Deciphering Key Pharmacological Pathways of Qingdai Acting on Chronic Myeloid Leukemia Using a Network Pharmacology-Based Strategy.
Qingdai, a traditional Chinese medicine (TCM) used for the treatment of chronic myeloid leukemia (CML) with good efficacy, has been used in China for decades. However, due to the complexity of traditional Chinese medicinal compounds, the pharmacological mechanism of Qingdai needs further research. In this study, we investigated the pharmacological mechanisms of Qingdai in the treatment of CML using network pharmacology approaches. First, components in Qingdai that were selected by pharmacokinetic profiles and biological activity predicted putative targets based on a combination of 2D and 3D similarity measures with known ligands. Then, an interaction network of Qingdai putative targets and known therapeutic targets for the treatment of chronic myeloid leukemia was constructed. By calculating the 4 topological features (degree, betweenness, closeness, and coreness) of each node in the network, we identified the candidate Qingdai targets according to their network topological importance. The composite compounds of Qingdai and the corresponding candidate major targets were further validated by a molecular docking simulation. Seven components in Qingdai were selected and 32 candidate Qingdai targets were identified; these were more frequently involved in cytokine-cytokine receptor interaction, cell cycle, p53 signaling pathway, MAPK signaling pathway, and immune system-related pathways, which all play important roles in the progression of CML. Finally, the molecular docking simulation showed that 23 pairs of chemical components and candidate Qingdai targets had effective binding. This network-based pharmacology study suggests that Qingdai acts through the regulation of candidate targets to interfere with CML and thus regulates the occurrence and development of CML.


Morphological quantitative changes in the number of lymphocytes, macrophages and plasma cells in the uterus and lymph nodes of rats exposed to the systemic administration of BCG.
This study was designed to investigate the effect of systemic administration of BCG on the morphological quantitative changes in the number of lymphocytes, macrophages and plasma cells in the uterus and lymph nodes of rats. Thirty female virgin Wistar Albino rats, aging 6 months and weighing between 200-250 g, were assigned to the two experimental groups; BCG treated and controls (n = 15). BCG group received 0.1 ml BCG in tail skin and control group received 0.1 ml saline at the same place. Two weeks after injections, rats in both groups were anesthesized with a high dose of ether and decapitated. Uterus and ileocecal lymph nodes were processed to determine a napthyl acid esterase (ANAE)-positive T lymphocytes and macrophages. The plasma cells were stained with the methyl green-pyronin method. It was found that the numbers of T lymphocytes, macrophages and plasma cells in the uterus and the ileocecal lymph nodes of BCG treated group significantly increased indicating the presence of an immune response to the systemic BCG administration. It was concluded that the systemic administration of BCG increases humoral and cellular immunity in endometrium, myometrium and regional lymph nodes. The immune deficiency system plays an important role in the pathogenesis of endometriosis. Therefore, the endometriosis might be prevented by using periodical administration of BCG. However, further experimental and clinical studies associated with these issue are required.


Integrative analysis reveals selective 9p24.1 amplification, increased PD-1 ligand expression, and further induction via JAK2 in nodular sclerosing Hodgkin lymphoma and primary mediastinal large B-cell lymphoma.
Classical Hodgkin lymphoma (cHL) and mediastinal large B-cell lymphoma (MLBCL) are lymphoid malignancies with certain shared clinical, histologic, and molecular features. Primary cHLs and MLBCLs include variable numbers of malignant cells within an inflammatory infiltrate, suggesting that these tumors escape immune surveillance. Herein, we integrate high-resolution copy number data with transcriptional profiles and identify the immunoregulatory genes, PD-L1 and PD-L2, as key targets at the 9p24.1 amplification peak in HL and MLBCL cell lines. We extend these findings to laser-capture microdissected primary Hodgkin Reed-Sternberg cells and primary MLBCLs and find that programmed cell death-1 (PD-1) ligand/9p24.1 amplification is restricted to nodular sclerosing HL, the cHL subtype most closely related to MLBCL. Using quantitative immunohistochemical methods, we document the association between 9p24.1 copy number and PD-1 ligand expression in primary tumors. In cHL and MLBCL, the extended 9p24.1 amplification region also included the Janus kinase 2 (JAK2) locus. Of note, JAK2 amplification increased protein expression and activity, specifically induced PD-1 ligand transcription and enhanced sensitivity to JAK2 inhibition. Therefore, 9p24.1 amplification is a disease-specific structural alteration that increases both the gene dosage of PD-1 ligands and their induction by JAK2, defining the PD-1 pathway and JAK2 as complementary rational therapeutic targets.


High-resolution analysis of intrahost genetic diversity in dengue virus serotype 1 infection identifies mixed infections.
Little is known about the rate at which genetic variation is generated within intrahost populations of dengue virus (DENV) and what implications this diversity has for dengue pathogenesis, disease severity, and host immunity. Previous studies of intrahost DENV variation have used a low frequency of sampling and/or experimental methods that do not fully account for errors generated through amplification and sequencing of viral RNAs. We investigated the extent and pattern of genetic diversity in sequence data in domain III (DIII) of the envelope (E) gene in serial plasma samples (n = 49) taken from 17 patients infected with DENV type 1 (DENV-1), totaling some 8,458 clones. Statistically rigorous approaches were employed to account for artifactual variants resulting from amplification and sequencing, which we suggest have played a major role in previous studies of intrahost genetic variation. Accordingly, nucleotide sequence diversities of viral populations were very low, with conservative estimates of the average levels of genetic diversity ranging from 0 to 0.0013. Despite such sequence conservation, we observed clear evidence for mixed infection, with the presence of multiple phylogenetically distinct lineages present within the same host, while the presence of stop codon mutations in some samples suggests the action of complementation. In contrast to some previous studies we observed no relationship between the extent and pattern of DENV-1 genetic diversity and disease severity, immune status, or level of viremia.


Dishabituation and sensitization emerge as separate processes during development in Aplysia.
Until recently, dishabituation and sensitization have commonly been considered to reflect a unitary process: Sensitization refers to a general facilitation produced by strong or noxious stimuli that enhances subsequent responding; dishabituation has been thought to represent a special instance of sensitization in which the facilitation is simply superimposed on a habituated response level. The unitary process hypothesis was based on the observation that both decremented and nondecremented responses are facilitated by a common noxious or strong stimulus. However, this observation does not rule out the possibility that dishabituation and sensitization could reflect separate processes that are activated in parallel by a strong stimulus. Recent cellular experiments by Hochner et al. (1986) suggest that this, in fact, occurs in the sensory neurons of the gill withdrawal reflex in Aplysia. A developmental analysis of learning in the marine mollusc Aplysia permits a direct behavioral test of this hypothesis. If dishabituation and sensitization reflect a unitary process then they should emerge at the same time ontogenetically. On the other hand, if they reflect different processes, then they might emerge according to different ontogenetic timetables. In the present study we examined the temporal emergence of dishabituation and sensitization in the defensive siphon withdrawal reflex in 3 stages of juvenile Aplysia: stage 11, early stage 12, and late stage 12. Animals received one of 2 kinds of training: Dishabituation training, in which the effect of strong tail shock on habituated responses were observed, and Sensitization training, in which the effect of strong tail shock on nondecremented responses was observed. We found that, while dishabituation was present in all stages examined, sensitization did not emerge until several weeks later, in late stage 12. These results were confirmed and extended in a group of animals that were tested twice: first in stage 11, when they showed no sensitization, and again 13 weeks later, in late stage 12, when they then showed significant sensitization. Our analysis of nondecremented responses prior to the emergence of sensitization also revealed an unexpected inhibitory component of tail shock that produces reflex depression. Moreover, there was a clear progression in the net effects of tail shock during development: reflex depression was produced in stages 11 and early stage 12, followed by a transition to reflex facilitation (sensitization) in late stage 12. Finally, when sensitization emerged in late stage 12, the process of dishabituation showed a significant increase compared with previous developmental stages.(ABSTRACT TRUNCATED AT 400 WORDS)


Correlation of the distribution of Rickettsia conorii, microscopic lesions, and clinical features in South African tick bite fever.
Three South African patients with severe Rickettsia conorii infection had complicated courses of illness with 2 fatal cases and 1 with gangrene of multiple digits. Immunofluorescent organisms of R. conorii were demonstrated in vascular endothelium of brain, leptomeninges, renal glomerular arterioles and capillaries, renal arteries and veins, myocardial capillaries and arteries, pulmonary alveolar capillaries, pancreatic septa, splenic arterioles, and dermis. Rickettsiae were also observed in hepatic sinusoidal lining cells, splenic and lymph node macrophages, and the blood vessels of the partially viable zone of the amputated digits. Pathologic lesions included cerebral and cerebellar perivascular mononuclear leukocytes, mild mononuclear leptomeningitis, glomerular arteriolitis, vascular and perivascular mononuclear cell-rich inflammatory foci in the kidney, pancreas, skin, and myocardium, hepatocellular necrosis, and pulmonary edema. The sites of lesions and rickettsiae showed strong topographical correlation. Thrombi and hemorrhage occurred in a minority of the sites of vascular injury. Rickettsiae were the apparent direct cause of meningoencephalitis, peripheral gangrene, and other foci of vascular injury. Fatal R. conorii infection with disseminated organ involvement emphasizes the pathogenic potential of this disease.


Nanotexaphyrin: One-Pot Synthesis of a Manganese Texaphyrin-Phospholipid Nanoparticle for Magnetic Resonance Imaging.
The discovery and synthesis of novel multifunctional organic building blocks for nanoparticles is challenging. Texaphyrin macrocycles are capable and multifunctional chelators. However, they remain elusive as building blocks for nanoparticles because of the difficulty associated with synthesis of texaphyrin constructs capable of self-assembly. A novel manganese (Mn)-texaphyrin-phospholipid building block is described, along with its one-pot synthesis and self-assembly into a Mn-nanotexaphyrin. This nanoparticle possesses strong resilience to manganese dissociation, structural stability, in vivo bio-safety, and structure-dependent T1 and T2 relaxivities. Magnetic resonance imaging (MRI) contrast enhanced visualization of lymphatic drainage is demonstrated with respect to proximal lymph nodes on the head and neck VX-2 tumors of a rabbit. Synthesis of 17 additional metallo-texaphyrin building blocks suggests that this novel one-pot synthetic procedure for nanotexaphyrins may lead to a wide range of applications in the field of nanomedicines.


