Histone deacetylase inhibition alters dendritic cells to assume a tolerogenic phenotype and ameliorates arthritis in SKG mice.
The purpose of this study was to elucidate the effects of histone deacetylase inhibition on the phenotype and function of dendritic cells and on arthritis in SKG mice. Arthritis was induced in SKG mice by zymosan A injection. Trichostatin A, a histone deacetylase inhibitor, was administered and its effects on arthritis were evaluated by joint swelling and histological evaluation. Interleukin-17 production in lymph node cells was determined by an enzyme-linked immunosorbent assay (ELISA). Foxp3 expression in lymph node cells and the phenotypes of splenic dendritic cells were examined by fluorescence-activated cell sorting (FACS). Bone marrow-derived dendritic cells (BM-DC) were generated with granulocyte macrophage colony-stimulating factor. The effects of trichostatin A on cell surface molecules, cytokine production, indoleamine 2,3-dioxygenase (IDO) expression and T cell stimulatory capacity were examined by FACS, ELISA, quantitative real-time polymerase chain reaction and Western blot, and the allo-mixed lymphocyte reaction, respectively. Trichostatin A, when administered before the onset of arthritis, prevented SKG mice from getting arthritis. Trichostatin A treatment also showed therapeutic effects on arthritis in SKG mice, when it was administered after the onset of arthritis. Trichostatin A treatment reduced Th17 cells and induced regulatory T cells in lymph node, and also decreased co-stimulatory molecule expression on splenic dendritic cells in vivo. In vitro, trichostatin A markedly suppressed zymosan A-induced interleukin-12 and interleukin-6 production by BM-DC and up-regulated IDO expression at mRNA and protein levels. Trichostatin A-treated BM-DC also showed less T cell stimulatory capacity. Histone deacetylase inhibition changes dendritic cells to a tolerogenic phenotype and ameliorates arthritis in SKG mice.


Motor neuron disease and frontotemporal dementia: One, two, or three diseases?
The relationship between motor neurone disease (MND) and frontotemporal dementia (FTD) has been a topic of scientific exploration for over hundred years. A connection between both diseases was first postulated in 1932 and has been strengthened by a steady stream of case reports since then. By the late 20th century, the link between both diseases was firmly established, with the resulting condition often referred to as MND/FTD. Several strands of evidence support the notion of an MND/FTD overlap. First, a small but well-documented group of patients present with a full-blown FTD, associated with MND. Second, subtle but characteristic changes in frontal-executive functions and social cognition have been described in non-demented MND patients, often in association with frontal atrophy/hypoactivity on neuroimaging. Third, amyotrophic features have been documented in patients primarily diagnosed with FTD. Moreover, the same genetic defect can lead to FTD and MND phenotypes in different members of the same family. However, as the current research is moving toward a more fine-grained evaluation, an increasingly complex picture begins to emerge. Some features, such as psychotic symptoms or severe language deficits (particularly in comprehension and verb processing), seem to occur more often in MND/dementia than in the classical FTD. On the basis of the review of 100 years of literature as well as 10 years of clinical experience of longitudinal follow-up of MND/dementia patients, this review argues in favor of MND/dementia (or, more precisely, MND/dementia/aphasia) as a separate clinical entity, not sufficiently explained by a combination of MND and FTD.


Impact of aromatase genetic variation on hormone levels and global outcome after severe TBI.
Although experimental traumatic brain injury (TBI) studies support estradiol as a neuroprotectant and potent stimulator of neuroplasticity, clinical studies suggest a negative association between endogenous estradiol profiles and mortality/poor outcomes. However, no studies have evaluated associations with cerebral spinal fluid (CSF) hormone profiles and aromatase gene (cytochrome P450 [CYP]19A1) variability on clinical TBI outcomes. We evaluated 110 adults with severe TBI. Average and daily estradiol, testosterone, and estradiol/testosterone ratios (E2:T) were measured using CSF and serum samples and compared to healthy controls. Eighteen tagging and four functional single-nucleotide polymorphisms (SNPs) for CYP19A1 were genotyped and compared to hormones, acute mortality, and Glasgow Outcome Scale (GOS) scores 6 months post-TBI. TBI subjects had lower CSF estradiol over time versus controls. CSF testosterone was initially high, but declined over time. E2/T ratios were initially low, compared to controls, but rose over time. Higher mean E2/T ratio in bivariate analysis was associated with lower mortality (p=0.019) and better GOS-6 scores (p=0.030). rs2470152 influenced CSF E2/T ratio and also serum and CSF testosterone (p≤0.05 all comparisons). Multiple-risk SNPs rs2470152, rs4646, and rs2470144 were associated with worse GOS-6 scores (p≤0.05, all comparisons), and those with>1 risk SNP variant had a higher risk for poor outcome, compared with those with ≤1 risk variant. TBI results in low CSF estradiol and dynamic CSF testosterone and E2/T ratio. In contrast to clinical serum hormone studies, higher CSF E2/T ratio was associated with better outcome. Further, genetic variation in CYP19A1 influences both hormone dynamics and outcome post-TBI.


Influence of acute coffee consumption on postprandial oxidative stress.
Coffee has been reported to be rich in antioxidants, with both acute and chronic consumption leading to enhanced blood antioxidant capacity. High-fat feeding is known to result in excess production of reactive oxygen and nitrogen species, promoting a condition of postprandial oxidative stress. We tested the hypothesis that coffee intake following a high-fat meal would attenuate the typical increase in blood oxidative stress during the acute postprandial period. On 3 different occasions, 16 men and women consumed a high-fat milk shake followed by either 16 ounces of caffeinated or decaffeinated coffee or bottled water. Blood samples were collected before and at 2 and 4 hours following intake of the milk shake and analyzed for triglycerides (TAG), malondialdehyde (MDA), hydrogen peroxide (H2O2), and Trolox equivalent antioxidant capacity (TEAC). Values for TAG and MDA (P < 0.001), as well as for H2O2 (P < 0.001), increased significantly following milk shake consumption, with values higher at 4 hours compared with 2 hours post consumption for TAG and H2O2 (P < 0.05). TEAC was unaffected by the milk shake consumption. Coffee had no impact on TAG, MDA, H2O2, or TEAC, with no condition or interaction effects noted for any variable (P > 0.05). Acute coffee consumption following a high-fat milk shake has no impact on postprandial oxidative stress.


A spatio-temporal model of Notch signalling in the zebrafish segmentation clock: conditions for synchronised oscillatory dynamics.
In the vertebrate embryo, tissue blocks called somites are laid down in head-to-tail succession, a process known as somitogenesis. Research into somitogenesis has been both experimental and mathematical. For zebrafish, there is experimental evidence for oscillatory gene expression in cells in the presomitic mesoderm (PSM) as well as evidence that Notch signalling synchronises the oscillations in neighbouring PSM cells. A biological mechanism has previously been proposed to explain these phenomena. Here we have converted this mechanism into a mathematical model of partial differential equations in which the nuclear and cytoplasmic diffusion of protein and mRNA molecules is explicitly considered. By performing simulations, we have found ranges of values for the model parameters (such as diffusion and degradation rates) that yield oscillatory dynamics within PSM cells and that enable Notch signalling to synchronise the oscillations in two touching cells. Our model contains a Hill coefficient that measures the co-operativity between two proteins (Her1, Her7) and three genes (her1, her7, deltaC) which they inhibit. This coefficient appears to be bounded below by the requirement for oscillations in individual cells and bounded above by the requirement for synchronisation. Consistent with experimental data and a previous spatially non-explicit mathematical model, we have found that signalling can increase the average level of Her1 protein. Biological pattern formation would be impossible without a certain robustness to variety in cell shape and size; our results possess such robustness. Our spatially-explicit modelling approach, together with new imaging technologies that can measure intracellular protein diffusion rates, is likely to yield significant new insight into somitogenesis and other biological processes.


Efficacy of a Community Health Worker-Based Intervention in Improving Dietary Habits Among Community-Dwelling Older People: A Controlled, Crossover Trial in Japan.
Background. Community health workers (CHWs), often called "health promotion volunteers" in Japan, are individuals who act as a natural helping resource in the community. Aim. This study tested the efficacy of a CHW-based intervention to improve dietary habits among community-dwelling older people in Japan, using a controlled, crossover design. Method. Seventy-eight people aged 65 to 74 years with poor dietary variety living in four administrative districts in Hikone City (Shiga Prefecture, Japan) were nonrandomly allocated to an immediate-intervened group (IIG; n = 41) or a delayed-intervened group (DIG; n = 37). Participants joined a biweekly, four-session program (120 minutes/session), comprising "CHW drama-style lectures," "group discussion among participants and CHWs," "tasting of dishes," and "take-home practical activities." For the initial 2-month period, the IIG received the intervention and the DIG did not. The groups were crossed over for the subsequent 2-month period. The primary outcome measure was participants' dietary variety score (score range: 0-10). Results. The dietary variety score in the IIG significantly increased in the initial 2-month period compared with the DIG (effect size 1.60 points; 95% confidence interval: 0.75, 2.45). The intervention had a similar effect in the DIG in the subsequent 2-month period. Moreover, an analysis within the IIG showed that the intervention effects persisted for at least 2 months after the intervention. Conclusions. The CHW-based intervention improved dietary habits among older people. Our findings provide evidence that a CHW-based natural helping approach is a possible solution to promote healthy aging in the community.


Bone Tissue Properties Measurement by Reference Point Indentation in Glucocorticoid-Induced Osteoporosis.
Glucocorticoids, widely used in inflammatory disorders, rapidly increase bone fragility and, therefore, fracture risk. However, common bone densitometry measurements are not sensitive enough to detect these changes. Moreover, densitometry only partially recognizes treatment-induced fracture reductions in osteoporosis. Here, we tested whether the reference point indentation technique could detect bone tissue property changes early after glucocorticoid treatment initiation. After initial laboratory and bone density measurements, patients were allocated into groups receiving calcium + vitamin D (Ca+D) supplements or anti-osteoporotic drugs (risedronate, denosumab, teriparatide). Reference point indentation was performed on the cortical bone layer of the tibia by a handheld device measuring bone material strength index (BMSi). Bone mineral density was measured by dual-energy X-ray absorptiometry (DXA). Although Ca+D-treated patients exhibited substantial and significant deterioration, risedronate-treated patients exhibited no significant change, and both denosumab- and teriparatide-treated participants exhibited significantly improved BMSi 7 weeks after initial treatment compared with baseline; these trends remained stable for 20 weeks. In contrast, no densitometry changes were observed during this study period. In conclusion, our study is the first to our knowledge to demonstrate that reference point indentation is sensitive enough to reflect changes in cortical bone indentation after treatment with osteoporosis therapies in patients newly exposed to glucocorticoids.


