Transcriptome Sequencing (RNAseq) Enables Utilization of Formalin-Fixed, Paraffin-Embedded Biopsies with Clear Cell Renal Cell Carcinoma for Exploration of Disease Biology and Biomarker Development.
Formalin-fixed, paraffin-embedded (FFPE) tissues are an underused resource for molecular analyses. This proof of concept study aimed to compare RNAseq results from FFPE biopsies with the corresponding RNAlater® (Qiagen, Germany) stored samples from clear cell renal cell carcinoma (ccRCC) patients to investigate feasibility of RNAseq in archival tissue. From each of 16 patients undergoing partial or full nephrectomy, four core biopsies, such as two specimens with ccRCC and two specimens of adjacent normal tissue, were obtained with a 16g needle. One normal and one ccRCC tissue specimen per patient was stored either in FFPE or RNAlater®. RNA sequencing libraries were generated applying the new Illumina TruSeq® Access library preparation protocol. Comparative analysis was done using voom/Limma R-package. The analysis of the FFPE and RNAlater® datasets yielded similar numbers of detected genes, differentially expressed transcripts and affected pathways. The FFPE and RNAlater datasets shared 80% (n = 1106) differentially expressed genes. The average expression and the log2 fold changes of these transcripts correlated with R2 = 0.97, and R2 = 0.96, respectively. Among transcripts with the highest fold changes in both datasets were carbonic anhydrase 9 (CA9), neuronal pentraxin-2 (NPTX2) and uromodulin (UMOD) that were confirmed by immunohistochemistry. IPA revealed the presence of gene signatures of cancer and nephrotoxicity, renal damage and immune response. To simulate the feasibility of clinical biomarker studies with FFPE samples, a classifier model was developed for the FFPE dataset: expression data for CA9 alone had an accuracy, specificity and sensitivity of 94%, respectively, and achieved similar performance in the RNAlater dataset. Transforming growth factor-ß1 (TGFB1)-regulated genes, epithelial to mesenchymal transition (EMT) and NOTCH signaling cascade may support novel therapeutic strategies. In conclusion, in this proof of concept study, RNAseq data obtained from FFPE kidney biopsies are comparable to data obtained from fresh stored material, thereby expanding the utility of archival tissue specimens.


Treatment of pain associated with deep endometriosis: alternatives and evidence.
Pain is the most evident clinical manifestation of deep infiltrating endometriosis (DIE). Several hormonal and immunologic mechanisms are markedly altered in DIE compared with superficial peritoneal and ovarian endometriosis, and may explain its most aggressive behavior and the presence of severe pain symptoms. Hormonal therapies, such as combined hormonal contraceptives and progestogens, should be regarded as first-line treatment, as they are efficacious, safe, and well tolerated. Gonadotropin-releasing hormone agonists may be used in patients with symptoms persisting after the administration of first-line therapies. Scanty literature is available for danazol treatment in patients with DIE and, however, it has become less popular due to the high rates of androgenic adverse events (AEs). The partial relief of pain that often is achieved with available therapies and its recurrence after the suspension of the treatment have brought to the development of new therapies (such as aromatase inhibitors, oral GnRH antagonists) that are currently under investigation. Surgical excision of DIE should be considered in patients with pain symptoms persisting after first-line hormonal therapies. The benefits of surgery in terms of pain improvement should be always balanced with the risk of intraoperative complications and for this reason surgical cases should be referred to tertiary centers for the treatment of DIE. A multidisciplinary approach is mandatory in patients with DIE involving the bowel and/or the urinary tract.


Transcriptome characterization and polymorphism detection between subspecies of big sagebrush (Artemisia tridentata).
Big sagebrush (Artemisia tridentata) is one of the most widely distributed and ecologically important shrub species in western North America. This species serves as a critical habitat and food resource for many animals and invertebrates. Habitat loss due to a combination of disturbances followed by establishment of invasive plant species is a serious threat to big sagebrush ecosystem sustainability. Lack of genomic data has limited our understanding of the evolutionary history and ecological adaptation in this species. Here, we report on the sequencing of expressed sequence tags (ESTs) and detection of single nucleotide polymorphism (SNP) and simple sequence repeat (SSR) markers in subspecies of big sagebrush. cDNA of A. tridentata sspp. tridentata and vaseyana were normalized and sequenced using the 454 GS FLX Titanium pyrosequencing technology. Assembly of the reads resulted in 20,357 contig consensus sequences in ssp. tridentata and 20,250 contigs in ssp. vaseyana. A BLASTx search against the non-redundant (NR) protein database using 29,541 consensus sequences obtained from a combined assembly resulted in 21,436 sequences with significant blast alignments (≤ 1e⁻¹⁵). A total of 20,952 SNPs and 119 polymorphic SSRs were detected between the two subspecies. SNPs were validated through various methods including sequence capture. Validation of SNPs in different individuals uncovered a high level of nucleotide variation in EST sequences. EST sequences of a third, tetraploid subspecies (ssp. wyomingensis) obtained by Illumina sequencing were mapped to the consensus sequences of the combined 454 EST assembly. Approximately one-third of the SNPs between sspp. tridentata and vaseyana identified in the combined assembly were also polymorphic within the two geographically distant ssp. wyomingensis samples. We have produced a large EST dataset for Artemisia tridentata, which contains a large sample of the big sagebrush leaf transcriptome. SNP mapping among the three subspecies suggest the origin of ssp. wyomingensis via mixed ancestry. A large number of SNP and SSR markers provide the foundation for future research to address questions in big sagebrush evolution, ecological genetics, and conservation using genomic approaches.


Unprecedented Phthalocyanines Bearing Eight Di-butylamino Peripheral Substituents: Synthesis, Spectroscopy, and Structure.
Unprecedented 2,3,9,10,16,17,23,24-octakis(di-butylamino)phthalocyanine compounds M{Pc[N(C4H9)2]8} (M = 2H, Mg, Cu, Zn) (1-4) were prepared and structurally characterized on the basis of single-crystal X-ray diffraction analysis, representing the first structurally characterized alkylamino-substituted phthalocyanine examples. These novel phthalocyanine derivatives have also been characterized by a wide range of spectroscopic methods including MALDI-TOF mass spectra, NMR, electronic absorption, and IR spectroscopy in addition to elemental analysis. Their electrochemistry was also studied by cyclic voltammetry.


