Efficient DNP NMR of membrane proteins: sample preparation protocols, sensitivity, and radical location.
Although dynamic nuclear polarization (DNP) has dramatically enhanced solid-state NMR spectral sensitivities of many synthetic materials and some biological macromolecules, recent studies of membrane-protein DNP using exogenously doped paramagnetic radicals as polarizing agents have reported varied and sometimes surprisingly limited enhancement factors. This motivated us to carry out a systematic evaluation of sample preparation protocols for optimizing the sensitivity of DNP NMR spectra of membrane-bound peptides and proteins at cryogenic temperatures of ~110 K. We show that mixing the radical with the membrane by direct titration instead of centrifugation gives a significant boost to DNP enhancement. We quantify the relative sensitivity enhancement between AMUPol and TOTAPOL, two commonly used radicals, and between deuterated and protonated lipid membranes. AMUPol shows ~fourfold higher sensitivity enhancement than TOTAPOL, while deuterated lipid membrane does not give net higher sensitivity for the membrane peptides than protonated membrane. Overall, a ~100 fold enhancement between the microwave-on and microwave-off spectra can be achieved on lipid-rich membranes containing conformationally disordered peptides, and absolute sensitivity gains of 105-160 can be obtained between low-temperature DNP spectra and high-temperature non-DNP spectra. We also measured the paramagnetic relaxation enhancement of lipid signals by TOTAPOL and AMUPol, to determine the depths of these two radicals in the lipid bilayer. Our data indicate a bimodal distribution of both radicals, a surface-bound fraction and a membrane-bound fraction where the nitroxides lie at ~10 Å from the membrane surface. TOTAPOL appears to have a higher membrane-embedded fraction than AMUPol. These results should be useful for membrane-protein solid-state NMR studies under DNP conditions and provide insights into how biradicals interact with phospholipid membranes.


Human stroma and epithelium co-culture in a microfluidic model of a human prostate gland.
The prostate is a walnut-sized gland that surrounds the urethra of males at the base of the bladder comprising a muscular portion, which controls the release of urine, and a glandular portion, which secretes fluids that nourish and protect sperms. Here, we report the development of a microfluidic-based model of a human prostate gland. The polydimethylsiloxane (PDMS) microfluidic device, consisting of two stacked microchannels separated by a polyester porous membrane, enables long-term in vitro cocultivation of human epithelial and stromal cells. The porous separation membrane provides an anchoring scaffold for long-term culturing of the two cell types on its opposite surfaces allowing paracrine signaling but not cell crossing between the two channels. The microfluidic device is transparent enabling high resolution bright-field and fluorescence imaging. Within this coculture model of a human epithelium/stroma interface, we simulated the functional development of the in vivo human prostate gland. We observed the successful differentiation of basal epithelial cells into luminal secretory cells determined biochemically by immunostaining with known differentiation biomarkers, particularly androgen receptor expression. We also observed morphological changes where glandlike mounds appeared with relatively empty centers reminiscent of prostatic glandular acini structures. This prostate-on-a-chip will facilitate the direct evaluation of paracrine and endocrine cross talk between these two cell types as well as studies associated with normal vs disease-related events such as prostate cancer.


Population genetic analysis of Helicobacter pylori by multilocus enzyme electrophoresis: extensive allelic diversity and recombinational population structure.
Genetic diversity and relationships in 74 Helicobacter pylori isolates recovered from patients assigned to distinct clinical categories were estimated by examination of allelic variation in six genes encoding metabolic housekeeping enzymes by multilocus enzyme electrophoresis. Seventy-three distinct allele profiles, representing multilocus chromosomal genotypes, were identified. All six loci were highly polymorphic, with an average of 11.2 alleles per locus. The mean genetic diversity in the sample was 0.735, a value that exceeds the level of diversity recorded in virtually all bacterial species studied by multilocus enzyme electrophoresis. A high frequency of occurrence of null alleles (lack of enzyme activity) was identified and warrants further investigation at the molecular level. Lack of linkage disequilibrium (nonrandom association (of alleles over loci) indicates that horizontal transfer and recombination of metabolic enzyme genes have contributed to the generation of chromosomal diversity in H. pylori. In this sample of isolates, there was no statistically significant association of multilocus enzyme electrophoretic types or cluster of related chromosomal types and disease category.


Impairment of EEG desynchronisation before and during movement and its relation to bradykinesia in Parkinson's disease.
It has been suggested that the basal ganglia act to release cortical elements from idling (alpha) rhythms so that they may become coherent in the gamma range, thereby binding together those distributed activities necessary for the effective selection and execution of a motor act. This hypothesis was tested in 10 patients with idiopathic Parkinson's disease. Surface EEG was recorded during self paced squeezing of the hand and elbow flexion performed separately, simultaneously, or sequentially. Recordings were made after overnight withdrawal of medication and, again, 1 hour after levodopa. The medication related improvement in EEG desynchronisation (in the 7.5-12.5 Hz band) over the 1 second before movement and during movement were separately correlated with the improvement in movement time for each electrode site. Correlation coefficients (r) > 0.632 were considered significant (p<0.05). Improvement in premovement desynchronisation correlated with reduction in bradykinesia over the contralateral sensorimotor cortex and supplementary motor area in flexion and squeeze, respectively. However, when both movements were combined either simultaneously or sequentially, this correlation shifted anteriorly, to areas overlying prefrontal cortex. Improvement in EEG desynchronisation during movement only correlated with reduction in bradykinesia in two tasks. Correlation was seen over the supplementary motor area during flexion, and central prefrontal and ipsilateral premotor areas during simultaneous flex and squeeze. The results are consistent with the idea that the basal ganglia liberate frontal cortex from idling rhythms, and that this effect is focused and specific in so far as it changes with the demands of the task. In particular, the effective selection and execution of more complex tasks is associated with changes over the prefrontal cortex.


The PI3K/Akt Pathway Regulates Oxygen Metabolism via Pyruvate Dehydrogenase (PDH)-E1α Phosphorylation.
Inhibition of the PI3K/Akt pathway decreases hypoxia within SQ20B human head and neck cancer xenografts. We set out to understand the molecular mechanism underlying this observation. We measured oxygen consumption using both a Clark electrode and an extracellular flux analyzer. We made these measurements after various pharmacologic and genetic manipulations. Pharmacologic inhibition of the PI3K/mTOR pathway or genetic inhibition of Akt/PI3K decreased the oxygen consumption rate (OCR) in vitro in SQ20B and other cell lines by 30% to 40%. Pharmacologic inhibition of this pathway increased phosphorylation of the E1α subunit of the pyruvate dehydrogenase (PDH) complex on Ser293, which inhibits activity of this critical gatekeeper of mitochondrial respiration. Expressing wild-type PTEN in a doxycycline-inducible manner in a cell line with mutant PTEN led to an increase in PDH-E1α phosphorylation and a decrease in OCR. Pretreatment of SQ20B cells with dichloroacetate (DCA), which inhibits PDH-E1α phosphorylation by inhibiting dehydrogenase kinases (PDK), reversed the decrease in OCR in response to PI3K/Akt/mTOR inhibition. Likewise, introduction of exogenous PDH-E1α that contains serine to alanine mutations, which can no longer be regulated by phosphorylation, also blunted the decrease in OCR seen with PI3K/mTOR inhibition. Our findings highlight an association between the PI3K/mTOR pathway and tumor cell oxygen consumption that is regulated in part by PDH phosphorylation. These results have important implications for understanding the effects of PI3K pathway activation in tumor metabolism and also in designing cancer therapy trials that use inhibitors of this pathway.


Acute Chikungunya and persistent musculoskeletal pain following the 2006 Indian epidemic: a 2-year prospective rural community study.
Chikungunya virus (CHIKV) data from population studies are sparse. During the 2006 epidemic, 509 clinical cases (43% attack rate) were identified in a village survey (West India); laboratory investigations demonstrated normal blood cell counts, elevated acute-phase reactants [erythrocyte sedimentation rate, C-reactive protein and interleukin-6 (IL-6)] and excluded malaria and dengue. Acute CHIKV was characterized by high fever, severe peripheral polyarthralgias, axial myalgias and intense fatigue in over 90% of cases; skin rash (34%) and headache (19%) were uncommon. There were 49% and 62% of survey cases seropositive for IgM (rapid assay) and IgG (immunofluorescence) anti-CHIKV antibodies, respectively. Sixty-five percent of cases recovered within 4 weeks. None of the cases died. Of the population, 4·1% and 1·6% suffered from persistent rheumatic pains, predominantly non-specific, at 1 and 2 years, respectively. Chronic inflammatory arthritis was uncommon (0·3% at 1 year) although serum IL-6 often remained elevated in chronic cases. A larger population study is required to describe post-CHIKV rheumatism and its prognosis.


Bariatric surgery in morbidly obese adolescents: a systematic review and meta-analysis.
Pubmed, Embase, and Cochrane were systematically reviewed for available evidence on bariatric surgery in adolescents. Thirty-seven included studies evaluated the effect of laparoscopic adjustable gastric banding (LAGB), Roux-en-Y gastric bypass (RYGB), or laparoscopic sleeve gastrectomy (LSG) in patients ≤18 years old. Fifteen of 37 studies were prospective, including one RCT. Mean body mass index (BMI) loss after LAGB was 11.6 kg/m(2) (95% CI 9.8-13.4), versus 16.6 kg/m(2) (95% CI 13.4-19.8) after RYGB and 14.1 kg/m(2) (95% CI 10.8-17.5) after LSG. Two unrelated deaths were reported after 495 RYGB procedures. All three bariatric procedures result in substantial weight loss and improvement of comorbidity with an acceptable complication rate, indicating that surgical intervention is applicable in appropriately selected morbidly obese adolescents.


