Suppression of pulmonary CYP2A13 expression by carcinogen-induced lung tumorigenesis in a CYP2A13-humanized mouse model.
CYP2A13 is a human cytochrome P450 (P450) enzyme important in the bioactivation of the tobacco-specific lung procarcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). CYP2A13 expression levels vary dramatically among lung biopsy samples from patients, presumably owing in part to a suppression of CYP2A13 expression by disease-associated inflammation. Here, we determined whether CYP2A13 expression in the lungs of CYP2A13-humanized mice is suppressed by the presence of lung tumors. Tissues from an NNK lung tumor bioassay were examined. CYP2A13-humanized mice (95-100%) had multiple lung tumors at 16 weeks after NNK (30 or 50 mg/kg) treatment; whereas only ∼9% of saline-treated CYP2A13-humanized mice had lung tumor (∼1/lung). Mice with lung tumors, from the NNK-treated groups, were used for dissecting adjacent tumor-free lung tissues; whereas mice without visible lung tumors, from the saline-treated group, were used as controls. Compared with the controls, the levels of CYP2A13 protein and mRNA were both reduced significantly (by ≥50%) in the NNK-treated groups. The levels of mouse CYP2B10 and CYP2F2 mRNAs were also significantly lower in the dissected normal lung tissues from tumor-bearing mice than in lungs from the control mice. Pulmonary tissue levels of three proinflammatory cytokines, tumor necrosis factor alpha, interferon gamma, and interleukin-6, were significantly higher in the tumor-bearing mice than in the controls, indicating occurrence of low-grade lung inflammation at the time of necropsy. Taken together, these findings support the hypothesis that CYP2A13 levels in human lungs can be suppressed by disease-associated inflammation in tissue donors, a scenario causing underestimation of CYP2A13 levels in healthy lungs.


A multi-ancestry genome-wide study incorporating gene-smoking interactions identifies multiple new loci for pulse pressure and mean arterial pressure.
Elevated blood pressure (BP), a leading cause of global morbidity and mortality, is influenced by both genetic and lifestyle factors. Cigarette smoking is one such lifestyle factor. Across five ancestries, we performed a genome-wide gene-smoking interaction study of mean arterial pressure (MAP) and pulse pressure (PP) in 129 913 individuals in stage 1 and follow-up analysis in 480 178 additional individuals in stage 2. We report here 136 loci significantly associated with MAP and/or PP. Of these, 61 were previously published through main-effect analysis of BP traits, 37 were recently reported by us for systolic BP and/or diastolic BP through gene-smoking interaction analysis and 38 were newly identified (P < 5 × 10-8, false discovery rate < 0.05). We also identified nine new signals near known loci. Of the 136 loci, 8 showed significant interaction with smoking status. They include CSMD1 previously reported for insulin resistance and BP in the spontaneously hypertensive rats. Many of the 38 new loci show biologic plausibility for a role in BP regulation. SLC26A7 encodes a chloride/bicarbonate exchanger expressed in the renal outer medullary collecting duct. AVPR1A is widely expressed, including in vascular smooth muscle cells, kidney, myocardium and brain. FHAD1 is a long non-coding RNA overexpressed in heart failure. TMEM51 was associated with contractile function in cardiomyocytes. CASP9 plays a central role in cardiomyocyte apoptosis. Identified only in African ancestry were 30 novel loci. Our findings highlight the value of multi-ancestry investigations, particularly in studies of interaction with lifestyle factors, where genomic and lifestyle differences may contribute to novel findings.


Non-high density lipoprotein cholesterol versus low density lipoprotein cholesterol as a discriminating factor for myocardial infarction.
Serum total cholesterol (TC) and LDL cholesterol (LDL-C) have been used as major laboratory measures in clinical practice to assess cardiovascular risk in the general population and disease management as well as prognosis in patients. However, some studies have also reported the use of non-HDL cholesterol (non-HDL-C). As non-HDL-C can be calculated by subtracting HDL-C from TC, both of which do not require fasting blood sample in contrast to LDL-C which requires fasting blood sample, we aimed to compare non-HDL-C with LDL-C as a predictor of myocardial infarction (MI). This hospital based cross sectional study was undertaken among 51 cases of MI and equal number of controls. MI was diagnosed based on the clinical history, ECG changes and biochemical parameters. 5 mL of fasting blood sample was collected from each research participant for the analysis of lipid profile. Non-HDL-C was calculated by using the equation; Non-HDL-C = TC - HDL-C. Statistical analysis was performed using SPSS 14.0. 42 MI cases were dyslipidemic in contrast to 20 dyslipidemic subjects under control group. The differences in the median values of each lipid parameter were statistically significant between MI cases and controls. The lipid risk factors most strongly associated with MI were HDL-C (OR 5.85, 95% CI 2.41-14.23, P value = 0.000) followed by non-HDL-C (OR 3.77, 95% CI 1.64-8.66, P value = 0.002), LDL-C/HDL-C (OR 3.38, 95% CI 1.44-7.89, P value = 0.005), TC/HDL-C (OR 2.93, 95% CI 1.36-7.56, P value = 0.026), LDL-C (OR 2.70, 95% CI 1.20-6.10, P value = 0.017), TC (OR 2.68, 95% CI 1.04-6.97, P value = 0.042) and Tg (OR 2.54, 95% CI 1.01-6.39, P value = 0.047). Area under the receiver operating curve was greater for non-HDL-C than for LDL-C. Non-HDL-C was also found to be more sensitive and specific than LDL-C for MI. HDL-C and non-HDL-C are better discriminating parameters than LDL-C for MI. Thus, we can simply perform test for HDL-C and non-HDL-C both of which do not require fasting blood sample rather than waiting for fasting blood sample to measure LDL-C.


Effect of three different cultivars of Lepidium meyenii (Maca) on learning and depression in ovariectomized mice.
Lepidium meyenii Walp. (Brassicaceae), known as Maca, is a Peruvian hypocotyl growing exclusively between 4000 and 4500 m altitude in the central Peruvian Andes, particularly in Junin plateau and is used traditionally to enhance fertility. Maca is a cultivated plant and different cultivars are described according to the color of the hypocotyls. The study aimed to elucidate the effect of Yellow, Red and Black Maca on cognitive function and depression in ovariectomized (OVX) mice. In all experiments OVX mice were treated during 21 days and divided in four groups: control group, Yellow Maca, Red Maca and Black Maca. Latent learning was assessed using the water finding task and the antidepressant activity of the three varieties of Maca was evaluated using the forced swimming test. Animals were sacrificed at the end of each treatment and the uterus were excised and weighed. Black Maca was the variety that showed the best response in the water finding task, particularly in the trained mice. The three varieties were effective to reduce finding latency in non trained and trained mice (P < 0.05). In the force swimming test, all varieties assessed reduced the time of immobility and increased uterine weight in OVX mice. Black Maca appeared to have more beneficial effects on latent learning in OVX mice; meanwhile, all varieties of Maca showed antidepressant activity.


Dynamics of plasma gonadotropin and sex steroid release in polycystic ovarian disease after pituitary-ovarian inhibition with an analog of gonadotropin-releasing hormone.
To assess the dynamics of the suppression and recovery of plasma gonadotropins and sex steroids during and after inhibition of pituitary-ovarian function by a long-acting agonist GnRH-analog (GnRH-A), eight patients with polycystic ovarian disease were treated with 12 micrograms/kg X day GnRH-A for 56 consecutive days. In response to GnRH-A, these patients had a sharp and pronounced decline of their initially elevated immunoreactive LH and bioactive LH (bioLH) levels. Plasma immunoreactive FSH levels declined more rapidly than did bioLH, but the FSH decline was less sustained. Plasma testosterone, androstenedione, and estrone (E1) levels also declined during GnRH-A administration. The pattern of plasma androgen decrease resembled that of bioLH. There was a positive correlation between bioLH and the two androgens (r = 0.85; P less than 0.05, by Spearman's rank correlation, for both hormones). Cessation of GnRH-A administration was followed by prompt progressive increases in gonadotropin and androgen concentrations to pretreatment values. FSH recovered faster than bioLH. BioLH plasma concentrations reached pretreatment values by day 28. The recovery of plasma androstenedione and testosterone levels correlated positively with that of bioLH. Although plasma E1 levels were higher during the recovery period than during treatment, they never reached the concentrations found during the basal period, whereas estradiol concentrations were slightly but not significantly higher than those in the basal period. As a consequence, the E1 to estradiol ratio, very high in the basal period, approximated unity during recovery. These data indicate that hyperandrogenism in polycystic ovarian disease is gonadotropin dependent and accompanied by a relative abundance of LH bioactivity basally and during GnRH-A administration. Thus, the relative increase in bioLH secretion appears to be independent of the rate of gonadotropin secretion and the circulating sex steroid concentrations.


Molecular pathology of tumors of the central nervous system.
Since the update of the 4th edition of the WHO Classification of Central Nervous System (CNS) Tumors published in 2016, particular molecular characteristics are part of the definition of a subset of these neoplasms. This combined 'histo-molecular' approach allows for a much more precise diagnosis of especially diffuse gliomas and embryonal CNS tumors. This review provides an update of the most important diagnostic and prognostic markers for state-of-the-art diagnosis of primary CNS tumors. Defining molecular markers for diffuse gliomas are IDH1/IDH2 mutations, 1p/19q codeletion and mutations in histone H3 genes. Medulloblastomas, the most frequent embryonal CNS tumors, are divided into four molecularly defined groups according to the WHO 2016 Classification: wingless/integrated (WNT) signaling pathway activated, sonic hedgehog (SHH) signaling pathway activated and tumor protein p53 gene (TP53)-mutant, SHH-activated and TP53-wildtype, and non-WNT/non-SHH-activated. Molecular characteristics are also important for the diagnosis of several other CNS tumors, such as RELA fusion-positive subtype of ependymoma, atypical teratoid rhabdoid tumor (AT/RT), embryonal tumor with multilayered rosettes, and solitary fibrous tumor/hemangiopericytoma. Immunohistochemistry is a helpful alternative for further molecular characterization of several of these tumors. Additionally, genome-wide methylation profiling is a very promising new tool in CNS tumor diagnostics. Much progress has thus been made by translating the most relevant molecular knowledge into a more precise clinical diagnosis of CNS tumors. Hopefully, this will enable more specific and more effective therapeutic approaches for the patients suffering from these tumors.