Anti-atherosclerotic effect of a new synthetic functional oil containing mono- and diacylglycerol from corn oil.
Synthetic oil containing diacylglycerol and monoacylglycerol, called 'functional oil' (FO), was newly produced and evaluated for its putative anti-atherosclerotic potential by in vitro assays and in vivo test using hypercholesterolemic mice (C57BL/6). The FO revealed good inhibitory activities against both liver acyl-CoA:cholesterol acyltransferase and serum lipoprotein-associated phospholipase A2. The FO showed enhanced activities on lipoprotein interaction such as HDL particle rearrangement to produce different sizes of HDL species. In control mice, hypercholesterolemia was induced by consumption of high-cholesterol, high-fat (HCHF) diet that contained 1.25% cholesterol/15% fat/0.5% Na-cholate with or without 5% of corn oil. In experimental mice, 5% of the FO + HCHF diet was fed during the same period. After the 4-week administration of the diet, serum total cholesterol concentration of the FO-fed group decreased by 38 or 20% when compared to the HCHF diet control group or corn oil (99.9% of triacylglycerol) diet group, respectively. The percentage of HDL cholesterol to total cholesterol was 36% of HDL cholesterol in the FO-fed group, while the HCHF control group and corn oil-fed group showed 21 and 25%, respectively. These results indicate that the FO possesses a blood cholesterol-lowering effect in mouse model and inhibition effects against the atherogenic enzymes.


Angiotensin receptor gene polymorphisms and 2-year change in hyperintense lesion volume in men.
This longitudinal study examined the relationship between 2-year change in white matter hyperintense lesion (WML) volume and polymorphisms in genes coding for the angiotensin-II type 1 and type 2 receptors, AGTR1 A1166C and AGTR2 C3123A, respectively. 137 depressed and 94 non-depressed participants aged >or=60 years were enrolled. Standard clinical evaluations were performed on all participants and blood samples obtained for genotyping. 1.5-T MRI (magnetic resonance imaging) data were obtained at baseline and approximately 2 years later. These scans were processed using a semi-automated segmentation process, which allowed for the calculation of WML volume at each time point. Statistical models were tested for the relationship between change in WML volume and genotype, while also controlling for age, sex, diagnostic strata, baseline WML volume and comorbid cerebrovascular risk factors. In men, AGTR1 1166A allele homozygotes exhibited significantly less change in WML volume than 1166C carriers. We also found that men reporting hypertension (HTN) with the AGTR2 3123C allele exhibit less change in WML volume than hypertensive men with the 3123A allele, or men without HTN. There were no significant relationships between these polymorphisms and change in WML volume in women. No significant gene-gene or gene-depression interactions were observed. Our results parallel earlier observed gender differences of the relationship between other renin-angiotensin system polymorphisms and HTN. Further work is needed to determine whether these observed relationships are secondary to polymorphisms affecting response to antihypertensive medication, and whether antihypertensive medications can slow WML progression and lower the risk of morbidity associated with WMLs.


Endostatin has ATPase activity, which mediates its antiangiogenic and antitumor activities.
Endostatin is an endogenous angiogenesis inhibitor with broad-spectrum antitumor activities. Although the molecular mechanisms of endostatin have been extensively explored, the intrinsic biochemical characteristics of endostatin are not completely understood. Here, we revealed for the first time that endostatin embedded novel ATPase activity. Moreover, mutagenesis study showed that the ATPase activity of endostatin mutants positively correlated with effects on endothelial cell activities and tumor growth. E-M, an endostatin mutant with higher ATPase activity than that of wild-type (WT) endostatin, significantly increased endostatin-mediated inhibitory effects on endothelial cell proliferation, migration, tube formation, and adhesion. In vivo study showed that E-M displayed enhanced antitumor effects compared with WT. On the other hand, K96A, K96R, and E176A, endostatin mutants with lower ATPase activities than that of WT, showed reduced or comparable effects on targeting both in vitro endothelial cell activities and in vivo tumor angiogenesis and tumor growth. Furthermore, endostatin and its mutants exhibited distinct abilities in regulations of gene expression (Id1, Id3), cell signaling (Erk, p38, and Src phosphorylation), and intracellular ATP levels. Collectively, our study demonstrates that endostatin has novel ATPase activity, which mediates its antiangiogenic and antitumor activities, suggesting that construction of endostatin analogues with high ATPase activity may provide a new direction for the development of more potent antiangiogenic drugs.


TRUST-TECH-based expectation maximization for learning finite mixture models.
In spite of the initialization problem, the Expectation-Maximization (EM) algorithm is widely used for estimating the parameters of finite mixture models. Most popular model-based clustering techniques might yield poor clusters if the parameters are not initialized properly. To reduce the sensitivity of initial points, a novel algorithm for learning mixture models from multivariate data is introduced in this paper. The proposed algorithm takes advantage of TRUST-TECH (TRansformation Under STability-reTaining Equilibra CHaracterization) to compute neighborhood local maxima on likelihood surface using stability regions. Basically, our method coalesces the advantages of the traditional EM with that of the dynamic and geometric characteristics of the stability regions of the corresponding nonlinear dynamical system of the log-likelihood function. Two phases namely, the EM phase and the stability region phase, are repeated alternatively in the parameter space to achieve improvements in the maximum likelihood. The EM phase obtains the local maximum of the likelihood function and the stability region phase helps to escape out of the local maximum by moving towards the neighboring stability regions. The algorithm has been tested on both synthetic and real datasets and the improvements in the performance compared to other approaches are demonstrated. The robustness with respect to initialization is also illustrated experimentally.


Osmoregulatory modulation of thermal sweating in humans: reflex effects of drinking.
To gain better insight into the interaction between thermoregulation and osmoregulation, we examined the thermal sweating response to drinking in cell-dehydrated humans. Cell dehydration (CDH) was induced by infusion of a 3% NaCl solution, at 1.2 ml/kg, for 2 h; infusion of a 0.9% NaCl solution in a separate experiment served as a control (euhydrated condition, EH). After infusion, subjects were heated by immersion of the lower legs in 42 degrees C water at an ambient temperature of 28 degrees C for 90 min. Subjects drank 4.3 ml/kg of H2O (approximately 38 degrees C) at 60 min of heating. The 3% NaCl infusion increased plasma osmolality by 13.6 +/- 0.8 mosmol/kgH2O and plasma arginine vasopressin concentration ([AVP]) by 3.3 +/- 0.7 pg/ml. Neither variable was altered with 0.9% NaCl infusion. Before drinking, esophageal temperature (Tes) had increased by 0.91 +/- 0.08 degrees C in CDH and by 0.40 +/- 0.11 degrees C in EH. Local chest sweating rate (SRch) had increased by 0.67 +/- 0.08 and 0.63 +/- 0.07 mg.min-1.cm-2 in CDH and EH, respectively. Thus the change in SRch per unit rise in Tes was much lower in CDH than in EH. Drinking immediately increased SRch and reduced Tes in CDH, with a reduction in plasma [AVP] and thirst rating. Drinking did not change thermoregulatory and osmoregulatory responses in EH. These results suggest that the act of drinking itself eliminates, at least partially, an osmotic inhibitory input to the thermoregulatory center, as well as osmotic AVP secretion and thirst.


Nonparalytic botulinum molecules for the control of pain.
Local injections of botulinum toxins have been reported to be useful not only for the treatment of peripheral neuropathic pain and migraine but also to cause long-lasting muscle paralysis, a potentially serious side effect. Recently, a botulinum A-based molecule ("BiTox") has been synthesized that retains neuronal silencing capacity without triggering muscle paralysis. In this study, we examined whether BiTox delivered peripherally was able to reduce or prevent the increased nociceptive sensitivity found in animal models of inflammatory, surgical, and neuropathic pain. Plasma extravasation and edema were also measured as well as keratinocyte proliferation. No motor deficits were seen and acute thermal and mechanical nociceptive thresholds were unimpaired by BiTox injections. We found reduced plasma extravasation and inflammatory edema as well as lower levels of keratinocyte proliferation in cutaneous tissue after local BiTox injection. However, we found no evidence that BiTox was transported to the dorsal root ganglia or dorsal horn and no deficits in formalin-elicited behaviors or capsaicin or formalin-induced c-Fos expression within the dorsal horn. In contrast, Bitox treatment strongly reduced A-nociceptor-mediated secondary mechanical hyperalgesia associated with either complete Freund's adjuvant (CFA)-induced joint inflammation or capsaicin injection and the hypersensitivity associated with spared nerve injury. These results imply that although local release of neuromodulators from C-fibers was inhibited by BiTox injection, C-nociceptive signaling function was not impaired. Taken together with recent clinical data the results suggest that BiTox should be considered for treatment of pain conditions in which A-nociceptors are thought to play a significant role.


Pattern of fin rays along the antero-posterior axis based on their connection to distal radials.
Teleost paired fins are composed of two endoskeletal domains, proximal and distal radials, and an exoskeletal domain, the fin ray. The zebrafish pectoral fin displays elaborately patterned radials along the anteroposterior (AP) axis. Radials are considered homologous to tetrapod limb skeletons, and their patterning mechanisms in embryonic development are similar to those of limb development. Nevertheless, the pattern along the AP axis in fin rays has not been well described in the zebrafish pectoral fin, although several recent reports have revealed that fin ray development shares some cellular and genetic properties with fin/limb endoskeleton development. Thus, fin ray morphogenesis may involve developmental mechanisms for AP patterning in the fin/limb endoskeleton, and may have a specific pattern along the AP axis. We conducted detailed morphological observations on fin rays and their connection to distal radials by comparing intra- and inter-strain zebrafish specimens. Although the number of fin rays varied, pectoral fin rays could be categorized into three domains along the AP axis, according to the connection between the fin rays and distal radials; additionally, the number of fin rays varied in the posterior part of the three domains. This result was confirmed by observation of the morphogenesis process of fin rays and distal radials, which showed altered localization of distal radials in the middle domain. We also evaluated the expression pattern of lhx genes, which have AP patterning activity in limb development, in fin rays and during distal radial development and found these genes to be expressed during morphogenesis in both fin rays and distal radials. The fin ray and its connection to the endoskeleton are patterned along the AP axis, and the pattern along the AP axis in the fin ray and the radial connection is constructed by the developmental mechanism related to AP patterning in the limb/fin bud. Our results indicate the possibility that the developmental mechanisms of fin rays and their connection are comparable to those of the distal element of the limb skeleton.