Cardiac troponin testing and the simplified Pulmonary Embolism Severity Index. The SWIss Venous ThromboEmbolism Registry (SWIVTER).
A low simplified Pulmonary Embolism Severity Index (sPESI), defined as age ≤80 years and absence of systemic hypotension, tachycardia, hypoxia, cancer, heart failure, and lung disease, identifies low-risk patients with acute pulmonary embolism (PE). It is unknown whether cardiac troponin testing improves the prediction of clinical outcomes if the sPESI is not low. In the prospective Swiss Venous Thromboembolism Registry, 369 patients with acute PE and a troponin test (conventional troponin T or I, highly sensitive troponin T) were enrolled from 18 hospitals. A positive test result was defined as a troponin level above the manufacturers assay threshold. Among the 106 (29%) patients with low sPESI, the rate of mortality or PE recurrence at 30 days was 1.0%. Among the 263 (71%) patients with high sPESI, 177 (67%) were troponin-negative and 86 (33%) troponin-positive; the rate of mortality or PE recurrence at 30 days was 4.6% vs. 12.8% (p=0.015), respectively. Overall, risk assessment with a troponin test (hazard ratio [HR] 3.39, 95% confidence interval [CI] 1.38-8.37; p=0.008) maintained its prognostic value for mortality or PE recurrence when adjusted for sPESI (HR 5.80, 95%CI 0.76-44.10; p=0.09). The combination of sPESI with a troponin test resulted in a greater area under the receiver-operating characteristic curve (HR 0.72, 95% CI 0.63-0.81) than sPESI alone (HR 0.63, 95% CI 0.57-0.68) (p=0.023). In conclusion, although cardiac troponin testing may not be required in patients with a low sPESI, it adds prognostic value for early death and recurrence for patients with a high sPESI.


Postpolypectomy Fever, a rare adverse event of polypectomy: nested case-control study.
Although postpolypectomy fever (PPF) without colon perforation or hemorrhage is rare, its incidence and risk factors have not been investigated. The objective of this study was to analyze the incidence and risk factors for PPF among inpatients. Seven patients with PPF were matched with 70 patients without PPF from a total of 3,444 patients who underwent colonoscopic polypectomy. The PPF incidence during index hospitalization after colonoscopy was calculated, and univariate and multivariate analyses were performed to calculate the adjusted odds ratios (ORs) for risk factors. PPF without bleeding or perforation in the colon occurred in seven patients (0.2%). The median age was 58 years for cases and 61 years for controls. The median interval from polypectomy to occurrence of fever was 7 hours, and the median duration of fever was 9 hours. Polyp size >2 cm (adjusted OR, 1.08; 95% confidence interval [CI], 1.01 to 1.15; p=0.02) and hypertension (adjusted OR, 14.40; 95% CI, 1.23 to 180.87; p=0.03) were associated with a significantly increased risk of PPF. PPF increased the length of hospitalization. Although the crude incidence of PPF is low, PPF may prolong hospitalization. Risk factors for PPF include hypertension and large polyps.


Manipulation of adsorbed atoms and creation of new structures on room-temperature surfaces with a scanning tunneling microscope.
A general method of manipulating adsorbed atoms and molecules on room-temperature surfaces with the use of a scanning tunneling microscope is described. By applying an appropriate voltage pulse between the sample and probe tip, adsorbed atoms can be induced to diffuse into the region beneath the tip. The field-induced diffusion occurs preferentially toward the tip during the voltage pulse because of the local potential energy gradient arising from the interaction of the adsorbate dipole moment with the electric field gradient at the surface. Depending upon the surface and pulse parameters, cesium (Cs) structures from one nanometer to a few tens of nanometers across have been created in this way on the (110) surfaces of gallium arsenide (GaAs) and indium antimonide (InSb), including structures that do not naturally occur.


Serum insulin-like growth factor I (IGF-I) and IGF-binding protein 3 levels are increased in central precocious puberty: effects of two different treatment regimens with gonadotropin-releasing hormone agonists, without or in combination with an antiandrogen (cyproterone acetate).
Central precocious puberty (CPP) is characterized by early activation of the pituitary-gonadal axis, which leads to increased growth velocity and development of secondary sexual characteristics. It is generally believed that gonadal sex steroids stimulate pulsatile GH secretion, which, in turn, stimulates insulin-like growth factor I (IGF-I) and IGF-binding protein 3 (IGFBP-3) production. However, little is known about GH, IGF-I, and IGFBP-3 serum levels in children with precocious puberty. Treatment of CPP by GnRH agonists has become the treatment of choice. However, the effect of long term treatment with GnRH in combination with an antiandrogen (cyproterone acetate) to block the possible effect of adrenal androgens has not previously been evaluated. We, therefore, studied 40 patients with idiopathic CPP that were treated for 24 months with either GnRH analog (Buserelin) in combination with cyproterone acetate (Androcur; n = 23) or with long acting GnRH analog (Decapeptyl Depot; n = 17). We found that serum IGF-I levels were increased before treatment in both groups (mean +/- SE, 446 +/- 35 and 391 +/- 35 micrograms/L; P < 0.0001, respectively) compared to those in normal age-matched prepubertal children. Similarly, IGFBP-3 levels were significantly elevated (4675 +/- 209 and 4305 +/- 162 micrograms/L, respectively; P < 0.0001) in the two groups. Treatment with GnRH analog in combination with cyproterone acetate significantly decreased height velocity and serum IGF-I and IGFBP-3 levels to normal levels after 2 yr of treatment (P < 0.0001). Serum IGF-I levels remained unchanged during monthly im treatment with long acting GnRH analog, whereas IGFBP-3 levels significantly increased during the first year of this treatment despite unmeasurable estradiol levels. Thus, in both groups, the molar ratio between IGF-I and IGFBP-3 (i.e. free biologically active IGF-I) declined concomitantly with a decrease in growth velocity. Serum levels of IGF-I and IGFBP-3 (expressed as the SD score for bone age), but not those of estradiol, correlated with height velocity before and during treatment (r = 0.34; P < 0.0001 and r = 0.24; P = 0.003, respectively). Six of the patients with a subnormal GH response to clonidine had similar IGF-I and IGFBP-3 serum levels and growth velocity compared to the other 34 girls with CPP and a normal GH response.(ABSTRACT TRUNCATED AT 400 WORDS)