First Report of Cucurbit chlorotic yellows virus Infecting Muskmelon and Cucumber in Sudan.
In summer 2009, a survey for virus diseases in cucurbits was conducted in open fields and plastichouses in Khartoum State, the most important growing area for cucurbits in Sudan. Chlorosis and yellowing symptoms on middle and lower leaves were observed on many muskmelon (Cucumis melo L.) plants grown in open fields in the Assilat agricultural scheme and on approximately 80% of the cucumber (Cucumis sativus L.) plants grown in plastichouses in Khartoum North. Large populations of whiteflies (Bemisia tabaci L.) were present in both locations. Leaf symptoms that were observed were similar to those caused by Cucurbit chlorotic yellows virus (CCYV), a recently described new Crinivirus species infecting cucurbits in Japan (4), indicating presence of this virus previously only reported from Japan, Taiwan (2), and China (1). Samples from seven symptomatic muskmelon leaves were collected from individual plants grown in different open fields in Assilat and from a symptomatic cucumber plant grown in a plastichouse. Total RNA was extracted from these samples with the RNeasy Plant Mini Kit (QIAGEN, Hilden, Germany) to amplify putative CCYV sequences with primers (Crini-s2 5'-CATTCCTACCTGTTTAGCCA and Crini-as2 5'-TGCACTTATAATCTGCTGGTAC) designed from CCYV sequences available at GenBank. A 353-bp DNA fragment of the HSP70 gene was amplified by reverse transcription (RT)-PCR for all samples. Further analysis by direct sequencing of two PCR products showed 99 to 100% nt sequence identity to Asian CCYV isolates. Amplification of the coat protein sequence with the primer pair (CCYV-CPs 5'-ATGGAGAAGACTGACAATAAACAA and CCYV-CPas 5'-TTTACTACAACCTCCCGGTG) followed by cloning and sequencing yielded a 760-bp fragment having 99% nucleotide sequence identity to all Asian isolates. For confirmation, dsRNA preparations of symptomatic muskmelon tissue (collected in June 2010) were made, showing dsRNA patterns typical for criniviruses after separation on agarose gels. This dsRNA was used as template for random RT-PCR followed by sequencing of the cloned PCR products (3). Comparison with sequences available at GenBank revealed that cDNA sequences from dsRNA also were 99 to 100% identical to the CCYV genome sequence (AB523788.1). Whitefly transmission of the virus was confirmed by giving a population of B. tabaci an acquisition access period of 24 h and a further 24 h on muskmelon and cucumber seedlings. Symptoms were observed after 5 to 7 days, and the presence of CCYV was confirmed by RT-PCR. In conclusion, symptoms, RT-PCR, and dsRNA sequencing results confirm the presence and establishment of CCYV in cucurbit crops in Sudan. It is remarkable that the sequences obtained from the Sudanese samples show only negligible sequence differences from Asian isolates. Because of the large whitefly vector populations, the spread of CCYV to neighboring countries in Africa and potentially southern Europe, or wherever cucurbits are grown, can be expected. To our knowledge, this is the first report of CCYV in Sudan and outside Eastern Asia. The sequences obtained in this study have been submitted to GenBank under Accession Nos. JF807053 to JF807055. References: (1) Q. S. Gu et al. Plant Dis. 95:73, 2011. (2) L. H. Huang et al. Plant Dis. 94:1168, 2010. (3) W. Menzel et al. Arch. Virol. 154:1343, 2009. (4) M. Okuda et al. Phytopathology 100:560, 2010.


Quantum phases of Bose-Fermi mixtures in optical lattices.
We analyze the ground-state phase diagram of an ultracold Bose-Fermi mixture placed in an optical lattice. The quantum phases involve pairing of fermions and one or several bosons. Depending on the physical parameters these composites can form a Fermi liquid, a density wave, a superfluid or a domain insulator. We determine by means of a mean-field formalism the phase boundaries for finite tunneling, and analyze the experimental feasibility of these sort of phases.


In utero exposure to diethylstilbestrol (DES) or bisphenol-A (BPA) increases EZH2 expression in the mammary gland: an epigenetic mechanism linking endocrine disruptors to breast cancer.
Diethylstilbestrol (DES) and bisphenol-A (BPA) are estrogen-like endocrine-disrupting chemicals that induce persistent epigenetic changes in the developing uterus. However, DES exposure in utero is also associated with an increased risk of breast cancer in adult women. Similarly, fetal exposure to BPA induces neoplastic changes in mammary tissue of mice. We hypothesized that epigenetic alterations would precede the increased risk of breast neoplasia after in utero exposure to endocrine disruptors. Enhancer of Zeste Homolog 2 (EZH2) is a histone methyltransferase that has been linked to breast cancer risk and epigenetic regulation of tumorigenesis. We examined the effect of BPA and DES on EZH2 expression and function in MCF-7 cells and in mammary glands of mice exposed in utero. DES and BPA treatment approximated human exposure. EZH2 functional activity was assessed by measuring histone H3 trimethylation. Treatment of MCF-7 cells with DES or BPA led to a 3- and 2-fold increase in EZH2 mRNA expression, respectively (p < 0.05) as well as increased EZH2 protein expression. Mice exposed to DES in utero showed a >2-fold increase in EZH2 expression in adult mammary tissue compared with controls (p < 0.05). EZH2 protein was elevated in mammary tissue of mice exposed to DES or BPA. Histone H3 trimethylation was increased in MCF-7 cells treated with BPA or DES. Similarly, mice exposed to BPA or DES in utero showed increased mammary histone H3 trimethylation. Developmental programming of EZH2 is a novel mechanism by which in utero exposure to endocrine disruptors leads to epigenetic regulation of the mammary gland.


The Neural Basis of Vocal Pitch Imitation in Humans.
Vocal imitation is a phenotype that is unique to humans among all primate species, and so an understanding of its neural basis is critical in explaining the emergence of both speech and song in human evolution. Two principal neural models of vocal imitation have emerged from a consideration of nonhuman animals. One hypothesis suggests that putative mirror neurons in the inferior frontal gyrus pars opercularis of Broca's area may be important for imitation. An alternative hypothesis derived from the study of songbirds suggests that the corticostriate motor pathway performs sensorimotor processes that are specific to vocal imitation. Using fMRI with a sparse event-related sampling design, we investigated the neural basis of vocal imitation in humans by comparing imitative vocal production of pitch sequences with both nonimitative vocal production and pitch discrimination. The strongest difference between these tasks was found in the putamen bilaterally, providing a striking parallel to the role of the analogous region in songbirds. Other areas preferentially activated during imitation included the orofacial motor cortex, Rolandic operculum, and SMA, which together outline the corticostriate motor loop. No differences were seen in the inferior frontal gyrus. The corticostriate system thus appears to be the central pathway for vocal imitation in humans, as predicted from an analogy with songbirds.


Focal macular electroretinograms after intravitreal injections of bevacizumab for age-related macular degeneration.
To evaluate the changes in the best-corrected visual acuity (BCVA), macular thickness, and focal macular electroretinograms (FMERGs) after three intravitreal injections of bevacizumab for a choroidal neovascularization (CNV) associated with age-related macular degeneration (AMD). The medical records of 18 eyes of 18 patients who had received three consecutive monthly intravitreal injections of bevacizumab were retrospectively studied. The BCVA, macular thickness determined by optical coherence tomography (OCT), and FMERGs were measured before the first injection, and 10 days after each of the intravitreal bevacizumab injections. The number of eyes with improvement in BCVA after the first injection was one (6%), after the second injection was four (22%), and after the third injection was five (28%). The number of eyes with reduction in macular thickness was 4 (33%), 8 (44%), and 10 (56%) after each of the three injections. The number of eyes with increase in b-wave amplitude of the FMERGs was 7 (38%), 6 (33%), and 10 (56%) after each of the three each injections. The mean macular thickness was significantly thinner after the first injection, and the mean BCVA was significantly improved after the second injection. The mean amplitude and implicit time of the b-wave of the FMERGs were significantly improved only after the third injection (P<0.05). All parameters improved but the best was after the third injection, indicating that three monthly intravitreous injections with bevacizumab may be an effective treatment regimen for AMD.


The protective effects of carvacrol and thymol against paracetamol-induced toxicity on human hepatocellular carcinoma cell lines (HepG2).
Acetaminophen (APAP) overdose could induce liver damage and lead to acute liver failure. The treatment of APAP overdoses could be improved by new therapeutic strategies. Thymus spp., which has many beneficial effects and has been used in folk medicine, is one such potential strategy. In the present study, the hepatoprotective activity of the main constituents of Thymus spp., carvacrol and thymol, were evaluated in light of APAP-induced hepatotoxicity. We hoped to understand the hepatoprotective mechanism of these agents on the antioxidant system and pro-inflammatory cytokines in vitro. Dose-dependent effects of thymol and carvacrol (25, 50, and 100 µM) were tested on cultured HepG2 cells. N-Acetylcysteine (NAC) was tested as positive control. We showed that APAP inhibited HepG2 cell growth by inducing inflammation and oxidative stress. Incubating APAP-exposed HepG2 cells with carvacrol and thymol for 24 h ameliorated this inflammation and oxidative stress. We also evaluated alanine transaminase and lactate dehydrogenase levels of HepG2 cells. We found that thymol and carvacrol protected against APAP-induced toxicity in HepG2 cells by increasing antioxidant activity and reducing pro-inflammatory cytokines, such as tumor necrosis factor α and interleukin 1β. Taking together high-dose thymol and carvacrol treatment has an effect close to NAC treatment in APAP toxicity, but thymol has better treatment effect than carvacrol.