Control of gene expression in trypanosomes.
Trypanosomes are protozoan agents of major parasitic diseases such as Chagas' disease in South America and sleeping sickness of humans and nagana disease of cattle in Africa. They are transmitted to mammalian hosts by specific insect vectors. Their life cycle consists of a succession of differentiation and growth phases requiring regulated gene expression to adapt to the changing extracellular environment. Typical of such stage-specific expression is that of the major surface antigens of Trypanosoma brucei, procyclin in the procyclic (insect) form and the variant surface glycoprotein (VSG) in the bloodstream (mammalian) form. In trypanosomes, the regulation of gene expression is effected mainly at posttranscriptional levels, since primary transcription of most of the genes occurs in long polycistronic units and is constitutive. The transcripts are processed by transsplicing and polyadenylation under the influence of intergenic polypyrimidine tracts. These events show some developmental regulation. Untranslated sequences of the mRNAs seem to play a prominent role in the stage-specific control of individual gene expression, through a modulation of mRNA abundance. The VSG and procyclin transcription units exhibit particular features that are probably related to the need for a high level of expression. The promoters and RNA polymerase driving the expression of these units resemble those of the ribosomal genes. Their mutually exclusive expression is ensured by controls operating at several levels, including RNA elongation. Antigenic variation in the bloodstream is achieved through DNA rearrangements or alternative activation of the telomeric VSG gene expression sites. Recent discoveries, such as the existence of a novel nucleotide in telomeric DNA and the generation of point mutations in VSG genes, have shed new light on the mechanisms and consequences of antigenic variation.


Setting research priorities to reduce global mortality from preterm birth and low birth weight by 2015.
This paper aims to identify health research priorities that could improve the rate of progress in reducing global neonatal mortality from preterm birth and low birth weight (PB/LBW), as set out in the UN's Millennium Development Goal 4. We applied the Child Health and Nutrition Research Initiative (CHNRI) methodology for setting priorities in health research investments. In the process coordinated by the World Health Organization in 2007-2008, 21 researchers with interest in child, maternal and newborn health suggested 82 research ideas that spanned across the broad spectrum of epidemiological research, health policy and systems research, improvement of existing interventions and development of new interventions. The 82 research questions were then assessed for answerability, effectiveness, deliverability, maximum potential for mortality reduction and the effect on equity using the CHNRI method. The top 10 identified research priorities were dominated by health systems and policy research questions (eg, identification of LBW infants born at home within 24-48 hours of birth for additional care; approaches to improve quality of care of LBW infants in health facilities; identification of barriers to optimal home care practices including care seeking; and approaches to increase the use of antenatal corticosteriods in preterm labor and to improve access to hospital care for LBW infants). These were followed by priorities for improvement of the existing interventions (eg, early initiation of breastfeeding, including feeding mode and techniques for those unable to suckle directly from the breast; improved cord care, such as chlorhexidine application; and alternative methods to Kangaroo Mother Care (KMC) to keep LBW infants warm in community settings). The highest-ranked epidemiological question suggested improving criteria for identifying LBW infants who need to be cared for in a hospital. Among the new interventions, the greatest support was shown for the development of new simple and effective interventions for providing thermal care to LBW infants, if KMC is not acceptable to the mother. The context for this exercise was set within the MDG4, requiring an urgent and rapid progress in mortality reduction from low birth weight, rather than identifying long-term strategic solutions of the greatest potential. In a short-term context, the health policy and systems research to improve access and coverage by the existing interventions, coupled with further research to improve effectiveness, deliverability and acceptance of existing interventions, and epidemiological research to address the key gaps in knowledge, were all highlighted as research priorities.


A causal role for brain-derived neurotrophic factor in the homeostatic regulation of sleep.
Slow-wave activity (SWA), the EEG power between 0.5 and 4 Hz during non-rapid eye movement (NREM) sleep, is one of the best characterized markers of sleep need, because it increases as a function of preceding waking duration and decreases during sleep, but the underlying mechanisms remain unknown. We hypothesized that SWA is high at sleep onset because it reflects the occurrence, during the previous waking period, of widespread synaptic potentiation in cortical and subcortical areas. Consistent with this hypothesis, we recently showed that the more rats explore, the stronger is the cortical expression of BDNF during wakefulness, and the larger is the increase in SWA during the subsequent sleep period. There is compelling evidence that BDNF plays a causal role in synaptic potentiation, and exogenous application of BDNF in vivo is sufficient to induce long-term increases in synaptic strength. We therefore performed cortical unilateral microinjections of BDNF in awake rats and measured SWA during the subsequent sleep period. SWA during NREM sleep was higher in the injected hemisphere relative to the contralateral one. The effect was reversible within 2 h, and did not occur during wakefulness or rapid eye movement sleep. Asymmetries in NREM SWA did not occur after vehicle injections. Furthermore, microinjections, during wakefulness, of a polyclonal anti-BDNF antibody or K252a, an inhibitor of BDNF TrkB receptors, led to a local SWA decrease during the following sleep period. These effects were also reversible and specific for NREM sleep. These results show a causal link between BDNF expression during wakefulness and subsequent sleep regulation.


Morphogen regulation of stem cell plasticity in intestinal regeneration and carcinogenesis.
The intestinal epithelium is a tissue with high cell turnover, supported by adult intestinal stem cells. Intestinal homeostasis is underpinned by crypt basal columnar stem cells, marked by expression of the LGR5 gene. However, recent research has demonstrated considerable stem cell plasticity following injury, with dedifferentiation of a range of other intestinal cell populations, induced by a permissive microenvironment in the regenerating mucosa. The regulation of this profound adaptive cell reprogramming response is the subject of current research. There is a demonstrable contribution from disruption of key homeostatic signaling pathways such as wingless-related integration site and bone morphogenetic protein, and an emerging signaling hub role for the mechanoreceptor transducers Yes-associated protein 1/transcriptional coactivator with PDZ-binding motif, negatively regulated by the Hippo pathway. However, a number of outstanding questions remain, including a need to understand how tissues sense damage, and how pathways intersect to mediate dynamic changes in the stem cell population. Better understanding of these pathways, associated functional redundancies, and how they may be both enhanced for recovery of inflammatory diseases, and co-opted in neoplasia development, may have significant clinical implications, and could lead to development of more targeted molecular therapies which target individual stem or stem-like cell populations.


Fine structure analysis of the yeast centrin, Cdc31p, identifies residues specific for cell morphology and spindle pole body duplication.
Centrin/Cdc31p is a Ca2+-binding protein related to calmodulin found in the MTOC of diverse organisms. In yeast, Cdc31p localizes to the SPB where it interacts with Kar1p and is required for SPB duplication. Recent findings suggest that centrin also functions elsewhere in the cell. To dissect the functions of Cdc31p, we generated cdc31 mutations chosen only for temperature sensitivity, but otherwise unbiased as to phenotype. Three phenotypes of the cdc31 mutants, temperature sensitivity, G2/M arrest, and cell lysis, were not well correlated, indicating that the mutations may differentially affect Cdc31p's interactions with other proteins. Alleles near the C-terminal region exhibited high G2/M arrest and genetic interactions with kar1-Delta17, suggesting that this region modulates an SPB-related function. Alleles causing high lysis and reduced Kic1p kinase activity mapped to the middle of the gene, suggesting disruption of a KIC1-like function and defects in activating Kic1p. A third region conferred temperature sensitivity without affecting cell lysis or G2/M arrest, suggesting that it defines a third function. Mutations in the C-terminal region were also defective for interaction with Kic1p. Mapping the alleles onto a predicted structure of Cdc31p, we have identified surfaces likely to be important for interacting with both Kar1p and Kic1p.


ADCK4-Associated Glomerulopathy Causes Adolescence-Onset FSGS.
Hereditary defects of coenzyme Q10 biosynthesis cause steroid-resistant nephrotic syndrome (SRNS) as part of multiorgan involvement but may also contribute to isolated SRNS. Here, we report 26 patients from 12 families with recessive mutations in ADCK4. Mutation detection rate was 1.9% among 534 consecutively screened cases. Patients with ADCK4 mutations showed a largely renal-limited phenotype, with three subjects exhibiting occasional seizures, one subject exhibiting mild mental retardation, and one subject exhibiting retinitis pigmentosa. ADCK4 nephropathy presented during adolescence (median age, 14.1 years) with nephrotic-range proteinuria in 44% of patients and advanced CKD in 46% of patients at time of diagnosis. Renal biopsy specimens uniformly showed FSGS. Whereas 47% and 36% of patients with mutations in WT1 and NPHS2, respectively, progressed to ESRD before 10 years of age, ESRD occurred almost exclusively in the second decade of life in ADCK4 nephropathy. However, CKD progressed much faster during adolescence in ADCK4 than in WT1 and NPHS2 nephropathy, resulting in similar cumulative ESRD rates (>85% for each disorder) in the third decade of life. In conclusion, ADCK4-related glomerulopathy is an important novel differential diagnosis in adolescents with SRNS/FSGS and/or CKD of unknown origin.


Pacemaker implantation in small hospitals: complication rates comparable to larger centres.
Some countries have a demography that makes it necessary to maintain relatively small pacemaker centres. We wanted to assess the quality of pacemaker surgery in two such hospitals. Through patient records we gathered information on ∼535 consecutive primary pacemaker implantations in two small pacemaker centres with 30 and 80 annual operations, respectively. All patients were followed for 3 years. All complications documented in the patient records were registered. Furthermore, we performed a non-systematic literature search comparing our data with reports from major centres published over the last 10 years.We found 72 complications in 64 (12.0%) of the patients, the most common being bleeding, lead failure, and pneumothorax. If minor bleedings without any consequences for the patients are excluded, the number of complications was 46 in 40 patients (7.5%). We had to reoperate on 5.2% of the patients. There was no statistically significant difference in complication rates between the two hospitals. Education candidates generated statistically significant more complications than experienced doctors (13.7 vs. 7.1%, P < 0.05). There are no generally accepted norms of complication rates in pacemaker surgery. However, we found no indications that our centres have a rate of complications that is unacceptably high.