Stage-by-stage change in DNA methylation status of Dnmt1 locus during mouse early development.
Methylation of DNA is involved in tissue-specific gene control, and establishment of DNA methylation pattern in the genome is thought to be essential for embryonic development. Three isoforms of Dnmt1 (DNA methyltransferase 1) transcripts, Dnmt1s, Dnmt1o, and Dnmt1p, are produced by alternative usage of multiple first exons. Dnmt1s is expressed in somatic cells. Dnmt1p is found only in pachytene spermatocytes, whereas Dnmt1o is specific to oocytes and preimplantation embryos. Here we determined that there is a tissue-dependent differentially methylated region (T-DMR) in the 5' region of Dnmt1o but not in that of the Dnmt1s/1p. The methylation status of the Dnmt1o T-DMR was distinctively different in the oocyte from that in the sperm and adult somatic tissues and changed at each stage from fertilization to blastocyst stage, suggesting that active methylation and demethylation occur during preimplantation development. The T-DMR was highly methylated in somatic cells and embryonic stem cells. Analysis using Dnmt-deficient embryonic stem cell lines revealed that Dnmt1, Dnmt3a, and Dnmt3b are each partially responsible for maintenance of methylation of Dnmt1o T-DMR. In particular, there are compensatory and cooperative roles between Dnmt3a and Dnmt3b. Thus, the regulatory region of Dnmt1o, but not of Dnmt1s/1p, appeared to be a target of DNA methylation. The present study also suggested that the DNA methylation status of the gene region dynamically changes during embryogenesis independently of the change in the bulk DNA methylation status.


Machine Learning Methods for Binary and Multiclass Classification of Melanoma Thickness From Dermoscopic Images.
Thickness of the melanoma is the most important factor associated with survival in patients with melanoma. It is most commonly reported as a measurement of depth given in millimeters (mm) and computed by means of pathological examination after a biopsy of the suspected lesion. In order to avoid the use of an invasive method in the estimation of the thickness of melanoma before surgery, we propose a computational image analysis system from dermoscopic images. The proposed feature extraction is based on the clinical findings that correlate certain characteristics present in dermoscopic images and tumor depth. Two supervised classification schemes are proposed: a binary classification in which melanomas are classified into thin or thick, and a three-class scheme (thin, intermediate, and thick). The performance of several nominal classification methods, including a recent interpretable method combining logistic regression with artificial neural networks (Logistic regression using Initial variables and Product Units, LIPU), is compared. For the three-class problem, a set of ordinal classification methods (considering ordering relation between the three classes) is included. For the binary case, LIPU outperforms all the other methods with an accuracy of 77.6%, while, for the second scheme, although LIPU reports the highest overall accuracy, the ordinal classification methods achieve a better balance between the performances of all classes.


Improved force field for molecular modeling of poly(3-hexylthiophene).
An ab initio-based improved force field is reported for poly(3-hexylthiophene) (P3HT) in the solid state, deriving torsional parameters and partial atomic charges from ab initio molecular structure calculations with explicit treatment of the hexyl side chains. The force field is validated by molecular dynamics (MD) simulations of solid P3HT with different molecular weights including calculation of structural parameters, mass density, melting temperature, glass transition temperature, and surface tension. At 300 K, the P3HT crystalline structure features planar backbones with non-interdigitated all-trans hexyl side chains twisted ~90° from the plane of the backbone. For crystalline P3HT with infinitely long chains, the calculated 300 K mass density (1.05 g cm(-3)), the melting temperature (490 K), and the 300 K surface tension (32 mN/m) are all in agreement with reported experimental values, as is the glass transition temperature (300 K) for amorphous 20-mers.


TAL effectors specificity stems from negative discrimination.
Transcription Activator-Like (TAL) effectors are DNA-binding proteins secreted by phytopathogenic bacteria that interfere with native cellular functions by binding to plant DNA promoters. The key element of their architecture is a domain of tandem-repeats with almost identical sequences. Most of the polymorphism is located at two consecutive amino acids termed Repeat Variable Diresidue (RVD). The discovery of a direct link between the RVD composition and the targeted nucleotide allowed the design of TAL-derived DNA-binding tools with programmable specificities that revolutionized the field of genome engineering. Despite structural data, the molecular origins of this specificity as well as the recognition mechanism have remained unclear. Molecular simulations of the recent crystal structures suggest that most of the protein-DNA binding energy originates from non-specific interactions between the DNA backbone and non-variable residues, while RVDs contributions are negligible. Based on dynamical and energetic considerations we postulate that, while the first RVD residue promotes helix breaks--allowing folding of TAL as a DNA-wrapping super-helix--the second provides specificity through a negative discrimination of matches. Furthermore, we propose a simple pharmacophore-like model for the rationalization of RVD-DNA interactions and the interpretation of experimental findings concerning shared affinities and binding efficiencies. The explanatory paradigm presented herein provides a better comprehension of this elegant architecture and we hope will allow for improved designs of TAL-derived biotechnological tools.


Testing platelet function.
Platelet function tests have been traditionally used to aid in the diagnosis and management of patients with bleeding problems. Given the role of platelets in atherothrombosis, several dedicated platelet function instruments are now available that are simple to use and can be used as point-of-care assays. These can provide rapid assessment of platelet function within whole blood without the requirement of sample processing. Some tests can be used to monitor antiplatelet therapy and assess risk of bleeding and thrombosis, although current guidelines advise against this. This article discusses the potential utility of tests/instruments that are available.


A multisource approach to improving epidemiologic estimates: application to global B-cell malignancies.
The compilation of comprehensive, worldwide epidemiologic data can inform hypotheses on cancer etiology and guide future drug development. These statistics are reported by a multitude of sources using varying methods; thus, compiling a complete database of these statistics is a challenge. To this end, this paper examined the usefulness of a novel, multisource approach-extracting data from the peer-reviewed literature, online reports, and query systems from cancer registries and health agencies and directly contacting cancer registry personnel-for building a comprehensive, multinational epidemiologic cancer database. The major B-cell malignancies were chosen as the cancer subtype to test this approach largely because their epidemiology has not been well characterized in the peer-reviewed literature. We found that a multisource approach yields a more comprehensive epidemiologic database than what would have been possible with the use of literature searches alone. In addition, our paper revealed that cancer registries vary considerably in their methodology, comprehensiveness, and ability to gather information on specific B-cell malignancy subtypes. Collectively, this paper demonstrates the feasibility and value of a multisource approach to gathering epidemiologic data.


Iron reduction as an adjuvant to interferon therapy in patients with chronic hepatitis C who have previously not responded to interferon: a multicenter, prospective, randomized, controlled trial.
Hepatic iron concentration has consistently been observed as being directly correlated with the response to interferon therapy in chronic hepatitis C virus (HCV). We therefore conducted a randomized, controlled trial comparing iron reduction by phlebotomy with iron reduction followed by retreatment with interferon in 96 patients with chronic hepatitis C who had previously not responded to a course of interferon. During the initial phase when all patients were undergoing phlebotomy, we found that serum alanine transaminase (ALT) activities decreased but by less than 50% from baseline in 67 patients (89%), decreased by more than 50% in 12 patients (13%) and became normal in 9 patients (9%) with no overall change in HCV-RNA levels. Subsequently no patient in either treatment group achieved a sustained virologic response. Improvements in necroinflammatory changes were noted in liver biopsy specimens in those patients receiving phlebotomy plus interferon (mean index 8.59 vs. 7.37, P <. 05). A slight but not statistically significant decrease in histologic activity index was noted in those subjects treated by phlebotomy alone (mean index 8.4 vs. 7.75, P not significant). We conclude that, although prior phlebotomy therapy does not improve the rate of sustained response to interferon retreatment, it does result in less liver injury manifested by a decrease in serum transaminase activity and a slight improvement in liver histopathology.


Repetitive TMS of the somatosensory cortex improves writer's cramp and enhances cortical activity.
Since the somatosensory system is believed to be affected in focal dystonia, we focused on the modulation of the primary somatosensory cortex (SI) induced by repetitive transcranial magnetic stimulation (rTMS) in order to improve symptoms of writer's cramp. Patients with writer's cramp (N=9 in the pilot study and N=11 in the advanced study) were treated with 30-minute 1 Hz real- or sham-rTMS of the SI cortex every day for 5 days. Before and after rTMS, 1.5 T fMRI was examined during simple hand movements. While in the pilot study the rTMS coil was navigated over the SI cortex with a maximum of blood oxygenation-level dependent (BOLD) signal induced by passive movement, patients in the advanced study had the coil above the postcentral sulcus. After real-rTMS, 4 pilot study patients and 10 advanced study patients experienced subjective and objective improvement in writing, while only minimal changes were observed after sham-rTMS. Patients involved in the active movement task exhibited a rTMS-induced BOLD signal increase bilaterally in the SI cortex, posterior parietal cortex and in the supplementary motor area (P<0.001 corrected). After sham-rTMS, no BOLD signal changes were observed. In conclusion, 1 Hz rTMS of the SI cortex can improve writer's cramp while increasing the cortical activity in both hemispheres. Handwriting improved in most patients, as well as the subjective benefit, and lasted for 2-3 weeks. The beneficial effects of rTMS paralleled the functional reorganization in the SI cortex and connected areas, reflecting the impact of somatosensory system on active motion control.


Netrin-1 simultaneously suppresses corneal inflammation and neovascularization.
To investigate the effect of netrin-1 on alkali burn-induced corneal inflammation and neovascularization. The expression of netrin-1 and its receptors UNC5A, UNC5B, UNC5C, UNC5D, adenosine 2b receptor (A2BAR), deleted in colorectal cancer (DCC), and neogenin in normal and alkali-burned rat cornea were determined by RT-PCR and/or Western blot analysis, or immunostaining. Topical netrin-1 protein was applied to treat rat corneal alkali-burn injury for 14 consecutive days, started right after the injury or 10 days postinjury. Corneal inflammation and neovascularization were observed under slit lamp microscope. The apoptosis of corneal cells was determined by terminal deoxynucleotidyl transferase-mediated nick end labeling assay. Corneal inflammatory cell infiltration was evaluated by immunostaining of anti-PMN and anti-ED1 antibodies. The expression of epidermal growth factor (EGF), vascular epidermal growth factor (VEGF), and pigment epithelium-derived factor (PEDF) in rat cornea was determined by Western blot analysis. Netrin-1 and its receptor UNC5B were expressed in normal rat corneal epithelium and stromal cells, and their expression decreased after corneal alkali burn. Exogenous netrin-1 administered on rat ocular surfaces resolved alkali burn-induced corneal inflammation, and also suppressed corneal neovascularization. Furthermore, netrin-1 could reverse neovascularization in alkali-burned cornea. The authors found that netrin-1 executed the functions through various mechanisms, including upregulating EGF expression, accelerating epithelial wound healing, inhibiting neutrophil and macrophage infiltration, reducing corneal cell apoptosis, and restoring the equilibrium of VEGF and PEDF in the wounded cornea. Netrin-1 could dampen inflammation, inhibit, and reverse neovascularization in alkali-burned cornea.