Tailored radical hysterectomy for locally advanced cervical cancer.
The survival and prognostic factors for locally advanced cervical cancer treated with nerve-sparing Okabayashi-Kobayashi radical hysterectomy have not been elucidated. We aimed to evaluate the oncological outcomes of those patients after radical hysterectomy with adjuvant chemotherapy. This retrospective cohort study was conducted from January 2002 to December 2011. Treatment was conducted at a single tertiary center in northern Japan. We used the Okabayashi-Kobayashi radical hysterectomy with lymphadenectomy. We applied unilateral nerve preservation for stage IIA/IIB cancer if there was a one-sided extension of the disease outside the cervix. Indication for adjuvant therapy was based on Sedlis criteria. High-risk was defined as evidence of lymph node metastasis, pathological parametrial invasion, and a positive/close surgical margin. The choice of adjuvant therapy was chemotherapy which consisted of paclitaxel and cisplatin. The study included 76 early-stage IB1 (≤4 cm) and IIA1 cervical cancer and 45 locally advanced stage IB2 (>4 cm), IIA2, and IIB disease treated consecutively. The median follow-up was 106 (range: 6-203) months. There were 18 (15%) patients with recurrence, with five of 76 in the early-stage (7%) and 13 of 45 in the locally advanced disease (29%) (P<0.001). For locally advanced cervical cancer, pT classification (P<0.001), lymph node metastasis (P=0.007), and histology (P=0.05) were associated with locoregional recurrence. The five-year locoregional recurrence rate in the locally advanced disease was 20% and 5% in the early-stage disease (P=0.01). The five-year disease-free survival in the locally advanced cervical cancer was 71% and 93% in the early-stage disease (P<0.001). The overall survival in locally advanced disease depended on the adeno-type histology and lymph node metastasis. The tailored use of nerve-sparing Okabayashi-Kobayashi radical hysterectomy with adjuvant chemotherapy based on tumor histology and lymph node metastasis may be a possible option as a treatment of locally advanced cervical cancer in selected patients.


Field succession studies and casework can help to identify forensically useful Diptera.
Fly development rates, and to a lesser extent succession data, can be used to provide an estimate of a minimum postmortem interval (mPMI). Yet, these data are most useful when a full account of species' ecology, seasonality, and distribution is known. We conducted succession experiments on human cadavers over different seasons near Sydney, Australia, to document forensically useful information, including the pre-appearance interval for carrion flies. We also compiled a detailed record of flies identified in casework collected in 156 cases distributed across New South Wales, Australia. We then compared the occurrence of fly species from both field and casework datasets to identify any consistencies or gaps to determine how useful species might be for forensic investigations. In the field experiments, we found differences in species diversity and abundance between seasons, as well as yearly variation between two winter seasons. Most fly species we recorded ovipositing showed a 2- or 3-day delay between adult arrival and oviposition in summer, with a longer delay in winter. Species that were previously encountered in casework, such as Calliphora augur (Fabricius, 1775) and Calliphora ochracea Schiner, 1868, were confirmed as forensically useful, with their colonization behavior and seasonal preferences documented here. Although not encountered in casework, we confirmed Hemipyrellia fergusoni Patton, 1925 as a primary colonizer of human cadavers. Our study emphasizes the need to link field and casework data for a complete understanding of all aspects of a carrion fly's ecology to assist forensic investigators in mPMI estimations.


Genome-wide expression analysis offers new insights into the origin and evolution of Physcomitrella patens stress response.
Changes in the environment, such as those caused by climate change, can exert stress on plant growth, diversity and ultimately global food security. Thus, focused efforts to fully understand plant response to stress are urgently needed in order to develop strategies to cope with the effects of climate change. Because Physcomitrella patens holds a key evolutionary position bridging the gap between green algae and higher plants, and because it exhibits a well-developed stress tolerance, it is an excellent model for such exploration. Here, we have used Physcomitrella patens to study genome-wide responses to abiotic stress through transcriptomic analysis by a high-throughput sequencing platform. We report a comprehensive analysis of transcriptome dynamics, defining profiles of elicited gene regulation responses to abiotic stress-associated hormone Abscisic Acid (ABA), cold, drought, and salt treatments. We identified more than 20,000 genes expressed under each aforementioned stress treatments, of which 9,668 display differential expression in response to stress. The comparison of Physcomitrella patens stress regulated genes with unicellular algae, vascular and flowering plants revealed genomic delineation concomitant with the evolutionary movement to land, including a general gene family complexity and loss of genes associated with different functional groups.


Is priesthood an adaptive strategy? : Evidence from a historical Irish population.
This study examines the socioeconomic and familial background of Irish Catholic priests born between 1867 and 1911. Previous research has hypothesized that lack of marriage opportunities may influence adoption of celibacy as part of a religious institution. The present study traced data from Irish seminary registries for 46 Catholic priests born in County Limerick, Ireland, using 1901 Irish Census returns and Land Valuation records. Priests were more likely to originate from landholding backgrounds, and with landholdings greater in size and wealth than the local average. Priests were found to originate from families with more sons than the national average, but with similar numbers of daughters. These findings are discussed in relation to competition for resources and lineage survival strategies.


[Lumbosacral facet syndrome: functional and organic disorders of lumbosacral facet joints].
Disorders of lumbosacral (LS) facet joints are manifested by low back pain with or without referred leg pain and decreased mobility of the LS spine (LS facet syndrome). The most frequent causes of LS facet syndrome are functional disorders (functional blockade or dysfunction of facet joint=reversible restriction of facet joint movements caused by meniscoid entrapment) and degenerative changes of facet joints while the others are less frequent (spondyloarthropathies, infection, tuberculosis, synovial cyst, injury). Although it has been proven that the facet joints are one of the most frequent sources of chronic low back pain (15-45%), the fact is that the facet syndrome has been frequently overlooked in patients with chronic low back pain. Following are the main reasons for explaining why the facet syndrome has been overlooked in patients with chronic low back pain: 1. Facet joints disorders are manifested by non-specific clinical picture, 2.Diagnosis of facet syndrome cannot be established by either the conventional clinical examination or radiological examinations, 3. A very small number of doctors are practicing manual functional examination which can establish the diagnosis of facet joint dysfunction and 4. Diagnostic anesthetic block which can confirm the facet syndrome diagnosis is not a widely accessible method. There is a lack of research in frequency of facet syndrome in patients with acute low back pain. Chronic mechanical low back pain caused by dysfunction of several structurally unchanged facet joints can have the same features as the inflammatory pain which can result in misdiagnosis of spondyloarthropathy. Acute LS facet syndrome caused by dysfunction of facet joints responds very well to manual therapy. The most important therapeutic options in patients with chronic LS facet syndrome caused by degenerative changes and/or dysfunction of facet joints are manual therapy, kinesiotherapy (flexion exercises), therapeutic blocks and radiofrequency denervation. The article describes etiopathogenesis, clinical manifestations, diagnosis, differential diagnosis and therapy of LS facet syndrome with emphasis on functional disorders of facet joints that can cause particular diagnostic and therapeutic problems.


Resting-state BOLD signal variability is associated with individual differences in metacontrol.
Numerous studies demonstrate that moment-to-moment neural variability is behaviorally relevant and beneficial for tasks and behaviors requiring cognitive flexibility. However, it remains unclear whether the positive effect of neural variability also holds for cognitive persistence. Moreover, different brain variability measures have been used in previous studies, yet comparisons between them are lacking. In the current study, we examined the association between resting-state BOLD signal variability and two metacontrol policies (i.e., persistence vs. flexibility). Brain variability was estimated from resting-state fMRI (rsfMRI) data using two different approaches (i.e., Standard Deviation (SD), and Mean Square Successive Difference (MSSD)) and metacontrol biases were assessed by three metacontrol-sensitive tasks. Results showed that brain variability measured by SD and MSSD was highly positively related. Critically, higher variability measured by MSSD in the attention network, parietal and frontal network, frontal and ACC network, parietal and motor network, and higher variability measured by SD in the parietal and motor network, parietal and frontal network were associated with reduced persistence (or greater flexibility) of metacontrol (i.e., larger Stroop effect or worse RAT performance). These results show that the beneficial effect of brain signal variability on cognitive control depends on the metacontrol states involved. Our study highlights the importance of temporal variability of rsfMRI activity in understanding the neural underpinnings of cognitive control.


Induction of preneoplastic lung lesions in guinea pigs by cigarette smoke inhalation and their exacerbation by high dietary levels of vitamins C and E.
The development of effective chemopreventive agents against cigarette smoke-induced lung cancer could be greatly facilitated by the availability of suitable laboratory animal models. Here we report that male Hartley guinea pigs treated with cigarette smoke by inhalation twice a day for 28 days developed preneoplastic lung lesions, including bronchial hyperplasia, dysplasia and squamous metaplasia, analogous to those found in human smokers. The lesions were accompanied by increased expression of proliferating cell nuclear antigen and activation of the serine/threonine kinase Akt in the bronchial epithelium. In contrast, no lung lesions were found in guinea pigs ('sham smoked') that were submitted to identical procedures but without cigarettes. Compared with a diet low in vitamin C (50 p.p.m.) and vitamin E (15 p.p.m.), a diet high in vitamin C (4000 p.p.m.) and vitamin E (40 p.p.m.) significantly increased the incidence of these lesions. The inclusion of 1,4-phenylenebis(methylene)selenocyanate (p-XSC), a synthetic chemopreventive organoselenium compound, in the high vitamin C-high vitamin E diet at a level of 15 p.p.m. as selenium appeared to decrease the lesion incidence. Administration of (-)-epigallocatechin gallate, a powerful green tea polyphenolic antioxidant, at 560 p.p.m. in the drinking water had no effect. As in human smokers, levels of ascorbate in blood plasma, lung, liver and the adrenal glands were significantly decreased by cigarette smoke inhalation. These results identify a relevant in vivo laboratory model of cigarette smoke-induced lung cancer, suggest that p-XSC may have activity as a chemopreventive agent against cigarette smoke-induced lung lesions and provide additional evidence that very high dietary levels of certain antioxidants can have co-carcinogenic activity in cigarette smoke-induced lung cancer.


Synergy between myogenic and non-myogenic cells in a 3D tissue-engineered craniofacial skeletal muscle construct.
In vitro skeletal muscle engineering involves the culture of isolated primary myogenic cells in an environment conducive to the formation of a three-dimensional (3D) tissue construct capable of generating force. Isolated human myogenic cells have been used to study cell-cell interactions, permitting identification of functions intrinsic to skeletal muscle in two dimensions (2D). However, the independent contribution of human myogenic and non-myogenic cell types that comprise skeletal muscle to myogenic cell differentiation, force generation and matrix remodelling has yet to be established in 3D. The objective of this study was to use isolated human myogenic and non-myogenic muscle-derived cells (MDC) seeded in 3D collagen constructs to engineer a biomimetic craniofacial skeletal construct. The aim was to purify the two subpopulations of myogenic and non-myogenic cells from human masseter muscle and quantitate myogenic cell differentiation, force generation and matrix remodelling of the 3D collagen construct. The results showed that both the heterogeneous mixture of cells and the purified myogenic cell population expressed myogenin, indicative of myogenic cell differentiation. Further, there was a synergistic effect as the heterogeneous co-culture of myogenic and non-myogenic cells generated the highest peak force and expressed the most MMP-2 mRNA compared to isolated individual cell populations.