NO inhibitory constituents as potential anti-neuroinflammatory agents for AD from Blumea balsamifera.
Our continuous search for new nitric oxide (NO) inhibitory substances as anti-neuroinflammatory agents for AD resulted in the isolation of one new labdane diterpenoid and three new guaiane sesquiterpenoids, as well as ten known compounds from Blumea balsamifera. Their structures were elucidated by NMR spectroscopic data analysis and the time-dependent density functional theory (TDDFT) electronic circular dichroism (ECD) calculations. The anti-neuroinflammatory effects were examined by inhibiting NO release in LPS-induced murine microglial BV-2 cells. The possible mechanism of NO inhibition of some bioactive compounds was also investigated using molecular docking, which revealed the interactions of bioactive compounds with the iNOS protein.


MHC polymorphism in Caribbean African green monkeys.
African green monkeys (AGM) are among the most widely used nonhuman primate models used in various fields of medical research. One species of AGM that originated from West Africa, Chlorocebus sabaeus, was introduced three centuries ago in the Caribbean islands. We present here a systematic study of the major histocompatibility complex (MHC) polymorphism of Caribbean AGM which is currently frequently used as an animal model. We studied 54 animals originated from Barbados (N=25) or Saint Kitts (N=29). The MHC polymorphism was characterized by means of 17 MHC microsatellites spread across MHC and DRB genotyping by DGGE sequencing. We defined nine frequent MHC haplotypes of which two were found in the two insular populations suggesting either past exchanges between the two populations or a common origin of the founders of the two populations. By the analysis of a previously described EST library, we characterized 38 MHC cDNA sequences (17 class I and 21 class II). In conclusion, we characterized for the first time the MHC polymorphism of Barbados and Saint Kitts AGM. We found a restricted polymorphism due to a founding effect, which is responsible for a strong bottleneck. The poorness of MHC polymorphism observed in the Caribbean AGM populations is similar to that observed in the Mauritian cynomolgus macaque population.


Hybrid structural model of the complete human ESCRT-0 complex.
The human Hrs and STAM proteins comprise the ESCRT-0 complex, which sorts ubiquitinated cell surface receptors to lysosomes for degradation. Here we report a model for the complete ESCRT-0 complex based on the crystal structure of the Hrs-STAM core complex, previously solved domain structures, hydrodynamic measurements, and Monte Carlo simulations. ESCRT-0 expressed in insect cells has a hydrodynamic radius of RH = 7.9 nm and is a 1:1 heterodimer. The 2.3 Angstroms crystal structure of the ESCRT-0 core complex reveals two domain-swapped GAT domains and an antiparallel two-stranded coiled-coil, similar to yeast ESCRT-0. ESCRT-0 typifies a class of biomolecular assemblies that combine structured and unstructured elements, and have dynamic and open conformations to ensure versatility in target recognition. Coarse-grained Monte Carlo simulations constrained by experimental RH values for ESCRT-0 reveal a dynamic ensemble of conformations well suited for diverse functions.


Aberrant recombination events in B cell lines derived from a kappa-deficient human.
We have analyzed the structure of Ig kappa chain genes in B cell lines derived from a human individual who cannot synthesize any kappa chains, and whose Igs all contain lambda chains (1). We have characterized secondary DNA recombination events at two kappa alleles which have undergone misaligned V-J recombinations. One such secondary recombination has joined the flanking sequences of a V kappa and a J kappa 2 gene segment as if it were the reciprocal product of a V-J kappa 2 recombination, and resulted in the displacement of the recombined VJ kappa 1 gene segments from the C kappa locus. The non-rearranged form of the V kappa fragment which had recombined with the J kappa 2 flank was cloned. Nucleotide sequencing of this fragment identified a V kappa gene that differed by at least 38% from all previously sequenced human V kappa genes. The other V-J kappa segment analyzed has undergone a secondary recombination at a different site from that described above, at a site within the intervening sequence between the J kappa and C kappa gene segments, similar to the location of secondary recombinations which have occurred in lambda + B cell lines from mice and humans (2,3). These results prove that multiple recombinations can occur at one J kappa-C kappa locus.


Short-term and long-term cardiovascular risk, metabolic syndrome and HIV in Tanzania.
To compare short-term and long-term cardiovascular disease (CVD) risk scores and prevalence of metabolic syndrome in HIV-infected adults receiving and not receiving antiretroviral therapy (ART) to HIV-negative controls. A cross-sectional study including 151 HIV-infected, ART-naive, 150 HIV-infected on ART and 153 HIV-negative adults. Traditional cardiovascular risk factors were determined by standard investigations. The primary outcome was American College of Cardiology/American Heart Association Atherosclerotic CVD (ASCVD) Risk Estimator lifetime CVD risk score. Secondary outcomes were ASCVD 10-year risk, Framingham risk scores, statin indication and metabolic syndrome. Compared with HIV-negative controls, more HIV-infected adults on ART were classified as high lifetime CVD risk (34.7% vs 17.0%, p<0.001) although 10-year risk scores were similar, a trend which was similar across multiple CVD risk models. In addition, HIV-infected adults on ART had a higher prevalence of metabolic syndrome versus HIV-negative controls (21.3% vs 7.8%, p=0.008), with two common clusters of risk factors. More than one-quarter (28.7%) of HIV-infected Tanzanian adults on ART meet criteria for statin initiation. HIV-infected ART-treated individuals have high lifetime cardiovascular risk, and this risk seems to develop rapidly in the first 3-4 years of ART as does the development of clusters of metabolic syndrome criteria. These data identify a new subgroup of low short-term/high-lifetime risk HIV-infected individuals on ART who do not currently meet criteria for CVD risk factor modification but require further study.


