how wide buffers should be. We present evidence from a six-year, landscape-scale experiment testing the impacts of clearcutting, buffer width, and hydroperiod on body size and condition and biomass of breeding adults for two amphibian species at 11 vernal pools in the northeastern United States. We randomly assigned treatments (i.e., reference, 100m buffer, 30m buffer) across pools, clearcut to create buffers, and captured all spotted salamanders and wood frogs. Clearcuts strongly and negatively impacted size, condition, and biomass, but wider buffers mitigated effect magnitude and duration. Among recaptured individuals, for example, 30m-treatment salamanders were predicted to be about 9.5 mm shorter than, while 100m-treatment salamanders did not differ in length from, reference-treatment salamanders. Similarly, among recaptured frogs, mean length in the 30m treatment was predicted to decrease by about 1 mm/year, while in the 100m and reference treatments, length was time-invariant. Some, but not all, metrics recovered with time. For example, female new-captured and recaptured salamanders were predicted, respectively and on average, to weigh 4.5 and 7 g less in the 30m versus reference treatment right after the cut. While recaptured-female mass was predicted to recover by 9.5 years post-cut, new-captured-female mass did not recover. Hydroperiod was an important mediator: in the 100m treatment, cutting predominately affected pools that were stressed hydrologically. Overall, salamanders and female frogs were impacted more than male frogs. Our results highlight the importance of individualized metrics like body size, which can reveal sublethal effects and illuminate mechanisms by which habitat disturbance impacts wildlife populations. Individualized metrics thus provide critical insights that complement species occurrence and abundance-based population assessments.


Idiopathic adhesive capsulitis: long-term results of conservative treatment.
Adhesive capsulitis, also termed "frozen shoulder," is controversial by definition and diagnostic criteria that are not sufficiently understood. The clinical course of this condition is considered self-limiting and is divided into three clinical phases. Several treatment methods for adhesive capsulitis have been reported in the literature, none of which has proven superior to others. To evaluate the long-term follow-up of patients with idiopathic adhesive capsulitis who were treated conservatively. We conducted a long-term follow-up (range 5.5-16 years, mean 9.2 years) of 54 patients suffering from idiopathic adhesive capsulitis. All patients were treated with physical therapy and non-steroidal anti-inflammatory drugs. An increased statistically significant improvement (P < 0.00001) was found between the first and last visits to the clinic in all measured movement directions: elevation and external and internal rotation. Conservative treatment (physical therapy and NSAIDs) is a good long-term treatment regimen for idiopathic adhesive capsulitis.


Reduced lung function in a chronic asthma model is associated with prolonged inflammation, but independent of peribronchial fibrosis.
In asthma, mechanisms contributing to chronicity remain to be determined. Recent models of sensitisation with prolonged airway allergen challenges reproduce typical features of chronic asthma. However, the interplay between inflammation, structural changes and lung function is poorly understood. This study was performed to delineate functional, structural and immunological airway changes after cessation of long term challenges to elucidate factors contributing to the development of prolonged lung function changes. Mice sensitised systemically were consecutively challenged intranasally with ovalbumin for two or eight weeks. After the end of challenges, lung function, airway inflammation, features of airway remodelling, local T-cell cytokines and systemic ovalbumin-specific antibodies were monitored. Long term challenges resulted in airway hyperresponsiveness lasting 2 weeks and reduced baseline lung function for 6 weeks after their cessation. In contrast, these changes resolved within one week after short term challenges. Prolonged transforming growth factor beta (TGF-beta)1 production and marked peribronchial fibrosis were only induced by long term challenges. Importantly, fibrosis became apparent only after the onset of lung function changes and outlasted them. Further, long term challenges led to prolonged and intense airway inflammation with marked lymphocytosis, but moderate eosinophilia, sustained IL-5 production and ovalbumin-specific IgG2a antibodies, the latter suggesting a Th1 component to the immune response. In contrast, following short term challenges airway inflammation was dominated by eosinophils and associated with a strong, but transient IL-13 response. Prolonged lung function changes after long term allergen challenges seem to develop and resolve independently of the persistent peribronchial fibrosis. They are more closely associated with intense airway inflammation, marked lymphocytosis, prolonged IL-5 and TGF-beta1 production in the airways and a Th1 immune response.