Doxorubicin and daunorubicin induce processing and release of interleukin-1β through activation of the NLRP3 inflammasome.
Anthracyclines including doxorubicin and daunorubicin are commonly used for the treatment of both hematologic and solid tumors. Dose related adverse effects often limit the effectiveness of anthracyclines in chemotherapy. Drug-related systemic inflammation mediated by interleukin-1beta (IL-1β) has been implicated in contributing to these adverse effects. The molecular mechanisms underlying anthracycline-mediated expression and IL-1β release are not understood. Elucidating the molecular basis by which anthracyclines upregulate IL-1β activity may present opportunities to decrease the inflammatory consequences of these drugs. Here we demonstrate that doxorubicin induces a systemic increase in IL-1β and other inflammatory cytokines, chemokines and growth factors including TNF-α, IL-6, CXCL1/Gro-α, CCL2/MCP-1, granulocyte colony stimulating factor (GCSF), and CXCL10/IP-10. Studies with IL-1R-deficient mice demonstrate that IL-1 signaling plays a role in doxorubicin-induced increases in IL-6 and GCSF. In vitro studies with doxorubicin and daunorubicin failed to induce expression of proIL-1β in unprimed murine bone marrow-derived macrophages (BMDM) but enhanced the expression of proIL-1β in BMDM that had previously been primed with LPS. Furthermore, doxorubicin and daunorubicin induced the processing and release of IL-1β from LPS-primed BMDM by providing danger signals that lead to assembly and activation of the inflammasome. The release of IL-1β required the expression of ASC, caspase-1, and NLRP3, demonstrating that doxorubicin and daunorubicin-induced inflammation is mediated by the NLRP3 inflammasome. As with other agents that induce activation of the NLRP3 inflammasome, the ability of doxorubicin to provide proinflammatory danger signals was inhibited by co-treatment of cells with ROS inhibitors or by incubating cells in high extracellular potassium. These studies suggest that proinflammatory responses to anthracycline chemotherapeutic agents are mediated, at least in part, by promoting the processing and release of IL-1β, and that some of the adverse inflammatory consequences that complicate chemotherapy with anthracyclines may be reduced by suppressing the actions of IL-1β.


Chronobiological study of the relationship between dyspnoea and airway obstruction in symptomatic asthmatic subjects.
1. We performed a chronobiological study of the relationship between peak expiratory flow rate (PEFR) and magnitude of dyspnoea in 35 symptomatic asthmatic patients to determine how accurately asthmatic subjects assess spontaneous airway obstruction and whether this accuracy varies throughout the 24 h period and depends on characteristics of the subjects or asthma. 2. At 07.00, 11.00, 15.00, 19.00 and 23.00 hours on 8 consecutive days in their ordinary environment and under their usual drug regimen, the subjects first rated their dyspnoea with a visual analogue scale and immediately after recorded their PEFR. 3. The linear regression coefficients between dyspnoea score and PEFR for the 35 subjects were continuously distributed between -0.93 and +0.21, with most r values ranging from -0.9 to -0.5. The group median r values calculated at each time point showed that strongest correlation between dyspnoea score and PEFR occurred at 7 h. which coincided with the lowest PEFR values. 4. We chose r = -0.7 (r2 = 0.5) as a limit to distinguish good perceivers (-1 less than r less than or equal to -0.7) from bad perceivers of airway obstruction. These two populations differed only by a higher variability of both PEFR and dyspnoea score in good than in bad perceivers, but not for severity, duration or treatment of asthma, absolute value of dyspnoea score, age or sex. 5. Our study quantified an important intra- and between-subject variability in the accuracy of perception of spontaneous airway obstruction in asthmatic subjects which seems to be unrelated to most characteristics of asthma.


Effects of light-emitting diode irradiation on RANKL-induced osteoclastogenesis.
Bone homeostasis is maintained by a balance between osteoblastic bone formation and osteoclastic bone resorption, where intracellular reactive oxygen species (ROS) are crucial mediators of osteoclastogenesis. Recently, low-level light therapy (LLLT), a form of laser medicine used in various clinical fields, was shown to alleviate oxidative stress by scavenging intracellular ROS. The present study aimed to investigate the impact of 635 nm irradiation from a light-emitting diode (LED) on osteoclastogenesis from receptor activator of nuclear factor kappa-B (NF-κB) ligand (RANKL)-stimulated mouse bone marrow-derived macrophages (BMMs). The effects of LED irradiation on osteoclastogenesis were assessed in tartrate-resistant acid phosphatase (TRAP), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), cell viability, and resorption pit formation, respectively. Quantitative real-time polymerase chain reaction (qPCR) and Western blot analyses were also performed to assess mRNA expression of osteoclastogenesis-related genes and phosphorylation of extracellular signal-regulated kinase 1/2 (ERK 1/2), p38, and c-Jun-N-terminal kinase (JNK). NF-κB activity was assayed by luciferase reporter assay and Intracellular ROS generation was investigated by the 2',7'-dichlorodihydrofluorescein diacetate (H2 DCF-DA) detection method. LED irradiation significantly inhibited RANKL-mediated osteoclast differentiation from BMMs and mRNA expression of TRAP, osteoclast-associated immunoglobulin-like receptor (OSCAR), and dendrocyte-expressed seven-transmembrane protein (DC-STAMP). Exposure to LED light likewise significantly decreased RANKL-facilitated NF-κB activity, p38 and ERK phosphorylation and intracellular ROS generation, and increased gene expression of nuclear factor E2-related factor 2 (Nrf2). Taken together, the results presented herein show that LED irradiation downregulates osteoclastogenesis by reducing ROS production. Therefore, LED irradiation/LLLT might be useful as an alternative, conservative approach to osteoporosis management.


Diagnostic reliability considerations of specific IgE determination.
Total IgE determination constitutes a good method for the screening of atopic diseases, though its actual value is controversial because normal values of total IgE do not exclude the existence of atopic disease, and high values of total IgE are not pathognomonic of atopy by themselves. The first step in identifying an atopic individual as such, after doing his anamnesis, can be carried out by means of total IgE determination. Most atopic individuals have high IgE values, but a normal result must be carefully interpreted: age and season-related variations must be considered. In general, atopic patients with IgE values greater than 1000 Ul/ml, always have positive specific IgE against some allergen. Antigen-specific IgE will be the next step in the in vitro identification of the responsible allergen. Nowadays, there are more than 400 characterized allergens available for in vitro diagnostic tests and several useful methodologies for specific IgE determination. Specific IgE results obtained with the different methods vary significantly, with absolute agreement in 55-65% of the cases, differences in one IgE class in 20-30% of the cases and differences in more than two classes in 5-10%. The specificity of the anti-IgE antibody used in the assay is of critical importance because any contaminant antibody can render unspecific results. On the other hand, it must be pointed out that there is a compromise between specificity and sensitivity, such that an increase in the sensitivity of a technique leads to a decrease in its specificity. It cannot be said that there is one method which is better than the others; it is better to examine them individually, allergen by allergen. Thus, specific IgE determination varies depending on the type of allergen. In general terms, for inhalant allergens, specificity and sensitivity of the methods are within the range of 85-95%, but these values (especially the specificity) decrease in the case of food allergens, and they are still lower when the allergen is a beta-lactamic drug. There is a good correlation between clinical history and specific IgE against inhalant allergens, and a lower correlation in the case of food allergens. Due to the fact that most food allergens are not standardized, the definitive diagnosis of food hypersensitivity is achieved by means of provocation tests. Nevertheless, negative specific IgE (7-18% of the cases) does not rule out a sensitization against the tested allergen, and a positive specific IgE without symptoms must be carefully interpreted because it can be due to a low degree of sensitization, unable to express clinical symptoms at this moment, but useful in the future as a guide on the disease course. In the evolutive period of the disease, specific IgE levels can be modified in a natural way (in beta-lactam allergy, 50% of the cases with specific IgE become negative after a year), or as an effect of the treatment (e.g., after immunotherapy in the case of Hymenoptera venom allergy), or it can remain positive for a long time, as in the case of pollinosis. On the other hand, the cutoff of the method, and subsequently the range of values to be considered as positive, will depend on the allergen studied. While inhalant allergens (with the exception of some molds) offer relatively high mean values of specific IgE, food and drug allergens yield less significant values. In general, a class greater than 2 is interpreted as clinically significant, class 1 as dubious or negative (depending on the allergen) and class 0 as negative. In the interpretation of the results, the possible presence of IgG and anti-IgE antibodies, capable of modifying the results, will be taken into account. When comparing the diagnostic reliability of specific IgE with respect to other allergologic diagnostic methods, we find a significant and positive correlation of this technique with skin tests (but never greater than 90-95%) and with the histamine release test.


Disc herniation caused by a viscoelastic nucleus after total lumbar disc replacement-a case report.
Degenerative disc disease (DDD) is highly prevalent. If conservative treatment fails, spinal fusion procedures are commonly performed. Total disc replacement (TDR) might be a surgical option for a distinct subset of patients with DDD. Several prostheses have been or are still available. Despite some promising initial clinical results, there is still limited experience with hardware-related adverse events. This report highlights an unreported complication after TDR with a viscoelastic device. Literature about long-term outcome and safety of this particular TDR is scarce. Hence, there exists limited experience with TDR-related complications with such a failure mode. We report a 34-year-old male presented to us with an acute S1 radiculopathy on the right. His past medical history was significant for prior TDR at the level L5/S1 at another hospital 2 years prior to this acute episode. Imaging studies revealed an intraspinal mass compromising the right S1 nerve root. This mass mimicked a disc herniation and sequestrectomy was performed. Intraoperatively, the prolapsed sequester turned out to be part of the viscoelastic nucleus of the disc prosthesis. Interbody fusion combined with posterior instrumentation was ultimately performed. The patient did well afterwards, but is currently (2 years later) developing adjacent segment disease with facet syndromes. Since TDR might be beneficial for certain patients, spine surgeons should be aware of potential device-related complications.