Current Promises and Limitations of Combined Virtual Reality and Functional Magnetic Resonance Imaging Research in Humans: A Commentary on Huffman and Ekstrom (2019).
Real-world navigation requires movement of the body through space, producing a continuous stream of visual and self-motion signals, including proprioceptive, vestibular, and motor efference cues. These multimodal cues are integrated to form a spatial cognitive map, an abstract, amodal representation of the environment. How the brain combines these disparate inputs and the relative importance of these inputs to cognitive map formation and recall are key unresolved questions in cognitive neuroscience. Recent advances in virtual reality technology allow participants to experience body-based cues when virtually navigating, and thus it is now possible to consider these issues in new detail. Here, we discuss a recent publication that addresses some of these issues (D. J. Huffman and A. D. Ekstrom. A modality-independent network underlies the retrieval of large-scale spatial environments in the human brain. Neuron, 104, 611-622, 2019). In doing so, we also review recent progress in the study of human spatial cognition and raise several questions that might be addressed in future studies.


Flooding and mental health: a systematic mapping review.
Floods are the most common type of global natural disaster. Floods have a negative impact on mental health. Comprehensive evaluation and review of the literature are lacking. To systematically map and review available scientific evidence on mental health impacts of floods caused by extended periods of heavy rain in river catchments. We performed a systematic mapping review of published scientific literature in five languages for mixed studies on floods and mental health. PUBMED and Web of Science were searched to identify all relevant articles from 1994 to May 2014 (no restrictions). The electronic search strategy identified 1331 potentially relevant papers. Finally, 83 papers met the inclusion criteria. Four broad areas are identified: i) the main mental health disorders-post-traumatic stress disorder, depression and anxiety; ii] the factors associated with mental health among those affected by floods; iii) the narratives associated with flooding, which focuses on the long-term impacts of flooding on mental health as a consequence of the secondary stressors; and iv) the management actions identified. The quantitative and qualitative studies have consistent findings. However, very few studies have used mixed methods to quantify the size of the mental health burden as well as exploration of in-depth narratives. Methodological limitations include control of potential confounders and short-term follow up. Floods following extreme events were excluded from our review. Although the level of exposure to floods has been systematically associated with mental health problems, the paucity of longitudinal studies and lack of confounding controls precludes strong conclusions. We recommend that future research in this area include mixed-method studies that are purposefully designed, using more rigorous methods. Studies should also focus on vulnerable groups and include analyses of policy and practical responses.


One-step synthesis of hierarchical [B]-ZSM-5 using cetyltrimethylammonium bromide as mesoporogen.
One-step facile synthesis of boron containing ZSM-5 microspheres is developed using 1,6-diaminohexane as the structure-directing agent and cetyltrimethylammonium bromide as the mesoporogen. High boron incorporation up to Si/B ratio of 38 is achieved and evidenced by the stretching vibrations of B-O-Si at 667 cm-1 and 917 cm-1 using Fourier-transform infrared spectra. The morphology of the crystals resembles berry-like spheres with sizes approximately 15 μm, which is composed of aggregated nanocrystals having sizes around 450 nm, is observed using scanning electron microscopy. The textural properties, i.e. the surface areas and pore volumes are investigated using N2 adsorption at -196 °C. t-plot micropore volume of 0.11 cm3/g and mesopore volume of 0.14 cm3/g are obtained applying this synthesis method for mesopores having pore diameters within the range of 2-10 nm.


Acute consciousness disorders in intensive care medicine - value of its grading for prognostic conclusion.
To understand consciousness we have to understand the mechanism of its function, which is to effectively organize sensory inputs from our environment. Consciousness is the basic, essential outcome of the process of organizing these sensory inputs, resulting in cognitive, mental, emotional, executive, instinctual or other marginally aware states. This reciprocal process of the CNS implies that organization is an act, which precedes consciousness, i.e. preconscious function. Most scientific explanations portray consciousness as an "emergent property" of classical computer-like activities in the brains neural networks. Doctors at ICU work daily with patients with altered human consciousness. Therefore, they must recognize and manage it skilfully and use adequate approaches for definite solutions. We observed a series of patients with traumatic and non traumatic brain injuries admitted to the ICU. The quality of life of these patients during the course of intensive care was very elementary and the final outcome GCS (oGCS) for future life was defined as a comatose state or apallic state, very rarely was it restored to premorbid condition as far as lucidity, attention, cognition, and executive functions. We found that a significant oGCS increase in relation to condition at admission or intake GCS (iGCS) in the group with 184 patients total (p<0.00001), in cardio-pulmonary resuscitation (CPR), traumatic brain injury (TBI) subgroups (p<0.00001) and in spontaneous haemorrhage (SH) (p<0.05) represents the only basic prerequisite for further improvement. It is not easy to find good therapeutic approaches after traumatic and non traumatic brain injury. A statistically significant oGCS increase in relation to iGCS due to quite intense medical care and keeping disclosed the state of unconsciousness with further probable evolution through the following possible ways: death in fluent comatose state, delirium and awakening, delirium ending in death, direct awakening from comatose state. Therefore significantly increased oGCS is the only basic prerequisite for pragmatically optimal "quality of life" in the course of later life. We raise general questions for both scientists and clinicians that will assist in their efforts to understand the basic endogenous conscious biological processes, their pathological changes and the links between them.


Failure to proliferate and mitotic arrest of CDK11(p110/p58)-null mutant mice at the blastocyst stage of embryonic cell development.
The CDK11(p110) protein kinases are part of large-molecular-weight complexes that also contain RNA polymerase II, transcriptional elongation factors, and general pre-mRNA splicing factors. CDK11(p110) isoforms may therefore couple transcription and pre-mRNA splicing by their effect(s) on certain proteins required for these processes. The CDK11(p58) kinase isoform is generated from the CDK11(p110) mRNA through the use of an internal ribosome entry site in a mitosis-specific manner, suggesting that this kinase may regulate the cell cycle during mitosis. The in vivo role and necessity of CDK11(p110/p58) kinase function during mammalian development were examined by generating CDK11(p110/p58)-null mice through targeted disruption of the corresponding gene using homologous recombination. While heterozygous mice develop normally, disruption of both CDK11(p110/p58) alleles results in early embryonic lethality due to apoptosis of the blastocyst cells between 3.5 and 4 days postcoitus. Cells within these embryos exhibit both proliferative defect(s) and a mitotic arrest. These results are consistent with the proposed cellular functions of the CDK11(p110/p58) kinases and confirm that the CDK11(p110/p58) kinases are essential for cellular viability as well as normal early embryonic development.