Contingency management of toothbrushing behavior in a summer camp for children.
The control of toothbrushing behavior by contingency management was studied with eight, 10 to 12-yr-old boys at a summer camp for children. This behavior occurred infrequently during baseline conditions with and without verbal instructions to the subjects to brush their teeth. Toothbrushing behavior was maintained at a high level when the behavior was required as a prerequisite for the opportunity to swim (contingency management). When, after 22 days, the contingency arrangement between toothbrushing and swimming was terminated, toothbrushing behavior returned to a low level.


NetMHCIIpan-2.0 - Improved pan-specific HLA-DR predictions using a novel concurrent alignment and weight optimization training procedure.
Binding of peptides to Major Histocompatibility class II (MHC-II) molecules play a central role in governing responses of the adaptive immune system. MHC-II molecules sample peptides from the extracellular space allowing the immune system to detect the presence of foreign microbes from this compartment. Predicting which peptides bind to an MHC-II molecule is therefore of pivotal importance for understanding the immune response and its effect on host-pathogen interactions. The experimental cost associated with characterizing the binding motif of an MHC-II molecule is significant and large efforts have therefore been placed in developing accurate computer methods capable of predicting this binding event. Prediction of peptide binding to MHC-II is complicated by the open binding cleft of the MHC-II molecule, allowing binding of peptides extending out of the binding groove. Moreover, the genes encoding the MHC molecules are immensely diverse leading to a large set of different MHC molecules each potentially binding a unique set of peptides. Characterizing each MHC-II molecule using peptide-screening binding assays is hence not a viable option. Here, we present an MHC-II binding prediction algorithm aiming at dealing with these challenges. The method is a pan-specific version of the earlier published allele-specific NN-align algorithm and does not require any pre-alignment of the input data. This allows the method to benefit also from information from alleles covered by limited binding data. The method is evaluated on a large and diverse set of benchmark data, and is shown to significantly out-perform state-of-the-art MHC-II prediction methods. In particular, the method is found to boost the performance for alleles characterized by limited binding data where conventional allele-specific methods tend to achieve poor prediction accuracy. The method thus shows great potential for efficient boosting the accuracy of MHC-II binding prediction, as accurate predictions can be obtained for novel alleles at highly reduced experimental costs. Pan-specific binding predictions can be obtained for all alleles with know protein sequence and the method can benefit by including data in the training from alleles even where only few binders are known. The method and benchmark data are available at http://www.cbs.dtu.dk/services/NetMHCIIpan-2.0.


Growth differences of male and female Göttingen minipigs during ad libitum feeding: a pilot study.
Even though minipigs have been used in biomedical research for nearly half a century now, no specific nutrient requirements are available. For that reason a series of studies into the nutrient requirements of Göttingen minipigs were carried out. Firstly, a pilot study was carried out to determine the ad libitum feed intake (FI) during growth, as a reference for later feed restriction studies. Four male and four female minipigs were fed two types of diet, one standard pig diet (20.6% crude protein; 11.7% crude fat; 13.5 mJ/kg DM metabolizable energy) and one diet specially designed for minipigs (12.0% crude protein; 2.9% crude fat; 11.9 MJ/kg DM metabolizable energy). When fed ad libitum for 13 weeks, female Göttingen minipigs developed a significantly (P<0.05) higher body weight (BW) than males (27.4 vs 16.6 kg) on either diet. The large difference in growth between male and female Göttingen minipigs did not appear to be the result from differences in metabolizable energy intake. Metabolizable energy intake of male and female Göttingen minipigs could be predicted by ME=1877 kJxBW(0.61). Both male and female Göttingen minipigs became obese when fed ad libitum, defined by relative backfat thickness. Relative backfat thickness ranged from 5 to 13 cm/100 kg. Females had thicker relative backfat layers than males. Remarkably, no large changes in haematology and clinical chemistry occurred in ad libitum fed Göttingen minipigs as compared to reference values, and no abnormalities other than enlarged fat reserves were observed at necropsy. Apparently, Göttingen minipigs do not restrain FI voluntarily, and restricted feeding is therefore indicated to prevent obesity.


Safety and efficacy of topical diclofenac sodium gel for knee osteoarthritis in elderly and younger patients: pooled data from three randomized, double-blind, parallel-group, placebo-controlled, multicentre trials.
NSAIDs used for the treatment of osteoarthritis (OA) have dose-related risks for gastrointestinal, cardiovascular and renal adverse events (AEs), particularly in elderly patients. Topical NSAIDs reduce systemic NSAID exposure and may mitigate these risks. To evaluate the safety and efficacy of topical diclofenac sodium 1% gel (DSG) versus vehicle in patients aged 25-64 or ≥65 years who have been diagnosed with knee OA. Pooled data from three 12-week, randomized, double-blind, parallel-group, multicentre trials. US primary care, internal medicine, orthopaedic and rheumatology practices. Aged ≥25 years with mild to moderate (Kellgren-Lawrence grade 1-3) knee OA. After a 1-week analgesic washout, patients applied 4 g of DSG or vehicle four times daily to one knee. Rescue paracetamol (acetaminophen) up to 4 g/day was allowed. Key efficacy outcomes common to the three trials were Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) pain (0-20) and physical function (0-68) subscales, global rating of disease (GRD; 100-mm visual analogue scale [VAS]) and pain on movement (POM; 100-mm VAS). ANOVA was used to compare efficacy outcome differences (DSG vs vehicle) by age (25-64 or ≥65 years). A flare design was used that defined a subset of patients who experienced increased pain during the washout period (modified efficacy subpopulation [MES]). The MES included both patients aged 25-64 (n = 602) and ≥65 (n = 374) years. Patients in each age group applied >90% of scheduled doses. Among patients aged 25-64 years, the improvement from baseline to week 12 (least squares mean [standard error]) was greater for DSG versus vehicle for WOMAC pain (-5.8 [0.3] vs -4.7 [0.3], p = 0.007), WOMAC physical function (-17.9 [0.9] vs -14.2 [0.9], p = 0.002), GRD (-29.5 [1.6] vs -23.8 [1.6], p = 0.01) and POM (-37.3 [1.8] vs -29.0 [1.8], p < 0.001). Among patients aged ≥65 years, the improvements from baseline for most efficacy outcome scores were significantly greater with DSG versus vehicle: WOMAC pain (-5.3 [0.3] vs -4.1 [0.4], p = 0.02), WOMAC physical function (-15.5 [1.1] vs -11.0 [1.1], p = 0.004) and POM (-33.7 [2.2] vs -26.4 [2.2], p = 0.02). The efficacy of DSG did not differ significantly between patients aged 25-64 years and ≥65 years: WOMAC pain (p = 0.85), WOMAC physical function (p = 0.70), GRD (p = 0.86) and POM (p = 0.81). The incidence of any AE was greater with DSG than with vehicle among patients aged 25-64 years (56.6% vs 50.8%) and ≥65 years (55.8% vs 43.9%). Treatment-related application site dermatitis was more common with DSG compared with vehicle in both younger (4.0% vs 0.7%, respectively) and older (5.8% vs 0.4%, respectively) patients and was the main reason for the difference in treatment-related AEs between the DSG and vehicle groups. Gastrointestinal AEs were infrequent among patients treated with DSG and similar to incidence rates with vehicle in both age groups. DSG was effective and generally well tolerated in adults regardless of age. These data support the topical application of DSG for relief of OA knee pain in elderly and younger patients. Clinicaltrials.gov registration numbers NCT00171626, NCT00171678, NCT00426621.


A biotin enrichment strategy identifies novel carbonylated amino acids in proteins from human plasma.
Protein carbonylation is an irreversible protein oxidation correlated with oxidative stress, various diseases and ageing. Here we describe a peptide-centric approach for identification and characterisation of up to 14 different types of carbonylated amino acids in proteins. The modified residues are derivatised with biotin-hydrazide, enriched and characterised by tandem mass spectrometry. The strength of the method lies in an improved elution of biotinylated peptides from monomeric avidin resin using hot water (95°C) and increased sensitivity achieved by reduction of analyte losses during sample preparation and chromatography. For the first time MS/MS data analysis utilising diagnostic biotin fragment ions is used to pinpoint sites of biotin labelling and improve the confidence of carbonyl peptide assignments. We identified a total of 125 carbonylated residues in bovine serum albumin after extensive in vitro metal ion-catalysed oxidation. Furthermore, we assigned 133 carbonylated sites in 36 proteins in native human plasma protein samples. The optimised workflow enabled detection of 10 hitherto undetected types of carbonylated amino acids in proteins: aldehyde and ketone modifications of leucine, valine, alanine, isoleucine, glutamine, lysine and glutamic acid (+14Da), an oxidised form of methionine - aspartate semialdehyde (-32Da) - and decarboxylated glutamic acid and aspartic acid (-30Da). Proteomic tools provide a promising way to decode disease mechanisms at the protein level and help to understand how carbonylation affects protein structure and function. The challenge for future research is to identify the type and nature of oxidised residues to gain a deeper understanding of the mechanism(s) governing carbonylation in cells and organisms and assess their role in disease.