Reduction of ischemic injury by sublingual nitroglycerin in patients with acute myocardial infarction.
The effect of sublingual nitroglycerin (NTG) on myocardial ischemic injury was evaluated in eleven patients with acute anterior myocardial infarction. Precordial 35-lead ST-segment maps were obtained in each patient immediately before and 3-10 minutes after 0.4 mg sublingual NTG. The following measurements were made from each ST map: N-ST (number of leads showing ST elevation greater than 1mm), sigmaST (total ST elevation in all leads), ST (average ST-segment elevation in those leads with less than 1mm elevation). Following 0.4 mg sublingual NTG evidence of myocardial ischemic injury as assessed by ST-segment mapping decreased in association with reduction of heart rate X systolic blood pressure product (10.80 X 10(3) to 9.49 X 10(3), P less than 0.001). Group mean values diminished significantly for N-ST (18.1 to 14.4, P less than 0.001), sigma ST (37.9 to 30.1 P less than 0.005) and ST (1.7 to 1.4, P less than 0.001). Evaluation performed by the technique of precordial ST-segment mapping suggests that sublingual nitroglycerin in a commonly employed clinical dose is associated with evidence of reduced ischemic cardiac injury in patients with acute myocardial infarction. This effect appears to be related to reduction of myocardial oxygen demand by the nitrate.


Chitosan/PEI patch releasing EGF and the EGFR gene for the regeneration of the tympanic membrane after perforation.
Damage to the eardrum causes acute pain and can lead to chronic otitis media if it develops into chronic tympanic membrane (TM) perforations. Chronic TM perforations are usually treated with surgical methods such as tympanoplasty and myringoplasty. However, these surgeries are not only complicated and difficult but also cost a lot of money. Our research team developed chitosan patches (E-CPs) that release epidermal growth factor (EGF) as a patch therapy to replace surgical methods. However, there was a limitation in the healing ratio of the treatment compared to the surgical methods. In this study, we developed EGF and epidermal growth factor receptor (EGFR) gene-releasing polyethyleneimine (PEI)/chitosan patches (EErP-CPs) to increase the regeneration of TM perforations. The addition of PEI increased the adhesion and migration ability of TM cells on the patches. The simultaneous release of the EGF and the EGFR gene further enhanced TM cell proliferation, adhesion and migratory ability. It was confirmed that the EGF protein and EGFR gene were released for 30 days; however, EGF was released and increased TM cell viability almost immediately after treatment and EGFR took a minimum of 3 days before showing its effect on improved cell viability. It was also shown that EErP-CPs are more hydrophilic and have more positive charge than E-CP because of added amine groups from PEI. In conclusion, the developed EErP-CPs resulted in the improved healing of TM perforations and can potentially be applied to the regeneration of both chronic and acute tympanic membrane perforations.


A multidimensional measure of animal ethics orientation - Developed and applied to a representative sample of the Danish public.
We present a questionnaire-based measure of four animal ethics orientations. The orientations, which were developed in light of existing empirical studies of attitudes to animal use and ethical theory, are: animal rights, anthropocentrism, lay utilitarianism, and animal protection. The two latter orientations can be viewed as variants of animal welfarism. Three studies were conducted in Denmark in order to identify the hypothesised orientations, evaluate their concurrent validity, and report their prevalence and relevance in animal-related opinion formation and behaviour. Explorative factor analysis (Study 1) and confirmative factor analysis (Study 2) successfully identified the four orientations. Study 2 revealed good measurement invariance, as there was none or very modest differential item functioning across age, gender, living area, and contrasting population segments. Evaluation of concurrent validity in Study 2 found that the orientations are associated with different kinds of behaviour and opinion when the human use of animals is involved in the hypothesised directions. In Study 3, a representative population study, the animal protection orientation proved to be most prevalent in the Danish population, and as in study 2 the four orientations were associated with different behaviours and opinions. Remarkably, the animal protection orientation does not lead to increased animal welfare-friendly meat consumption, the main reason for this being non-concern about the current welfare status of farm animals. We argue that the developed measure covers a wide range of diversity in animal ethics orientations that is likely to exist in a modern society such as Denmark and can be used in future studies to track changes in the orientations and to understand and test hypotheses about the sources and justifications of people's animal-related opinions and behaviours.


Use of psychotropic medications by youths in therapeutic foster care and group homes.
This article examines the use of psychotropic medications among youths in residential community-based placements. Data are from a study funded by the National Institute of Mental Health of therapeutic foster care (June 1999 through May 2001) and group homes (January through June 2001). Data were collected from staff by means of in-person interviews. Many youths in both settings received psychotropic medications, and approximately one-half were taking multiple psychotropic medications. After the authors controlled for demographic and clinical factors, the youths in group homes were nearly twice as likely as the youths in therapeutic foster care to receive medication. However, residential setting was not related to polypharmacology. Additional work is needed to study the appropriateness of use and implications of such patterns for research on intervention outcomes.


Predominant promotion by tacrolimus of chondrogenic differentiation to proliferating chondrocytes.
Tacrolimus (FK506) has been used as a therapeutic drug beneficial for the treatment of rheumatoid arthritis in humans. In this study, we investigated the effects of FK506 on cellular differentiation in cultured chondrogenic cells. Culture with FK506 led to a significant and concentration-dependent increase in Alcian blue staining for matrix proteoglycan at 0.1 to 1,000 ng/ml, but not in alkaline phosphatase (ALP) activity, in ATDC5 cells, a mouse pre-chondrogenic cell line, cultured for 7 to 28 days, while the non-steroidal anti-inflammatory drug indomethacin significantly decreased Alcian blue staining in a concentration-dependent manner, without altering ALP activity. FK506 significantly increased the expression of mRNA for both type II and type X collagen, but not for osteopontin, in ATDC5 cells. Similar promotion was seen in chondrogenic differentiation in both mouse metatarsals and chondrocytes cultured with FK506. However, FK506 failed to significantly affect transcriptional activity of the reporter construct for either sry-type HMG box 9 (Sox9) or runt-related transcription factor-2 (Runx2), which are both transcription factors responsible for chondrocytic maturation as a master regulator. These results suggest that FK506 may predominantly promote cellular differentiation into proliferating chondrocytes through a mechanism not relevant to the transactivation by either Sox9 or Runx2 in chondrogenic cells.


Are Serum Metal Ion Levels a Concern at Mid-term Followup of Revision Knee Arthroplasty With a Metal-on-metal Hinge Design?
Elevated serum levels of chromium and cobalt ions in metal-on-metal (MoM) bearing surfaces is a well-known phenomenon in THA. However, few studies have addressed this issue in complex primary and revision knee arthroplasty using a MoM hinged mechanism, and no study, to our knowledge, has investigated knees with MoM hinges in patients without megaprostheses (tumor prostheses). We analyzed a series of patients who received MoM hinged revision knee prostheses and asked: (1) What are the serum metal ion levels at short-term followup? (2) Is there any correlation between metal ion levels and the Knee Society Score (KSS) at this followup? Between 2013 and 2017, we performed 198 revision knee arthroplasties, of which 32 (17.7%) were performed with a latest-generation MoM hinge knee design. In addition, three complex primary TKAs utilizing the same design were included in this study. The device features a metal-on-polyethylene bearing with a MoM hinge. During that period, our general indications for using a hinge were single-stage and two-stage revision surgeries, revisions with large bone defects, and primary TKA with > 20° mechanical malalignment or collateral ligament insufficiency. Of the 35 patients who received this device, 23 patients (65% of the overall group who received this implant; 11 males, 12 females) were available for followup at a median of 28 months (range, 13-61 months), and the remaining 12 (35%) patients were lost to followup. Our rationale for reporting before the more typical 2-year minimum was the finding of elevated serum ion levels with unclear clinical significance. Median age at the time of surgery was 68 years (range, 52-84 years). None of the patients included in the study had other implants with MoM bearings. Serum ion levels of chromium (III) and cobalt were assessed using mass spectrometry. Ion levels > 5 ppb were considered elevated. Clinical outcome was assessed using the original KSS. Median chromium serum level was 6.3 ppb (range, 0.6-31.9 ppb) and median cobalt serum level was 10.5 (range, 1.0-47.5 ppb). Of the 23 patients, 16 had elevated serum ion levels. There was a moderate correlation between KSS and both chromium (p = 0.029, r = 0.445) and cobalt (p = 0.012, r = 0.502) levels. Elevated metal ion levels and radiolucent lines are common after surgery with this MoM hinge design at short-term followup, and we believe this finding is of great concern. Although no patient has yet been revised, these patients will be closely monitored. We recommend that serum ion analysis become a routine part of followup after any hinge TKA in an attempt to better understand the potential consequences of this phenomenon. Level IV, therapeutic study.


Reconstructive challenges of complex battlefield injury.
Throughout our current conflicts in both Iraq and Afghanistan, the likelihood of surviving a battlefield injury has been near 90% according to reports provided by the US Army Medical Research and Materiel Command. This is the highest survival rate recorded in modern combat. Advancements in protective equipment, rapid evacuation of casualties from point of injury to both stabilization and definitive care, improved medic and corpsmen personnel, more readily available resuscitation, and forward positioned surgical units have all significantly decreased troop mortality. Along with this phenomenon of increased survival, however, has come a surge in the number and significance of injuries seen by the medical community. As a result, orthopaedists have been forced to confront reconstructive challenges and difficult decisions in an often highly motivated patient population with high functional expectations. The objective of this article is to outline a few examples of difficult challenges encountered in battlefield trauma care and discuss the treatments utilized.


ERP differences of pre-lexical processing between dyslexic and non-dyslexic children.
The present Event-Related Potential (ERP) study aimed to investigate group differences in the early processing stages of 36 dyslexic and 24 non-dyslexic 8-12 year old children performing a lexical decision (word/pseudoword judgment) task. Our data showed larger amplitudes of negative-going waveforms in non-dyslexic children than dyslexic children over occipital/occipitotemporal electrodes at about 220 ms after stimulus onset. This electrophysiological response has previously been identified in adult readers and labeled as the N170 component. Notably, as reflected by the topographic maps children irrespective of group processed the linguistic stimuli bilaterally and we did not observe any differences in ERP parameters in words and pseudowords within groups. Contrarily, behavioral responses indicate that words were more quickly recognized than pseudowords irrespective of group. By applying post-hoc ROI analyses based on a source estimation approach (sLORETA) we observed that non-dyslexic participants, when compared to dyslexic children, demonstrated significantly stronger current density over the left hemispheric inferior temporal lobe when processing pseudowords. We concluded that impaired reading is reflected by the decreased amplitude of the early lexical component N170. The lack of a left hemispheric processing preference in both groups and similar activation for words and pseudowords can be considered a lack of reading experience and less established reading system in children. Our results indicate that dyslexic children commit fewer specialized neuronal circuits for processing print and confirm the reasoning that acquiring reading skills requires cortical reorganization over occipitotemporal regions.