Renal sodium excretion in sons of hypertensive parents.
The objective of this study was to evaluate whether renal excretion of sodium is impaired and whether tubular reabsorption of sodium is increased in normotensive white men with a familial predisposition to develop essential hypertension. We compared 11 normotensive sons of two hypertensive parents (SOHT) with 11 normotensive sons of two normotensive parents (SONT); renal sodium handling was assessed after 1 week of low-sodium diet (10 mmol/d) and after 1 week of high-sodium diet (200 mmol/d). The SOHT were on average 5.5 years older than the SONT (46.9 +/- 5.2 [SD] vs 41.4 +/- 4.1, P = .012). On the sixth day of each diet, mean urinary sodium excretion did not differ between the two groups (12.9 +/- 6.3 vs 12.7 +/- 6.7 mmol/d on low-sodium diet, P = .930; 197 +/- 25 vs 200 +/- 27 mmol/d on high-sodium diet, P = .817). On the seventh day of each diet, baseline means for filtered load of sodium, absolute excretion of sodium, fractional excretion of sodium (an index of total tubular sodium reabsorption), and fractional excretion of lithium (an inverse index of proximal tubular sodium reabsorption) also did not differ between the groups. To assess renal sodium handling under non-steady-state conditions, we infused 2 L normal saline intravenously over a 2-hour period. The means for absolute excretion of sodium, fractional excretion of sodium, and fractional excretion of lithium increased from baseline, but the increases did not differ in magnitude between the groups.(ABSTRACT TRUNCATED AT 250 WORDS)


Differential distribution of competence for panplacodal and neural crest induction to non-neural and neural ectoderm.
It is still controversial whether cranial placodes and neural crest cells arise from a common precursor at the neural plate border or whether placodes arise from non-neural ectoderm and neural crest from neural ectoderm. Using tissue grafting in embryos of Xenopus laevis, we show here that the competence for induction of neural plate, neural plate border and neural crest markers is confined to neural ectoderm, whereas competence for induction of panplacodal markers is confined to non-neural ectoderm. This differential distribution of competence is established during gastrulation paralleling the dorsal restriction of neural competence. We further show that Dlx3 and GATA2 are required cell-autonomously for panplacodal and epidermal marker expression in the non-neural ectoderm, while ectopic expression of Dlx3 or GATA2 in the neural plate suppresses neural plate, border and crest markers. Overexpression of Dlx3 (but not GATA2) in the neural plate is sufficient to induce different non-neural markers in a signaling-dependent manner, with epidermal markers being induced in the presence, and panplacodal markers in the absence, of BMP signaling. Taken together, these findings demonstrate a non-neural versus neural origin of placodes and neural crest, respectively, strongly implicate Dlx3 in the regulation of non-neural competence, and show that GATA2 contributes to non-neural competence but is not sufficient to promote it ectopically.


Unusual effects of monocarboxylic acids on the structure and on the transport and mechanical properties of chitosan films.
The purpose of this study was to study the transport of monocarboxylic acids in chitosan films, since this is important for understanding and predicting the drying kinetics of chitosan from aqueous solutions. Despite the wealth of data on chitosan films prepared from aqueous monocarboxylic acid solutions, this transport has not been reported. Chitosan films were exposed to formic, acetic, propionic and butyric acid vapours, it was found that the rate of uptake decreased with increasing molecular size. The equilibration time was unexpectedly long, especially for propionic and butyric acid, nine months. A clear two-stage uptake curve was observed for propionic acid. Evidently, the rate of uptake was determined by acid-induced changes in the material. X-ray diffraction and infrared spectroscopy indicated that the structure of the chitosan acetate and buffered chitosan films changed during exposure to acid and during the subsequent drying. The dried films previously exposed to the acid showed less crystalline features than the original material and a novel repeating structure possibly involving acid molecules. The molar mass of the chitosan decreased on exposure to acid but tensile tests revealed that the films were always ductile. The films exposed to acid vapour (propionic and butyric acid) for the longest period of time were insoluble in the size-exclusion chromatography eluent, and they were also the most ductile/extensible of all samples studied.


Focal congenital hyperinsulinism managed by medical treatment: a diagnostic algorithm based on molecular genetic screening.
Congenital hyperinsulinism (CHI) requires rapid diagnosis and treatment to avoid irreversible neurological sequelae due to hypoglycaemia. Aetiological diagnosis is instrumental in directing the appropriate therapy. Current diagnostic algorithms provide a complete set of diagnostic tools including (i) biochemical assays, (ii) genetic facility and (iii) state-of-the-art imaging. They consider the response to a therapeutic diazoxide trial an early, crucial step before proceeding (or not) to specific genetic testing and eventually imaging, aimed at distinguishing diffuse vs focal CHI. However, interpretation of the diazoxide test is not trivial and can vary between research groups, which may lead to inappropriate decisions. Objective of this report is proposing a new algorithm in which early genetic screening, rather than diazoxide trial, dictates subsequent clinical decisions. Two CHI patients weaned from parenteral glucose infusion and glucagon after starting diazoxide. No hypoglycaemia was registered during a 72-h continuous glucose monitoring (CGMS), or hypoglycaemic episodes were present for no longer than 3% of 72-h. Normoglycaemia was obtained by low-medium dose diazoxide combined with frequent carbohydrate feeds for several years. We identified monoallelic, paternally inherited mutations in KATP channel genes, and (18) F-DOPA PET-CT revealed a focal lesion that was surgically resected, resulting in complete remission of hypoglycaemia. Although rare, some patients with focal lesions may be responsive to diazoxide. As a consequence, we propose an algorithm that is not based on a 'formal' diazoxide response but on genetic testing, in which patients carrying paternally inherited ABCC8 or KCNJ11 mutations should always be subjected to (18) F-DOPA PET-CT.


Pericardial mesothelioma presenting as a suspected ST-elevation myocardial infarction.
Primary cardiac and pericardial tumors are rare entities with an autopsy frequency of 0.001-0.03%. Metastases to the heart and pericardium are much more common than primary tumors. Malignant pericardial mesotheliomas account for up to 50% of primary pericardial tumors. We report the case of a 75-year-old woman with hypertension, dyslipidemia and atrial fibrillation who went to the emergency department due to nonspecific thoracic discomfort of over six hours duration associated with syncope. Physical examination revealed a low-amplitude arrhythmic pulse, no heart murmurs and no signs of pulmonary congestion. The ECG revealed atrial fibrillation with ST-segment elevation in V2-V6, I and aVL. The patient was transferred for emergent coronary angiography, which revealed a long stenosis in the mid-distal portion of the left anterior descending artery. The echocardiogram showed a large pericardial effusion with diffuse thickening of the myocardium. Due to worsening hemodynamics, cardiac rupture was suspected and the patient underwent urgent sternotomy and pericardiotomy with drainage of a large quantity of hematic fluid. The surgeons then identified a large, unresectable tumor occupying the distal half of the anterior portion of the heart. This is, to our knowledge, the first case report of primary pericardial mesothelioma presenting with suspected ST-elevation myocardial infarction. In this case, direct observation of the tumor led to biopsy and the final diagnosis. These are highly malignant tumors and when diagnosed are usually already at an advanced stage.


Mucosal dysplasia in ileal pelvic pouches after restorative proctocolectomy.
Inflammation, villous atrophy, colonic metaplasia, and dysplasia have been observed within the mucosa of ileal pelvic pouches after restorative proctocolectomy. This study was designed to determine the prevalence of mucosal dysplasia in ileal pouch and any associated risk factors. Prospectively registered patients having restorative proctocolectomy were recruited. A cross-sectional study was performed using a questionnaire focusing on disease history, functional results, and pouchitis after surgery. Participants underwent screening endoscopic pouch examination using sigmoidoscopy. Mucosal biopsies were taken from six specific locations in the pouch from proximal ileal-pouch (inflow) to ileoanal anastomosis. All biopsies were performed under strict surveillance protocol regardless of patients' symptoms. Biopsies were interpreted by two pathologists unaware of each other's report. A total of 138 patients completed the protocol. Colectomy specimens from restorative proctocolectomy showed chronic ulcerative colitis in 118 (85.6 percent), familial adenomatous polyposis in 10 (7.2 percent), Crohn's colitis in 2 (1.4 percent), and indeterminate colitis in 8 (5.8 percent) patients. Twenty-two patients (18.3 percent) had dysplasia and eight (6.7 percent) had invasive cancer found in colectomy specimens after restorative proctocolectomy. Median interval between proctocolectomy and pouch biopsy was 5.4 years. Inflammatory changes were present in a majority of specimens, but these did not correlate with clinical history of pouchitis. No villous atrophy was identified. Pouch biopsies from only one patient were indefinite for dysplasia. Subsequent biopsies were negative. Clinical and microscopic evidence of ileal-pouch inflammation is common. Ileal-pouch mucosal dysplasia is uncommon, occurring in only 1 of 138 patients. Villous atrophy and colonic metaplasia were not observed in this series. Routine pouch surveillance with biopsies may not be warranted.


Staphylococcus aureus and MRSA colonization rates among personnel and dogs in a small animal hospital: association with nosocomial infections.
The genetic relationship of thirty Methicillin-resistant S. aureus (MRSA) isolates derived from the nasal cavities of canine patients hospitalized (n = 7), veterinary personnel (n = 20), and environmental sources (n = 3) sampled during a 20-month investigation period, were analyzed in this study. Genetic relatedness of all MRSA isolates was investigated involving commonly used typing techniques: Pulsed-field gel electrophoresis (PFGE), Multilocus sequence typing (MLST), spa typing, PCR for detection of Panton-Valentine leukocidine (PVL) genes and staphylococcal cassette chromosome mec-typing (SCCmec). Analysis of typing results revealed a certain predominant (72%) genotype: PFGE type IMT-A, ST22, spa type t032, SCCmecIV. This genotype has been reported previously (Walter et al., 2008c) being the predominant PFGE type associated with MRSA-positive clinical specimens, mostly from wound infections, derived from small and exotic animals of that facility. Furthermore, occasionally high rates in nasal colonization of veterinary personnel (18 of 88: 20%) in one of three personal screening periods were accompanied by a sudden rise of MRSA infections in small animals. Our data strongly indicate that high rates of colonized veterinary staff lead to an increase of nosocomial infections in small animal hospitals. We therefore recommend the introduction of surveillance of nosocomial infections especially in surgical veterinary hospitals.