β1 integrin signaling maintains human epithelial progenitor cell survival in situ and controls proliferation, apoptosis and migration of their progeny.
β1 integrin regulates multiple epithelial cell functions by connecting cells with the extracellular matrix (ECM). While β1 integrin-mediated signaling in murine epithelial stem cells is well-studied, its role in human adult epithelial progenitor cells (ePCs) in situ remains to be defined. Using microdissected, organ-cultured human scalp hair follicles (HFs) as a clinically relevant model for studying human ePCs within their natural topobiological habitat, β1 integrin-mediated signaling in ePC biology was explored by β1 integrin siRNA silencing, specific β1 integrin-binding antibodies and pharmacological inhibition of integrin-linked kinase (ILK), a key component of the integrin-induced signaling cascade. β1 integrin knock down reduced keratin 15 (K15) expression as well as the proliferation of outer root sheath keratinocytes (ORSKs). Embedding of HF epithelium into an ECM rich in β1 integrin ligands that mimic the HF mesenchyme significantly enhanced proliferation and migration of ORSKs, while K15 and CD200 gene and protein expression were inhibited. Employing ECM-embedded β1 integrin-activating or -inhibiting antibodies allowed to identify functionally distinct human ePC subpopulations in different compartments of the HF epithelium. The β1 integrin-inhibitory antibody reduced β1 integrin expression in situ and selectively enhanced proliferation of bulge ePCs, while the β1 integrin-stimulating antibody decreased hair matrix keratinocyte apoptosis and enhanced transferrin receptor (CD71) immunoreactivity, a marker of transit amplifying cells, but did not affect bulge ePC proliferation. That the putative ILK inhibitor QLT0267 significantly reduced ORSK migration and proliferation and induced massive ORSK apoptosis suggests a key role for ILK in mediating the ß1 integrin effects. Taken together, these findings demonstrate that ePCs in human HFs require β1 integrin-mediated signaling for survival, adhesion, and migration, and that different human HF ePC subpopulations differ in their response to β1 integrin signaling. These insights may be exploited for cell-based regenerative medicine strategies that employ human HF-derived ePCs.


Lack of peroxisomal catalase affects heat shock response in Caenorhabditis elegans.
Exact mechanisms of heat shock-induced lifespan extension, although documented across species, are still not well understood. Here, we show that fully functional peroxisomes, specifically peroxisomal catalase, are needed for the activation of canonical heat shock response and heat-induced hormesis in Caenorhabditis elegans Although during heat shock, the HSP-70 chaperone is strongly up-regulated in the WT and in the absence of peroxisomal catalase (ctl-2(ua90)II), the small heat shock proteins display modestly increased expression in the mutant. Nuclear foci formation of HSF-1 is reduced in the ctl-2(ua90)II mutant. In addition, heat-induced lifespan extension, observed in the WT, is absent in the ctl-2(ua90)II strain. Activation of the antioxidant response and pentose phosphate pathway are the most prominent changes observed during heat shock in the WT worm but not in the ctl-2(ua90)II mutant. Involvement of peroxisomes in the cell-wide cellular response to transient heat shock reported here gives new insight into the role of organelle communication in the organism's stress response.


Laparoscopic technique for performing duodenal switch with gastric reduction.
The duodenal switch procedure with gastric reduction (DS) is a hybrid procedure for morbid obesity that combines moderate intake restriction with moderate malabsorption. This report describes the laparoscopic hand-assisted technique for the duodenal switch procedure (LapDS). Restriction is achieved via a greater curvature gastrectomy, reducing gastric capacity to 120 ml. The malabsorptive component is constructed by dividing the duodenum 4 cm distal to the pylorus and anastomosing the proximal duodenum to the distal 250 cm of ileum. The biliopancreatic limb is anastomosed to create a 100 cm common channel. Laparoscopic cholecystectomy, cholangiogram, liver biopsy and appendectomy are performed in conjunction with DS. 345 LapDS procedures (27 lap-assisted; 318 hand-assisted) were performed between September 1999 and February 2002. There were 299 women and 46 men with a mean age of 43 years (range 19-67 years). Mean BMI was 50 (range 36-118 kg/m(2)). Mean operating time was 201 minutes (range 105-480). The median length of hospital stay was 3.0 days (range 2-22 days, excluding one outlier). There were 7 conversions to open laparotomy, 14 reoperations, and 21 readmissions. There were 3 pulmonary emboli, 2 deep venous thromboses, and 4 perioperative proximal anastomotic strictures. There were no deaths. Mean percent excess weight loss at 6, 18, and 24 months was 51%, 89%, and 91%, respectively. Laparoscopic assisted duodenal switch procedure can be performed safely with acceptable operative times and without excess morbidity or mortality.


Intermediate honeycomb ordering to trigger oxygen redox chemistry in layered battery electrode.
Sodium-ion batteries are attractive energy storage media owing to the abundance of sodium, but the low capacities of available cathode materials make them impractical. Sodium-excess metal oxides Na2MO3 (M: transition metal) are appealing cathode materials that may realize large capacities through additional oxygen redox reaction. However, the general strategies for enhancing the capacity of Na2MO3 are poorly established. Here using two polymorphs of Na2RuO3, we demonstrate the critical role of honeycomb-type cation ordering in Na2MO3. Ordered Na2RuO3 with honeycomb-ordered [Na(1/3)Ru(2/3)]O2 slabs delivers a capacity of 180 mAh g(-1) (1.3-electron reaction), whereas disordered Na2RuO3 only delivers 135 mAh g(-1) (1.0-electron reaction). We clarify that the large extra capacity of ordered Na2RuO3 is enabled by a spontaneously ordered intermediate Na1RuO3 phase with ilmenite O1 structure, which induces frontier orbital reorganization to trigger the oxygen redox reaction, unveiling a general requisite for the stable oxygen redox reaction in high-capacity Na2MO3 cathodes.


Use of an aggressive MCF-7 cell line variant, TMX2-28, to study cell invasion in breast cancer.
An estrogen receptor-negative variant of the MCF-7 breast cancer cell line, TMX2-28, was used as a model in which to study breast cancer cell invasion. Using a reconstituted basement membrane (Matrigel) assay to evaluate cell invasion, we determined that TMX2-28 cells are more invasive than MCF-7 cells and that the invasiveness of TMX2-28 is similar to that of the aggressive MDA-MB-231 breast cancer cell line. TMX2-28 cells displayed a rounded, epithelial cell-like morphology, suggesting an amoeboid mode of cell invasion, in contrast to the mesenchymal mode of invasion characteristic of spindle-shaped, fibroblast-like MDA-MB-231 cells. Using real-time reverse transcription-PCR, we found that mitogen-inducible gene 2 (MIG2) is expressed at a 17-fold higher level in TMX2-28 cells than in nonaggressive MCF-7 cells and that MIG2 mRNA levels are low in the nontumorigenic human mammary epithelial cell line, 184. We determined that MIG2 plays a role in cell invasion by using small interfering RNA (siRNA) to suppress the expression of MIG2 mRNA levels in TMX2-28 cells. TMX2-28 cell invasion was reduced by 48% when the cells were transfected with siRNAs targeting MIG2, relative to cells transfected with siRNAs against glyceraldehyde-3-phosphate dehydrogenase. Finally, MIG2 expression was evaluated in reductive mammoplasty and breast tumor tissue. Although all 21 normal tissues from reduction mammoplasty showed immunoreactivity for MIG2, ranging from weak (62%) to strong (24%), only half of the 34 formalin-fixed breast tumors showed immunoreactivity for MIG2. Of these 17 positive cases, 10 were considered to overexpress MIG2 (moderate to strong staining). Examination of 30 frozen breast tumors supported the finding that MIG2 is overexpressed in a subset of breast cancers. We suggest that MIG2's normal regulation and function are disrupted in breast cancer.


The immunogenicity of different insulins in several animal species.
The immunologic properties of homologous and heterologous insulins have been investigated. Pigs, dogs, cows, sheep, goats, rabbits, guinea pigs, and rats were immunized with different hormone preparations alone or in combination with complete Freund adjuvant. The results obtained provide convincing evidence that in pigs homologous insulin cannot produce specific antibodies, whereas heterologous insulin can. Because the insulins of dogs and pigs have identical amino acid sequences, no antigenicity of porcine insulin in dogs could be observed either. In cattle, sheep, and goats, not only heterologous but also homologous insulins stimulated antibody production. Sheep and goats proved to be excellent reactor animals. Most of the small laboratory animals developed specific antibodies against insulin after hyperimmunization. In rabbits, not only the groups injected with nonchromatographed bovine insulin but also those hyperimmunized with single-component bovine insulin responded with a high serum level of specific antibodies. The data suggest that highly purified insulin preparations have not less antigenic activity than nonchromatographed insulin. Immunologically, des-Phe-B1-insulin acted exactly like the original hormone. Histologic examination of the pancreases of 45 pigs, 22 of which had high antibody titers, did not reveal insulitis. The results of the present paper point out that the production of specific antibodies is essentially a question of species specificity.


Animal disease outbreak control: the use of crisis management tools.
In this era of globalisation the effective control of animal disease outbreaks requires powerful crisis management tools. In the 1990s software packages for different sectors of the government and agricultural industry began to be developed. In 2004, as a special application for tracking the movement of animals and animal products, the European Union developed the Trade Control and Expert System (TRACES) on the basis of its predecessor, the ANImal MOvement (ANIMO) project. The nationwide use of the ANIMO system by the veterinary authorities in Germany marked the beginning of the development in 1993 of a computerised national animal disease reporting system--the TierSeuchenNachrichten (TSN)--using the ANIMO hardware and software components. In addition to TRACES and TSN the third pillar for the management of animal disease outbreaks and crises in Germany is the national cattle and swine database--called Herkunftssicherungs- und Informationssystem für Tiere. A high degree of standardisation is necessary when integrating the different solutions at all levels of government and with the private sector. In this paper, the authors describe the use of these tools on the basis of their experience and in relation to what we can do now and what we should opt for in the future.


Oxidative damage of aromatic dipeptides by the environmental oxidants NO2˙ and O3.
Irreversible oxidative damage at both aromatic side chains and dipeptide linkage occurs in the aromatic N- and C-protected dipeptides 7-11 upon exposure to the environmental pollutants NO2˙ and O3. The reaction proceeds through initial oxidation of the aromatic ring by in situ generated NO3˙, or by NO2˙, respectively, which leads to formation of nitroaromatic products. The indole ring in Phe-Trp undergoes oxidative cyclization to a pyrroloindoline. An important reaction pathway for dipeptides with less oxidisable aromatic side chains proceeds through fragmentation of the peptide bond with concomitant acyl migration. This process is likely initiated by an ionic reaction of the amide nitrogen with the NO2˙ dimer, N2O4.