Inverse enzyme isotope effects in human purine nucleoside phosphorylase with heavy asparagine labels.
Transition path-sampling calculations with several enzymes have indicated that local catalytic site femtosecond motions are linked to transition state barrier crossing. Experimentally, femtosecond motions can be perturbed by labeling the protein with amino acids containing 13C, 15N, and nonexchangeable 2H. A slowed chemical step at the catalytic site with variable effects on steady-state kinetics is usually observed for heavy enzymes. Heavy human purine nucleoside phosphorylase (PNP) is slowed significantly (kchemlight/kchemheavy = 1.36). An asparagine (Asn243) at the catalytic site is involved in purine leaving-group activation in the PNP catalytic mechanism. In a PNP produced with isotopically heavy asparagines, the chemical step is faster (kchemlight/kchemheavy = 0.78). When all amino acids in PNP are heavy except for the asparagines, the chemical step is also faster (kchemlight/kchemheavy = 0.71). Substrate-trapping experiments provided independent confirmation of improved catalysis in these constructs. Transition path-sampling analysis of these partially labeled PNPs indicate altered femtosecond catalytic site motions with improved Asn243 interactions to the purine leaving group. Altered transition state barrier recrossing has been proposed as an explanation for heavy-PNP isotope effects but is incompatible with these isotope effects. Rate-limiting product release governs steady-state kinetics in this enzyme, and kinetic constants were unaffected in the labeled PNPs. The study suggests that mass-constrained femtosecond motions at the catalytic site of PNP can improve transition state barrier crossing by more frequent sampling of essential catalytic site contacts.


What do urban college students really think about health insurance? A qualitative study.
Objective: To determine barriers and opportunities to health insurance enrollment among an undergraduate students at a large urban university. Participants: Participants were 31 college students enrolled in 4-year and community colleges in the City University of New York (CUNY), and six health services and insurance enrollment specialists who facilitate and assist in the health insurance enrollment process for CUNY students. Methods: Focus groups were conducted with students and in-depth interviews with key informants in May 2017. Results: The research revealed important insights into how students perceive and value insurance and yielded recommendations for the university to improve enrollment of its students. Conclusions: Many colleges can increase student enrollment in health insurance by informing and educating students about the process. Improving enrollment processes can increase insurance rates and improve student population health.


Health-related quality of life of patients with advanced breast cancer treated with everolimus plus exemestane versus placebo plus exemestane in the phase 3, randomized, controlled, BOLERO-2 trial.
The randomized, controlled BOLERO-2 (Breast Cancer Trials of Oral Everolimus) trial demonstrated significantly improved progression-free survival with the use of everolimus plus exemestane (EVE + EXE) versus placebo plus exemestane (PBO + EXE) in patients with advanced breast cancer who developed disease progression after treatment with nonsteroidal aromatase inhibitors. This analysis investigated the treatment effects on health-related quality of life (HRQOL). Using the European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire-Core 30 (EORTC QLQ-C30) questionnaire, HRQOL was assessed at baseline and every 6 weeks thereafter until disease progression and/or treatment discontinuation. The 30 items in 15 subscales of the QLQ-C30 include global health status wherein higher scores (range, 0-100) indicate better HRQOL. This analysis included a protocol-specified time to definitive deterioration (TDD) analysis at a 5% decrease in HRQOL versus baseline, with no subsequent increase above this threshold. The authors report additional sensitivity analyses using 10-point minimal important difference decreases in the global health status score versus baseline. Treatment arms were compared using the stratified log-rank test and Cox proportional hazards model adjusted for trial stratum (visceral metastases, previous hormone sensitivity), age, sex, race, baseline global health status score and Eastern Cooperative Oncology Group performance status, prognostic risk factors, and treatment history. Baseline global health status scores were found to be similar between treatment groups (64.7 vs 65.3). The median TDD in HRQOL was 8.3 months with EVE + EXE versus 5.8 months with PBO + EXE (hazard ratio, 0.74; P = .0084). At the 10-point minimal important difference, the median TDD with EVE + EXE was 11.7 months versus 8.4 months with PBO + EXE (hazard ratio, 0.80; P = .1017). In patients with advanced breast cancer who develop disease progression after treatment with nonsteroidal aromatase inhibitors, EVE + EXE was associated with a longer TDD in global HRQOL versus PBO + EXE.


Characterization and expression analysis of genes directing galactomannan synthesis in coffee.
Galactomannans act as storage reserves for the seeds in some plants, such as guar (Cyamopsis tetragonoloba) and coffee (Coffea arabica and Coffea canephora). In coffee, the galactomannans can represent up to 25 % of the mass of the mature green coffee grain, and they exert a significant influence on the production of different types of coffee products. The objective of the current work was to isolate and characterize cDNA encoding proteins responsible for galactomannan synthesis in coffee and to study the expression of the corresponding transcripts in the developing coffee grain from C. arabica and C. canephora, which potentially exhibit slight galactomannan variations. Comparative gene expression analysis was also carried out for several other tissues of C. arabica and C. canephora. cDNA banks, RACE-PCR and genome walking were used to generate full-length cDNA for two putative coffee mannan synthases (ManS) and two galactomannan galactosyl transferases (GMGT). Gene-specific probe-primer sets were then generated and used to carry out comparative expression analysis of the corresponding genes in different coffee tissues using quantitative RT-PCR. Two of the putative galactomannan biosynthetic genes, ManS1 and GMGT1, were demonstrated to have very high expression in the developing coffee grain of both Coffea species during endosperm development, consistent with our proposal that these two genes are responsible for the production of the majority of the galactomannans found in the grain. In contrast, the expression data presented indicates that the ManS2 gene product is probably involved in the synthesis of the galactomannans found in green tissue. The identification of genes implicated in galactomannan synthesis in coffee are presented. The data obtained will enable more detailed studies on the biosynthesis of this important component of coffee grain and contribute to a better understanding of some functional differences between grain from C. arabica and C. canephora.


Derivation and external validation of a novel risk score for prediction of 30-day mortality after percutaneous coronary intervention.
Early mortality after percutaneous coronary intervention (PCI) is relatively rare. Current risk prediction models for this event are outdated. We sought to derive a 30-day mortality risk score after PCI. The score was derived from a pooled database of 21 randomised clinical trials using a logistic regression model incorporating clinical and angiographic variables. The score was validated in a separate unrestricted study population, the Assessment of Dual AntiPlatelet Therapy With Drug Eluting Stents (ADAPT-DES) registry. Of 32,882 eligible patients, 75% had data for all 19 variables used for score derivation. The independent predictors of 30-day mortality were age, presentation with ACS, diabetes mellitus, use of first-generation drug-eluting stents, left main or left anterior descending artery lesion, prior myocardial infarction (MI), and suboptimal flow in the artery before or after PCI. The median [interquartile range] score in the derivation cohort was 5 [3, 6] and overall mortality was 0.49%, ranging from 0.08% to 1.64% with scores of 0-16. The 30-day mortality rate was approximately tenfold higher in patients with a score at or above versus below the median of 5 (0.86% versus 0.08%, p<0.0001). Discrimination in both cohorts was very good (C statistic=0.848 and 0.828, respectively), and calibration was satisfactory. A novel risk score incorporating eight readily available clinical and angiographic variables had high discrimination for 30-day death after PCI across a wide range of clinical scenarios.


A segment interaction analysis of proximal-to-distal sequential segment motion patterns.
The purpose of this study was to examine the motion-dependent interaction between adjacent lower extremity segments during the actions of kicking and the swing phases of running and walking. This was done to help explain the proximal-to-distal sequential pattern of segment motions typically observed in these activities and to evaluate general biomechanical principles used to explain this motion pattern. High speed film data were collected for four subjects performing each skill. Equations were derived which expressed the interaction between segments in terms of resultant joint moments at the hip and knee and several interactive moments which were functions of gravitational forces or kinematic variables. The angular motion-dependent interaction between the thigh and leg was found to play a significant role in determining the sequential segment motion patterns observed in all three activities. The general nature of this interaction was consistent across all three movements except during phases in which there were large differences in the knee angle. Support was found for the principle of summation of segment speeds, whereas no support was found for the principle of summation of force or for general statements concerning the effect of negative thigh acceleration on positive leg acceleration. The roles played by resultant joint moments in producing the observed segment motion sequences are discussed.


Landscape rules predict optimal superhighways for the first peopling of Sahul.
Archaeological data and demographic modelling suggest that the peopling of Sahul required substantial populations, occurred rapidly within a few thousand years and encompassed environments ranging from hyper-arid deserts to temperate uplands and tropical rainforests. How this migration occurred and how humans responded to the physical environments they encountered have, however, remained largely speculative. By constructing a high-resolution digital elevation model for Sahul and coupling it with fine-scale viewshed analysis of landscape prominence, least-cost pedestrian travel modelling and high-performance computing, we create over 125 billion potential migratory pathways, whereby the most parsimonious routes traversed emerge. Our analysis revealed several major pathways-superhighways-transecting the continent, that we evaluated using archaeological data. These results suggest that the earliest Australian ancestors adopted a set of fundamental rules shaped by physiological capacity, attraction to visually prominent landscape features and freshwater distribution to maximize survival, even without previous experience of the landscapes they encountered.


The electron microscopy of the choroid plexus.
1. The choroid plexus of the rat has been studied in detail by electron microscopy. Samples from the frog, rabbit, and cat have also been examined without noting significant differences. 2. The surface of the ependymal epithelium is covered by pedicels of variable size. There is reason for thinking of these structures as labile. They may actually pinch off and contribute to the secretory product. In any case, the surface area is vastly increased by their presence. Polypoid border seems an apt term to apply to this type of surface. 3. There is also a great expansion of the basal surface of ependymal cells. In the vicinity of cell junctions this surface is deeply infolded, and continuous with elaborate interdigitations of the lateral intercellular surfaces. Analogous infolding of the basal cell surface is known to exist in other epithelia also noted for their water transport (kidney tubules, salivary gland, and ciliary body). 4. Pretreatment of rats with diamox, an agent known to block cerebro-spinal fluid production, did not produce an important morphological change in the features of the ependyma, or any other part of the choroid plexus. 5. Capillaries of the choroid plexus have a very attenuated endothelium. This is seen to be fenestrated. It is thought this probably represents the condition in life, and is not simply a fixation artefact. 6. Pial cells tend to interpose sheets of cytoplasm between the capillaries and ependyma. The sheets are not continuous, however, and so would not constitute a serious diffusion barrier. These cells belong to the reticuloendothelial system, and undergo shape changes, and probably increase in number, when the system is stimulated by the repeated injection of trypan blue.