The antidepressant agomelatine blocks the adverse effects of stress on memory and enables spatial learning to rapidly increase neural cell adhesion molecule (NCAM) expression in the hippocampus of rats.
Agomelatine, a novel antidepressant with established clinical efficacy, acts as a melatonin receptor agonist and 5-HT(2C) receptor antagonist. As stress is a significant risk factor in the development of depression, we sought to determine if chronic agomelatine treatment would block the stress-induced impairment of memory in rats trained in the radial-arm water maze (RAWM), a hippocampus-dependent spatial memory task. Moreover, since neural cell adhesion molecule (NCAM) is known to be critically involved in memory consolidation and synaptic plasticity, we evaluated the effects of agomelatine on NCAM, and polysialylated NCAM (PSA-NCAM) expression in rats given spatial memory training with or without predator stress. Adult male rats were pre-treated with agomelatine (10 mg/kg i.p., daily for 22 d), followed by a single day of RAWM training and memory testing. Rats were given 12 training trials and then they were placed either in their home cages (no stress) or near a cat (predator stress). Thirty minutes later the rats were given a memory test trial followed immediately by brain extraction. We found that: (1) agomelatine blocked the predator stress-induced impairment of spatial memory; (2) agomelatine-treated stressed, as well as non-stressed, rats exhibited a rapid training-induced increase in the expression of synaptic NCAM in the ventral hippocampus; and (3) agomelatine treatment blocked the water-maze training-induced decrease in PSA-NCAM levels in both stressed and non-stressed animals. This work provides novel observations which indicate that agomelatine blocks the adverse effects of stress on hippocampus-dependent memory and activates molecular mechanisms of memory storage in response to a learning experience.


Rescue of degenerating neurons and cells by stem cell released molecules: using a physiological renormalization strategy.
Evidence suggests that adult stem cell types and progenitor cells act collectively in a given tissue to maintain and heal organs, such as muscle, through a release of a multitude of molecules packaged into exosomes from the different cell types. Using this principle for the development of bioinspired therapeutics that induces homeostatic renormalization, here we show that the collection of molecules released from four cell types, including mesenchymal stem cells, fibroblast, neural stem cells, and astrocytes, rescues degenerating neurons and cells. Specifically, oxidative stress induced in a human recombinant TDP-43- or FUS-tGFP U2OS cell line by exposure to sodium arsenite was shown to be significantly reduced by our collection of molecules using in vitro imaging of FUS and TDP-43 stress granules. Furthermore, we also show that the collective secretome rescues cortical neurons from glutamate toxicity as evidenced by increased neurite outgrowth, reduced LDH release, and reduced caspase 3/7 activity. These data are the first in a series supporting the development of stem cell-based exosome systems therapeutics that uses a physiological renormalization strategy to treat neurodegenerative diseases.


Alveolar tissue inflammation in asthma.
As physiologic and autopsy evidence suggests that peripheral airways and parenchyma are involved in asthma, we hypothesized that significant alveolar tissue inflammation is present in patients with stable, chronic asthma. Eleven patients with nocturnal asthma (NA) and 10 patients with non-nocturnal asthma (NNA) were studied. Each subject underwent two bronchoscopies with proximal airway endobronchial and distal alveolar tissue transbronchial biopsy in a random order at 4:00 P.M. and 4:00 A.M. Morphometric analysis was used to determine the number per volume (Nv) of inflammatory cells. Between-group comparisons showed that the Nv of eosinophils was greater in the NA alveolar tissue 4:00 A.M. compared with the subjects with NNA (40.2 x 10(3) [26.4-57.1 x 10(3), IQ] versus 15.7 x 10(3) [2.1-35.2 x 10(3), IQ], p = 0.05). In regard to the airway biopsies, no difference in the inflammatory and epithelial cells between the two groups was seen at either time. The NA group exhibited greater eosinophils and macrophages in the alveolar tissue at 4:00 A.M. compared with 4:00 P.M. (40.2 x 10(3) [26.4-57.1 x 10(3), IQ] versus 10.3 x 10(3) [2.7-16.8 x 10(3), IQ], p = 0.016 for eosinophils and 215.1 x 10(3) [129.9-356.1 x 10(3), IQ] versus 166.3 x 10(3) [150.7-212.6 x 10(3), IQ], p = 0.031 for macrophages). Only alveolar tissue eosinophils, not proximal airway tissue eosinophils, correlated with the nocturnal decrement in lung function (r = -0.54, p = 0.03). These findings suggest that eosinophils and macrophages accumulate to a greater extent in the alveolar tissue and these changes contribute more to the variation in lung function compared with inflammation in the more proximal tissue.


Hypoxia-inducible factors mediate coordinated RhoA-ROCK1 expression and signaling in breast cancer cells.
Overexpression of Rho kinase 1 (ROCK1) and the G protein RhoA is implicated in breast cancer progression, but oncogenic mutations are rare, and the molecular mechanisms that underlie increased ROCK1 and RhoA expression have not been determined. RhoA-bound ROCK1 phosphorylates myosin light chain (MLC), which is required for actin-myosin contractility. RhoA also activates focal adhesion kinase (FAK) signaling. Together, these pathways are critical determinants of the motile and invasive phenotype of cancer cells. We report that hypoxia-inducible factors coordinately activate RhoA and ROCK1 expression and signaling in breast cancer cells, leading to cell and matrix contraction, focal adhesion formation, and motility through phosphorylation of MLC and FAK. Thus, intratumoral hypoxia acts as an oncogenic stimulus by triggering hypoxia-inducible factor → RhoA → ROCK1 → MLC → FAK signaling in breast cancer cells.


Small-Scale High-Fidelity Simulation for Mass Casualty Incident Readiness.
This content can be used for trauma centers, emergency medicine residency programs, and emergency nursing. Mass casualty incidents (MCI) are becoming increasingly common and are occurring in locations that have not experienced them previously which adds to the challenge of readiness for emergency departments (EDs). Sporadic occurrences and limited resources add to the complexity of preparing for such an event. In advance of a large gathering in our metropolitan area, we developed and conducted a simulation to better prepare not only our residents, but our MCI planning committee, registered nurses (RNs) and emergency room technicians (ERTs) for an MCI.Emergency medicine is at the forefront of any hospital's response to an MCI. These events stretch the resources and force EDs to function differently than usual.1 Responding effectively is crucial to minimizing the morbidity and mortality of our patients while maximizing use of available resources. We can improve our level-headedness, efficiency, and department and hospital-level planning through simulation. This has particular implications for residency training with effects on education, preparedness, and wellness. The learners will (1) recognize state of mass casualty exercise as evidenced by verbalization or triaging by START (Simple Triage and Rapid Treatment) criteria, (2) triage several patients, including critically ill or peri-arrest acuities, according to START criteria, (3) recognize the need to limit care based on available resources, as evidenced by verbal orders or communication of priorities to team, and (4) limit emergency resuscitation, given limited resources, by only providing treatments and employing diagnostics that do not deplete limited time, staffing, and space inappropriately. A small-scale, high-fidelity simulation was created to replicate the pace and acuity of patients presenting in an MCI. Three critically injured patients with multiple gunshot wounds, represented by high-fidelity manikins with moulaged wounds, were presented over a 6-minute span. The team was allowed 10 minutes total to conduct life-saving measures, targeted evaluation, and disposition of the patients. The simulation was then adapted for use in a second institution's simulation center to replicate and validate the objectives given a different system. The learners were immediately verbally debriefed and feedback of the simulation, fidelity and appropriateness of the experience solicited. Unprompted, several of the learners volunteered that the efficacy of the experience was highly educational and valuable. Anonymized digital feedback was requested in the form of an online survey and was generally positive.The educational content was created by experts in simulation medicine and validated by content experts in the fields of Emergency Medicine, Trauma Surgery and Emergency Nursing. After the scenario ended, the learners were taken to a second room for debriefing by a trauma surgeon, an emergency medicine attending, and the nurse trauma educator. The actors were able to participate as secondary learners and were rotated out of simulation duties to participate in the debriefing. After this twenty-minute educational debrief, the learners were brought back to the simulation bay and were given a similar scenario. After this iteration, the team debriefed a second time. This hour schedule of cases and debrief was repeated a total of four times with a total of twelve individual learners. Suggestions and verbal feedback were noted for incorporation into appropriate committees or hospital departments. No formal assessment was done and inclusion was strictly on a voluntary basis. An evaluation of the session (on a Likert scale of 1-5) had six respondents which showed an average of 5 on how educational the session was, 4.8 on how realistic the session was, and 4.8 on how effective the session was. Simulation allows participants to safely gain practical experience in MCI management. The experience was well-received, and the learners verbalized increased confidence should they encounter an MCI in the future. We developed this simulation to give residents and nurses first-hand experience performing under high-stress, resource-limited conditions. We also had other learners observing the process which allowed for productive debriefing and planning for improvement. The ideas generated from this ultimately became part of the hospital's MCI response plan. The main takeaways were triage strategy and limited resource management. Mass casualty incident, mass gathering, penetrating trauma, high-fidelity simulation, team-based simulation, trauma center, hospital response planning.


Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system.
A simple and robust method for targeted mutagenesis in zebrafish has long been sought. Previous methods generate monoallelic mutations in the germ line of F0 animals, usually delaying homozygosity for the mutation to the F2 generation. Generation of robust biallelic mutations in the F0 would allow for phenotypic analysis directly in injected animals. Recently the type II prokaryotic clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated proteins (Cas) system has been adapted to serve as a targeted genome mutagenesis tool. Here we report an improved CRISPR/Cas system in zebrafish with custom guide RNAs and a zebrafish codon-optimized Cas9 protein that efficiently targeted a reporter transgene Tg(-5.1mnx1:egfp) and four endogenous loci (tyr, golden, mitfa, and ddx19). Mutagenesis rates reached 75-99%, indicating that most cells contained biallelic mutations. Recessive null-like phenotypes were observed in four of the five targeting cases, supporting high rates of biallelic gene disruption. We also observed efficient germ-line transmission of the Cas9-induced mutations. Finally, five genomic loci can be targeted simultaneously, resulting in multiple loss-of-function phenotypes in the same injected fish. This CRISPR/Cas9 system represents a highly effective and scalable gene knockout method in zebrafish and has the potential for applications in other model organisms.