Activation of notch signaling in a xenograft model of brain metastasis.
The potential of metastasis can be predicted from clinical features like tumor size, histologic grade, and gene expression patterns. We examined the whole-genome transcriptomic profile of a xenograft model of breast cancer to understand the characteristics of brain metastasis. Variants of the MDA-MB-435 cell were established from experimental brain metastases. The LvBr2 variant was isolated from lesions in a mouse injected in the left ventricle of the heart, and these cells were used for two cycles of injection into the internal carotid artery and selection of brain lesions, resulting in the Br4 variant. To characterize the different metastatic variants, we examined the gene expression profile of MDA-MB-435, LvBr2, and Br4 cells using microarrays. We could identify 2,016 differentially expressed genes in Br4 by using the F test. Various metastasis-related genes and a number of genes related to angiogenesis, migration, tumorigenesis, and cell cycle were differentially expressed by the Br4 cells. Notably, the Notch signaling pathway was activated in Br4, with increased Jag2 mRNA, activated Notch intracellular domain, and Notch intracellular domain/CLS promoter-luciferase activity. Br4 cells were more migratory and invasive than MDA-MB-435 cells in collagen and Matrigel Transwell assays, and the migration and invasion of Br4 cells were significantly inhibited by inactivation of Notch signaling using DAPT, a gamma-secretase inhibitor, and RNA interference-mediated knockdown of Jagged 2 and Notch1. Taken together, these results suggest that we have isolated variants of a human cancer cell line with enhanced brain metastatic properties, and the activation of Notch signaling might play a crucial role in brain metastasis.


Relative Reactivity and Bioavailability of Mercury Sorbed to or Coprecipitated with Aged Iron Sulfides.
The potential for inorganic mercury (Hg) to be converted to methylmercury depends, in part, on the chemical form of Hg and its bioavailability to anaerobic microorganisms that can methylate Hg. In anaerobic settings, Hg can be associated with sulfide phases, including ferrous iron sulfide (FeS), which can sorb or be coprecipitated with Hg. The objective of this study was to determine if the aging state of FeS alters the Hg coordination environment as well as the reactivity and bioavailability of sorbed and coprecipitated Hg species. FeS particles were synthesized with and without Hg2+ and aged in anaerobic conditions for multiple time frames spanning from 1 h to 1 month. For FeS particles synthesized without Hg, Hg2+ was subsequently sorbed to the FeS for 1 day. Analysis of Hg speciation of these materials by X-ray absorption near edge spectroscopy revealed a predominance of four-coordinate Hg-S species in the sorbed Hg-FeS solids and a mixture of two- and four-coordinate Hg-S in the coprecipitated Hg-FeS. The leaching potential of the Hg was assessed by exposing the particles to a solution of dissolved glutathione (a thiolate-based Hg chelator). As expected, the sorbed Hg-FeS released more soluble Hg compared to the coprecipitated Hg-FeS. However, when these particles were exposed to Desulfovibrio desulfuricans ND132 (a known Hg methylator), more Hg was methylated from the coprecipitated Hg-FeS than the sorbed Hg-FeS, consistent with expectations from the Hg-S coordination state and inconsistent with the selective leaching results. Overall, these results suggest that the bioavailability of particulate Hg cannot be easily discerned by its leaching potential into bulk solution. Rather, bioavailability entails more subtle interactions at particle-cell interfaces and perhaps correlates with the local Hg-S coordination state in the particles.


Outcome of haematopoietic stem cell transplantation in dyskeratosis congenita.
Dyskeratosis congenita (DC) is a genetic multisystem disorder with frequent involvement of the bone marrow. Haematopoietic stem cell transplantation (HSCT) is the only definitive cure to restore haematopoiesis, even though it cannot correct other organ dysfunctions. We collected data on the outcome of HSCT in the largest cohort of DC (n = 94) patients ever studied. Overall survival (OS) and event-free survival (EFS) at 3 years after HSCT were 66% and 62%, respectively. Multivariate analysis showed better outcomes in patients aged less than 20 years and in patients transplanted from a matched, rather than a mismatched, donor. OS and EFS curves tended to decline over time. Early lethal events were infections, whereas organ damage and secondary malignancies appeared afterwards, even a decade after HSCT. A non-myeloablative conditioning regimen appeared to be most advisable. Organ impairment present before HSCT seemed to favour the development of chronic graft-versus-host disease and T-B immune deficiency appeared to enhance pulmonary fibrosis. According to the present data, HSCT in DC is indicated in cases of progressive marrow failure, whereas in patients with pre-existing organ damage, this should be carefully evaluated. Further efforts to investigate treatment alternatives to HSCT should be encouraged.


Pneumothorax in a weight lifter: the importance of vigilance.
A 27-year-old weight lifter developed mild chest pain and dyspnea after 'bouncing' a 250-lb barbell off his chest. A plain radiograph revealed a large, right-sided pneumothorax with collapse of the lung. He underwent an emergency tube thoracostomy. One month later, he resumed lifting without recurrence. This case report demonstrates the need to be responsive to dyspnea and chest pain in healthy, young athletes. Pneumothorax in sports is uncommon, but cases of spontaneous and trauma-induced pneumothorax have been reported. Initial symptoms may be minimal, but prompt recognition can help prevent respiratory and cardiovascular compromise. Treatment depends in part on the size of the pneumothorax.


Human neonatal hepatocyte transplantation induces long-term rescue of unconjugated hyperbilirubinemia in the Gunn rat.
Crigler-Najjar type 1 disease is a rare inherited metabolic disease characterized by high levels of unconjugated bilirubin due to the complete absence of hepatic uridine diphosphoglucuronate-glucuronosyltransferase activity. Hepatocyte transplantation (HT) has been proposed as an alternative treatment for Crigler-Najjar syndrome, but it is still limited by the quality and the low engraftment and repopulation ability of the cells used. Because of their attachment capability and expression of adhesion molecules as well as the higher proportion of hepatic progenitor cells, neonatal hepatocytes may have an advantage over adult cells. Adult or neonatal hepatocytes were transplanted into Gunn rats, a model for Crigler-Najjar disease. Engraftment and repopulation were studied and compared by immunofluorescence (IF). Additionally, the serum bilirubin levels, the presence of bilirubin conjugates in rat serum, and the expression of uridine diphosphate glucuronosyltransferase 1 family polypeptide A1 (UGT1A1) in rat liver samples were also analyzed. Here we show that neonatal HT results in long-term correction in Gunn rats. In comparison with adult cells, neonatal cells showed better engraftment and repopulation capability 3 days and 6 months after transplantation, respectively. Bilirubinemia decreased in the transplanted animals during the whole experimental follow-up (6 months). Bilirubin conjugates were also present in the serum of the transplanted animals. Western blots and IF confirmed the presence and expression of UGT1A1 in the liver. This work is the first to demonstrate the advantage of using neonatal hepatocytes for the treatment of Crigler-Najjar in vivo.


Breast and cervical cancer screening practices among disabled women aged 40-75: does quality of the experience matter?
Women with disabilities (WWD) face significant barriers accessing healthcare, which may affect rates of routine preventive services. We examined the relationship between disability status and routine breast and cervical cancer screening among middle-aged and older unmarried women and the differences in reported quality of the screening experience. Data were from a 2003-2005 cross-sectional survey of 630 unmarried women in Rhode Island, 40-75 years of age, stratified by marital status (previously vs. never married) and partner gender (women who partner with men exclusively [WPM] vs. women who partner with women exclusively or with both women and men [WPW]). WWD were more likely than those without a disability to be older, have a high school education or less, have household incomes <$30,000, be unemployed, and identify as nonwhite. In addition, WWD were less likely to report having the mammogram or Pap test procedure explained and more likely to report that the procedures were difficult to perform. After adjustment for important demographic characteristics, we found no differences in cancer screening behaviors by disability status. However, the quality of the cancer screening experience was consistently and significantly associated with likelihood of routine cancer screening. Higher quality of cancer screening experience was significantly associated with likelihood of having routine breast and cervical cancer screening. Further studies should explore factors that affect quality of the screening experience, including facility characteristics and interactions with medical staff.


Neural correlates of personal space intrusion.
A parietal-frontal network in primates is thought to support many behaviors occurring in the space around the body, including interpersonal interactions and maintenance of a particular "comfort zone" or distance from other people ("personal space"). To better understand this network in humans, we used functional MRI to measure the responses to moving objects (faces, cars, simple spheres) and the functional connectivity of two regions in this network, the dorsal intraparietal sulcus (DIPS) and the ventral premotor cortex (PMv). We found that both areas responded more strongly to faces that were moving toward (vs away from) subjects, but did not show this bias in response to comparable motion in control stimuli (cars or spheres). Moreover, these two regions were functionally interconnected. Tests of activity-behavior associations revealed that the strength of DIPS-PMv connectivity was correlated with the preferred distance that subjects chose to stand from an unfamiliar person (personal space size). In addition, the magnitude of DIPS and PMv responses was correlated with the preferred level of social activity. Together, these findings suggest that this parietal-frontal network plays a role in everyday interactions with others.


Characteristics of 44 cervical cancers diagnosed following Pap-negative, high risk HPV-positive screening in routine clinical practice.
To characterize the cervical cancers diagnosed following a Pap-negative, high risk human papillomavirus (HPV)-positive (Pap-/HPV+) screen in routine clinical practice. Using data from Kaiser Permanente Northern California, we investigated the cases of cervical cancer diagnosed between January, 2003 and January, 2009 following Pap-/HPV+ screen. Two cervical specimens were routinely collected for cervical cancer screening, one for conventional cytology and the other for high risk HPV testing using Hybrid Capture 2 (Qiagen). Forty-four women (median age at diagnosis=44years) were diagnosed with primary invasive cervical cancer with a recent history of one or more Pap-/HPV+ screens. Twenty-six women had one Pap-/HPV+ screen preceding the diagnosis of cancer, 15 had two, and three had three. There were 16 squamous cancers, one small cell cancer, 24 adenocarcinomas, 2 adenosquamous carcinomas, and one case with separate invasive squamous and adenocarcinoma. FIGO Stage was IA in 11 women, IB in 31 women and IIA in 2 women. Treatment included a pelvic node dissection in 30, 2 (6.7%) of whom had positive nodes. HPV testing contributes to early cervical cancer diagnosis detection in women with negative Pap tests. Most women in this cohort have early stage, node negative, treatable and potentially curable disease. Adenocarcinoma predominated as might be expected because cytology misses these cancers and their precursors. The majority of cancers were diagnosed following a single Pap-/HPV+ screen, suggesting that effective triage to colposcopy of women with a Pap-/HPV+ screen would be preferable to retesting in one year as currently recommended.