Changes in the community structure of the symbiotic microbes of wild amphibians from the eastern edge of the Tibetan Plateau.
Environment has a potential effect on the animal symbiotic microbiome. Here, to study the potential relationship of the symbiotic microbiomes of wild amphibians with altitude, we collected the gut and skin samples from frogs (nine species) and the environmental samples (water and soil samples) from the Leshan Mountains (altitude: 360-410 m) and Gongga Mountains (altitude: 3340-3989 m) on the eastern edge of the Tibetan Plateau. Bufo gargarizans (Bg) samples were collected from both the Leshan and Gongga mountain regions (Bg was the only species sampled on both mountains). The DNA extracted from each sample was performed high-throughput sequencing (MiSeq) of bacterial 16S rRNA gene amplicons. High relative abundance of Caulobacteraceae and Sphingomonadaceae was found in skin samples from both Bg and the other high-altitude amphibians (nine species combined). High relative abundance of Coxiellaceae and Mycoplasmataceae was found in gut samples from both Bg and the other high-altitude amphibians. Furthermore, the alpha and beta diversities of skin and gut samples from Bg and the other amphibian species (nine species combined) were similar. In terms of the symbiotic microbial community, the low-altitude samples were less diverse and more similar to each other than the high-altitude samples were. We speculated that extreme high-altitude environments and host phylogeny may affect the amphibian microbiome. Despite the distinct microbial community differences between the skin and gut microbiomes, some functions were similar in the Bg and combined high-altitude samples. The Bg and high-altitude skin samples had higher oxidative stress tolerance and biofilm formation than the low-altitude skin samples. However, the opposite results were observed for the Bg and high-altitude gut samples. Further study is required to determine whether these characteristics favor high-altitude amphibian adaptation to extreme environments.


Evolution and Epidemic Spread of SARS-CoV-2 in Colombia: A Year into the Pandemic.
Current efforts to understand the epidemiology, transmission dynamics and emergence of novel SARS-CoV-2 variants worldwide has enabled the scientific community to generate critical information aimed at implementing disease surveillance and control measures, as well as to reduce the social, economic and health impact of the pandemic. Herein, we applied an epidemic model coupled with genomic analysis to assess the SARS-CoV-2 transmission dynamics in Colombia. This epidemic model allowed to identify the geographical distribution, Rt dynamics and predict the course of the pandemic considering current implementation of countermeasures. The analysis of the incidence rate per 100,000 inhabitants carried out across different regions of Colombia allowed visualizing the changes in the geographic distribution of cases. The cumulative incidence during the timeframe March 2020 to March 2021 revealed that Bogotá (8063.0), Quindío (5482.71), Amazonas (5055.68), Antioquia (4922.35) and Tolima (4724.41) were the departments with the highest incidence rate. The highest median Rt during the first period evaluated was 2.13 and 1.09 in the second period; with this model, we identified improving opportunities in health decision making related to controlling the pandemic, diagnostic testing capacity, case registration and reporting, among others. Genomic analysis revealed 52 circulating SARS-CoV-2 lineages in Colombia detected from 774 genomes sequenced throughout the first year of the pandemic. The genomes grouped into four main clusters and exhibited 19 polymorphisms. Our results provide essential information on the spread of the pandemic countrywide despite implementation of early containment measures. In addition, we aim to provide deeper phylogenetic insights to better understand the evolution of SARS-CoV-2 in light of the latent emergence of novel variants and how these may potentially influence transmissibility and infectivity.


Expression of Mycobacterium leprae genes from a Streptococcus mutans promoter in Escherichia coli K-12.
Genomic libraries of Mycobacterium leprae DNA partially digested with Pst I were constructed in the expression vector pYA626, which contains the promoter region from the Streptococcus mutans gene encoding aspartate beta-semialdehyde dehydrogenase, which is very efficiently expressed in Escherichia coli. We have detected several clones that complement a mutation in the citrate synthase gene of E. coli. Southern blot analysis demonstrated that the complementing DNA was M. leprae DNA. Sodium dodecyl sulfate/polyacrylamide gel analysis of polypeptides produced by minicells containing the citrate synthase-complementing recombinant molecules demonstrated the production of a 46-kDa polypeptide. When the citrate synthase-complementing fragment was cloned in pYA626 in the reverse orientation, the recombinant molecule was no longer able to complement the mutation in the citrate synthase gene and no longer produced the 46-kDa polypeptide. When the DNA fragment was cloned in the Pst I site of pHC79, so as to allow expression from the beta-lactamase promoter, the resulting recombinant failed to complement the mutation in the E. coli citrate synthase gene yet still produced the 46-kDa polypeptide, but in one-fourth the amount than when expressed from the S. mutans asd promoters. This demonstrates that M. leprae translational sequences can be recognized by E. coli translational machinery. Promoter expression vectors can be used to obtain expression of protein antigens to be used for early diagnosis of leprosy or components of a vaccine and proteins that are targets of potential antileprosy drugs.


Prognosis prediction of measurable enhancing lesion after completion of standard concomitant chemoradiotherapy and adjuvant temozolomide in glioblastoma patients: application of dynamic susceptibility contrast perfusion and diffusion-weighted imaging.
To assess the prognosis predictability of a measurable enhancing lesion using histogram parameters produced by the normalized cerebral blood volume (nCBV) and normalized apparent diffusion coefficient (nADC) after completion of standard concomitant chemoradiotherapy (CCRT) and adjuvant temozolomide (TMZ) medication in glioblastoma multiforme (GBM) patients. This study was approved by the institutional review board (IRB), and the requirement for informed consent was waived. A total of 59 patients with newly diagnosed GBM who received standard CCRT with TMZ and adjuvant TMZ for six cycles underwent perfusion-weighted and diffusion-weighted imaging. Twenty-seven patients had a measurable enhancing lesion and 32 patients lacked a measurable enhancing lesion based on the Response Assessment in Neuro-Oncology (RANO) criteria in the follow-up MRI, which was performed within 3 months after adjuvant TMZ therapy was completed. We measured the nCBV and nADC histogram parameters based on the measurable enhancing lesion. The progression free survival (PFS) was analyzed by the Kaplan-Meier method with the use of the log-rank test. The median PFS of patients lacking measurable enhancing lesion was longer than for those with measurable enhancing lesions (17.6 vs 3.3 months, P<.0001). There was a significant, positive correlation between the 99th percentile nCBV value of a measurable enhancing lesion and the PFS (P= .044, R(2)= .152). In addition, the median PFS was longer in patients with a 99th percentile nCBV value ≧ 4.5 than it was in those with a value <4.5 (4.4 vs 3.1 months, P = .036). We found that the nCBV value can be used for the prognosis prediction of a measurable enhancing lesion after the completion of standard treatment for GBM, wherein a high 99th percentile nCBV value (≧ 4.5) suggests a better PFS for GBM patients.


Maternal Prenatal External Locus of Control and Reduced Mathematical and Science Abilities in Their Offspring: A Longitudinal Birth Cohort Study.
A personality scale that identifies individuals' general attitude to what happens to them as largely a matter of luck or fate or of powerful others (externality) or whether they feel they can influence the consequences (internality) is known as locus of control (LOC). A continuous scale can distinguish those who are more external from those who are more internal. Lower scholastic achievement is associated with externality and higher achievement with internality, but little is known about the association of parental LOC on children's academic performance. Data collected within the Avon Longitudinal Study of Parents and Children (ALSPAC) are analyzed to assess associations between mothers' LOC orientation, measured during pregnancy, and their children's abilities in mathematics and science reasoning. We found that maternal external LOC is associated with lower scores for her child assessed by tests measuring mental arithmetic as well as understanding of mathematical and scientific concepts. Additionally, we determined the extent to which three separate sets of factors previously found to positively influence the developing child's ability mediate these findings: (a) perinatal and infant exposures, such as prenatal smoking, binge drinking, consumption of oily fish, and postnatal breast feeding; (b) parenting attitudes and strategies; and (c) the interface of the parents with their child's school. The three factors identify at least 50% of the mechanism by which maternal externality is associated with poor academic outcomes in her child and may be candidates for further investigation as possible intervention targets.


Silicon-based plasmonic waveguides.
We propose and comprehensively investigate Si-based plasmonic waveguides as a means to confine and manipulate photonic signals. The high refractive index of Si assures strong confinement and a very high level of photonic integration with achievable waveguide separations of the order of 10 nm and waveguide bends with 500 nm radius at telecommunication wavelengths, while using Al and Cu plasmonic material platforms, makes such waveguides fully compatible with existing CMOS fabrication processes. Their potential future in hybrid electronic/photonic chips is further reinforced as various configurations have been shown to compensate SPP propagation loss. The group velocity dispersion of such waveguides allows over 10 Tb/s signal transfer rates. The figures of merit allowing comparison of passive and active functionalities achievable with various waveguides have also been introduced.


A mouse model of hereditary hemorrhagic telangiectasia generated by transmammary-delivered immunoblocking of BMP9 and BMP10.
Hereditary hemorrhagic telangiectasia (HHT) is a potentially life-threatening genetic vascular disorder caused by loss-of-function mutations in the genes encoding activin receptor-like kinase 1 (ALK1), endoglin, Smad4, and bone morphogenetic protein 9 (BMP9). Injections of mouse neonates with BMP9/10 blocking antibodies lead to HHT-like vascular defects in the postnatal retinal angiogenesis model. Mothers and their newborns share the same immunity through the transfer of maternal antibodies during lactation. Here, we investigated whether the transmammary delivery route could improve the ease and consistency of administering anti-BMP9/10 antibodies in the postnatal retinal angiogenesis model. We found that anti-BMP9/10 antibodies, when intraperitoneally injected into lactating dams, are efficiently transferred into the blood circulation of lactationally-exposed neonatal pups. Strikingly, pups receiving anti-BMP9/10 antibodies via lactation displayed consistent and robust vascular pathology in the retina, which included hypervascularization and defects in arteriovenous specification, as well as the presence of multiple and massive arteriovenous malformations. Furthermore, RNA-Seq analyses of neonatal retinas identified an increase in the key pro-angiogenic factor, angiopoietin-2, as the most significant change in gene expression triggered by the transmammary delivery of anti-BMP9/10 antibodies. Transmammary-delivered BMP9/10 immunoblocking in the mouse neonatal retina is therefore a practical, noninvasive, reliable, and robust model of HHT vascular pathology.