Phylogenomics Reveals Three Sources of Adaptive Variation during a Rapid Radiation.
Speciation events often occur in rapid bursts of diversification, but the ecological and genetic factors that promote these radiations are still much debated. Using whole transcriptomes from all 13 species in the ecologically and reproductively diverse wild tomato clade (Solanum sect. Lycopersicon), we infer the species phylogeny and patterns of genetic diversity in this group. Despite widespread phylogenetic discordance due to the sorting of ancestral variation, we date the origin of this radiation to approximately 2.5 million years ago and find evidence for at least three sources of adaptive genetic variation that fuel diversification. First, we detect introgression both historically between early-branching lineages and recently between individual populations, at specific loci whose functions indicate likely adaptive benefits. Second, we find evidence of lineage-specific de novo evolution for many genes, including loci involved in the production of red fruit color. Finally, using a "PhyloGWAS" approach, we detect environment-specific sorting of ancestral variation among populations that come from different species but share common environmental conditions. Estimated across the whole clade, small but substantial and approximately equal fractions of the euchromatic portion of the genome are inferred to contribute to each of these three sources of adaptive genetic variation. These results indicate that multiple genetic sources can promote rapid diversification and speciation in response to new ecological opportunity, in agreement with our emerging phylogenomic understanding of the complexity of both ancient and recent species radiations.


Dependence of response properties on sparse connectivity in a spiking neuron model of the lateral geniculate nucleus.
We present a large-scale anatomically constrained spiking neuron model of the lateral geniculate nucleus (LGN), which operates solely with retinal input, relay cells, and interneurons. We show that interneuron inhibition and sparse connectivity between LGN cells could be key factors for explaining a number of observed classical and extraclassical response properties in LGN of monkey and cat. Among them are 1) weak orientation tuning, 2) contrast invariance of spatial frequency tuning in the absence of cortical feedback, 3) extraclassical surround suppression, and 4) orientation tuning of extraclassical surround suppression. The model also makes two surprising predictions: 1) a possible pinwheel-like spatial organization of orientation preference in the parvo layers of monkey LGN, much like what is seen in V1, and 2) a stimulus-induced trend (bias) in the orientation and phase preference of surround suppression, originating from the stimulus discontinuity between center and surround gratings rather than from specific circuitry.


Blood Component Therapy and Coagulopathy in Trauma: A Systematic Review of the Literature from the Trauma Update Group.
Traumatic coagulopathy is thought to increase mortality and its treatment to reduce preventable deaths. However, there is still uncertainty in this field, and available literature results may have been overestimated. We searched the MEDLINE database using the PubMed platform. We formulated four queries investigating the prognostic weight of traumatic coagulopathy defined according to conventional laboratory testing, and the effectiveness in reducing mortality of three different treatments aimed at contrasting coagulopathy (high fresh frozen plasma/packed red blood cells ratios, fibrinogen, and tranexamic acid administration). Randomized controlled trials were selected along with observational studies that used a multivariable approach to adjust for confounding. Strict criteria were adopted for quality assessment based on a two-step approach. First, we rated quality of evidence according to the Grading of Recommendations Assessment, Development and Evaluation (GRADE) criteria. Then, this rating was downgraded if other three criteria were not met: high reporting quality according to shared standards, absence of internal methodological and statistical issues not detailed by the GRADE system, and absence of external validity issues. With few exceptions, the GRADE rating, reporting and methodological quality of observational studies was "very low", with frequent external validity issues. The only two randomized trials retrieved were, instead, of high quality. Only weak evidence was found for a relation between coagulopathy and mortality. Very weak evidence was found supporting the use of fibrinogen administration to reduce mortality in trauma. On the other hand, we found high evidence that the use of 1:1 vs. 1:2 high fresh frozen plasma/packed red blood cells ratios failed to obtain a 12% mortality reduction. This does not exclude lower mortality rates, which have not been investigated. The use of tranexamic acid in trauma was supported by "high" quality evidence according to the GRADE classification but was downgraded to "moderate" for external validity issues. Tranexamic acid is effective in reducing mortality in trauma. The other transfusion practices we investigated have been inadequately studied in the literature, as well as the independent association between mortality and coagulopathy measured with traditional laboratory testing. Overall, in this field of research literature quality is poor.


The lesser of two evils: a contextual view of the English case of the conjoined twins.
In 2000 conjoined twin girls were born in Manchester, England. They were joined in such a way that it was impossible to separate them to enable both twins to survive. Their bodies were fused at the lower abdomen and they shared an aorta and a bladder. Their arms and legs were at right angles to their conjoined trunk. The situation of the conjoined twins is of supreme importance as a private tragedy for their family. The resulting litigation also presents an important landmark in English law. In deciding the fate of the two children, the Court of Appeal provided an authoritative review, analysis and application of family law and medical law with regard to neonates. The most significant legal legacy of the case, however, may well be in the field of criminal law by way of the court's interpretation and application of the defences of necessity and self-defence.


Body condition score as a selection tool for targeted selective treatment-based nematode control strategies in Merino ewes.
Sheep nematode control utilising refugia-based strategies have been shown to delay anthelmintic resistance, but the optimal indices to select individuals to be left untreated under extensive sheep grazing conditions are not clear. This experiment tested the hypothesis that high body condition can indicate ability of mature sheep to better cope with worms and therefore remain untreated in a targeted treatment programme. Adult Merino ewes from flocks on two private farms located in south-west Western Australia (Farm A, n = 271, and Farm B, n = 258) were measured for body condition score (BCS), body weight and worm egg counts (WEC) on four occasions between May and December (pre-lambing, lamb marking, lamb weaning and post-weaning). Half of the ewes in each flock received anthelmintic treatments to suppress WEC over the experimental period and half remained untreated (unless critical limits were reached). Response to treatment was analysed in terms of BCS change and percentage live weight change. No effect of high or low initial WEC groups was shown for BCS response, and liveweight responses were inconsistent. A relatively greater BCS response to treatment was observed in ewes in low BCS pre-lambing compared to better-conditioned ewes on one farm where nutrition was sub-optimal and worm burdens were high. Sheep in low body condition pre-lambing were more than three times more likely to fall into a critically low BCS (<2.0) if left untreated. Recommendations can be made to treat ewes in lower BCS and leave a proportion of the higher body condition sheep untreated in a targeted selective treatment programme, to provide a population of non-resistant worms to delay the development of resistance.


[Causes of deaths in Portugal and challenges in prevention].
In Portugal, the hope of living, in terms of probability, either at birth or at 65 years old, has reached levels that represent huge gains and certainly reflect the successful initiatives developed in many sectors in the last 30 years. In 2008-2010, the average life expectancy at birth was 79.20 years for men and women. By contrast, in 1980, the average life expectancy at birth was 71.78 years. Furthermore, premature mortality, defined as occurring before 70 years of age, expressed as rate, represents the likelihood of dying before this age, which is presently of 24.3%, meaning that one in four dies Portuguese citizen before reaching the age of 70. Main cause of deaths and the number of years in the Portuguese population that theoretically stops living when you die before the age of 70 (potential years lost), for both sexes, includes the following: traffic accidents with motor vehicles - 22,550 years; malignant tumor of trachea, bronchus and lung - 19,768 years, vascular brain diseases - 16,070 years, self-injury caused intentionally - 14,753 years. It's important to analyze these figures and its evolution in order to allow an understanding of their impact and implement effective corrective measures. The contribution of actions and programs to prevent some of the above causes of deaths and potential years lost is still to be objectively interpreted but it is well known the importance of reducing risks factors as tobacco and alcohol consumption, encouraging balanced nutrition in calories and composition, promoting regular physical exercise and improving information and health education on the reduction of many chronic non-communicable diseases, what in turn, reflects in the decline of premature mortality. It is essential to analyze the causes that anticipate the death of the Portuguese population and the cause of these causes, that is, their risk factors and design programs that reduce the exposure of citizens to avoidable risks, which is the essence of all public health programmes.


Efficacy of human Lyme disease vaccine formulations in a mouse model.
Although immunization with recombinant outer surface protein A (OspA) appears to protect mice against infection by the agent of Lyme disease, all reported experiments have involved formulations that would not be suitable for use in humans or have not used realistic challenges. This study was designed to determine whether vaccines prepared and used in a phase I human trial, including one currently being used for a phase II trial in sites with endemic Borrelia burgdorferi, conferred protection in the C3H/HeJ mouse model. The challenge was ticks collected from a major site of the trial. None of the vaccinated mice became infected or developed disease, whereas 60% of unvaccinated mice became infected. Spirochetes were destroyed within the guts of virtually all recovered challenge ticks. These preparations of recombinant OspA effectively induced immunity to protect mice from Lyme disease when bitten by ticks collected from a field trial site.


Pregnenolone for cognition and mood in dual diagnosis patients.
Mood and substance-use disorders are both associated with cognitive deficits. Patients with mood and substance-use disorders have poorer cognition than patients with only a mood disorder. Pregnenolone may have beneficial effects on mood and cognition. In a proof-of-concept investigation, 70 participants with bipolar disorder or recurrent major depressive disorder and history of substance abuse/dependence (abstinent for > or =14days prior to enrollment) were randomly assigned to receive pregnenolone (titrated to 100mg/day) or placebo for 8weeks. Participants were assessed using the Mini International Neuropsychiatric Interview, Hamilton Rating Scale for Depression (HRSD), Young Mania Rating Scale (YMRS), Rey Auditory Verbal Learning Test (RAVLT), Trail Making Test (TMT-B), and Stroop Test. Mood was assessed bi-weekly, while cognition was evaluated at baseline, and weeks 4 and 8. Groups were compared using a random regression analysis that used all of the available data. The pregnenolone group showed trends toward greater improvement, relative to placebo, on the HRSD and YMRS. A post hoc analysis of completers found a statistically significant reduction in HRSD scores with pregnenolone as compared to placebo. Pregnenolone appeared to be safe and well tolerated. Findings suggest that pregnenolone use may be associated with some improvement in manic and depressive symptoms, but not cognition in depressed patients with a history of substance use. Larger trials examining the impact of pregnenolone on mood in more narrowly defined populations may be warranted.