Second recorded case of human infection by Echinococcus oligarthrus.
The paper reviews a previously published case of hydatid disease in the human heart of a Brazilian person who died of tetanus. Based on present knowledge about the distinguishing characteristics of Echinococcus granulosus, E. vogeli, and E. oligarthrus, it was recognized that the infection was due to E. oligarthrus, mainly based the morphologic features of the hooklets of the protoscolex. This is the second human infection due to E. oligarthrus and the first showing wall features of cysts. Therefore, some human infections of polycystic hydatid disease observed outside the range of the bush dog, the only definitive host of E. vogeli (Panama to Northern Argentina), may be due to E. oligarthrus rather than to E. vogeli.


A seed-specific Brassica napus oleosin promoter interacts with a G-box-specific protein and may be bi-directional.
In Brassica napus, oleosins are expressed at high levels in the seed during the latter stages of embryo development. The cis-acting regulatory properties of an 872 bp promoter fragment of a B. napus oleosin gene were examined by analysis of beta-glucuronidase (GUS) expression in transgenic tobacco plants containing an oleosin promoter-GUS transcriptional fusion. The reporter gene was expressed at high levels only in seeds, specifically in embryo and endosperm tissue and regulated throughout seed development. These data demonstrate that oleosin gene transcription is regulated in a tissue-specific and temporally regulated manner and clearly indicate that oleosin protein expression is co-ordinated primarily at the transcriptional level. Oleosin mRNA was shown to be abscisic acid (ABA) inducible and an ABA-response element in the oleosin promoter was shown to be bound by a protein factor in a sequence-specific manner. Sequence analysis of the oleosin promoter has identified several other putative cis-acting sequences which may direct oleosin gene expression. The presence of a large open reading frame in the bottom strand of the oleosin promoter (ORF2) which encodes a polypeptide similar to the ethylene-induced E4 gene of tomato is reported. A PCR-generated DNA probe containing the ORF2 sequence hybridised with a 1.4 kb transcript in total RNA extracts of a variety of tissues, including leaves and germinated seed cotyledons. This finding suggests that the oleosin gene promoter directs transcription in both directions. It is the first report of a bi-directional nuclear gene promoter in plants.


Feeding behavior and performance of lambs are influenced by flavor diversity.
This study determined whether early experiences of sheep with the same feed, but presented in multiple or single flavors, would influence intake, the profile of hormones involved in feed intake regulation, and the subsequent acceptability of novel feeds. Thirty-five 2-mo-old lambs were randomly assigned to 5 treatments (7 lambs/treatment). Lambs in 1 treatment (the diversity treatment) were simultaneously fed an unflavored plain ration of alfalfa (control) and barley (75:25; as-fed basis) and the same ration mixed (0.2%) with 1 of 3 flavors: 1) sweet, 2) umami, or 3) bitter. The other 4 treatments (monotonous diets) received only 1 of the 4 rations. All animals were fed their respective rations from 0800 to 1600 h for 60 d. On d 55, intake was recorded every 30 min for 8 h. On d 58, blood samples from lambs were collected at 1 h prefeeding and at 30, 60, 210, 300, and 540 min postfeeding. Preference tests were conducted by simultaneously offering novel feeds: 1) high-energy feed, 2) high-protein feed, 3) beet pulp mixed with phytochemicals, or 4) low-quality feed. Lambs in the diversity treatment consumed more feed than did lambs in the other treatments (P < 0.001). Lambs in the diversity treatment consumed equivalent amounts of plain and umami feeds, with a greater amount (P < 0.001) of the umami feed being consumed than the bitter and sweet feeds. Lambs in the diversity treatment tended to grow faster than did lambs in the other treatments (P=0.06). On d 55, lambs in the diversity treatment showed decreased (P < 0.05) feed intake compared with lambs in the other treatments during the 2 peaks of food consumption (30 and 270 min from feeding) and showed a trend for the least plasma concentrations of ghrelin (P=0.06). In contrast, lambs in the diversity treatment consumed more feed than did lambs exposed to monotonous flavors at 60, 90, 120, and 180 min from feeding (P < 0.05). Lambs in the diversity treatment also showed the least concentrations of cholecystokinin and glucagon-like peptide 1 (P < 0.001). There were trends for the greatest concentrations of leptin (P=0.14) and IGF-1 (P=0.16) in the diversity treatment, and for the least concentration of leptin in the bitter treatment (P=0.14). Previous experience with flavored feeds affected the preference of lambs for high-energy and low-quality feeds, and for beet pulp mixed with phytochemicals (treatment × feed × day effect; P < 0.05). Thus, exposure to diverse flavors has the potential to increase feed intake and induce a more even consumption of feed across time by reducing peaks and nadirs of intake compared with exposure to monotonous rations. Flavor diversity may also influence the initial acceptability of and preference for novel feeds.