What cancer patients actually know regarding medical cannabis? A cross-sectional survey with a critical analysis of the current attitudes.
In Italy medical cannabis is a prescription drug since 1998. Even though it could not be considered a therapy as such, it is indicated as a symptomatic treatment also in cancer patients, to cure iatrogenic nausea/vomiting and chronic pain. We conducted a knowledge survey about medical cannabis among cancer patients referred to two outpatient cancer care centers and a home care service. From February to April 2018, 232 patient were enrolled; 210 patients were on active disease-oriented treatment (90.5%), while 22 (9.5%) not. Eighty-one percent of the patients have heard about medical cannabis, but only 2% from healthcare professionals. Thirty-four percent of responders thought about using cannabis to treat one or more of their own health problems, especially pain (55%). Despite that, 18% of the participants believe that medical cannabis could have negative effects on their own symptoms. Patients with high educational level better knew cannabis (odds ratio = 3.52; 95% confidence interval: 1.07-11.53), and medical cannabis (odds ratio = 3.21; 95% confidence interval: 1.48-6.98), when compared to patient with low educational level. Patients who were on active disease-oriented treatment better knew medical cannabis (odds ratio = 3.91; 95% confidence interval: 1.26-12.11) compared to "out of treatment" patients. Metastatic patients were less informed about medical cannabis compared to patients on adjuvant treatment. Our survey shows that most of Italian cancer patients know medical cannabis and a third of them have considered using cannabis to treat one (or more) of their own health problems. In the same time, they are poorly informed and do not tend to ask for information about medical cannabis to healthcare professionals.


Missed nursing care in newborn units: a cross-sectional direct observational study.
Improved hospital care is needed to reduce newborn mortality in low/middle-income countries (LMIC). Nurses are essential to the delivery of safe and effective care, but nurse shortages and high patient workloads may result in missed care. We aimed to examine nursing care delivered to sick newborns and identify missed care using direct observational methods. A cross-sectional study using direct-observational methods for 216 newborns admitted in six health facilities in Nairobi, Kenya, was used to determine which tasks were completed. We report the frequency of tasks done and develop a nursing care index (NCI), an unweighted summary score of nursing tasks done for each baby, to explore how task completion is related to organisational and newborn characteristics. Nursing tasks most commonly completed were handing over between shifts (97%), checking and where necessary changing diapers (96%). Tasks with lowest completion rates included nursing review of newborns (38%) and assessment of babies on phototherapy (15%). Overall the mean NCI was 60% (95% CI 58% to 62%), at least 80% of tasks were completed for only 14% of babies. Private sector facilities had a median ratio of babies to nurses of 3, with a maximum of 7 babies per nurse. In the public sector, the median ratio was 19 babies and a maximum exceeding 25 babies per nurse. In exploratory multivariable analyses, ratios of ≥12 babies per nurse were associated with a 24-point reduction in the mean NCI compared with ratios of ≤3 babies per nurse. A significant proportion of nursing care is missed with potentially serious effects on patient safety and outcomes in this LMIC setting. Given that nurses caring for fewer babies on average performed more of the expected tasks, addressing nursing is key to ensuring delivery of essential aspects of care as part of improving quality and safety.


Diversity of aggregation motifs in gold(i) dithiocarboxylate complexes.
Neutral, homoleptic gold(i) dithiocarboxylate complexes with different-sized Me, iPr, Mes, Tip, Mes* and Dmp substituents at the dithiocarboxylate unit were synthesized by reacting the corresponding dithiocarboxylic acid with potassium dicyanoaurate(i). They were characterized by IR spectroscopy and elemental analyses and their structures were determined by single crystal X-ray diffraction. The dinuclear complexes with an intramolecular d10-d10 type aurophilic interaction aggregate through additional intermolecular aurophilic interactions in different binding motifs. The molecular structures and the patterns of aggregation in the solid state are presented and discussed with respect to the influence of the substituent size.


Asthmatic airway epithelium is intrinsically inflammatory and mitotically dyssynchronous.
Asthma is an inflammatory condition for which anti-inflammatory glucocorticoids are the standard of care. However, similar efficacy has not been shown for agents targeting inflammatory cells and pathways. This suggests a noninflammatory cell contributor (e.g., epithelium) to asthmatic inflammation. Herein, we sought to define the intrinsic and glucocorticoid-affected properties of asthmatic airway epithelium compared with normal epithelium. Human primary differentiated normal and asthmatic airway epithelia were cultured in glucocorticoid-free medium beginning at -48 hours. They were pulsed with dexamethasone (20 nM) or vehicle for 2 hours at -26, -2, +22, and +46 hours. Cultures were mechanically scrape-wounded at 0 hours and exposed continuously to bromodeoxyuridine (BrdU). Cytokine secretions were analyzed using cytometric bead assays. Wound regeneration/mitosis was analyzed by microscopy and flow cytometry. Quiescent normal (n = 3) and asthmatic (n = 6) epithelia showed similar minimal inflammatory cytokine secretion and mitotic indices. After wounding, asthmatic epithelia secreted more basolateral TGF-β1, IL-10, IL-13, and IL-1β (P < 0.05) and regenerated less efficiently than normal epithelia (+48 h wound area reduction = [mean ± SEM] 50.2 ± 7.5% versus 78.6 ± 7.7%; P = 0.02). Asthmatic epithelia showed 40% fewer BrdU(+) cells at +48 hours (0.32 ± 0.05% versus 0.56 ± 0.07% of total cells; P = 0.03), and those cells were more dyssynchronously distributed along the cell cycle (52 ± 10, 25 ± 4, 23 ± 7% for G1/G0, S, and G2/M, respectively) than normal epithelia (71 ± 1, 12 ± 2, and 17 ± 2% for G1/G0, S, and G2/M, respectively). Dexamethasone pulses improved asthmatic epithelial inflammation and regeneration/mitosis. In summary, we show that inflammatory/fibrogenic cytokine secretions are correlated with dyssynchronous mitosis upon injury. Intermittent glucocorticoids simultaneously decreased epithelial cytokine secretions and resynchronized mitosis. These data, generated in an airway model lacking inflammatory cells, support the concept that epithelium contributes to asthmatic inflammation.