Data partitions, Bayesian analysis and phylogeny of the zygomycetous fungal family Mortierellaceae, inferred from nuclear ribosomal DNA sequences.
Although the fungal order Mortierellales constitutes one of the largest classical groups of Zygomycota, its phylogeny is poorly understood and no modern taxonomic revision is currently available. In the present study, 90 type and reference strains were used to infer a comprehensive phylogeny of Mortierellales from the sequence data of the complete ITS region and the LSU and SSU genes with a special attention to the monophyly of the genus Mortierella. Out of 15 alternative partitioning strategies compared on the basis of Bayes factors, the one with the highest number of partitions was found optimal (with mixture models yielding the best likelihood and tree length values), implying a higher complexity of evolutionary patterns in the ribosomal genes than generally recognized. Modeling the ITS1, 5.8S, and ITS2, loci separately improved model fit significantly as compared to treating all as one and the same partition. Further, within-partition mixture models suggests that not only the SSU, LSU and ITS regions evolve under qualitatively and/or quantitatively different constraints, but that significant heterogeneity can be found within these loci also. The phylogenetic analysis indicated that the genus Mortierella is paraphyletic with respect to the genera Dissophora, Gamsiella and Lobosporangium and the resulting phylogeny contradict previous, morphology-based sectional classification of Mortierella. Based on tree structure and phenotypic traits, we recognize 12 major clades, for which we attempt to summarize phenotypic similarities. M. longicollis is closely related to the outgroup taxon Rhizopus oryzae, suggesting that it belongs to the Mucorales. Our results demonstrate that traits used in previous classifications of the Mortierellales are highly homoplastic and that the Mortierellales is in a need of a reclassification, where new, phylogenetically informative phenotypic traits should be identified, with molecular phylogenies playing a decisive role.


Toll-like receptor 3 activation drives the inflammatory response in oxygen-induced retinopathy in rats.
Ischaemia is one of the most important causes of blindness. The purpose of this study was to investigate the potential role and mechanisms by which toll-like receptor 3 (TLR3) influences the progression of the inflammatory response in a rat model of oxygen-induced retinopathy (OIR). OIR rat models were successfully established and received single intravitreal injections of polyinosine-polycytidylic acid (poly (I:C)) and anti-TLR3 antibody, respectively, on postnatal day 17 (P17). Pathological retinal neovascularisation was evaluated by haematoxylin and eosin staining and immunohistochemistry with Isolectin B4 FITC (fluorescein isothyocyanate). Retinal expressions of TLR3 and nuclear factor kappa B (NF-κB) were measured using real-time PCR, immunohistochemistry and western blot. Furthermore, interleukin-6 (IL-6) and tumour necrosis factor α (TNFα) expression levels were assessed with real-time PCR and ELISA. Both gene and protein expression levels of TLR3 and NF-κB were significantly elevated in the retinas of OIR rats compared to the controls. Increased IL-6 and TNFα expression levels were also observed in the retinas of OIR rats. Furthermore, TLR3 signalling pathway components, including NF-κB and IL-6/TNFα, were markedly upregulated upon stimulation with poly(I:C). In addition, the pre-treatment of TLR3 neutralising antibody in OIR models significantly decreased TLR3 and NF-κB expressions, as well as related inflammatory factors IL-6/TNFα expression. Our results suggest that upregulation of the TLR3 signalling pathway is involved in the pro-inflammatory response in OIR rat retinas.


Evaluation of serum levels of zinc, copper, iron, and zinc/copper ratio in cutaneous leishmaniasis.
The purpose of this study was to evaluate the levels of zinc (Zn), copper (Cu), iron (Fe) and zinc/ copper ratio in the serum of patients with cutaneous leishmaniasis in Qom Province, center of Iran. Serum levels of zinc and copper were determined by flame atomic absorption spectrophotometer and serum iron concentration was measured by using an Auto Analyzer. The study group consisted of 60 patients with cutaneous leishmaniasis and the control group of 100 healthy volunteers from the same area who were not exposed to cutaneous leishmaniasis. There were no statistically significant differences in age and body mass index between the two groups. Serum Zn (P< 0.001) and Fe (P< 0.05) levels were lower in patients with cutaneous leishmaniasis than the control group. We also found serum Cu concentration (P< 0.05) in the patient group was significantly higher than that of the control group. However, zinc/ copper ratio (P< 0.001) was lower in patients with cutaneous leishmaniasis than in the control group. Our data indicated that Zn/Cu ratio was significantly lower in patients with CL as compared to the controls. Earlier reports suggest that, this ratio imbalance could be a useful marker for immune dysfunction in leishmaniasis. There was also strong association of Zn, Cu and Fe with CL. It suggests the use of blood zinc, copper, iron concentration and the copper/zinc ratio (Zn/Cu), as a means for estimating the prognosis of CL.


Additive value of "otosclerosis-weighted" images for the CT diagnosis of fenestral otosclerosis.
Background Otosclerotic foci are usually seen as minute low-density lesions and this may be attributed to relatively low sensitivity on visual assessment using computed tomography (CT). Otosclerotic foci can be detected by using the accurate region of interest (ROI) setting, while small ROI settings by less-experienced radiologists may result in false negative findings. Purpose To evaluate the diagnostic ability of our proposed method ("otosclerosis-weighted" imaging [OWI]), which is based on reversing the density, compared with conventional CT (CCT) imaging alone. Material and Methods Temporal bone CTs of consecutive patients with otosclerosis were analyzed. Gender- and age-matched control participants were also included. All CT images were obtained using a 64-detector row scanner. OWI was obtained by extracting the temporal bone region using the threshold technique and reversing the density (black to white). Four independent radiologists took part in two reading sessions. In the first session, the observers read only CCT imaging. In the second session, they read OWI along with the CCT imaging. Sensitivity was assessed for the four readers. Results Thirty temporal bones of 25 patients with otosclerosis (3 men, 22 women; mean age, 53.9 ± 9.0 years) and 30 temporal bones of 30 control participants (4 men, 26 women; mean age, 44.0 ± 16.2 years) were included. For all observers, reading with a combination of the two methods was associated with a higher sensitivity (63.3-80.0%) than with conventional CT images alone (30.0-60.0%; P < 0.05, each). Conclusion Application of our proposed method based on threshold value may help detect foci of fenestral otosclerosis.


Melittin induces PTCH1 expression by down-regulating MeCP2 in human hepatocellular carcinoma SMMC-7721 cells.
Hepatocellular carcinoma (HCC) has a high mortality rate worldwide and still remains to be a noticeable public health problem. Therefore, new remedies are urgently needed. Melittin, a major component of bee venom, is known to suppress cell growth in various cancers including HCC. However, the mechanism of the anticancer effect of melittin on HCC has not been fully elucidated. It has been reported that Methyl-CpG binding protein 2 (MeCP2) plays a key role in tumor proliferation, apoptosis, migration and invasion. In the present study, we found the high expression of MeCP2 in human HCC tissues and in the SMMC-7721 cell line. MeCP2 silencing inhibited cell proliferation, while over-expression of MeCP2 promoted cell growth in SMMC-7721 cells. It indicates that MeCP2 may be an attractive target for human HCC. We further found that melittin could inhibit cell proliferation by reducing MeCP2 expression in vitro. Interestingly, the inhibitory effect of melittin on cell proliferation was due to a delay in G0/G1 cell cycle progression, without influencing cell apoptosis. Next, we investigated the potential molecular mechanisms and found that MeCP2 could modulate Shh signaling in SMMC-7721 cells. Further study indicates that melittin may induce the demethylation of PTCH1 promoter, resulting in the increased expression of PTCH1. Furthermore, the expression of Shh and GLI1 was significantly lowered upon treatment of melittin. These results suggest that melittin can block Shh signaling in vitro. In short, these results indicate that melittin inhibits cell proliferation by down-regulating MeCP2 through Shh signaling in SMMC-7721 cells.


Molecular specialization of astrocyte processes at nodes of Ranvier in rat optic nerve.
The HNK-1 and L2 monoclonal antibodies are thought to recognize identical or closely associated carbohydrate epitopes on a family of neural plasma membrane glycoproteins, including myelin-associated glycoprotein, the neural cell adhesion molecule, and the L1 and J1 glycoproteins, all of which have been postulated to play a part in mediating cell-cell interactions in the nervous system. We have used these two antibodies in immunofluorescence and immunogold-electron microscopic studies of semithin and ultrathin frozen sections of adult rat optic nerve, respectively, and we show that they bind mainly to astrocyte processes around nodes of Ranvier. Most other elements of the nerve, including astrocyte cell bodies and large astrocytic processes, are not labeled by the antibodies. To our knowledge, this is the first demonstration that perinodal astrocyte processes are biochemically specialized. We provide evidence that one of the HNK-1+/L2+ molecules concentrated around perinodal astrocyte processes is the J1 glycoprotein; our findings, taken together with previously reported observations, suggest that the other known HNK-1+/L2+ molecules are not concentrated on these processes. Since anti-J1 antibodies previously have been shown to inhibit neuron to astrocyte adhesion in vitro, we hypothesize that J1 may play an important part in the axon-glial interactions that presumably are involved in the assembly and/or maintenance of nodes of Ranvier.


Multicapillary Flow Reactor: Synthesis of 1,2,5-Thiadiazepane 1,1-Dioxide Library Utilizing One-Pot Elimination and Inter-/Intramolecular Double aza-Michael Addition Via Microwave-Assisted, Continuous-Flow Organic Synthesis (MACOS).
A microwave-assisted, continuous-flow organic synthesis (MACOS) protocol for the synthesis of functionalized 1,2,5-thiadiazepane 1,1-dioxide library, utilizing a one-pot elimination and inter-/intramolecular double aza-Michael addition strategy is reported. The optimized protocol in MACOS was utilized for scale-out and further extended for library production using a multicapillary flow reactor. A 50-member library of 1,2,5-thiadiazepane 1,1-dioxides was prepared on a 100- to 300-mg scale with overall yields between 50 and 80% and over 90 % purity determined by proton nuclear magnetic resonance (1H-NMR) spectroscopy.