Adverse cardiorespiratory events following primary vaccination of very low birth weight infants.
To examine the relationship between primary vaccination of preterm infants and prevalence ratios of associated factors for unwanted cardiorespiratory events, following the recommendation of the Centers for Disease Control and Prevention Advisory Committee on Immunization Practices for immunization of preterm infants at 2 months of chronological age. Two-year retrospective study of very low birth weight infants receiving their primary vaccination. Major cardiorespiratory events, such as apnea, bradycardia, SpO(2) desaturation, and minor adverse events, such as temperature instability, poor handling and local reactions, were recorded. Prevalence ratio with 95% confidence interval for associated factors between infants with and without cardiorespiratory events was calculated. Eighty neonates were studied (median [range] birth weight 970 g [428-1,490]), gestational age of 27.4 weeks (23.3-33). Adverse reactions occurred in 35 (44%): minor events in 19 (24%) patients, major events in 28 (35%). Infants with major events had significantly lower gestational age (p = 0.008) and a higher incidence of bronchopulmonary dysplasia (71% vs. 48%; p < 0.05). In very low birth weight infants with major events, O(2) desaturations before vaccination were 3.40 (1.41-8.23) times higher and treatment with methylxanthines for apnea and bradycardia syndrome was 8.05 (2.50-25.89) times higher compared to infants without major events. Major cardiorespiratory events occurred in over 1/3 of all very low birth weight infants after vaccination. Associated factors were low gestational age, bronchopulmonary dysplasia, methylxanthine treatment, and persisting O(2) desaturations before vaccination. Primary vaccination of very low birth weight infants should be performed under continuous monitoring of vital parameters.


Bacterial adhesion to bisphosphonate coated hydroxyapatite.
Staphylococcus aureus (S. aureus) is commonly associated with microbial infection of orthopaedic implants. Such infections often lead to osteomyelitis, which may result in failure of the implant due to localised bone destruction. Bacterial adhesion and subsequent colonisation of the device may occur as a consequence of contamination during surgery, or by seeding from a distant site through the blood circulation. Coating of the hydroxyapatite (HA) ceramic component of artificial hip joints with the bisphosphonates clodronate (C) and pamidronate (P) has been proposed as a means to minimise osteolysis and thereby prevent loosening of the implant. However, the effect of the bisphosphonate coating on bacterial adhesion to the HA materials must be determined before this approach can be implemented. In this study coated HA materials were incubated with the S. aureus and the number of adherent bacteria determined using the Modified Vortex Device (MVD) method. The number of bacteria adherent to the P coated HA material was significantly greater than that adherent to uncoated HA (60-fold increase) or to the C coated HA (90-fold increase). Therefore, even though earlier studies suggested that P bound to HA may improve osseointegration, the results presented would suggest that the use of this coating may be limited by the potential increased susceptibility of the coated device to infection.