Controlled release of bevacizumab through nanospheres for extended treatment of age-related macular degeneration.
Bevacizumab (Avastin(®)) has been used by ophthalmologists in many countries as an off-label drug for the treatment of wet age-related macular degeneration (AMD). Due to its short half-life necessitating frequent intravitreal injection, a method for sustained delivery is in need. We demonstrated that bevacizumab could be released in a sustained fashion over 90 days from nano- and microspheres fabricated from poly(DL-lactide-co-glycolide) and poly(ethylene glycol)-b-poly(D,L-lactic acid), respectively. The drug release rate could be adjusted by alteration of the drug/polymer ratio. The use of such nano- and microspheres as bevacizumab delivery vehicles may improve the treatment of wet AMD.


Interaction of adenoviral proteins with pRB and p53.
The transforming gene products of the small DNA tumor viruses subvert host cell growth control mechanisms by binding to specific cell regulatory proteins. These include the retinoblastoma gene product (pRB) and p53. One indication of the pivotal roles played by these regulatory products is the observation that they are each targeted consistently by viruses of several groups, by adenoviruses, the human papillomaviruses, and the papovaviruses. In adenovirus, pRB and p53 are targeted by the E1A and E1B genes, respectively. The genetic probes made possible by manipulation of the virus genes in vitro have helped to illuminate the pathways in which pRB and p53 function. E1A studies have contributed to our current understanding that the retinoblastoma product is one of a family of related proteins, which with associated cyclins and kinases can modulate the activity of the cellular E2F transcription factor. E1B studies have helped explore models of p53 function, including the suggestion that p53, probably through aspects of its transcription regulating activity, can initiate a pathway in which programmed cell death can be invoked to stop unrestricted cell proliferation.


Kinase domain activation of FGFR2 yields high-grade lung adenocarcinoma sensitive to a Pan-FGFR inhibitor in a mouse model of NSCLC.
Somatic mutations in FGFR2 are present in 4% to 5% of patients diagnosed with non-small cell lung cancer (NSCLC). Amplification and mutations in FGFR genes have been identified in patients with NSCLCs, and clinical trials are testing the efficacy of anti-FGFR therapies. FGFR2 and other FGFR kinase family gene alterations have been found in both lung squamous cell carcinoma and lung adenocarcinoma, although mouse models of FGFR-driven lung cancers have not been reported. Here, we generated a genetically engineered mouse model (GEMM) of NSCLC driven by a kinase domain mutation in FGFR2. Combined with p53 ablation, primary grade 3/4 adenocarcinoma was induced in the lung epithelial compartment exhibiting locally invasive and pleiotropic tendencies largely made up of multinucleated cells. Tumors were acutely sensitive to pan-FGFR inhibition. This is the first FGFR2-driven lung cancer GEMM, which can be applied across different cancer indications in a preclinical setting.


The Manual Stimulation of Acupuncture Points in the Treatment of Post-Traumatic Stress Disorder: A Review of Clinical Emotional Freedom Techniques.
Background: The manual stimulation of acupuncture points has been combined with components of cognitive and exposure therapies into a clinical and self-help approach known as Emotional Freedom Techniques (EFT). More than 40 clinical trials and four meta-analytic reviews of EFT treatments have demonstrated large effect sizes with a range of conditions, including pain, PTSD (in both civilian and military veteran populations), phobias, anxiety, and depression. Objective: This review describes the approach, with a focus on PTSD in veterans and service members, provides an overview of how EFT is most commonly applied, and outlines obstacles and cautions related to its implementation. Methods: Peer-reviewed clinical trials and meta-analyses of EFT in the treatment of PTSD are assessed to identify the characteristics of the approach that render it suitable for the treatment of PTSD. Results: The literature demonstrates that remediation of PTSD and comorbid conditions is typically accomplished within brief time frames, ranging from one session for phobias to between four and ten sessions for PTSD. Clinical EFT has been shown to regulate stress hormones and limbic function and to improve various neurologic markers of general health. The epigenetic effects of EFT include upregulation of immunity genes and downregulation of inflammation genes. Six dismantling studies have indicated that the acupressure component of EFT is an active ingredient and not placebo. Conclusions: Seven empirically supported strengths of the approach were identified that make it especially suitable for use with veterans and active military: (1) the depth and breadth of treatment effects; (2) the relatively brief timeframes required for successful treatment; (3) the low risk of adverse events; (4) the minimal training time required for the approach to be applied effectively; (5) the simultaneous reduction of physical and psychologic symptoms; (6) the utility and cost-effectiveness of clinical EFT in a large group format; and (7) the method's adaptability to online and telemedicine applications.


Infrared optical activity: electric field approaches in time domain.
Vibrational circular dichroism (VCD) spectroscopy provides detailed information about the absolute configurations of chiral molecules including biomolecules and synthetic drugs. This method is the infrared (IR) analogue of the more popular electronic CD spectroscopy that uses the ultraviolet and visible ranges of the electromagnetic spectrum. Because conventional electronic CD spectroscopy measures the difference in signal intensity, problems such as weak signal and low time-resolution can limit its utility. To overcome the difficulties associated with that approach, we have recently developed femtosecond IR optical activity (IOA) spectrometry, which directly measures the IOA free-induction-decay (FID), the impulsive chiroptical IR response that occurs over time. In this Account, we review the time-domain electric field measurement and calculation methods used to simultaneously characterize VCD and related vibrational optical rotatory dispersion (VORD) spectra. Although conventional methods measure the electric field intensity, this vibrational technique is based on a direct phase-and-amplitude measurement of the electric field of the chiroptical signal over time. This method uses a cross-polarization analyzer to carry out heterodyned spectral interferometry. The cross-polarization scheme enables us to selectively remove the achiral background signal, which is the dominant noise component present in differential intensity measurement techniques. Because we can detect the IOA FID signal in a phase-amplitude-sensitive manner, we can directly characterize the time-dependent electric dipole/magnetic dipole response function and the complex chiral susceptibility that contain information about the angular oscillations of charged particles. These parameters yield information about the VCD and VORD spectra. In parallel with such experimental developments, we have also calculated the IOA FID signal and the resulting VCD spectrum. These simulations use a quantum mechanical/molecular mechanical molecular dynamics (QM/MM MD) method and calculate the electric dipole/magnetic dipole cross-correlation function in the time domain. Although many quantum chemistry calculation approaches can only consider a limited number of geometry-optimized conformations of chiral molecules in a gas phase, this computational method includes the solute-solvent interactions and the inhomogeneous distributions of solute conformers in condensed phases. A subsequent Fourier transformation of the chiral response function produced a theoretical VCD spectrum in the entire mid-IR frequency range. Directly comparing theory and experiment, we demonstrate quantitative agreement between frequency-tunable femtosecond IOA measurements and QM/MM MD simulations of (1S)-β-pinene in CCl(4) solution. We anticipate that these direct IOA measurement and calculation methods will be applied to the studies of equilibrium chiroptical properties and structure determinations. These methods provide tools to investigate ultrafast structural dynamics of chiral systems with unprecedented time resolution.


Using brain imaging to extract the structure of complex events at the rational time band.
A functional magnetic resonance imaging (fMRI) study was performed in which participants performed a complex series of mental calculations that spanned about 2 min. An Adaptive Control of Thought--Rational (ACT-R) model [Anderson, J. R. How can the human mind occur in the physical universe? New York: Oxford University Press, 2007] was developed that successfully fit the distribution of latencies. This model generated predictions for the fMRI signal in six brain regions that have been associated with modules in the ACT-R theory. The model's predictions were confirmed for a fusiform region that reflects the visual module, for a prefrontal region that reflects the retrieval module, and for an anterior cingulate region that reflects the goal module. In addition, the only significant deviations to the motor region that reflects the manual module were anticipatory hand movements. In contrast, the predictions were relatively poor for a parietal region that reflects an imaginal module and for a caudate region that reflects the procedural module. Possible explanations of these poor fits are discussed. In addition, exploratory analyses were performed to find regions that might correspond to the predictions of the modules.


Alamethicin channels - modelling via restrained molecular dynamics simulations.
Alamethicin channels have been modelled as approximately parallel bundles of transbilayer helices containing between N = 4 and 8 helices per bundle. Initial models were generated by in vacuo restrained molecular dynamics (MD) simulations, and were refined by 60 ps MD simulations with water molecules present within and at the mouths of the central pore. The helix bundles were stabilized by networks of H-bonds between intra-pore water molecules and Gln-7 side-chains. Channel conductances were predicted on the basis of pore radius profiles, and suggested that the N = 4 bundle formed an occluded pore, whereas pores with N > or = 5 helices per bundle were open. Continuum electrostatics calculations suggested that the N = 6 pore is cation-selective, whereas pores with N > or = 7 helices per bundle were predicted to be somewhat less ion-selective.


Evaluation of short and tall stature in children.
Children and adolescents whose heights and growth velocities deviate from the normal percentiles on standard growth charts present a special challenge to physicians. Height that is less than the 3rd percentile or greater than the 97th percentile is deemed short or tall stature, respectively. A growth velocity outside the 25th to 75th percentile range may be considered abnormal. Serial height measurements over time documented on a growth chart are key in identifying abnormal growth. Short or tall stature is usually caused by variants of a normal growth pattern, although some patients may have serious underlying pathologies. A comprehensive history and physical examination can help differentiate abnormal growth patterns from normal variants and identify specific dysmorphic features of genetic syndromes. History and physical examination findings should guide laboratory testing.


Tunable solid-state laser based on modified polymethyl methacrylate with methanol doped with Pyrromethene 580.
Solid-state dye material was fabricated by doping a laser dye Pyrromethene 580 (PM580) into the polymer host polymethyl methacrylate. Methanol was also injected into the host to improve the optical properties. The broadband and narrowband laser performances of the sample were studied in this paper. When the selected solid-state dye PM580 was placed in a Shoshan-type oscillator, narrow linewidth operation with a tuning range of 52 nm and good photostability was demonstrated. The narrowband laser output slope efficiency of 42.7% was obtained, and the corresponding broadband laser slope efficiency was 66.0%. To the best of our knowledge, the narrowband slope efficiency and tunable range are the best under similar conditions so far. For broadband and narrowband lasers, the beam quality factors were estimated to be M(y)(2)=10.7 and 3.9 in the vertical direction.