Brain's reward circuits mediate itch relief. a functional MRI study of active scratching.
Previous brain imaging studies investigating the brain processing of scratching used an exogenous intervention mimicking scratching, performed not by the subjects themselves, but delivered by an investigator. In real life, scratching is a conscious, voluntary, controlled motor response to itching, which is directed to the perceived site of distress. In this study we aimed to visualize in real-time by brain imaging the core mechanisms of the itch-scratch cycle when scratching was performed by subjects themselves. Secondly, we aimed to assess the correlations between brain patterns of activation and psychophysical ratings of itch relief or pleasurability of scratching. We also compared the patterns of brain activity evoked by self-scratching vs. passive scratching. We used a robust tridimensional Arterial Spin Labeling fMRI technique that is less sensitive to motion artifacts: 3D gradient echo and spin echo (GRASE)--Propeller. Active scratching was accompanied by a higher pleasurability and induced a more pronounced deactivation of the anterior cingulate cortex and insula, in comparison with passive scratching. A significant involvement of the reward system including the ventral tegmentum of the midbrain, coupled with a mechanism deactivating the periaqueductal gray matter (PAG), suggests that itch modulation operates in reverse to the mechanism known to suppress pain. Our findings not only confirm a role for the central networks processing reward in the pleasurable aspects of scratching, but also suggest they play a role in mediating itch relief.


Cancers in children younger than age 16 years in Kenya.
To determine the number of cancers seen in six months in children aged below sixteen years and to look for any associations with breastfeeding, parental ages, smoking, water availability and consanguinity. Prospective case study from June 1997 through December 1997. Seven provincial hospitals and Kenyatta National Referral Hospital. Both sexes with tissue proven malignancy. By obtaining clinical information, and laboratory evaluatory results. Proportions of vital statistics, anatomic site, stage and histology of cancer, vaccination, breastfeeding, congenital abnormality, parental vital statistics, education, smoking, water availability status and consanguinity. One hundred and fifty seven cases were evaluated. Male to female (M:F) ratio was 1.5:1. Mean ages: patients 6.5, fathers 36.6, mothers 31.1 years. Histologic types were: Burkitt's lymphoma (BL) 45%, nephroblastoma 14%, Hodgkin's lymphoma 9.5%, acute lymphocytic leukaemia 7.6%, retinoblastoma 5.7%, acute myeloid leukaemia 5.1%. The age distribution was as follows: 0-5 years, 46.5%, 6-10, 39.5% and 11-16 years 14%. The birth places of the children were as follows: Nyanza 25%, Central 21%, Eastern 16%, Western 13%, Rift Valley 9%, Coast 8%, Nairobi 8% and North Eastern had less than one per cent. The ethnic distribution was; Luo 25%, Kikuyu 22%, Luhya 13%, Kamba 10%, Coastal 6%, Kalenjin 6%, others 19%. Fifty per cent of the cases studied were breastfed for over six months, 34% for 3-6 months, five per cent less than three months and eleven per cent were not breastfed. Eighty five per cent of the cases received complete vaccination. The modal number of siblings was four and only 49% of parents received formal education. No consanguinity was noted in the studied population. The abdomen and jaw were main anatomic sites involved by the tumours. This study showed that the commonest childhood tumour is Burkitt's lymphoma and nephroblastoma is the commonest solid tumour. While the results demonstrate geographic and ethnic variations, other factors studied do not appear to influence the characteristics of cancers in the cases investigated.


Navigating choice in multiple sclerosis management.
With the advent of modern immunotherapies for relapsing-remitting multiple sclerosis (RRMS) and the increasing amount of treatment options on the market, MS has evolved as a treatable disease. Yet, at the same time, new challenges for the treating neurologists arise. This review article covers some of these challenges, including when and how to start treatment, treatment monitoring, and finally considerations on what the increasing choice in treatment options brings to disease management and longer-term planning. Among others, these important issues comprise pregnancy, treatment sequencing, switching or even stopping treatment. The ultimate goal for navigating choices in RRMS management is to choose the right drug for the right patient at the right time Throughout the article, there is a strong focus on practical aspects and individual decision making in MS to meet the concept of personalized medicine.


Early gene expression along the animal-vegetal axis in sea urchin embryoids and grafted embryos.
The HE gene is the earliest strictly zygotic gene activated during sea urchin embryogenesis. It is transiently expressed in a radially symmetrical domain covering the animal-most two-thirds of the blastula. The border of this domain, which is orthogonal to the primordial animal-vegetal axis, is shifted towards the animal pole in Li+-treated embryos. Exogenous micromeres implanted at the animal pole of whole embryos, animal or vegetal halves do not modify the extent and localization of the HE expression domain. In grafted embryos or animal halves, the Li+ effect is not affected by the presence of ectopic micromeres at the animal pole. A Li+-induced shift of the border, similar to that seen in whole embryos, occurs in embryoids developing from animal halves isolated from 8-cell stage embryos or dissected from unfertilised eggs. Therefore, the spatial restriction of the HE gene is not controlled by the inductive cascade emanating from the micromeres and the patterning along the AV-axis revealed by Li+ does not require interactions between cells from the animal and vegetal halves. This suggests that maternal primary patterning in the sea urchin embryo is not limited to a small vegetal center but extends along the entire AV axis.


Effects on kidney disease, fertility and development in mice inheriting a protein-truncating Denys-Drash syndrome allele (Wt1tmT396).
Denys-Drash syndrome (DDS) is caused by heterozygous mutations of the Wilms' tumour suppressor gene, WT1, characterised by early-onset diffuse mesangial sclerosis often associated with male pseudohermaphroditism and/or Wilms' tumourigenesis. Previously, we reported that the Wt1tmT396 allele induces DDS kidney disease in mice. In the present study heterozygotes (Wt1tmT396/+) were generated on inbred (129/Ola), crossbred (B6/129) and MF1 second backcross (MF1-N2) backgrounds. Whereas male heterozygotes on each background were fertile, inbred heterozygous females were infertile. Kidney disease (proteinuria and sclerosis) was not congenital and developed significantly earlier in inbred mice, although with variable onset. Disease onset in MF1-N2 stocks occurred later in Wt1tmT396/+ mice than reported previously for Wt1R394W/+ mice, and while no kidney disease has been reported in B6/129 Wt1+/- mice, B6/129 Wt1tmT396/+ mice were affected. Offspring of both male and female B6/129 and MF1-N2 Wt1tmT396/+ mice developed kidney disease, but its incidence was significantly higher in offspring of female heterozygotes. Wt1tmT396/tmT396 embryos exhibited identical developmental abnormalities to those reported for Wt1-/- embryos. The results indicate that the Wt1 (tmT396) allele does not predispose to Wilms' tumourigenesis or male pseudohermaphroditism, its effect on kidney disease and female fertility depends on genetic background, stochastic factors may affect disease onset, and disease transmission is subject to a partial parent-of-origin effect. Since the Wt1tmT396 allele has no detectable intrinsic functional activity in vivo, and kidney disease progression is affected by the type of Wt1 mutation, the data support the view that DDS nephropathy results from a dominant-negative action rather than WT1 haploinsufficiency or gain-of-function.


Loops versus lines and the compression stiffening of cells.
Both animal and plant tissue exhibit a nonlinear rheological phenomenon known as compression stiffening, or an increase in moduli with increasing uniaxial compressive strain. Does such a phenomenon exist in single cells, which are the building blocks of tissues? One expects an individual cell to compression soften since the semiflexible biopolymer-based cytoskeletal network maintains the mechanical integrity of the cell and in vitro semiflexible biopolymer networks typically compression soften. To the contrary, we find that mouse embryonic fibroblasts (mEFs) compression stiffen under uniaxial compression via atomic force microscopy studies. To understand this finding, we uncover several potential mechanisms for compression stiffening. First, we study a single semiflexible polymer loop modeling the actomyosin cortex enclosing a viscous medium modeled as an incompressible fluid. Second, we study a two-dimensional semiflexible polymer/fiber network interspersed with area-conserving loops, which are a proxy for vesicles and fluid-based organelles. Third, we study two-dimensional fiber networks with angular-constraining crosslinks, i.e. semiflexible loops on the mesh scale. In the latter two cases, the loops act as geometric constraints on the fiber network to help stiffen it via increased angular interactions. We find that the single semiflexible polymer loop model agrees well with the experimental cell compression stiffening finding until approximately 35% compressive strain after which bulk fiber network effects may contribute. We also find for the fiber network with area-conserving loops model that the stress-strain curves are sensitive to the packing fraction and size distribution of the area-conserving loops, thereby creating a mechanical fingerprint across different cell types. Finally, we make comparisons between this model and experiments on fibrin networks interlaced with beads as well as discuss implications for single cell compression stiffening at the tissue scale.


Assay of methylmalonic acid in the serum of patients with cobalamin deficiency using capillary gas chromatography-mass spectrometry.
To determine the incidence of elevated levels of serum methylmalonic acid in patients with cobalamin deficiency, we utilized a new capillary gas chromatographic-mass spectrometric technique to measure methylmalonic acid in the serum of 73 patients with clinically confirmed cobalamin deficiency. Values ranged from 55 to 22,300 ng/ml, and 69 of the 73 patients had values above the normal range of 19-76 ng/ml as determined for 50 normal blood donors. In the cobalamin-deficient patients, serum methylmalonic acid was significantly correlated with the serum folate level and the degree of neurologic involvement. Some patients with pernicious anemia who were intermittently treated with cyanocobalamin were found to have elevated serum levels of methylmalonic acid while free of hematologic and neurologic abnormalities. A cobalamin-deficient patient is described with a normal serum cobalamin and an elevated serum methylmalonic acid. We conclude that the ability to measure methylmalonic acid in human serum will be useful in studies designed to determine the incidence of cobalamin deficiency in various patient populations.


Application of a statistical design to the optimization of culture medium for recombinant interferon-gamma production by Chinese hamster ovary cells.
The importance of serum-free medium components on the growth of Chinese hamster ovary (CHO) cells and production of recombinant human interferon(IFN)-gamma was investigated. The complexity of the medium led to the adoption of a statistical optimization approach based on a Plackett-Burman design. From this analysis a set of nutritional components was identified as important for cell growth and recombinant protein production. Glycine was identified as an important determinant of specific growth rate, whereas for cell production bovine serum albumin (BSA), phenylalanine and tyrosine were also identified as important. BSA, sodium pyruvate, glutamate, methionine, proline, histidine, hydroxyproline, tyrosine and phenylalanine were shown to be important for IFN-gamma production. Other medium components, such as insulin, arginine, aspartate and serine produced an inhibitory effect on both cell growth and IFN-gamma production. The effect of the stimulatory nutrients as a whole group was tested by increasing their concentration in the medium. A significant improvement in specific cell growth rate, cell production and IFN-gamma production (up to 45%) was achieved on both shake-flask and fermentor cultures. An increase in the medium concentration of the negative variables had only a small inhibitory effect (approximately 10%) on the same parameters. Analysis of the effects of the group of stimulatory amino acids and BSA on CHO cell growth showed that the effect of the former was independent of BSA.