Low-frequency modes of aqueous alkali halide solutions: an ultrafast optical Kerr effect study.
A detailed picture of aqueous solvation of ions is central to the understanding of diverse phenomena in chemistry and biology. In this work, we report polarization resolved THz time domain measurements of the Raman spectral density of a wide range of aqueous salt solutions. In particular, the isotropic Raman spectral density reveals the frequency of the hydrogen bond formed between the halide ion and water. The frequency of this mode is measured for the series Cl(-), Br(-), and I(-) as a function of concentration, cation size, and charge. The frequencies extrapolated to zero concentration permit an estimation of the force constant of the mode, which is found to decrease with increasing halide mass and to be similar to the force constant associated with the water-water hydrogen bond. This result is consistent with recent calculations. The extrapolation of the frequency of the chloride hydrogen bond to zero concentration reveals a dependence of the frequency on the nature of the cation. This is ascribed to an interaction between the solvated anion and cation even at the lowest concentration studied here (<0.15 M). It is suggested that this behavior reflects the influence of the electric field of the cation on the hydrogen bond of an adjacent anion. Such interactions should be taken into account when modeling experimental data recorded at concentrations of ions in excess of 0.1 M. These measurements of the isotropic Raman spectral density are compared with those for the anisotropic response, which reflects the frequencies of the full range of hydrogen bonds in aqueous salt solutions. The anisotropic spectral density recovered can be modeled in terms of a concentration-dependent population of water-water H-bonds with a frequency unaffected by the ions, the halide-water hydrogen bonds, and a low-frequency collision-induced contribution.


Help-seeking for mental health issues in deployed Canadian Armed Forces personnel at risk for moral injury.
Objective: Potentially morally injurious experiences (PMIE) (events that transgress an individual's subjective moral standards) have been associated with psychologically distressing moral emotions such as shame and guilt. Military leaders and clinicians have feared that those with PMIEs may be less likely to seek help due to the withdrawing nature of shame/guilt; however, to date, help-seeking patterns of military personnel with PMIEs has not been explored. Our objective is to address this research gap. Method: Data from a nationally-representative mental health survey of active Canadian military personnel were analysed. To assess the association between exposure to three PMIEs and past-year help-seeking across different provider categories (i.e. professionals, para-professionals (those delegated with mental health advisory tasks but are not licenced to practice as medical professionals), non-professionals), a series of logistic regressions were conducted, controlling for exposure to other deployment and non-deployment-related psychological trauma, psychiatric variables, military factors, and sociodemographic variables. Analytical data frame included only personnel with a history of Afghanistan deployment (N = 4854). Results: Deployed members exposed to PMIEs were more likely to seek help from their family doctor/general practitioner (OR = 1.72; 95%CI = 1.25-2.36), paraprofessionals (OR = 1.72; 95%CI = 1.25-2.36), and non-professionals (OR = 1.44; 95%CI = 1.06-1.95) in comparison to members not exposed to PMIEs. Those exposed to PMIEs were also more likely to seek professional care from the civilian health care system (OR = 1.94; 95%CI = 1.27-2.96). Conclusion: Contrary to long-held, but untested, assumptions regarding the impact of PMIEs on help-seeking, we found those with PMIEs are more likely to seek help from gatekeeper professionals (i.e. general practitioners), para-professionals, and non-professionals rather than specialized mental health professionals (e.g. psychologists). Increased utilization of civilian professionals raises concerns that active military members may be avoiding military health services. Clinically, this highlights the need to increase awareness of moral injury to ensure that actively serving military members are provided with appropriate advice and treatment.


Statistics teaching in medical school: opinions of practising doctors.
The General Medical Council expects UK medical graduates to gain some statistical knowledge during their undergraduate education; but provides no specific guidance as to amount, content or teaching method. Published work on statistics teaching for medical undergraduates has been dominated by medical statisticians, with little input from the doctors who will actually be using this knowledge and these skills after graduation. Furthermore, doctor's statistical training needs may have changed due to advances in information technology and the increasing importance of evidence-based medicine. Thus there exists a need to investigate the views of practising medical doctors as to the statistical training required for undergraduate medical students, based on their own use of these skills in daily practice. A questionnaire was designed to investigate doctors' views about undergraduate training in statistics and the need for these skills in daily practice, with a view to informing future teaching. The questionnaire was emailed to all clinicians with a link to the University of East Anglia Medical School. Open ended questions were included to elicit doctors' opinions about both their own undergraduate training in statistics and recommendations for the training of current medical students. Content analysis was performed by two of the authors to systematically categorize and describe all the responses provided by participants. 130 doctors responded, including both hospital consultants and general practitioners. The findings indicated that most had not recognised the value of their undergraduate teaching in statistics and probability at the time, but had subsequently found the skills relevant to their career. Suggestions for improving undergraduate teaching in these areas included referring to actual research and ensuring relevance to, and integration with, clinical practice. Grounding the teaching of statistics in the context of real research studies and including examples of typical clinical work may better prepare medical students for their subsequent career.


Developing a co-culture system for effective megakaryo/thrombopoiesis from umbilical cord blood hematopoietic stem/progenitor cells.
Platelet transfusion can be a life-saving procedure in different medical settings. Thus, there is an increasing demand for platelets, of which shelf-life is only 5 days. The efficient ex vivo biomanufacturing of platelets would allow overcoming the shortages of donated platelets. We exploited a two-stage culture protocol aiming to study the effect of different parameters on the megakaryo/thrombopoiesis ex vivo. In the expansion stage, human umbilical cord blood (UCB)-derived CD34(+)-enriched cells were expanded in co-culture with human bone marrow mesenchymal stromal cells (BM-MSCs). The megakaryocytic commitment and platelet generation were studied, considering the impact of exogenous addition of thrombopoietin (TPO) in the expansion stage and a cytokine cocktail (Cyt) including TPO and interleukin-3 in the differentiation stage, with the use of different culture medium formulations, and in the presence/absence of BM-MSCs (direct versus non-direct cell-cell contact). Our results suggest that an early megakaryocytic commitment, driven by TPO addition during the expansion stage, further enhanced megakaryopoiesis. Importantly, the results suggest that co-culture with BM-MSCs under serum-free conditions combined with Cyt addition, in the differentiation stage, significantly improved the efficiency yield of megakaryo/thrombopoiesis as well as increasing %CD41, %CD42b and polyploid content; in particular, direct contact of expanded cells with BM-MSCs, in the differentiation stage, enhanced the efficiency yield of megakaryo/thrombopoiesis, despite inhibiting their maturation. The present study established an in vitro model for the hematopoietic niche that combines different biological factors, namely, the presence of stromal/accessory cells and biochemical cues, which mimics the BM niche and enhances an efficient megakaryo/thrombopoiesis process ex vivo.


Non-histone chromosomal proteins. Evidence for their role in mediating the binding of histones to deoxyribonucleic acid during the cell cycle.
By selective dissociation of histones with the ionic detergent sodium deoxycholate, we have demonstrated that these basic chromosomal polypeptides, which are effective inhibitors of transcription, are more tenaciously bound to DNA in mitotic than in S-phase chromatin. Evidence is presented which suggests that cell-cycle-stage-specific non-histone chromosomal proteins can account for such variations in the association of histones with DNA. When chromatin is reconstituted with DNA and histones are pooled from S-phase and mitotic cells and either S-phase or mitotic non-histone chromosomal proteins, a preferential extraction of histones with sodium deoxycholate from chromatin reconstituted with S-phase rather than mitotic non-histone chromosomal proteins is observed. In contrast, the extractability of histones with sodium deoxycholate from nucleohistone complexes reconstituted with DNA pooled from S-phase and mitotic cells and either S-phase or mitotic histones is identical. Since non-histone chromosomal proteins rather than histones are responsible for the differences in chromatin template activity during S-phase and mitosis, we propose that non-histone chromosomal proteins may modify gene expression during the cell cycle by mediating the binding of histones to DNA.


Investigation of the effect of the efficiency of noise at different intensities on the DNA of the newborns.
Hearing loss can occur in newborns exposed to high-level noise; noise exposure can cause more physiological stress and can lead to DNA damage. This study was designed to determine DNA damage in newborn rats exposed to sound at different concentrations. For this purpose, 28 newborn (3-6 days old) rats were divided into four groups of 7 rats in each group (Control and Groups of 40 decibel (dB), 70 dB, and 110 dB]. In the experimental groups, 40 dB, 70 dB, and 110 dB (7.5-15 kHz) of sound was applied to the experimental groups for 30 min a day for 7 days. DNA damage levels in the serums obtained from this study were determined by the enzyme-linked immunosorbent assay (ELISA) method. According to this, it was determined that DNA damage in the group exposed to 110 dB showed a statistically significant increase (P < 0.05) compared to the compared to the control, 40 dB, and 70 dB groups. Related to the subject, it was concluded that DNA damage may occur in newborns exposed to 110 dB or higher sound in neonatal units, wards, and home environments with newborn babies. Mothers should be warned about this situation and noise should be kept under 110 dB volume in the environments with the newborns.