Measures of light in studies on light-driven plant plasticity in artificial environments.
Within-canopy variation in light results in profound canopy profiles in foliage structural, chemical, and physiological traits. Studies on within-canopy variations in key foliage traits are often conducted in artificial environments, including growth chambers with only artificial light, and greenhouses with and without supplemental light. Canopy patterns in these systems are considered to be representative to outdoor conditions, but in experiments with artificial and supplemental lighting, the intensity of artificial light strongly deceases with the distance from the light source, and natural light intensity in greenhouses is less than outdoors due to limited transmittance of enclosure walls. The implications of such changes in radiation conditions on canopy patterns of foliage traits have not yet been analyzed. We developed model-based methods for retrospective estimation of distance vs. light intensity relationships, for separation of the share of artificial and natural light in experiments with combined light and for estimation of average enclosure transmittance, and estimated daily integrated light at the time of sampling (Q(int,C)), at foliage formation (Q(int,G)), and during foliage lifetime (Q(int,av)). The implications of artificial light environments were analyzed for altogether 25 studies providing information on within-canopy gradients of key foliage traits for 70 species × treatment combinations. Across the studies with artificial light, Q(int,G) for leaves formed at different heights in the canopy varied from 1.8- to 6.4-fold due to changing the distance between light source and growing plants. In experiments with combined lighting, the share of natural light at the top of the plants varied threefold, and the share of natural light strongly increased with increasing depth in the canopy. Foliage nitrogen content was most strongly associated with Q(int,G), but photosynthetic capacity with Q(int,C), emphasizing the importance of explicit description of light environment during foliage lifetime. The reported and estimated transmittances of enclosures varied between 0.27 and 0.85, and lack of consideration of the reduction of light compared with outdoor conditions resulted in major underestimation of foliage plasticity to light. The study emphasizes that plant trait vs. light relationships in artificial systems are not directly comparable to natural environments unless modifications in lighting conditions in artificial environments are taken into account.


Measuring temporal, spectral and spatial changes in electrophysiological brain network connectivity.
The topic of functional connectivity in neuroimaging is expanding rapidly and many studies now focus on coupling between spatially separate brain regions. These studies show that a relatively small number of large scale networks exist within the brain, and that healthy function of these networks is disrupted in many clinical populations. To date, the vast majority of studies probing connectivity employ techniques that compute time averaged correlation over several minutes, and between specific pre-defined brain locations. However, increasing evidence suggests that functional connectivity is non-stationary in time. Further, electrophysiological measurements show that connectivity is dependent on the frequency band of neural oscillations. It is also conceivable that networks exhibit a degree of spatial inhomogeneity, i.e. the large scale networks that we observe may result from the time average of multiple transiently synchronised sub-networks, each with their own spatial signature. This means that the next generation of neuroimaging tools to compute functional connectivity must account for spatial inhomogeneity, spectral non-uniformity and temporal non-stationarity. Here, we present a means to achieve this via application of windowed canonical correlation analysis (CCA) to source space projected MEG data. We describe the generation of time-frequency connectivity plots, showing the temporal and spectral distribution of coupling between brain regions. Moreover, CCA over voxels provides a means to assess spatial non-uniformity within short time-frequency windows. The feasibility of this technique is demonstrated in simulation and in a resting state MEG experiment where we elucidate multiple distinct spatio-temporal-spectral modes of covariation between the left and right sensorimotor areas.


Local endothelial DNA repair deficiency causes aging-resembling endothelial-specific dysfunction.
We previously identified genomic instability as a causative factor for vascular aging. In the present study, we determined which vascular aging outcomes are due to local endothelial DNA damage, which was accomplished by genetic removal of ERCC1 (excision repair cross-complementation group 1) DNA repair in mice (EC-knockout (EC-KO) mice). EC-KO showed a progressive decrease in microvascular dilation of the skin, increased microvascular leakage in the kidney, decreased lung perfusion, and increased aortic stiffness compared with wild-type (WT). EC-KO showed expression of DNA damage and potential senescence marker p21 exclusively in the endothelium, as demonstrated in aorta. Also the kidney showed p21-positive cells. Vasodilator responses measured in organ baths were decreased in aorta, iliac and coronary artery EC-KO compared with WT, of which coronary artery was the earliest to be affected. Nitric oxide-mediated endothelium-dependent vasodilation was abolished in aorta and coronary artery, whereas endothelium-derived hyperpolarization and responses to exogenous nitric oxide (NO) were intact. EC-KO showed increased superoxide production compared with WT, as measured in lung tissue, rich in endothelial cells (ECs). Arterial systolic blood pressure (BP) was increased at 3 months, but normal at 5 months, at which age cardiac output (CO) was decreased. Since no further signs of cardiac dysfunction were detected, this decrease might be an adaptation to prevent an increase in BP. In summary, a selective DNA repair defect in the endothelium produces features of age-related endothelial dysfunction, largely attributed to loss of endothelium-derived NO. Increased superoxide generation might contribute to the observed changes affecting end organ perfusion, as demonstrated in kidney and lung.


Pharmacological rescue of trafficking-impaired ATP-sensitive potassium channels.
ATP-sensitive potassium (KATP) channels link cell metabolism to membrane excitability and are involved in a wide range of physiological processes including hormone secretion, control of vascular tone, and protection of cardiac and neuronal cells against ischemic injuries. In pancreatic β-cells, KATP channels play a key role in glucose-stimulated insulin secretion, and gain or loss of channel function results in neonatal diabetes or congenital hyperinsulinism, respectively. The β-cell KATP channel is formed by co-assembly of four Kir6.2 inwardly rectifying potassium channel subunits encoded by KCNJ11 and four sulfonylurea receptor 1 subunits encoded by ABCC8. Many mutations in ABCC8 or KCNJ11 cause loss of channel function, thus, congenital hyperinsulinism by hampering channel biogenesis and hence trafficking to the cell surface. The trafficking defects caused by a subset of these mutations can be corrected by sulfonylureas, KATP channel antagonists that have long been used to treat type 2 diabetes. More recently, carbamazepine, an anticonvulsant that is thought to target primarily voltage-gated sodium channels has been shown to correct KATP channel trafficking defects. This article reviews studies to date aimed at understanding the mechanisms by which mutations impair channel biogenesis and trafficking and the mechanisms by which pharmacological ligands overcome channel trafficking defects. Insight into channel structure-function relationships and therapeutic implications from these studies are discussed.


Nestly--a framework for running software with nested parameter choices and aggregating results.
The execution of a software application or pipeline using various combinations of parameters and inputs is a common task in bioinformatics. In the absence of a specialized tool to organize, streamline and formalize this process, scientists must write frequently complex scripts to perform these tasks. We present nestly, a Python package to facilitate running tools with nested combinations of parameters and inputs. nestly provides three components. First, a module to build nested directory structures corresponding to choices of parameters. Second, the nestrun script to run a given command using each set of parameter choices. Third, the nestagg script to aggregate results of the individual runs into a CSV file, as well as support for more complex aggregation. We also include a module for easily specifying nested dependencies for the SCons build tool, enabling incremental builds. Source, documentation and tutorial examples are available at http://github.com/fhcrc/nestly. nestly can be installed from the Python Package Index via pip; it is open source (MIT license).


Direct interactions of coxsackievirus B3 with immune cells in the splenic compartment of mice susceptible or resistant to myocarditis.
In vitro replication of coxsackievirus B3 (CVB3) in cells of the immune system derived from uninfected adolescent A/J and C57BL/6J mice and replication of CVB3 in and association with immune cells from spleens of infected animals in vivo were assessed. Nonstimulated or mitogen-stimulated spleen cells were minimally permissive for viral replication during an 8-h period. Three days postinfection (p.i.), CVB3 RNA was localized in vivo to B cells and follicular dendritic cells of germinal centers in both A/J and C57BL/6J mice; however, extrafollicular localization was greater in C57BL/6J mice (P = 0.0054). Although the pattern of CVB3 RNA localization was different, the total load of infections virus (PFU per milligram of tissue) was not different. Splenic CVB3 titers (PFU per milligram of tissue) in both strains were maximal at day 3 or 4 p.i. and were back to baseline by day 7 p.i., with most infectious virus being non-cell associated. CVB3 titers (PFU per milligram of tissue) correlated directly with in situ hybridization positivity in splenic follicles and extrafollicular regions in both murine strains; however, follicular hybridization intensity was greater in A/J mice at day 5 p.i. (P = 0.021). Flow cytometric analysis demonstrated that 50.4% of total spleen cells positive for CVB3 antigen were B cells and 69.6% of positive splenic lymphocytes were B cells. Myocardial virus load in C57BL/6J mice was significantly lower than that in A/J mice at days 4 and 5 p.i. These data indicate that CVB3 replicates in murine splenocytes in vitro and in B cells and extrafollicular cells in vivo.


Clonal evolution in chronic lymphocytic leukemia detected by fluorescence in situ hybridization and conventional cytogenetics after stimulation with CpG oligonucleotides and interleukin-2: a prospective analysis.
Chronic lymphocytic leukemia (CLL) patients may acquire new chromosome abnormalities during the course of their disease. Clonal evolution (CE) has been detected by conventional chromosome banding (CBA), several groups also confirmed CE with fluorescence in situ hybridization (FISH). At present, there are minimal prospective data on CE frequency determined using a combination of both methods. Therefore, the aim of our study was to prospectively assess CE frequency using a combination of FISH and CBA after stimulation with CpG oligonucleotides and interleukin-2. Between 2008 and 2012, we enrolled 140 patients with previously untreated CLL in a prospective trial evaluating CE using FISH and CBA after stimulation. Patients provided baseline and regular follow-up peripheral blood samples for testing. There was a median of 3 cytogenetic examinations (using both methods) per patient. CE was detected in 15.7% (22/140) of patients using FISH, in 28.6% (40/140) using CBA, and in 34.3% (48/140) of patients by combining both methods. Poor-prognosis CE (new deletion 17p, new deletion 11q or new complex karyotype) was detected in 15% (21/140) of patients and was significantly associated with previous CLL treatment (p=0.013). CBA provides more complex information about cytogenetic abnormalities in CLL patients than FISH and confirms that many patients can acquire new abnormalities during the course of their disease in a relatively short time period.


Targeting the reversibly oxidized protein tyrosine phosphatase superfamily.
Controlled production of reactive oxygen species leads to reversible oxidation of protein tyrosine phosphatases (PTPs) and has emerged as an important tier of regulation over phosphorylation-dependent signal transduction. We present a modified cysteinyl-labeling assay that detects reversible oxidation of members of each of the different PTP subclasses. Here, we describe the methods for enriching reversibly oxidized PTPs from complex protein extracts, illustrating the procedure in IMR90 fibroblasts.