Changes in spectral sensitivity of multiplier phototubes resulting from changes in temperature.
The change in cathode spectral sensitivity of Westinghouse WX 4582 (S-11) and RCA 1P21(S-4) multiplier phototubes was measured across the visible spectrum, i.e., from 400 nm to 700 nm, for various temperature changes, both increases and decreases. Two methods were used for these measurements, viz., an adaptation of the Hardy spectrophotometer and the use of an environmental chamber. A decrease in temperature usually caused an increase in sensitivity in the short wavelength part of the spectrum and a decrease (as much as 90% at 700 nm) in the long wavelength part of the spectrum, with the crossover point (no appreciable change of sensitivity with change of temperature) at about 590 nm. An increase in temperature was accompanied by a reversal of spectral sensitivity changes, i.e., a decrease in the short wavelength part of spectrum and an increase in the long wavelength part of the spectrum. The change in cathode sensitivity varied with different types of phototubes and with phototubes of the same type.


Platelet-derived growth factor is a potent biologic response modifier of T cells.
Freshly isolated lymph node (LN) cells cultured in serum-containing medium were restricted to produce primarily interleukin 2 (IL-2) subsequent to T cell activation. Only minimal amounts of IL-4, IL-5, or interferon gamma (IFN-gamma) were produced under these conditions. Similar populations of LN cells cultured in serum-free medium were able to produce a variety of lymphokines after T cell activation, with the relative quantities of each species being dependent upon the lymphoid organ source of the lymphocytes. A similar relationship in the patterns of lymphokines produced by activated T cell hybridomas maintained under serum-free conditions was also observed, whereas activation in serum-supplemented media resulted in a predominant restriction to the secretion of IL-2. Additional studies determined that the entity in serum responsible for restricting T cell function in vitro was platelet-derived growth factor (PDGF). The PDGF-BB isoform was established to be the most active in the regulation of T cell function, enhancing IL-2 while depressing the production of IL-4, IL-5, and IFN-gamma at concentrations below 1 ng/ml. PDGF-AB was also found to be quite active, however, this isoform of PDGF was incapable of influencing IFN-gamma production at the concentrations tested. PDGF-AA was very weakly active. It therefore appears that PDGF, acting primarily through a beta receptor subunit (either alpha/beta- or beta/beta-type receptors) is able to influence profoundly the behavior of T cells, with some of its modulatory effects exhibiting isoform specificity. This is reflected by an enhancement in the production of IL-2, while simultaneously depressing the secretion of IL-4, IL-5, and IFN-gamma (PDGF-BB only) after T cell activation. Kinetic studies, where cell supernatants were analyzed both 24 and 48 h after T cell activation, suggested that "desensitization" to PDGF influences can occur naturally in vitro. Those species of lymphokines that were inhibited by PDGF over the first 24 h after activation could be produced at normal levels over the subsequent 24-h period. Finally, lymphokines maintained in the presence of PDGF-BB for greater than 24 h before their activation lost sensitivity to this growth factor. These cells regained responsiveness to PDGF after an additional incubation period in PDGF-free medium. Collectively, our data imply that the pattern of T cell lymphokines produced, plus the kinetics of their production after activation, are being controlled by the potent serum growth factor PDGF.(ABSTRACT TRUNCATED AT 400 WORDS)


Transformation of Arabidopsis thaliana with Agrobacterium tumefaciens.
Transformed Arabidopsis thaliana plants have been produced by a modified leaf disk transformation-regeneration method. Leaf pieces from sterilely grown plants were precultured for 2 days and inoculated with an Agrobacterium tumefaciens strain containing an avirulent Ti (tumor-inducing) plasmid with a chimeric gene encoding hygromycin resistance. After cocultivation for 2 days, the leaf pieces were placed on a medium that selects for hygromycin resistance. Shoots regenerated within 3 months and were excised, rooted, and transferred to soil. Transformation was confirmed by opine production, hygromycin resistance, and DNA blot hybridization of both primary transformants and progeny. This process for producing transgenic Arabidopsis plants should enhance the usefulness of the species for experimental biology.


Mitogen-activated protein kinase-dependent interleukin-1α intracrine signaling is modulated by YopP during Yersinia enterocolitica infection.
Yersinia enterocolitica is a food-borne pathogen that preferentially infects the Peyer's patches and mesenteric lymph nodes, causing an acute inflammatory reaction. Even though Y. enterocolitica induces a robust inflammatory response during infection, the bacterium has evolved a number of virulence factors to limit the extent of this response. We previously demonstrated that interleukin-1α (IL-1α) was critical for the induction of gut inflammation characteristic of Y. enterocolitica infection. More recently, the known actions of IL-1α are becoming more complex because IL-1α can function both as a proinflammatory cytokine and as a nuclear factor. In this study, we tested the ability of Y. enterocolitica to modulate intracellular IL-1α-dependent IL-8 production in epithelial cells. Nuclear translocation of pre-IL-1α protein and IL-1α-dependent secretion of IL-8 into the culture supernatant were increased during infection with a strain lacking the 70-kDa virulence plasmid compared to the case during infection with the wild type, suggesting that Yersinia outer proteins (Yops) might be involved in modulating intracellular IL-1α signaling. Infection of HeLa cells with a strain lacking the yopP gene resulted in increased nuclear translocation of pre-IL-1α and IL-1α-dependent secretion of IL-8 similar to what is observed with bacteria lacking the virulence plasmid. YopP is a protein acetylase that inhibits mitogen-activated protein kinase (MAP kinase)- and NF-κB-dependent signal transduction pathways. Nuclear translocation of pre-IL-1α and IL-1α-dependent secretion of IL-8 in response to Yersinia enterocolitica infection were dependent on extracellular signal-regulated kinase (ERK) and p38 MAP kinase signaling but independent of NF-κB. These data suggest that Y. enterocolitica inhibits intracellular pre-IL-1α signaling and subsequent proinflammatory responses through inhibition of MAP kinase pathways.


Home and Birth Center Birth in the United States: Time for Greater Collaboration Across Models of Care.
There has been a small, but significant, increase in community births (home and birth-center births) in the United States in recent years. The rate increased by 20% from 2004 to 2008, and another 59% from 2008 to 2012, though the overall rate is still low at less than 2%. Although the United States is not the only country with a large majority of births occurring in the hospital, there are other high-resource countries where home and birth-center birth are far more common and where community midwives (those attending births at home and in birth centers) are far more central to the provision of care. In many such countries, the differences in perinatal outcomes between hospital and community births are small, and there are lower rates of maternal morbidity in the community setting. In the United States, perinatal mortality appears to be higher for community births, though there has yet to be a national study comparing outcomes across settings that controls for planned place of birth. Rates of intervention, including cesarean delivery, are significantly higher in hospital births in the United States. Compared with the United States, countries that have higher rates of community births have better integrated systems with clearer national guidelines governing risk criteria and planned birth location, as well as transfer to higher levels of care. Differences in outcomes, systems, approaches, and client motivations are important to understand, because they are critical to the processes of person-centered care and to risk reduction across all birth settings.


Individual capacity-building approaches in a global pharmaceutical systems strengthening program: a selected review.
Medicines use related challenges such as inadequate adherence, high levels of antimicrobial resistance and preventable adverse drug reactions have underscored the need to incorporate pharmaceutical services to help achieve desired treatment outcomes, and protect patients from inappropriate use of medicines. This situation is further constrained by insufficient numbers of pharmaceutical personnel and inappropriate skill mix. Studies have addressed individual capacity building approaches of logistics, supply chain or disease specific interventions but few have documented those involving such pharmacy assistants/professionals, or health workers/professionals charged with improving access and provision of pharmaceutical services. We examined how different training modalities have been employed and adapted to meet country-specific context and needs by a global pharmaceutical systems strengthening program in collaboration with a country's Ministry of Health and local stakeholders. Structured, content analysis of training approaches from twelve selected countries and a survey among conveniently selected trainees in Bangladesh and Ethiopia. Case-based learning, practice and feedback, and repetitive interventions such as post-training action plan, supportive supervision and mentoring approaches are effective, evidence-based training techniques. In Ethiopia and Bangladesh, over 94% of respondents indicated that they have improved or developed skills or competencies as a result of the program's training activities. Supportive supervision structures and mentorship have been institutionalized with appropriate management structures. National authorities have been sensitized to secure funding from domestic resources or from the global fund grants for post-training follow-up initiatives. The Pharmaceutical Leadership Development Program is an effective, case-based training modality that motivates staff to develop quality-improvement interventions and solve specific challenges. Peer-to-peer learning mechanisms than traditional didactic methods was a preferred intervention among high level government officials both within country and between countries. Interventions must involve local institutions in the design and delivery of content for both pre-service and in-service training as well as web-based methods where feasible. Such efforts would meet the changing demand in the pharmaceutical system, and promote the ownership of the human capacity development interventions. The cost-effective partnership with universities demonstrate that competency based pre-service training will prepare the future pharmaceutical workforce with a critical foundation of knowledge and skills required to meet the growing demand for patient-centered pharmaceutical services in resource-constrained countries.


Imaging appearances of the sternum and sternoclavicular joints.
The sternum and sternoclavicular joints--critical structures of the anterior chest wall--may be affected by various anatomic anomalies and pathologic processes, some of which require treatment. Pectus excavatum and pectus carinatum are common congenital anomalies that are usually benign but may warrant surgical treatment if they cause compression of vital internal structures. By contrast, developmental variants such as the sternal foramen are asymptomatic and do not require further evaluation or treatment. Arthritides of the sternoclavicular joint (osteoarthritis, septic arthritis, and seronegative arthropathies) are common and must be differentiated before an appropriate management method can be selected. The recognition of complications of sternotomy (eg, sternal dehiscence, secondary osteomyelitis) is critical to avoid life-threatening sequelae such as acute mediastinitis. Likewise, the detection of sternal fractures and sternoclavicular dislocations is important, especially where they impinge on vital structures. In addition, sternal malignancies (most commonly, metastases and chondrosarcoma) must be distinguished from benign neoplasms. To achieve accurate and timely diagnoses that facilitate appropriate treatment, radiologists must be familiar with the appearances of these normal anatomic variants and diseases of the sternum.