The challenge of viral snRNPs.
Some gammaherpesviruses encode nuclear noncoding RNAs (ncRNAs) that assemble with host proteins. Their conservation and abundance implies that they serve important functions for the virus. This paper focuses on our studies of three classes of nuclear noncoding herpesvirus RNAs. (1) EBERs 1 and 2 are expressed by Epstein-Barr virus in latent infection of human B lymphocytes. Recent studies revealed three sites on EBER1 that associate with ribosomal protein L22. In addition, heterokaryon assays have definitively shown that both EBERs are confined to the nucleus, arguing that their contribution to viral latency is purely nuclear. (2) HSURs 1-7 are U RNAs encoded by Herpesvirus saimiri, which causes aggressive T-cell leukemias and lymphomas. Comparison of monkey T cells transformed with wild-type or mutant virus lacking HSURs 1 and 2 revealed significant changes in host mRNAs implicated in T-cell signaling. (3) PAN is a 1-kb polyadenylated RNA that accumulates in the nucleus of Kaposi's sarcoma-associated herpesvirus lytically infected cells. A novel element, the ENE, is essential for its high accumulation. Recent results indicate that the ENE functions to counteract poly(A)-dependent RNA degradation, which we propose contributes to nuclear surveillance of mRNA transcripts in mammalian cells. Continuing studies of these viral RNAs will provide insights into both cellular and viral gene expression.


Non-dipper status and left ventricular hypertrophy as predictors of incident chronic kidney disease.
We have hypothesized that non-dipper status and left ventricular hypertrophy (LVH) are associated with the development of chronic kidney disease (CKD) in non-diabetic hypertensive patients. This study included 102 patients with an estimated glomerular filtration rate (eGFR) ≥ 60 mL/min/1.73 m(2). Ambulatory blood pressure monitoring and echocardiography were performed at the beginning of the study, and the serum creatinine levels were followed. During the average follow-up period of 51 months, CKD developed in 11 patients. There was a significant difference in the incidence of CKD between dippers and non-dippers (5.0% vs 19.0%, P < 0.05). Compared to patients without CKD, patients with incident CKD had a higher urine albumin/creatinine ratio (52.3 ± 58.6 mg/g vs 17.8 ± 29.3 mg/g, P < 0.01), non-dipper status (72.7% vs 37.4%, P < 0.05), the presence of LVH (27.3% vs 5.5%, P < 0.05), and a lower serum HDL-cholesterol level (41.7 ± 8.3 mg/dL vs 50.4 ± 12.4 mg/dL, P < 0.05). Based on multivariate Cox regression analysis, non-dipper status and the presence of LVH were independent predictors of incident CKD. These findings suggest that non-dipper status and LVH may be the therapeutic targets for preventing the development of CKD in non-diabetic hypertensive patients.


Regression analysis using dependent Polya trees.
Many commonly used models for linear regression analysis force overly simplistic shape and scale constraints on the residual structure of data. We propose a semiparametric Bayesian model for regression analysis that produces data-driven inference by using a new type of dependent Polya tree prior to model arbitrary residual distributions that are allowed to evolve across increasing levels of an ordinal covariate (e.g., time, in repeated measurement studies). By modeling residual distributions at consecutive covariate levels or time points using separate, but dependent Polya tree priors, distributional information is pooled while allowing for broad pliability to accommodate many types of changing residual distributions. We can use the proposed dependent residual structure in a wide range of regression settings, including fixed-effects and mixed-effects linear and nonlinear models for cross-sectional, prospective, and repeated measurement data. A simulation study illustrates the flexibility of our novel semiparametric regression model to accurately capture evolving residual distributions. In an application to immune development data on immunoglobulin G antibodies in children, our new model outperforms several contemporary semiparametric regression models based on a predictive model selection criterion.


Surface dose reduction from bone interface in kilovoltage X-ray radiation therapy: a Monte Carlo study of photon spectra.
This study evaluated the dosimetric impact of surface dose reduction due to the loss of backscatter from the bone interface in kilovoltage (kV) X-ray radiation therapy. Monte Carlo simulation was carried out using the EGSnrc code. An inhomogeneous phantom containing a thin water layer (0.5-5 mm) on top of a bone (thickness = 1 cm) was irradiated by a clinical 105 kVp photon beam produced by a Gulmay D3225 X-ray machine. Field sizes of 2, 5, and 10 cm diameter and source-to-surface distance of 20 cm were used. Surface doses for different phantom configurations were calculated using the DOSXYZnrc code. Photon energy spectra at the phantom surface and bone were determined according to the phase-space files at the particle scoring planes which included the multiple crossers. For comparison, all Monte Carlo simulations were repeated in a phantom with the bone replaced by water. Surface dose reduction was found when a bone was underneath the water layer. When the water thickness was equal to 1 mm for the circular field of 5 cm diameter, a surface dose reduction of 6.3% was found. The dose reduction decreased to 4.7% and 3.4% when the water thickness increased to 3 and 5 mm, respectively. This shows that the impact of the surface dose uncertainty decreased while the water thickness over the bone increased. This result was supported by the decrease in relative intensity of the lower energy photons in the energy spectrum when the water layer was with and over the bone, compared to without the bone. We concluded that surface dose reduction of 7.8%-1.1% was found when the water thickness increased from 0.5-5 mm for circular fields with diameters ranging from 2-10 cm. This decrease of surface dose results in an overestimation of prescribed dose at the patient's surface, and might be a concern when using kV photon beam to treat skin tumors in sites such as forehead, chest wall, and kneecap.