SveDem, the Swedish Dementia Registry - a tool for improving the quality of diagnostics, treatment and care of dementia patients in clinical practice.
The Swedish Dementia Registry (SveDem) was developed with the aim to improve the quality of diagnostic work-up, treatment and care of patients with dementia disorders in Sweden. SveDem is an internet based quality registry where several indicators can be followed over time. It includes information about the diagnostic work-up, medical treatment and community support (www.svedem.se). The patients are diagnosed and followed-up yearly in specialist units, primary care centres or in nursing homes. The database was initiated in May 2007 and covers almost all of Sweden. There were 28 722 patients registered with a mean age of 79.3 years during 2007-2012. Each participating unit obtains continuous online statistics from its own registrations and they can be compared with regional and national data. A report from SveDem is published yearly to inform medical and care professionals as well as political and administrative decision-makers about the current quality of diagnostics, treatment and care of patients with dementia disorders in Sweden. SveDem provides knowledge about current dementia care in Sweden and serves as a framework for ensuring the quality of diagnostics, treatment and care across the country. It also reflects changes in quality dementia care over time. Data from SveDem can be used to further develop the national guidelines for dementia and to generate new research hypotheses.


Vigorous Appraisal of Fluoride on Industrial Proponent in Thermal Power Plant over Anthropoid Biosphere Using F(-) Ion-Selective Electrode.
This study was conducted to analyze the impact of fluoride in the anthropogenic condition in an industrial region promoting and affecting the health of the workers. Fluoride is toxic to humans in high concentrations, such as can occur in persons working in fluoride-containing mineral industries like aluminum industries. When workers are exposed to fluoride-containing minerals, they can suffer from a variety of health problems, such as dental disease. This paper presents the relationship of different clinical conditions correlated against the fluoride level. Contributing clinical aspects, such as morbidity, dysentery, overcrowding, and skin disease, are also studied to assess the consequences of fluoride upon consistent exposure. The relationship between pH and hardness of water with fluoride was measured, and then spatial maps were generated. The investigations resulted in a conclusion that hardness of water had a more pronounced impact on the level of fluoride concentration as compared with pH. Water with more hardness contains more fluoride concentration (25 mg/ml) as compared with soft water (4 mg/ml). This paper also revealed the concentration of fluoride content in the bodies of aluminum plant workers, which varied from 0.06 to 0.17 mg/L of blood serum in the case of pot room workers and 0.01 to 0.04 mg/L in the case of non-pot room workers. In fingernails, it varied from 0.09 to 3.77 mg/L and 0.39 to 1.15 mg/L in the case of pot room and non-pot room workers, respectively. In urine, it varied from 0.53 to 9.50 mg/L in pot room workers and 0.29 to 1.80 mg/L in non-pot room workers. This paper concluded that water was safe for drinking purposes if it has a low hardness (60-140 mg/ml) and pH (7.1-7.4).


Values for digestible indispensable amino acid scores (DIAAS) for some dairy and plant proteins may better describe protein quality than values calculated using the concept for protein digestibility-corrected amino acid scores (PDCAAS).
An experiment was conducted to compare values for digestible indispensable amino acid scores (DIAAS) for four animal proteins and four plant proteins with values calculated as recommended for protein digestibility-corrected amino acid scores (PDCAAS), but determined in pigs instead of in rats. Values for standardised total tract digestibility (STTD) of crude protein (CP) and standardised ileal digestibility (SID) of amino acids (AA) were calculated for whey protein isolate (WPI), whey protein concentrate (WPC), milk protein concentrate (MPC), skimmed milk powder (SMP), pea protein concentrate (PPC), soya protein isolate (SPI), soya flour and whole-grain wheat. The PDCAAS-like values were calculated using the STTD of CP to estimate AA digestibility and values for DIAAS were calculated from values for SID of AA. Results indicated that values for SID of most indispensable AA in WPI, WPC and MPC were greater (P<0·05) than for SMP, PPC, SPI, soya flour and wheat. With the exception of arginine and tryptophan, the SID of all indispensable AA in SPI was greater (P<0·05) than in soya flour, and with the exception of threonine, the SID of all indispensable AA in wheat was less (P<0·05) than in all other ingredients. If the same scoring pattern for children between 6 and 36 months was used to calculate PDCAAS-like values and DIAAS, PDCAAS-like values were greater (P<0·05) than DIAAS values for SMP, PPC, SPI, soya flour and wheat indicating that PDCAAS-like values estimated in pigs may overestimate the quality of these proteins.


Formation and cultivation of Borrelia burgdorferi spheroplast-L-form variants.
As clinical persistence of Borrelia burgdorferi in patients with active Lyme borreliosis occurs despite obviously adequate antibiotic therapy, in vitro investigations of morphological variants and atypical forms of B. burgdorferi were undertaken. In an attempt to learn more about the variation of B. burgdorferi and the role of atypical forms in Lyme borreliosis, borreliae isolated from antibiotically treated and untreated patients with the clinical diagnosis of definite and probable Lyme borreliosis and from patient specimens contaminated with bacteria were investigated. Furthermore, the degeneration of the isolates during exposure to penicillin G in vitro was analysed. Morphological analysis by darkfield microscopy and scanning electron microscopy revealed diverse alterations. Persisters isolated from a great number of patients (60-80%) after treatment with antibiotics had an atypical form. The morphological alterations in culture with penicillin G developed gradually and increased with duration of incubation. Pleomorphism, the presence of elongated forms and spherical structures, the inability of cells to replicate, the long period of adaptation to growth in MKP-medium and the mycoplasma-like colonies after growth in solid medium (PMR agar) suggest that B. burgdorferi produce spheroplast-L-form variants. With regard to the polyphasic course of Lyme borreliosis, these forms without cell walls can be a possible reason why Borrelia survive in the organism for a long time (probably with all beta-lactam antibiotics) [corrected] and the cell-wall-dependent antibody titers disappear and emerge after reversion.


Serological detection of circulating Angiostrongylus vasorum antigen and specific antibodies in dogs from central and northern Italy.
The most frequently employed method for the diagnosis of Angiostrongylus vasorum in dogs is the detection of first stage larvae (L1) in faeces. The sensitivity of coproscopy, however, is limited in case of low parasite load, intermittent larval excretion, and during pre-patency. An epidemiological survey on dogs was conducted applying serological methods in two Italian regions where angiostrongylosis is endemic in foxes. 265 dog serum samples from Tuscany (central Italy - site A) and 447 from Liguria (north-western Italy - site B) were tested with a sandwich-ELISA for detection of circulating antigen, and with an ELISA using A. vasorum adult somatic antigen purified by monoclonal antibodies for specific antibody detection. During previous examinations dogs naturally infected with Leishmania infantum (n=149), Dirofilaria immitis (n=40), Dirofilaria repens (n=30), Acanthocheilonema reconditum (n=27), Crenosoma vulpis (n=1), A. vasorum (n=2), Capillaria aerophila (n=35), Capillaria boehmi (n=3), Toxocara canis (n=68), Toxascaris leonina (n=5), hookworms (n=37) and Trichuris vulpis (n=39) were detected. Sera of these dogs were used to evaluate cross reactions. In site A, 2 dogs (0.8%) were seropositive for antibody and antigen detection and 4 (1.5%) for antibody detection only. From site B, 4 dogs (0.9%) were seropositive for both tests, while other 4 dogs (0.9%) for antigen detection only and 9 dogs (2%) for antibody detection only. Considering a subgroup of 347 dogs from site B which had also been tested with the Baermann technique, 2 (0.6%) were positive for both tests, 4 (1.2%) for antigen detection only and 9 (2.6%) for antibody detection only. The two dogs which were positive for both serological tests were also positive for A. vasorum L1 in the faeces. No significant difference in seropositivities was observed in the group of dogs with other proven parasitic infections. A. vasorum serology presents significant advantages (diagnosis before patency, single serum sample instead of repeated faecal samples, rapidity and affordability particularly in case of large number of samples) and it can be considered a valid alternative for diagnosis in individuals and in epidemiological studies.


Case Report: Cryptococcal meningitis in an immunocompetent patient in Nepal - challenges in diagnosis and treatment.
A 50 year old woman from Nepal had clinical features suggestive of meningitis. Cerebrospinal fluid (CSF) analysis was normal except for the presence of cryptococcal antigen. The inclusion of test for Cryptococcus in the CSF helped in making the diagnosis of cryptococcal meningitis in our patient who was apparently immunocompetent. Treatment with liposomal amphotericin B could not be started on time due financial constraints. The patient had a stroke and further deteriorated. Liposomal amphotericin B is stocked by the government of Nepal for free supply to patients with visceral leishmaniasis, but the policy does not allow the drug to be dispensed for other infections. The family members of our patient acquired the drug within a few days from a government center using their political connections and following administering the treatment the patient improved. This case demonstrates the utility of considering cryptococcal meningitis as a differential diagnosis, and including tests for Cryptococcus when dealing with immunocompetent patients presenting with meningitis. It also demonstrates the effects of the sociopolitical situation on health care delivery in low- and middle-income countries (LMICs) such as Nepal.


Kinematic differences between neutral and flat feet with and without symptoms as measured by the Oxford foot model.
Flatfoot is a common presentation in children. It is usually asymptomatic, though a small number of children experience pain. Foot function during flatfoot walking is rarely considered, yet as an activity that places significant demands on the feet, this could explain the differences in terms of symptoms. This paper investigates walking patterns in neutral and flat feet, with and without symptoms, to determine which kinematic parameters are associated with symptomatic flat feet. This is a retrospective study in which one hundred and six children between five and 18 years old were assessed by a physiotherapist for foot posture. Each foot was classified into one of four groups, giving 98 asymptomatic neutral, 47 asymptomatic mild flat, 29 asymptomatic flat, and 38 symptomatic flat feet with complete data for analysis. Using Plug-In-Gait and Oxford-Foot-Model markers, walking kinematics were measured, along with ground reaction forces. Median values of 14 lower limb joint angles were calculated at foot strike, midstance, and foot off. Each foot was treated as an independent sample. ANOVA and ANCOVA (with the speed-related variable relative stride length as the covariate) and post-hoc tests were used to assess whether angles differed between groups. The symptomatic flat feet showed significant differences from asymptomatic groups (most commonly the neutral feet) in terms of hip flexion, knee flexion and varus, hindfoot inversion-eversion, and forefoot abduction-adduction. Increased forefoot abduction occurred throughout stance phase in symptomatic flatfooted participants compared to all asymptomatic groups. The results suggest that foot motion in the transverse plane is closely associated with the presence of symptoms in flat feet and that this is accompanied by changes in the kinematics of the ankle, knee, and hip.