Low-pH-dependent changes in the conformation and oligomeric state of the prefusion form of herpes simplex virus glycoprotein B are separable from fusion activity.
The cellular requirements for activation of herpesvirus fusion and entry remain poorly understood. Low pH triggers change in the antigenic reactivity of the prefusion form of the herpes simplex virus (HSV) fusion protein gB in virions, both in vitro and during viral entry via endocytosis (S. Dollery et al., J. Virol. 84:3759-3766, 2010). However, the mechanism and magnitude of gB conformational change are not clear. Here we show that the conformation and oligomeric state of gB with mutations in the bipartite fusion loops were similarly altered despite the fusion-inactivating mutations. Together with previous studies, this suggests that fusion loop mutants undergo conformational changes but are defective for fusion because they fail to make productive contact with the outer leaflet of the host target membrane. A direct, reversible effect of low pH on the structure of gB was detected by fluorescence spectroscopy. A soluble form of gB containing cytoplasmic tail sequences (s-gB) was triggered by mildly acidic pH to undergo changes in tryptophan fluorescence emission, hydrophobicity, antigenic conformation, and oligomeric structure and thus resembled the prefusion form of gB in the virion. In contrast, soluble gB730, for which the postfusion crystal structure is known, was only marginally affected by pH using these measures. The results underscore the importance of using a prefusion form of gB to assess the activation and extent of conformation change. Further, acidic pH had little to no effect on the conformation or hydrophobicity of gD or on gD's ability to bind nectin-1 or HVEM receptors. Our results support a model in which endosomal low pH serves as a cellular trigger of fusion by activating conformational changes in the fusion protein gB.


ADAM12 induction by Twist1 promotes tumor invasion and metastasis via regulation of invadopodia and focal adhesions.
The Twist1 transcription factor promotes tumor invasion and metastasis by inducing epithelial-mesenchymal transition (EMT) and invadopodia-mediated extracellular matrix (ECM) degradation. The critical transcription targets of Twist1 for mediating these events remain to be uncovered. Here, we report that Twist1 strongly induces expression of a disintegrin and metalloproteinase 12 (ADAM12). We observed that the expression levels of Twist1 mRNA and ADAM12 mRNA are tightly correlated in human breast tumors. Knocking down ADAM12 blocked cell invasion in a 3D mammary organoid culture. Suppression of ADAM12 also inhibited Twist1-induced tumor invasion and metastasis in human breast tumor xenografts, without affecting primary tumor formation. Mechanistically, knockdown of ADAM12 in breast cancer cells significantly reduced invadopodia formation and matrix degradation, and simultaneously increased overall cell adhesion to the ECM. Live-imaging analysis showed that knockdown of ADAM12 significantly inhibited focal adhesion turnover. Mechanistically, both the disintegrin and metalloproteinase domains of ADAM12 are required for its function at invadopodia, whereas the metalloproteinase domain is dispensable for its function at focal adhesions. Taken together, these data suggest that ADAM12 plays a crucial role in tumor invasion and metastasis by regulating both invadopodia and focal adhesions.


Detection of sequence variation in the HVII region of the human mitochondrial genome in 689 individuals using immobilized sequence-specific oligonucleotide probes.
We have developed a rapid, immobilized probe-based assay for the detection of sequence variation in the hypervariable segment II (HVII) of the mitochondrial DNA (mtDNA) control region. Using a panel of 17 sequence-specific oligonucleotide (SSO) probes immobilized on nylon membrane strips, we typed 689 individuals from four population groups. The genetic diversity value for each population was calculated from the frequency data, and the frequencies of distinct "mitotypes" in each group were determined. We performed DNA sequence analysis of 129 samples to characterize the sequences associated with "blanks" (absence of probe signals) and weak probe signals. Out of 689 samples, we observed five heteroplasmic samples (excluding the variable C-stretch beginning at position 303) using the immobilized SSO probe panel. The SSO probe strips were used for the analysis of shed hairs and bloodstains from several criminal cases in Sweden, one of which is described here. We conclude that this mtDNA typing system is useful for human identification and significantly decreases casework turnaround time.


Interventions for emergency contraception.
Emergency contraception (EC) is using a drug or copper intrauterine device (Cu-IUD) to prevent pregnancy shortly after unprotected intercourse. Several interventions are available for EC. Information on the comparative effectiveness, safety and convenience of these methods is crucial for reproductive healthcare providers and the women they serve. To determine which EC method following unprotected intercourse is the most effective, safe and convenient to prevent pregnancy. The search included the Cochrane Controlled Trials Register, Popline, MEDLINE, PubMed, Biosis/EMBASE, Chinese biomedical databases and UNDP/UNFPA/WHO/World Bank Special Programme on Human Reproduction (HRP) emergency contraception database (July 2011). Content experts and pharmaceutical companies were contacted. Randomised controlled trials and controlled clinical trials including women attending services for EC following a single act of unprotected intercourse were eligible. Data on outcomes and trial characteristics were extracted in duplicate and independently by two review authors. Quality assessment was also done by two review authors independently. Meta-analysis results are expressed as risk ratio (RR) using a fixed-effect model with 95% confidence interval (CI). In the presence of statistically significant heterogeneity a random-effects model was applied. One hundred trials with 55,666 women were included. Most trials were conducted in China (86/100). Meta-analysis indicated that mid-dose mifepristone (25-50 mg) (20 trials; RR 0.64; 95% CI 0.45 to 0.92) or low-dose mifepristone (< 25 mg) (11 trials; RR 0.70; 95% CI 0.50 to 0.97) were significantly more effective than levonorgestrel (LNG), but the significance was marginal when only high-quality studies were included (4 trials; RR 0.70; 95% CI 0.49 to 1.01). Low-dose mifepristone was less effective than mid-dose mifepristone (25 trials; RR 0.73; 95% CI 0.55 to 0.97). This difference was not statistically significant when only high-quality trials were considered (6 trials; RR 0.75; 95% CI 0.50 to 1.10). Ulipristal acetate (UPA) appeared more effective (2 trials; RR 0.63) than LNG at a marginal level (P = 0.09) within 72 hours of intercourse.Regarding effectiveness in relation to the time of administration, women who took LNG within 72 hours of intercourse were significantly less likely to be pregnant than those who took it after 72 hours (4 trials; RR 0.51; 95% CI 0.31 to 0.84). It was not evident that the coitus-treatment time affected the effectiveness of mifepristone and UPA.Single-dose LNG (1.5 mg) showed similar effectiveness as the standard two-dose regimen (0.75 mg 12 h apart) (3 trials; RR 0.84; 95% CI 0.53 to 1.33). This conclusion was not modified by the time elapsed from intercourse to treatment administration.Mifepristone (all doses) (3 trials; RR 0.14; 95% CI 0.05 to 0.41) and LNG (5 trials; RR 0.54; 95% CI 0.36 to 0.80) were more effective than the Yuzpe regimen in preventing pregnancy. One trial compared gestrinone with mifepristone. No significant difference of effectiveness was identified in this trial (996 women; RR 0.75; 95% CI 0.32 to 1.76).All methods of EC were safe. Nausea and vomiting occurred with oestrogen-containing EC methods and progestogen and anti-progestogen methods caused changes in subsequent menses. LNG users were more likely to have a menstrual return before the expected date, but UPA users were more likely to have a menstrual return after the expected date. Menstrual delay was the main adverse effect of mifepristone and seemed to be dose-related. Intermediate-dose mifepristone (25-50 mg) was superior to LNG and Yuzpe regimens. Mifepristone low dose (< 25 mg) may be more effective than LNG (0.75 mg two doses), but this was not conclusive. UPA may be more effective than LNG. LNG proved to be more effective than the Yuzpe regimen. The copper IUD was the most effective EC method and was the only EC method to provide ongoing contraception if left in situ.


Reported estimates of adverse pregnancy outcomes among women with and without syphilis: a systematic review and meta-analysis.
To estimate probability of adverse pregnancy outcomes (APOs) among women with and without syphilis through a systematic review of published literatures. Chinese and English literatures were searched for studies assessing pregnancy outcomes in the presence of maternal syphilis through August 2013. The prevalence estimates were summarized and analyzed by meta-analysis. Fifty-four literatures involving 11398 syphilitic women and 43342 non-syphilitic women were included from 4187 records initially found. Among untreated mothers with syphilis, pooled estimates were 76.8% for all APOs, 36.0% for congenital syphilis, 23.2% for preterm, 23.4% for low birth weight, 26.4% for stillbirth or fetal loss, 14.9% for miscarriage and 16.2% for neonatal deaths. Among syphilitic mother receiving treatment only in the late trimester (>28 weeks), pooled estimates were 64.4% for APOs, 40.6% for congenital syphilis, 17.6% for preterm, 12.4% for low birth weight, and 21.3% for stillbirth or fetal loss. Among syphilitic mothers with high titers (≥1∶8), pooled estimates were 42.8% for all APOs, 25.8% for congenital syphilis, 15.1% for preterm, 9.4% for low birth weight, 14.6% for stillbirth or fetal loss and 16.0% for neonatal deaths. Among non-syphilitic mothers, the pooled estimates were 13.7% for all APOs, 7.2% for preterm birth, 4.5% for low birth weight, 3.7% for stillbirth or fetal loss, 2.3% for miscarriage and 2.0% for neonatal death. Begg's rank correlation test indicated little evidence of publication bias (P>0.10). Substantial heterogeneity was found across studies in the estimates of all adverse outcomes for both women with syphilis (I2 = 93.9%; P<0.0001) and women without syphilis (I2 = 94.8%; P<0.0001). Syphilis continues to be an important cause of substantial perinatal morbidity and mortality, which reminds that policy-makers charged with resource allocation that the elimination of mother-to-child transmission of syphilis is a public health priority.


[Economic impact of kidney patients with sepsis in hospital setting].
Introduction: Over the last decades, sepsis has become a real medical emergency, with a high mortality rate and often requiring admission to an intensive care unit. An increasing number of CKD patients contracts sepsis due to several clinical risk factors (use of catheters, immunosuppressive therapy, comorbidity, etc.) and is treated in Nephrology wards, generating additional costs that are not covered by hospital Diagnosis Related Groups (DRG) reimbursement. The aim of the study is to evaluate the costs of sepsis in one Nephrology Unit and to detect the mortality rate of CKD patients with sepsis. Methods: We conducted a retrospective study on a cohort of CKD patients admitted into one Nephrology Unit in 2017. CKD inpatients were divided in two groups: patients with sepsis (SP) and without (control group). Socio-demographic, clinical and therapeutic data, as well as routine biochemistry, were collected through a "sepsis form". SP were identified thanks to hospital discharge records (HDR). The hospital-related costs of a SP were obtained by summing up: (1) the average cost of an inpatient day of care for the average length of stay in the Nephrology Unit; (2) the average cost of the antimicrobial therapy, as recorded on the clinical folder. Results: Among the 408 CKD inpatients, 61 were septic. The overall average cost of a SP was 23.087,57 €; the average cost of the hospital stay and of the antimicrobial therapy was 19.364,98 € and 3.722,60 € respectively. The average length of stay in the Nephrology Unit was 16.7 days. The in-hospital mortality rate was 41.7%, with a 312% additional mortality rate. Conclusions: SP had an overall average cost three times higher than CKD inpatients without sepsis (9.290,79 €). This additional cost was due to a longer hospital stay (8.7 days more on average) and a higher cost of antimicrobial therapy per case (€ 221,24). A national multi-centre study is needed to confirm our data and to promote an adjustment of reimbursement tariff for DRG-sepsis, which is now applicable only to an ICU setting.