G Protein-coupled pH-sensing Receptor OGR1 Is a Regulator of Intestinal Inflammation.
A novel family of proton-sensing G protein-coupled receptors, including OGR1, GPR4, and TDAG8, was identified to be important for physiological pH homeostasis and inflammation. Thus, we determined the function of proton-sensing OGR1 in the intestinal mucosa. OGR1 expression in colonic tissues was investigated in controls and patients with IBD. Expression of OGR1 upon cell activation was studied in the Mono Mac 6 (MM6) cell line and primary human and murine monocytes by real-time PCR. Ogr1 knockout mice were crossbred with Il-10 deficient mice and studied for more than 200 days. Microarray profiling was performed using Ogr1 and Ogr1 (WT) residential peritoneal macrophages. Patients with IBD expressed higher levels of OGR1 in the mucosa than non-IBD controls. Treatment of MM6 cells with TNF, led to significant upregulation of OGR1 expression, which could be reversed by the presence of NF-κB inhibitors. Kaplan-Meier survival analysis showed a significantly delayed onset and progression of rectal prolapse in female Ogr1/Il-10 mice. These mice displayed significantly less rectal prolapses. Upregulation of gene expression, mediated by OGR1, in response to extracellular acidification in mouse macrophages was enriched for inflammation and immune response, actin cytoskeleton, and cell-adhesion gene pathways. OGR1 expression is induced in cells of human macrophage lineage and primary human monocytes by TNF. NF-κB inhibition reverses the induction of OGR1 expression by TNF. OGR1 deficiency protects from spontaneous inflammation in the Il-10 knockout model. Our data indicate a pathophysiological role for pH-sensing receptor OGR1 during the pathogenesis of mucosal inflammation.


Immunotherapy in Malignant Pleural Mesothelioma.
The only registered systemic treatment for malignant pleural mesothelioma (MPM) is platinum based chemotherapy combined with pemetrexed, with or without bevacizumab. Immunotherapy did seem active in small phase II trials. In this review, we will highlight the most important immunotherapy-based research performed and put a focus on the future of MPM. PD-(L)1 inhibitors show response rates between 10 and 29% in phase II trials, with a wide range in progression free (PFS) and overall survival (OS). However, single agent pembrolizumab was not superior to chemotherapy (gemcitabine or vinorelbine) in the recent published PROMISE-Meso trial in pre-treated patients. In small studies with CTLA-4 inhibitors there is evidence for response in some patients, but it fails to show a better PFS and OS compared to best supportive care in a randomized study. A combination of PD-(L)1 inhibitor with CTLA-4 inhibitor seem to have a similar response as PD-(L)1 monotherapy. The first results of combining durvalumab (PD-L1 blocking) with cisplatin-pemetrexed in the first line are promising. Another immune treatment is Dendritic Cell (DC) immunotherapy, which is recently tested in mesothelioma, shows remarkable anti-tumor activity in three clinical studies. The value of single agent checkpoint inhibitors is limited in MPM. There is an urgent need for biomarkers to select the optimal candidates for immunotherapy among MPM patients in terms of efficacy and tolerance. Results of combination checkpoint inhibitors with chemotherapy are awaiting.


EUROGIN 2011 roadmap on prevention and treatment of HPV-related disease.
The EUROGIN 2011 roadmap reviews the current burden of human papillomavirus (HPV)-related morbidity, as well as the evidence and potential practice recommendations regarding primary and secondary prevention and treatment of cancers and other disease associated with HPV infection. HPV infection causes ~600,000 cases of cancer of the cervix, vulva, vagina, anus and oropharynx annually, as well as benign diseases such as genital warts and recurrent respiratory papillomatosis. Whereas the incidence of cervical cancer has been decreasing over recent decades, the incidence of anal and oropharyngeal carcinoma, for which there are no effective screening programs, has been rising over the last couple of decades. Randomized trials have demonstrated improved efficacy of HPV-based compared to cytology-based cervical cancer screening. Defining the best algorithms to triage HPV-positive women, age ranges and screening intervals are priorities for pooled analyses and further research, whereas feasibility questions can be addressed through screening programs. HPV vaccination will reduce the burden of cervical precancer and probably also of invasive cervical and other HPV-related disease in women. Recent trials demonstrated that prophylactic vaccination also protects against anogenital HPV infection, anogenital intraepithelial lesions and warts associated with vaccine types, in males; and anal HPV infection and anal intraepithelial neoplasia in MSM. HPV-related oropharyngeal cancer could be treated less aggressively because of better survival compared to cancers of the oropharynx unrelated to HPV. Key findings in the field of cervical cancer prevention should now be translated in cost-effective strategies, following an organized approach integrating primary and secondary prevention, according to scientific evidence but adapted to the local situation with particular attention to regions with the highest burden of disease.


Patients' shame and attitudes toward discussing the results of literacy screening.
We investigated patients' willingness to have their reading ability documented in their medical records and the degree of shame and embarrassment associated with such disclosure. Structured interviews were conducted among a consecutive sample of 283 primary care patients at an urban public hospital. Patients' literacy was measured using the Rapid Estimate of Adult Literacy in Medicine (REALM). Self-report of degree of shame and embarrassment related to literacy skills was measured using an orally administered questionnaire. Fifty-one percent of patients had low literacy skills (< or =sixth grade) and 27.9% were assessed as having marginal literacy (seventh-eighth grade). Half (47.6%) of patients reading at or below the third-grade level admitted feeling ashamed or embarrassed about their difficulties reading, compared with 19.2% of those reading at the fourth-sixth-grade level and 6.5% of those reading at the seventh-eighth-grade level (p < 0.001). More than 90% of patients with low or marginal literacy reported it would be helpful for the doctor or nurse to know they did not understand some medical words. Patients with limited literacy were more likely to report feelings of shame as a result of disclosure (p < 0.05). Health care providers must recognize the potential shame patients might experience as a result of literacy screening.