Gender differences in factors predicting unsafe crossing decisions in adult pedestrians across the lifespan: a simulation study.
Adult pedestrian accident data has demonstrated that the risk of being killed or seriously injured varies with age and gender. A range of factors affecting road crossing choices of 218 adults aged 17-90+ were examined in a simulation study using filmed real traffic. With increasing age, women were shown to make more unsafe crossing decisions, to leave small safety margins and to become poorer at estimating their walking speed. However, the age effects on all of these were ameliorated by driving experience. Men differed from women in that age was not a major factor in predicting unsafe crossing decisions. Rather, reduced mobility was the key factor, leading them to make more unsafe crossings and delay longer in leaving the kerb. For men, driving experience did not predict unsafe road crossing decisions. Although male drivers were more likely to look both ways before crossing than male non-drivers, the impact of being a driver had a negative effect in terms of smaller safety margins and delay in leaving the kerb. The implications of the different predictor variables for men and women for unsafe road crossing are discussed and possible reasons for the differences explored.


Multiple mechanisms regulate imprinting of the mouse distal chromosome 7 gene cluster.
Genomic imprinting is an epigenetic process that results in the preferential silencing of one of the two parental copies of a gene. Although the precise mechanisms by which genomic imprinting occurs are unknown, the tendency of imprinted genes to exist in chromosomal clusters suggests long-range regulation through shared regulatory elements. We characterize a 800-kb region on the distal end of mouse chromosome 7 that contains a cluster of four maternally expressed genes, H19, Mash2, Kvlqt1, and p57(Kip2), as well as two paternally expressed genes, Igf2 and Ins2, and assess the expression and imprinting of Mash2, Kvlqt1, and p57(Kip2) during development in embryonic and extraembryonic tissues. Unlike Igf2 and Ins2, which depend on H19 for their imprinting, Mash2, p57(Kip2), and Kvlqt1 are unaffected by a deletion of the H19 gene region, suggesting that these more telomeric genes are not regulated by the mechanism that controls H19, Igf2, and Ins2. Mutations in human p57(Kip2) have been implicated in Beckwith-Wiedemann syndrome, a disease that has also been associated with loss of imprinting of IGF2. We find, however, that a deletion of the gene has no effect on imprinting within the cluster. Surprisingly, the three maternally expressed genes are regulated very differently by DNA methylation; p57(Kip2) is activated, Kvlqt1 is silenced, and Mash2 is unaffected in mice lacking DNA methyltransferase. We conclude that H19 is not a global regulator of imprinting on distal chromosome 7 and that the telomeric genes are imprinted by a separate mechanism(s).


Hdac6 deletion delays disease progression in the SOD1G93A mouse model of ALS.
Defects in axonal transport are thought to contribute to the pathogenesis of neurodegenerative disease. Because α-tubulin acetylation facilitates axonal transport, inhibition of the α-tubulin deacetylating enzymes, histone deacetylase 6 (Hdac6) and silent information regulator 2 (Sirt2), is thought to be an interesting therapeutic strategy for these conditions. Amyotrophic lateral sclerosis (ALS) is a one such rapidly progressive and fatal neurodegenerative disorder, in which axonal transport defects have been found in vitro and in vivo. To establish whether the inhibition of Hdac6 or Sirt2 may be of interest for ALS treatment, we investigated whether deleting Hdac6 or Sirt2 from the superoxide dismutase 1, SOD1(G93A) mouse affects the motor neuron degeneration in this ALS model. Deletion of Hdac6 significantly extended the survival of SOD1(G93A) mice without affecting disease onset, and maintained motor axon integrity. This protective effect was associated with increased α-tubulin acetylation. Deletion of Sirt2 failed to affect the disease course, but also did not modify α-tubulin acetylation. These findings show that Hdac6, rather than Sirt2, is a therapeutic target for the treatment of ALS. Moreover, Sirt2 appears not to be a major α-tubulin deacetylase in the nervous system.


Analysis of light emitting diode array lighting system based on human vision: normal and abnormal uniformity condition.
In this paper, condition for uniform lighting generated by light emitting diode (LED) array was systematically studied. To take human vision effect into consideration, contrast sensitivity function (CSF) was novelly adopted as critical criterion for uniform lighting instead of conventionally used Sparrow's Criterion (SC). Through CSF method, design parameters including system thickness, LED pitch, LED's spatial radiation distribution and viewing condition can be analytically combined. In a specific LED array lighting system (LALS) with foursquare LED arrangement, different types of LEDs (Lambertian and Batwing type) and given viewing condition, optimum system thicknesses and LED pitches were calculated and compared with those got through SC method. Results show that CSF method can achieve more appropriate optimum parameters than SC method. Additionally, an abnormal phenomenon that uniformity varies with structural parameters non-monotonically in LALS with non-Lambertian LEDs was found and analyzed. Based on the analysis, a design method of LALS that can bring about better practicability, lower cost and more attractive appearance was summarized.


AKR1D1 knockout mice develop a sex-dependent metabolic phenotype.
Steroid 5β-reductase (AKR1D1) plays important role in hepatic bile acid synthesis and glucocorticoid clearance. Bile acids and glucocorticoids are potent metabolic regulators, but whether AKR1D1 controls metabolic phenotype in vivo is unknown. Akr1d1-/- mice were generated on a C57BL/6 background. Liquid chromatography/mass spectrometry, metabolomic and transcriptomic approaches were used to determine effects on glucocorticoid and bile acid homeostasis. Metabolic phenotypes including body weight and composition, lipid homeostasis, glucose tolerance and insulin tolerance were evaluated. Molecular changes were assessed by RNA-Seq and Western blotting. Male Akr1d1-/- mice were challenged with a high fat diet (60% kcal from fat) for 20 weeks. Akr1d1-/- mice had a sex-specific metabolic phenotype. At 30 weeks of age, male, but not female, Akr1d1-/- mice were more insulin tolerant and had reduced lipid accumulation in the liver and adipose tissue yet had hypertriglyceridemia and increased intramuscular triacylglycerol. This phenotype was associated with sexually dimorphic changes in bile acid metabolism and composition but without overt effects on circulating glucocorticoid levels or glucocorticoid-regulated gene expression in the liver. Male Akr1d1-/- mice were not protected against diet-induced obesity and insulin resistance. In conclusion, this study shows that AKR1D1 controls bile acid homeostasis in vivo and that altering its activity can affect insulin tolerance and lipid homeostasis in a sex-dependent manner.


Alignment of the dye's molecular levels with the TiO(2) band edges in dye-sensitized solar cells: a DFT-TDDFT study.
We present a theoretical study of the lineup of the LUMO of Ru(II)-polypyridyl (N3 and N719) molecular dyes with the conduction band edge of a TiO(2) anatase nanoparticle. We use density functional theory (DFT) and the Car-Parrinello scheme for efficient optimization of the dye-nanoparticle systems, followed by hybrid B3LYP functional calculations of the electronic structure and time-dependent DFT (TDDFT) determination of the lowest vertical excitation energies. The electronic structure and TDDFT calculations are performed in water solution, using a continuum model. Various approximate procedures to compute the excited state oxidation potential of dye sensitizers are discussed. Our calculations show that the level alignment for the interacting nanoparticle-sensitizer system is very similar, within about 0.1 eV, to that for the separated TiO(2) and dye. The excellent agreement of our results with available experimental data indicates that the approach of this work could be used as an efficient predictive tool to help the optimization of dye-sensitized solar cells.


[Parental cancer--parents ways of coping, family functioning, and psychosocial adjustment of dependent children].
Children exposed to parental cancer have an increased risk of mental health problems. However, the parental illness itself and its features do not predict children's psychological adjustment. Parent- and family-related factors are more predictive for children's well-being and the incidence of psychopathological symptoms, respectively. This study focuses on parental ways of coping with illness from both, the ill and healthy parent's perspective, and the relationship with family functioning and children's adjustment. Results show a significant impact of parental coping styles on children's health-related quality of life and psychopathological symptoms and, furthermore, that this relationship is mediated by aspects of family functioning. This study support the importance of family systems approaches. Implications for further studies and practical issues are discussed.


Relationship between arterial distensibility and coronary atherosclerosis in angina patients.
Arterial stiffening is associated with an increased risk of cardiovascular disease. However, limited evidence exists on whether it also relates to subclinical atherosclerosis, thereby providing a non-invasive marker of the overall cardiovascular status. The aim of the present study was to provide information on arterial stiffening in angina patients in whom angiographic evaluation allowed quantification of coronary atherosclerosis. We studied 101 patients with angina from a large number admitted to our hospital for coronary angiography. In each patient, radial (RA), subdiaphragmatic aorta (AO) and carotid (CA) distensibility (Dist) were measured by an ultrasonic device, following ultrasonic exclusion of atherosclerotic lesions at these specific sites. Patients were classified into three groups according to the angiographic findings: (i) no significant coronary lesions (lumen obstruction < 50%, group A); (ii) one (group B); and (iii) two or three (group C) coronary vessels with hemodynamic significant plaques (lumen obstruction > 50%). Age, male prevalence, previous cardiovascular disease and interventions were progressively greater or more common from group A to C, whereas several other risk factors (plasma glucose, serum cholesterol, smoking, history of hypertension, etc.) did not differ between the three groups or between the group with single vessel (B) versus the group with multivessel disease (C). CA and AO Dist decreased progressively from group A to C with a significant relationship in the group as a whole between distensibility values and the number of diseased vessels. The progressive decrease in AO Dist from group A to C remained significant after adjustment for variables that showed between-group differences (such as gender, age and systolic blood pressure) and the ROC curve showed it to be a more sensitive and specific marker of coronary atherosclerosis than CA Dist. RA Dist was similar in the three groups and showed no relationship with the number of diseased vessels in the group as a whole. In patients with angina, AO and CA Dist are related to the severity of coronary atherosclerosis, with the relationship being better for alterations in aortic than in carotid mechanical properties. Large elastic artery (and in particular aortic) stiffening can thus be considered as a marker of the severity of coronary atherosclerosis, providing non-invasive obtainable information on the need to proceed with further clinical examinations.