Impact of relationship dynamics and gender roles in the protection of HIV discordant heterosexual couples: an exploratory study in the Puerto Rican context.
Most of the HIV/AIDS prevention efforts have not taken into consideration the context of the relationship and the gender constructs that influence relationship dynamics. These efforts have failed to view HIV prevention as a collaborative process between partners. Therefore, it is important to explore how relationship dynamics and gender constructs influence how men and women involved in an HIV discordant heterosexual relationship, visualize their role in the protection of their partners in order to design more effective prevention interventions. Five Puerto Rican HIV discordant heterosexual couples were interviewed via a qualitative semi-structured interview. The taped interviews were transcribed and analyzed using content analysis according to a set of defined categories. Women visualized their role as one of convincing their partners to use protection as well as being strong and firm in the demand of its use. Men viewed their role as one of being more supportive and willing to use protection, but recognized their resistance towards the use of condoms. Relationship dynamics such as communication and support promoted protection. Traditional and non-traditional gender roles were assumed by both men and women. Traditional gender roles inhibited protection but were also used in positive ways to promote it. Men showed a greater initiative to break with traditional gender norms. A positive relationship, marked by communication and support could serve as a facilitator in the protection and in the transformation of traditional gender norms. This points out to the need of viewing HIV/AIDS prevention as a collaborative rather than individualistic process.


Evaluation of self-collected cervicovaginal cell samples for human papillomavirus testing by polymerase chain reaction.
As human papillomavirus (HPV) becomes accepted as the central cause of cervical cancer, longitudinal studies are shifting focus away from causality to a more detailed investigation of the natural history of HPV infections. These studies commonly require repeated samples for HPV testing over several years, usually collected during a pelvic exam, which is inconvenient to the participants and costly to the study. To alleviate the inconvenience and cost of repeated clinic visits, it has been proposed that women collect cervicovaginal cells themselves, hopefully increasing participation in the natural history studies. We evaluated the technical feasibility of self-collection of cervicovaginal cells using a Dacron swab for HPV DNA detection. We compared the self-collected swab sample and two clinician-administered swab samples (one from the endocervix and another from the ectocervix) from a total of 268 women participating in a case-control study of adenocarcinoma and squamous cell carcinomas of the uterine cervix (111 cases and 157 controls). HPV DNA was detected and genotyped using an L1 consensus PCR assay. The overall agreement between the clinician- and self-collected swabs was excellent [88.1%; kappa = 0.73 (95% confidence interval (CI), 0.61-0.85)]. The correlation was highest between the two clinician-administered swabs [kappa = 0.81 (95% CI, 0.69-0.93)] but was still excellent when comparing either clinician-administered swab to the self-administered sample [kappa = 0.75 (95% CI, 0.63-0.87) and 0.67 (95% CI, 0.55-0.79) for ectocervix and endocervix, respectively]. The type-specific agreement between samples was higher for high-risk, or cancer-associated, HPV genotypes than for low risk, noncancer-associated HPV genotypes when comparing the self-administered swab sample to the clinician-administered swab sample (kappa = 0.78 for high-risk versus 0.66 for low-risk HPV infections, t = -1.45, P = 0.15). The decrease in agreement for low risk types was largely attributable to an increased detection of these types in the self-administered sample (McNemar's chi2 = 6.25, P = 0.01 for clinician- versus self-administered swab comparisons). The agreement did not vary significantly by age, menopausal status, case status, or clinic center. We have demonstrated that a self-collected Dacron swab sample of cervicovaginal cells is a technically feasible alternative to clinician-administered cervical cell collection in natural history studies of HPV and cervical cancer.


Effects of a bioactive olive pomace extract from Olea europaea on growth performance, gut function, and intestinal microbiota in broiler chickens.
The present study aims to investigate the effects of supplementing broiler diets with a bioactive olive pomace extract (OE) from Olea europaea on growth performance, digestibility, gut microbiota, bile acid composition, and immune response. To this end, three hundred and six 1-day-old broiler chickens (Ross 308) were housed in floor pens (6 pens/treatment, with 17 birds/pen). Animals were fed with a standard non-medicated starter diet for 21 D, and from 22 to 42 D of age with their respective experimental diet: a negative control with no additives (Control), a positive control with 100 ppm of monensin (Monensin) and the basal diet supplemented with 750 ppm of an OE (Lucta S.A., Spain). Feed intake and growth rate were monitored weekly throughout the trial. From 21 to 42 D of age, no significant differences in feed intake were observed among dietary treatments; however, lower average daily gain and higher feed conversion ratio (P < 0.05) was observed in birds fed the Control compared to Monensin and OE groups. Performance of birds fed OE or Monensin was similar throughout the trial. The apparent ileal digestibility of crude protein was higher in birds fed Monensin than Control treatment (P < 0.05). No significant changes on bacterial composition at a family level were observed in the caeca of birds fed the experimental diets. Moreover, no significant differences on plasma and intestinal bile acid composition were observed among treatments. Birds fed the OE showed a significant decrease of IL-8 expression in the ileum (P < 0.05). Additionally, the expression of TGF-β4, and Bu-1 was significantly upregulated (P < 0.01) in broilers fed the OE and Monensin diets compared to those fed the Control. In conclusion, the inclusion of 750 ppm of a bioactive olive pomace extract from Olea europaea in broiler chicken diets improved animal growth likely as result of its anti-inflammatory properties.


Natural epigenetic variation within and among six subspecies of the house sparrow, Passer domesticus.
Epigenetic modifications can respond rapidly to environmental changes and can shape phenotypic variation in accordance with environmental stimuli. One of the most studied epigenetic marks is DNA methylation. In the present study, we used the methylation-sensitive amplified polymorphism (MSAP) technique to investigate the natural variation in DNA methylation within and among subspecies of the house sparrow, Passer domesticus We focused on five subspecies from the Middle East because they show great variation in many ecological traits and because this region is the probable origin for the house sparrow's commensal relationship with humans. We analysed house sparrows from Spain as an outgroup. The level of variation in DNA methylation was similar among the five house sparrow subspecies from the Middle East despite high phenotypic and environmental variation, but the non-commensal subspecies was differentiated from the other four (commensal) Middle Eastern subspecies. Further, the European subspecies was differentiated from all other subspecies in DNA methylation. Our results indicate that variation in DNA methylation does not strictly follow subspecies designations. We detected a correlation between methylation level and some morphological traits, such as standardized bill length, and we suggest that part of the high morphological variation in the native populations of the house sparrow is influenced by differentially methylated regions in specific loci throughout the genome. We also detected 10 differentially methylated loci among subspecies and three loci that differentiated between commensal or non-commensal status. Therefore, the MSAP technique detected larger scale differences among the European and non-commensal subspecies, but did not detect finer scale differences among the other Middle Eastern subspecies.


Association Between Urbanicity and Dementia in China: A Population-Based Study.
This study investigated the relationship between urbanicity and dementia and predicted its nonlinear pattern among Chinese adults aged 50 years and older. This study used data from the Second National Sample Survey on Disability, which was implemented from April 1 to May 31, 2006 across China. Dementia status was determined by a 2-stage process: the combination of self-reports or family members' reports and an onsite medical diagnosis by experienced specialists based on the International Statistical Classification of Diseases and Related Health Problems 10th Revision Symptom Checklist for Mental Disorders. Logistic regression models were used to examine the relationship between urbanicity and dementia, and restricted polynomial spline regression models were plotted to examine the nonlinear exposure-response relationship of urbanicity and dementia. Logistic regression results showed that an increase of 10% in the degree of urbanization was associated with a 73% decrease in the odds of dementia after adjusting for covariates, particularly area-level socioeconomic variables. This observed association was stronger in the younger age group, and this age group difference was only present in women. Spline regression findings suggested a nonlinear exposure-response relationship between urbanicity and the odds of dementia. Areas with very high levels of urbanization were associated with increased odds of dementia. These findings highlight the necessity to properly examine the nuanced relationship between urbanicity and mental health, especially for women in the younger age group. Notably, there were increased odds of dementia at very high levels of urbanicity.


HSV-2-driven increase in the expression of α4β7 correlates with increased susceptibility to vaginal SHIV(SF162P3) infection.
The availability of highly susceptible HIV target cells that can rapidly reach the mucosal lymphoid tissues may increase the chances of an otherwise rare transmission event to occur. Expression of α4β7 is required for trafficking of immune cells to gut inductive sites where HIV can expand and it is expressed at high level on cells particularly susceptible to HIV infection. We hypothesized that HSV-2 modulates the expression of α4β7 and other homing receptors in the vaginal tissue and that this correlates with the increased risk of HIV acquisition in HSV-2 positive individuals. To test this hypothesis we used an in vivo rhesus macaque (RM) model of HSV-2 vaginal infection and a new ex vivo model of macaque vaginal explants. In vivo we found that HSV-2 latently infected RMs appeared to be more susceptible to vaginal SHIVSF162P3 infection, had higher frequency of α4β7high CD4+ T cells in the vaginal tissue and higher expression of α4β7 and CD11c on vaginal DCs. Similarly, ex vivo HSV-2 infection increased the susceptibility of the vaginal tissue to SHIVSF162P3. HSV-2 infection increased the frequencies of α4β7high CD4+ T cells and this directly correlated with HSV-2 replication. A higher amount of inflammatory cytokines in vaginal fluids of the HSV-2 infected animals was similar to those found in the supernatants of the infected explants. Remarkably, the HSV-2-driven increase in the frequency of α4β7high CD4+ T cells directly correlated with SHIV replication in the HSV-2 infected tissues. Our results suggest that the HSV-2-driven increase in availability of CD4+ T cells and DCs that express high levels of α4β7 is associated with the increase in susceptibility to SHIV due to HSV-2. This may persists in absence of HSV-2 shedding. Hence, higher availability of α4β7 positive HIV target cells in the vaginal tissue may constitute a risk factor for HIV transmission.