Novel cell-based method to detect Shiga toxin 2 from Escherichia coli O157:H7 and inhibitors of toxin activity.
Escherichia coli O157:H7 is a leading cause of food-borne illness. This human pathogen produces Shiga toxins (Stx1 and Stx2) which inhibit protein synthesis by inactivating ribosome function. The present study describes a novel cell-based assay to detect Stx2 and inhibitors of toxin activity. A Vero cell line harboring a destabilized variant (half-life, 2 h) of the enhanced green fluorescent protein (d2EGFP) was used to monitor the toxin-induced inhibition of protein synthesis. This Vero-d2EGFP cell line produced a fluorescent signal which could be detected by microscopy or with a plate reader. However, a greatly attenuated fluorescent signal was detected in Vero-d2EGFP cells that had been incubated overnight with either purified Stx2 or a cell-free culture supernatant from Stx1- and Stx2-producing E. coli O157:H7. Dose-response curves demonstrated that the Stx2-induced inhibition of enhanced green fluorescent protein fluorescence mirrored the Stx2-induced inhibition of overall protein synthesis and identified a picogram-per-milliliter threshold for toxin detection. To establish our Vero-d2EGFP assay as a useful tool for the identification of toxin inhibitors, we screened a panel of plant compounds for antitoxin activities. Fluorescent signals were maintained when Vero-d2EGFP cells were exposed to Stx1- and Stx2-containing medium in the presence of either grape seed or grape pomace extract. The antitoxin properties of the grape extracts were confirmed with an independent toxicity assay that monitored the overall level of protein synthesis in cells treated with purified Stx2. These results indicate that the Vero-d2EGFP fluorescence assay is an accurate and sensitive method to detect Stx2 activity and can be utilized to identify toxin inhibitors.


High dietary vitamin A (retinyl palmitate) and cellular immune functions in mice.
High dietary intakes (4000-650,000 IU/kg diet) of vitamin A (retinyl palmitate, RP) modified the functions of peritoneal macrophages (PM). The number of peritoneal exudated cells (PEC) obtained from CD-1 mice increased significantly at both 7 and 10 weeks after initiation of the RP diets. The percentage of PM in PEC showed no significant difference between dietary groups and was at levels of 55-60%. PM from mice fed high RP diets showed higher tumoricidal activities than PM from controls without any preincubation with macrophage activators. Enhancement of in vitro tumoricidal activity of PM increased with increasing contents of RP in the diets, reaching 30% lysis by PM isolated from mice fed the highest RP (650,000 IU/kg diet) diet. However, the in vitro activation of tumoricidal ability of PM by macrophage-activating factor (MAF) was inversely correlated with the dietary RP content. The tumoricidal activities of PM from mice fed the highest RP diet were not enhanced by MAF. However, these PM showed an increased ability to phagocytose SRBC and opsonized SRBC compared to controls. Splenocytes and thymocytes were incubated with [3H]thymidine immediately after isolation and their mitogenic activities were measured. Splenocytes, but not thymocytes, isolated from mice fed the highest RP diet had increased mitogenesis. On the other hand, NK activity was not affected by dietary RP intake. There was a similar lysis of target cells by both splenocytes and thymocytes from mice fed diets with various RP levels. IL-1 was produced from PM by incubation with LPS, and its production was assessed using the proliferation of normal mice thymocytes. Production of IL-1 in vitro showed about a two-fold increase using cells from mice fed the highest RP diet compared to controls. High RP diets induced increased phagocytic ability and tumoricidal activity of PM but did not enhance NK activity. These findings suggest that high RP diet may cause activation of PM.


The Optimisation of Pseudotyped Viruses for the Characterisation of Immune Responses to Equine Influenza Virus.
Pseudotyped viruses (PVs) produced by co-transfecting cells with plasmids expressing lentiviral core proteins and viral envelope proteins are potentially powerful tools for studying various aspects of equine influenza virus (EIV) biology. The aim of this study was to optimise production of equine influenza PVs. Co-transfection of the HAT protease to activate the haemagglutinin (HA) yielded a higher titre PV than TMPRSS2 with the HA from A/equine/Richmond/1/2007 (H3N8), whereas for A/equine/Newmarket/79 (H3N8), both proteases resulted in equivalent titres. TMPRSS4 was ineffective with the HA of either strain. There was also an inverse relationship between the amount of protease-expression plasmids and the PV titre obtained. Interestingly, the PV titre obtained by co-transfection of a plasmid encoding the cognate N8 NA was not as high as that generated by the addition of exogenous neuraminidase (NA) from Clostridium perfringens to allow the release of nascent PV particles. Finally, initial characterisation of the reliability of PV neutralisation tests (PVNTs) demonstrated good intra-laboratory repeatability. In conclusion, we have demonstrated that equine influenza PV production can be readily optimised to provide a flexible tool for studying EIV.


MRI of chondromyxoid fibroma.
Chondromyxoid fibroma is a rare benign primary bone tumor of cartilage. Despite a characteristic radiographic appearance, chondromyxoid fibroma with atypical radiographic findings may mimic more common tumors. To describe the MR findings of chondromyxoid fibroma. MR images of 19 histopathologically confirmed chondromyxoid fibromas were retrospectively analyzed for signal intensity, periosteal reaction, adjacent abnormal bone marrow and soft tissue signal, and patterns of contrast enhancement. All cases of chondromyxoid fibroma showed hypointense to intermediate signal intensity and internal hyperintense foci were observed in seven (37%) cases on T1-weighted images. On T2-weighted images, all lesions were hyperintense: peripheral intermediate signal band with central hyperintense signal in 11 (58%) of 19 lesions, whereas diffusely hyperintense with heterogeneous pattern in eight (42%). Periosteal reaction was observed in 11 (58%) of 19 cases. Adjacent abnormal bone marrow or soft tissue signal was observed in 12 (63%) or 14 (74%) of 19 cases, respectively. On contrast-enhanced T1-weighted images, peripheral nodular enhancement was observed in 69% (11/16) and diffuse contrast enhancement was observed in 31% (5/16) with homogeneous (n = 3) or heterogeneous (n = 2) patterns. Among the cases with peripheral nodular enhancement, the peripheral nodular enhancing portion generally corresponded to the peripheral intermediate signal band on T2-weighted images, although the peripheral enhancement was not as wide as a band of intermediate signal intensity. On the other hand, the central non-enhancing portion generally corresponded to the central hyperintense signal intensity on T2-weighted images. The helpful features of chondromyxoid fibroma are the peripheral intermediate signal band and central hyperintense signal on T2-weighted images, generally corresponding to the peripheral nodular enhancement and central non-enhancing portion on contrast-enhanced T1-weighted images, respectively.


Physical activity, energy expenditure and their correlates in two cohorts of Swedish subjects between adolescence and early adulthood.
To assess physical activity and energy expenditure and to identify their correlates during adolescence and early adulthood. In a cohort study, total energy expenditure (TEE) and physical activity level (PAL) were assessed at 15 and 21 y from a 7-day activity diary and predicted BMR. The influences on TEE and PAL of body size, gender, sexual maturity, physical activity habits, sitting time, region, social conditions, employment, food habits, smoking and alcohol intake were examined in multivariate analyses. 71 male and 89 female subjects, living in two socioeconomically different regions of Sweden, a university region and an industrial region. The university city of Uppsala and the industrial town of Trollhättan. At both 15 and 21 y, PAL and TEE were high, with gender, sitting time and physical activity habits as main correlates. At age 21 y, employment and the mother's educational level also appeared as significant correlates. The relations between the main variables and their correlates were more complex than at age 15 y, and the sitting time was reduced and the activity habits were changed. A reduction of daily sitting appears to be a major reason why high levels of physical activity and energy expenditure were maintained from 15 to 21 y of age in spite of changed and less frequent activity habits during this interval.


Testing association of rare genetic variants with resistance to three common antiseizure medications.
Drug resistance is a major concern in the treatment of individuals with epilepsy. No genetic markers for resistance to individual antiseizure medication (ASM) have yet been identified. We aimed to identify the role of rare genetic variants in drug resistance for three common ASMs: levetiracetam (LEV), lamotrigine (LTG), and valproic acid (VPA). A cohort of 1622 individuals of European descent with epilepsy was deeply phenotyped and underwent whole exome sequencing (WES), comprising 575 taking LEV, 826 LTG, and 782 VPA. We performed gene- and gene set-based collapsing analyses comparing responders and nonresponders to the three drugs to determine the burden of different categories of rare genetic variants. We observed a marginally significant enrichment of rare missense, truncating, and splice region variants in individuals who were resistant to VPA compared to VPA responders for genes involved in VPA pharmacokinetics. We also found a borderline significant enrichment of truncating and splice region variants in the synaptic vesicle glycoprotein (SV2) gene family in nonresponders compared to responders to LEV. We did not see any significant enrichment using a gene-based approach. In our pharmacogenetic study, we identified a slightly increased burden of damaging variants in gene groups related to drug kinetics or targeting in individuals presenting with drug resistance to VPA or LEV. Such variants could thus determine a genetic contribution to drug resistance.


Tau pathology in frontotemporal lobar degeneration with C9ORF72 hexanucleotide repeat expansion.
An expanded GGGGCC hexanucleotide repeat in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal lobar degeneration associated with TDP-43 pathology (FTLD-TDP). In addition to TDP-43-positive neuronal and glial inclusions, C9ORF72-linked FTLD-TDP has characteristic TDP-43-negative neuronal cytoplasmic and intranuclear inclusions as well as dystrophic neurites in the hippocampus and cerebellum. These lesions are immunopositive for ubiquitin and ubiquitin-binding proteins, such as sequestosome-1/p62 and ubiquilin-2. Studies examining the frequency of the C9ORF72 mutation in clinically probable Alzheimer's disease (AD) have found a small proportion of AD cases with the mutation. This prompted us to systematically explore the frequency of Alzheimer-type pathology in a series of 17 FTLD-TDP cases with mutations in C9ORF72 (FTLD-C9ORF72). We identified four cases with sufficient Alzheimer-type pathology to meet criteria for intermediate-to-high-likelihood AD. We compared AD pathology in the 17 FTLD-C9ORF72 to 13 cases of FTLD-TDP linked to mutations in the gene for progranulin (FTLD-GRN) and 36 cases of sporadic FTLD (sFTLD). FTLD-C9ORF72 cases had higher Braak neurofibrillary tangle stage than FTLD-GRN. Increased tau pathology in FTLD-C9ORF72 was assessed with thioflavin-S fluorescent microscopy-based neurofibrillary tangle counts and with image analysis of tau burden in temporal cortex and hippocampus. FTLD-C9ORF72 had significantly more neurofibrillary tangles and higher tau burden compared with FTLD-GRN. The differences were most marked in limbic regions. On the other hand, sFTLD and FTLD-C9ORF72 had a similar burden of tau pathology. These results suggest FTLD-C9ORF72 has increased propensity for tau pathology compared to FTLD-GRN, but not sFTLD. The accumulation of tau as well as lesions immunoreactive for ubiquitin and ubiquitin-binding proteins (p62 and ubiquilin-2) suggests that mutations in C9ORF72 may involve disrupted protein degradation that favors accumulation of multiple different proteins.