Economic impact of clinical decision support interventions based on electronic health records.
Unnecessary healthcare utilization, non-adherence to current clinical guidelines, or insufficient personalized care are perpetual challenges and remain potential major cost-drivers for healthcare systems around the world. Implementing decision support systems into clinical care is promised to improve quality of care and thereby yield substantial effects on reducing healthcare expenditure. In this article, we evaluate the economic impact of clinical decision support (CDS) interventions based on electronic health records (EHR). We searched for studies published after 2014 using MEDLINE, CENTRAL, WEB OF SCIENCE, EBSCO, and TUFTS CEA registry databases that encompass an economic evaluation or consider cost outcome measures of EHR based CDS interventions. Thereupon, we identified best practice application areas and categorized the investigated interventions according to an existing taxonomy of front-end CDS tools. Twenty-seven studies are investigated in this review. Of those, twenty-two studies indicate a reduction of healthcare expenditure after implementing an EHR based CDS system, especially towards prevalent application areas, such as unnecessary laboratory testing, duplicate order entry, efficient transfusion practice, or reduction of antibiotic prescriptions. On the contrary, order facilitators and undiscovered malfunctions revealed to be threats and could lead to new cost drivers in healthcare. While high upfront and maintenance costs of CDS systems are a worldwide implementation barrier, most studies do not consider implementation cost. Finally, four included economic evaluation studies report mixed monetary outcome results and thus highlight the importance of further high-quality economic evaluations for these CDS systems. Current research studies lack consideration of comparative cost-outcome metrics as well as detailed cost components in their analyses. Nonetheless, the positive economic impact of EHR based CDS interventions is highly promising, especially with regard to reducing waste in healthcare.


Fish-Oil-Derived DHA-mediated Enhancement of Apoptosis in Acute Lymphoblastic Leukemia Cells is Associated with Accumulation of p53, Downregulation of Survivin, and Caspase-3 Activation.
In acute lymphoblastic leukemia (ALL), resistance to chemotherapy is associated with inactivation of p53 and upregulation of survivin. Thus, targeting the p53 and survivin expression may provide an attractive strategy for ALL treatment. It has been shown that fish-oil-derived docosahexaenoic acid (DHA) activates several antitumorigenic mechanisms in tumor cells, but little is known regarding the role of DHA on modulating p53 and survivin expression in ALL cells. In this study, we investigated the alterations of the p53 and survivin expression and induction of apoptosis in DHA-treated Molt-4 cells that serve as a model for ALL cells. Molt-4 cells were treated with 50, 100, 150, and 200 μM DHA after which cell proliferation, survivin mRNA and protein levels, p53 protein level, caspase-3 activation, and apoptotic rates were evaluated by different cellular and molecular techniques. After 48- and 72-h treatments with DHA at concentrations ranging from 50 to 200 μM, cell proliferation rates were measured to be 80.5-44.4%, and 73.4-14.4%, respectively, compared to untreated cells. We also found that treatment for 48 h with 200 μM DHA resulted in 10.8- and 3.6-fold increase in p53 protein level and caspase-3 activation followed by 4.7-and 1.6-fold decrease in survivin mRNA and protein levels, respectively, compared to untreated cells. Treatment of cells with different concentrations of DHA dramatically increased the p53/survivin and caspase-3/survivin ratios by 2.8- to 16.9-fold and 3.3 to 5.6-fold increases, respectively, compared to untreated cells. A decrease in the number of cells ranging from 16% to 70% and an increase in the number of apoptotic cells ranging from 9.3% to 93% was also observed with increasing DHA concentrations. In conclusion, p53 and survivin may provide promising targets of DHA in ALL cells and this compound with high proapoptotic capacity represents the possibility of its therapeutic application for ALL treatment.


Relationship of mammographic density and gene expression: analysis of normal breast tissue surrounding breast cancer.
Previous studies of breast tissue gene expression have shown that the extratumoral microenvironment has substantial variability across individuals, some of which can be attributed to epidemiologic factors. To evaluate how mammographic density and breast tissue composition relate to extratumoral microenvironment gene expression, we used data on 121 patients with breast cancer from the population-based Polish Women's Breast Cancer Study. Breast cancer cases were classified on the basis of a previously reported, biologically defined extratumoral gene expression signature with two subtypes: an Active subtype, which is associated with high expression of genes related to fibrosis and wound response, and an Inactive subtype, which has high expression of cellular adhesion genes. Mammographic density of the contralateral breast was assessed using pretreatment mammograms and a quantitative, reliable computer-assisted thresholding method. Breast tissue composition was evaluated on the basis of digital image analysis of tissue sections. The Inactive extratumoral subtype was associated with significantly higher percentage mammographic density (PD) and dense area (DA) in univariate analysis (PD: P = 0.001; DA: P = 0.049) and in multivariable analyses adjusted for age and body mass index (PD: P = 0.004; DA: P = 0.049). Inactive/higher mammographic density tissue was characterized by a significantly higher percentage of stroma and a significantly lower percentage of adipose tissue, with no significant change in epithelial content. Analysis of published gene expression signatures suggested that Inactive/higher mammographic density tissue expressed increased estrogen response and decreased TGF-β signaling. By linking novel molecular phenotypes with mammographic density, our results indicate that mammographic density reflects broad transcriptional changes, including changes in both epithelia- and stroma-derived signaling.


NF-κB activation by equine arteritis virus is MyD88 dependent and promotes viral replication.
NF-κB, a family of transcription factors involved in different cell functions and immune responses is targeted by viruses. The mechanism of NF-κB signalling and its role in replication of EAV have not been investigated. We demonstrate that EAV infection in BHK-21 cells activates NF-κB, and this activation was found to be mediated through the MyD88 pathway. Infection of IKKβ(-/-) murine embryo fibroblasts (MEFs), which are deficient in NF-κB signalling, resulted in lower virus titre, less cytopathic effect, and reduced expression of viral proteins. These findings implicate the MyD88 pathway in EAV-induced NF-κB activation and suggest that NF-κB activation is essential for efficient replication of EAV.


Comparative effects of radial and focused extracorporeal shock wave therapies in coccydynia.
This study was conducted to compare the effects of radial and focused extracorporeal shock wave therapy (ESWT) in patients with coccydynia. In this prospective randomized double-blind study conducted between March 2021 and October 2021, 60 patients with coccydynia (50 males, 10 females; mean age: 35.9±12.0 years, range 18 to 65 years) were randomized into three groups (n=20) according to different wave types of ESWT: focused, radial, and sham. The Visual Analog Scale (VAS) was used for pain assessment, and the Oswestry Disability Index (ODI) was used for functional assessment in all patients before the treatment (baseline), after the completion of four sessions of treatment (fourth week), one month after the end of the treatment (eighth week), and three months after the end of the treatment (16th week). The mean body mass index of the participants was 26.2±3.0. Compared to baseline, the VAS scores at four weeks were reduced only in the radial ESWT group (p<0.05). Compared to baseline, the VAS and ODI scores at eight and 16 weeks were significantly reduced in both the focused and radial ESWT groups (p<0.05 for all). The radial ESWT group was significantly superior to the focused ESWT group in the comparisons between the groups at four weeks in the VAS values and at 16 weeks in the ODI scores (p<0.05 for all). Radial and focused ESWT are both effective in treating coccydynia compared to sham ESWT. However, radial ESWT may be more effective in the treatment of coccydynia.


Phenotypic spectrum in families with mesial temporal lobe epilepsy probands.
The traditional perception of mesial temporal lobe epilepsy (MTLE) as a predominantly acquired disorder is challenged due to emerging evidence of familial aggregation. In this study, we ascertained the extent of familial occurrence of epilepsy in MTLE patients, as well as phenotypic heterogeneity in affected relatives. We identified and reevaluated patients with MTLE, treated at Epilepsy Department for a period of two years. All eligible putatively affected relatives were asked to participate in the study. In addition to comprehensive epilepsy interview, they underwent EEG and MRI studies. 52 patients with MTLE were included; nine of them (17%) had at least one family member with epilepsy. Subsequently, we analyzed nine probands with MTLE and a total of 15 relatives with seizures. Among affected relatives, spectrums of clinical manifestations were observed. Typical MTL seizures were described in five individuals, while other types of focal or generalized tonic-clonic seizures were reported in other ten relatives. A total of seven individuals had febrile seizures. Hippocampal sclerosis was found in three probands and none of the relatives. Two of affected family members had a traumatic brain injury in addition to febrile seizures, prior to the occurrence of their epilepsy. We demonstrate that familiar occurrence of epilepsy and subsequently putative genetic background, accounts for a substantial proportion MTLE patients. In addition, we foreground the remarkable intra- and interfamilial phenotypic heterogeneity than usually described, displaying the complexity of the genotype-phenotype correlations.


Some international perspectives on legislation for the management of human-induced safety risks.
Legislation that governs the health and safety of communities near major-hazard installations in South Africa is largely based on existing legislation that had been developed in the United Kingdom and other European Union countries. The latter was developed as a consequence of several major human-induced technological disasters in Europe. The history of the evolution of health-and-safety legislation for the protection of vulnerable communities in European Union (EU) countries, France, Malaysia and the USA is explored through a literature survey. A concise comparison is drawn between EU countries, the USA and South Africa to obtain an exploratory view of whether current South-African legislation represents an optimum model for the protection of the health-and-safety of workers and communities near major-hazard installations. The authors come to the conclusion that South-African legislation needs revision as was done in the UK in 2011. Specific areas in the legislation that need revision are an overlap between occupational health and safety and environmental legislation, appropriate land-use planning for the protection of communities near major-hazard installations, the inclusion of vulnerability studies and the refinement of appropriate decision-making instruments such as risk assessment. This article is the first in a series that forms part of a broader study aimed at the development of an optimised model for the regulatory management of human-induced health and safety risks associated with hazardous installations in South Africa.