Sex reversal in zebrafish fancl mutants is caused by Tp53-mediated germ cell apoptosis.
The molecular genetic mechanisms of sex determination are not known for most vertebrates, including zebrafish. We identified a mutation in the zebrafish fancl gene that causes homozygous mutants to develop as fertile males due to female-to-male sex reversal. Fancl is a member of the Fanconi Anemia/BRCA DNA repair pathway. Experiments showed that zebrafish fancl was expressed in developing germ cells in bipotential gonads at the critical time of sexual fate determination. Caspase-3 immunoassays revealed increased germ cell apoptosis in fancl mutants that compromised oocyte survival. In the absence of oocytes surviving through meiosis, somatic cells of mutant gonads did not maintain expression of the ovary gene cyp19a1a and did not down-regulate expression of the early testis gene amh; consequently, gonads masculinized and became testes. Remarkably, results showed that the introduction of a tp53 (p53) mutation into fancl mutants rescued the sex-reversal phenotype by reducing germ cell apoptosis and, thus, allowed fancl mutants to become fertile females. Our results show that Fancl function is not essential for spermatogonia and oogonia to become sperm or mature oocytes, but instead suggest that Fancl function is involved in the survival of developing oocytes through meiosis. This work reveals that Tp53-mediated germ cell apoptosis induces sex reversal after the mutation of a DNA-repair pathway gene by compromising the survival of oocytes and suggests the existence of an oocyte-derived signal that biases gonad fate towards the female developmental pathway and thereby controls zebrafish sex determination.


Suppression of Heregulin-β1/HER2-Modulated Invasive and Aggressive Phenotype of Breast Carcinoma by Pterostilbene via Inhibition of Matrix Metalloproteinase-9, p38 Kinase Cascade and Akt Activation.
Invasive breast cancer is the major cause of death among females and its incidence is closely linked to HER2 (human epidermal growth factor receptor 2) overexpression. Pterostilbene, a natural analog of resveratrol, exerts its cancer chemopreventive activity similar to resveratrol by inhibiting cancer cell proliferation and inducing apoptosis. However, the anti-invasive effect of pterostilbene on HER2-bearing breast cancer has not been evaluated. Here, we used heregulin-β1 (HRG-β1), a ligand for HER3, to transactivate HER2 signaling. We found that pterostilbene was able to suppress HRG-β1-mediated cell invasion, motility and cell transformation of MCF-7 human breast carcinoma through down-regulation of matrix metalloproteinase-9 (MMP-9) activity and growth inhibition. In parallel, pterostilbene also inhibited protein and mRNA expression of MMP-9 driven by HRG-β1, suggesting that pterostilbene decreased HRG-β1-mediated MMP-9 induction via transcriptional regulation. Examining the signaling pathways responsible for HRG-β1-associated MMP-9 induction and growth inhibition, we observed that pterostilbene, as well as SB203580 (p38 kinase inhibitor), can abolish the phosphorylation of p38 mitogen-activated protein kinase (p38 kinase), a downstream HRG-β1-responsive kinase responsible for MMP-9 induction. In addition, HRG-β1-driven Akt phosphorylation required for cell proliferation was also suppressed by pterostilbene. Taken together, our present results suggest that pterostilbene may serve as a chemopreventive agent to inhibit HRG-β1/HER2-mediated aggressive and invasive phenotype of breast carcinoma through down-regulation of MMP-9, p38 kinase and Akt activation.


Evolutionary history and functional characterization of three large genes involved in sporulation in Bacillus cereus group bacteria.
The Bacillus cereus group of bacteria is a group of closely related species that are of medical and economic relevance, including B. anthracis, B. cereus, and B. thuringiensis. Bacteria from the Bacillus cereus group encode three large, highly conserved genes of unknown function (named crdA, crdB, and crdC) that are composed of 16 to 35 copies of a repeated domain of 132 amino acids at the protein level. Bioinformatic analysis revealed that there is a phylogenetic bias in the genomic distribution of these genes and that strains harboring all three large genes mainly belong to cluster III of the B. cereus group phylogenetic tree. The evolutionary history of the three large genes implicates gain, loss, duplication, internal deletion, and lateral transfer. Furthermore, we show that the transcription of previously identified antisense open reading frames in crdB is simultaneously regulated with its host gene throughout the life cycle in vitro, with the highest expression being at the onset of sporulation. In B. anthracis, different combinations of double- and triple-knockout mutants of the three large genes displayed slower and less efficient sporulation processes than the parental strain. Altogether, the functional studies suggest an involvement of these three large genes in the sporulation process.


Palliative care for intestinal obstruction in recurrent ovarian cancer: a multivariate analysis.
Bowel obstruction is the most common complication in patients with ovarian cancer. Management of this situation is controversial. The aim of our retrospective study was to determine the best approach for managing bowel obstruction in recurrent ovarian cancer. A retrospective analysis of data on 47 patients with intestinal obstruction by ovarian cancer was performed. Twenty-seven patients were submitted to surgery, with 21 intestinal procedures performed, 2 gastrostomy tubes placed, and 4 patients deemed inoperable. Twenty patients were managed medically with Octreotide (mean dosage of 0.48 mg/day), of which 1 patient required a nasogastric tube. Age, performance status, diagnosis of tumor to occlusion time, obstruction site, previous chemotherapy or radiotherapy, presence of ascites, or palpable masses were the variables analyzed. Student's t-test and Pearson chi-square test were used to compare the two different groups of treatment (surgical vs medical therapy). Disease-free-survival curves were plotted according to the Kaplan-Meier method and analyzed by the log-rank test. Cox's proportional hazards model was used for multivariate analysis. Values less than or equal to 0.05 were considered significant. The mean age of the patients was 58.7 years. Perioperative mortality and morbidity were both 22%. All patients died with minimal distress. Performance status results were significantly different between the patients submitted to surgery and patients treated with Octreotide (P= 0.03). No significant differences were found in the other variables analyzed. In multivariate analysis, only type of treatment emerges as a strong predictor of poor outcome (P < 0.001). Both surgery and Octreotide therapy are able to control distressing symptoms in end-stage ovarian cancer. Survival was significantly longer in the surgical group, and surgical palliation should be considered first in patients with good performance status.


Real-world performance and accuracy of stress echocardiography: the EVAREST observational multi-centre study.
Stress echocardiography is widely used to identify obstructive coronary artery disease (CAD). High accuracy is reported in expert hands but is dependent on operator training and image quality. The EVAREST study provides UK-wide data to evaluate real-world performance and accuracy of stress echocardiography. Participants undergoing stress echocardiography for CAD were recruited from 31 hospitals. Participants were followed up through health records which underwent expert adjudication. Cardiac outcome was defined as anatomically or functionally significant stenosis on angiography, revascularization, medical management of ischaemia, acute coronary syndrome, or cardiac-related death within 6 months. A total of 5131 patients (55% male) participated with a median age of 65 years (interquartile range 57-74). 72.9% of studies used dobutamine and 68.5% were contrast studies. Inducible ischaemia was present in 19.3% of scans. Sensitivity and specificity for prediction of a cardiac outcome were 95.4% and 96.0%, respectively, with an accuracy of 95.9%. Sub-group analysis revealed high levels of predictive accuracy across a wide range of patient and protocol sub-groups, with the presence of a resting regional wall motion abnormalitiy significantly reducing the performance of both dobutamine (P < 0.01) and exercise (P < 0.05) stress echocardiography. Overall accuracy remained consistently high across all participating hospitals. Stress echocardiography has high accuracy across UK-based hospitals and thus indicates stress echocardiography is being delivered effectively in real-world practice, reinforcing its role as a first-line investigation in the assessment of patients with stable chest pain.


Mechanism of cloned ATP-sensitive potassium channel activation by oleoyl-CoA.
Insulin secretion from pancreatic beta cells is coupled to cell metabolism through closure of ATP-sensitive potassium (KATP) channels, which comprise Kir6.2 and sulfonylurea receptor (SUR1) subunits. Although metabolic regulation of KATP channel activity is believed to be mediated principally by the adenine nucleotides, other metabolic intermediates, including long chain acyl-CoA esters, may also be involved. We recorded macroscopic and single-channel currents from Xenopus oocytes expressing either Kir6.2/SUR1 or Kir6. 2DeltaC36 (which forms channels in the absence of SUR1). Oleoyl-CoA (1 microM) activated both wild-type Kir6.2/SUR1 and Kir6.2DeltaC36 macroscopic currents, approximately 2-fold, by increasing the number and open probability of Kir6.2/SUR1 and Kir6.2DeltaC36 channels. It was ineffective on the related Kir subunit Kir1.1a. Oleoyl-CoA also impaired channel inhibition by ATP, increasing the Ki values for both Kir6.2/SUR1 and Kir6.2DeltaC36 currents by approximately 3-fold. Our results indicate that activation of KATP channels by oleoyl-CoA results from an interaction with the Kir6.2 subunit, unlike the stimulatory effects of MgADP and diazoxide which are mediated through SUR1. The increased activity and reduced ATP sensitivity of KATP channels by oleoyl-CoA might contribute to the impaired insulin secretion observed in non-insulin-dependent diabetes mellitus.


What mediates tree mortality during drought in the southern Sierra Nevada?
Severe drought has the potential to cause selective mortality within a forest, thereby inducing shifts in forest species composition. The southern Sierra Nevada foothills and mountains of California have experienced extensive forest dieback due to drought stress and insect outbreak. We used high-fidelity imaging spectroscopy (HiFIS) and light detection and ranging (LiDAR) from the Carnegie Airborne Observatory (CAO) to estimate the effect of forest dieback on species composition in response to drought stress in Sequoia National Park. Our aims were (1) to quantify site-specific conditions that mediate tree mortality along an elevation gradient in the southern Sierra Nevada Mountains, (2) to assess where mortality events have a greater probability of occurring, and (3) to estimate which tree species have a greater likelihood of mortality along the elevation gradient. A series of statistical models were generated to classify species composition and identify tree mortality, and the influences of different environmental factors were spatially quantified and analyzed to assess where mortality events have a greater likelihood of occurring. A higher probability of mortality was observed in the lower portion of the elevation gradient, on southwest- and west-facing slopes, in areas with shallow soils, on shallower slopes, and at greater distances from water. All of these factors are related to site water balance throughout the landscape. Our results also suggest that mortality is species-specific along the elevation gradient, mainly affecting Pinus ponderosa and Pinus lambertiana at lower elevations. Selective mortality within the forest may drive long-term shifts in community composition along the elevation gradient.