The mitochondrial genomes of two Pre-historic Hunter Gatherers in Sri Lanka.
Sri Lanka is an island in the Indian Ocean connected by the sea routes of the Western and Eastern worlds. Although settlements of anatomically modern humans date back to 48,000 years, to date there is no genetic information on pre-historic individuals in Sri Lanka. We report here the first complete mitochondrial sequences for Mesolithic hunter-gatherers from two cave sites. The mitochondrial haplogroups of pre-historic individuals were M18a and M35a. Pre-historic mitochondrial lineage M18a was found at a low prevalence among Sinhalese, Sri Lankan Tamils, and Sri Lankan Indian Tamil in the Sri Lankan population, whereas M35a lineage was observed across all Sri Lankan populations with a comparatively higher frequency among the Sinhalese. Both haplogroups are Indian derived and observed in the South Asian region and rarely outside the region.


MyoRing Implantation in Keratoconic Patients: 3 years Follow-up Data.
To evaluate long-term follow-up data on implantation of a full-ring intra-corneal implant (MyoRing) for management of keratoconus. A total of 40 keratoconic eyes of 37 consecutive patients who had undergone MyoRing implantation using the Pocket Maker microkeratome (Dioptex, GmbH, Linz, Austria) and completed 3 years of follow-up appointments were included in this retrospective study. Uncorrected distance visual acuity (UDVA), corrected distance visual acuity (CDVA), refraction and keratometry (K) readings were measured and evaluated preoperatively, and 3 years, postoperatively. No intraoperative complications were observed in this case series. Three years postoperatively, there was a significant improvement in UDVA, CDVA, K readings, spherical equivalent (SE), and manifest sphere and cylinder (P < 0.05 for all comparisons). UDVA was significantly improved from 1.14 ± 0.27 to 0.30 ± 0.21 LogMAR (P = 0.001), CDVA was also improved from 0.52 ± 0.23 to 0.18 ± 0.12 LogMAR (P = 0.001), SE was decreased by 4.35 diopters (D) and average keratometric values were reduced by 2.34 D (P = 0.001). Overall, 81% of subjects were moderately to highly satisfied 3 years after surgery and 64.90% agreed to have the fellow eye implanted with MyoRing. MyoRing implantation using the Pocket Maker microkeratome was found to be a minimally invasive procedure for improving visual acuity and refraction in the majority of the patients with keratoconus.


Identification of novel endogenous antisense transcripts by DNA microarray analysis targeting complementary strand of annotated genes.
Recent transcriptomic analyses in mammals have uncovered the widespread occurrence of endogenous antisense transcripts, termed natural antisense transcripts (NATs). NATs are transcribed from the opposite strand of the gene locus and are thought to control sense gene expression, but the mechanism of such regulation is as yet unknown. Although several thousand potential sense-antisense pairs have been identified in mammals, examples of functionally characterized NATs remain limited. To identify NAT candidates suitable for further functional analyses, we performed DNA microarray-based NAT screening using mouse adult normal tissues and mammary tumors to target not only the sense orientation but also the complementary strand of the annotated genes. First, we designed microarray probes to target the complementary strand of genes for which an antisense counterpart had been identified only in human public cDNA sources, but not in the mouse. We observed a prominent expression signal from 66.1% of 635 target genes, and 58 genes of these showed tissue-specific expression. Expression analyses of selected examples (Acaa1b and Aard) confirmed their dynamic transcription in vivo. Although interspecies conservation of NAT expression was previously investigated by the presence of cDNA sources in both species, our results suggest that there are more examples of human-mouse conserved NATs that could not be identified by cDNA sources. We also designed probes to target the complementary strand of well-characterized genes, including oncogenes, and compared the expression of these genes between mammary cancerous tissues and non-pathological tissues. We found that antisense expression of 95 genes of 404 well-annotated genes was markedly altered in tumor tissue compared with that in normal tissue and that 19 of these genes also exhibited changes in sense gene expression. These results highlight the importance of NAT expression in the regulation of cellular events and in pathological conditions. Our microarray platform targeting the complementary strand of annotated genes successfully identified novel NATs that could not be identified by publically available cDNA data, and as such could not be detected by the usual "sense-targeting" microarray approach. Differentially expressed NATs monitored by this platform may provide candidates for investigations of gene function. An advantage of our microarray platform is that it can be applied to any genes and target samples of interest.