Donor insemination: eugenic and feminist implications.
One concern regarding developments in genetics is that, when techniques such as genetic engineering become safe and affordable, people will use them for positive eugenics: to "improve" their offspring by enpowering them with exceptional qualities. Another is whether new reproductive technologies are being used to improve the condition of women or as the tools of a patriarchal system that appropriates female functions to itself and exploits women to further its own ends. Donor insemination is relevant to both of these issues. The degree to which people have used donor insemination in the past for positive eugenic purposes may give some insight into the likelihood of developing technologies being so used in the future. Donor insemination provides women with the opportunity to reproduce with only the most remote involvement of a man. To what degree do women take advantage of this to liberate themselves from male dominance? Through questionnaires and interviews, women who have used donor insemination disclosed their criteria for selecting sperm donors. The results are analyzed for the prevalence of positive eugenic criteria in the selection process and women's attitudes toward minimizing the male role in reproduction.


Life-Long Implications of Developmental Exposure to Environmental Stressors: New Perspectives.
The Developmental Origins of Health and Disease (DOHaD) paradigm is one of the most rapidly expanding areas of biomedical research. Environmental stressors that can impact on DOHaD encompass a variety of environmental and occupational hazards as well as deficiency and oversupply of nutrients and energy. They can disrupt early developmental processes and lead to increased susceptibility to disease/dysfunctions later in life. Presentations at the fourth Conference on Prenatal Programming and Toxicity in Boston, in October 2014, provided important insights and led to new recommendations for research and public health action. The conference highlighted vulnerable exposure windows that can occur as early as the preconception period and epigenetics as a major mechanism than can lead to disadvantageous "reprogramming" of the genome, thereby potentially resulting in transgenerational effects. Stem cells can also be targets of environmental stressors, thus paving another way for effects that may last a lifetime. Current testing paradigms do not allow proper characterization of risk factors and their interactions. Thus, relevant exposure levels and combinations for testing must be identified from human exposure situations and outcome assessments. Testing of potential underpinning mechanisms and biomarker development require laboratory animal models and in vitro approaches. Only few large-scale birth cohorts exist, and collaboration between birth cohorts on a global scale should be facilitated. DOHaD-based research has a crucial role in establishing factors leading to detrimental outcomes and developing early preventative/remediation strategies to combat these risks.


Predicting opioid misuse by chronic pain patients: a systematic review and literature synthesis.
Opioids can provide relief for people with chronic pain. However, a minority may develop aberrant drug behaviors. A critical issue is identifying "at-risk" patients. To synthesize the evidence of published strategies for identifying at-risk patients to guide clinicians' decisions and practices for prescribing opioid treatment for chronic pain patients (CPP). MEDLINE database search from 1966 to March 20, 2007, searching the bibliographies from all retrieved articles, and articles available in the authors' files. Studies were limited to human studies in the English language related to screening for predictors of aberrant drug behaviors in CPP who were prescribed long-term opioids. We included studies reviewing, developing measures, or investigating outcomes related to screening for aberrant opioid behaviors in CPP. We identified 6 published articles addressing clinician-based predictors of substance misuse of opioids and 9 published studies evaluating the predictive ability of clinical interviews and self-report measures for aberrant opioid behaviors in CPP. Several attempts have been made to develop procedures to identify at-risk patients including urine toxicology screening, structured interviews, observation, and self-report questionnaires. In general, the psychometric properties of the published questionnaires and interview protocols are weak; moreover, the samples included in the studies are often small and unrepresentative. Thus, none of them can be recommended for use with any confidence. Review of the published studies reveals that no one procedure or set of predictor variables is sufficient to identify CPP at-risk for opioid misuse or abuse. There is a scarcity of evidence regarding characteristics that predict aberrant behavior before beginning long-term opioids. Several predictors have been identified. Strong predictors include a personal history of illicit drug and alcohol abuse. Demographic factors have also been reported, but the results are not consistent. Prospective studies, especially ones with CPP who have not already been started on chronic opioid therapy, are needed.


Intracellular trafficking and cellular uptake mechanism of PHBV nanoparticles for targeted delivery in epithelial cell lines.
Nanotechnology is a science that involves imaging, measurement, modeling and a manipulation of matter at the nanometric scale. One application of this technology is drug delivery systems based on nanoparticles obtained from natural or synthetic sources. An example of these systems is synthetized from poly(3-hydroxybutyrate-co-3-hydroxyvalerate), which is a biodegradable, biocompatible and a low production cost polymer. The aim of this work was to investigate the uptake mechanism of PHBV nanoparticles in two different epithelial cell lines (HeLa and SKOV-3). As a first step, we characterized size, shape and surface charge of nanoparticles using dynamic light scattering and transmission electron microscopy. Intracellular incorporation was evaluated through flow cytometry and fluorescence microscopy using intracellular markers. We concluded that cellular uptake mechanism is carried out in a time, concentration and energy dependent way. Our results showed that nanoparticle uptake displays a cell-specific pattern, since we have observed different colocalization in two different cell lines. In HeLa (Cervical cancer cells) this process may occur via classical endocytosis pathway and some internalization via caveolin-dependent was also observed, whereas in SKOV-3 (Ovarian cancer cells) these patterns were not observed. Rearrangement of actin filaments showed differential nanoparticle internalization patterns for HeLa and SKOV-3. Additionally, final fate of nanoparticles was also determined, showing that in both cell lines, nanoparticles ended up in lysosomes but at different times, where they are finally degraded, thereby releasing their contents. Our results, provide novel insight about PHBV nanoparticles internalization suggesting that for develop a proper drug delivery system is critical understand the uptake mechanism.


Perspectives on health technology assessment: response from the patient's perspective.
Health technology assessment (HTA) involves values and judgments, but there have been few attempts to seek the views of members of the public or to ensure that they have access to the results. Patients and citizens can make an important contribution to HTA by determining priorities for assessment, designing and conducting assessments and appraisals, receiving and using the findings, and engaging in debates about policy priorities and rationing. Those responsible for HTA should make greater efforts to involve the public and ensure that the findings are accessible to patients for use when making treatment choices.


Placental infection by Zika virus in French Guiana.
To describe placental findings on prenatal ultrasound and anatomopathological examination in women with Zika virus (ZIKV) infection, and to assess their association with congenital ZIKV infection and severe adverse outcome, defined as fetal loss or congenital Zika syndrome (CZS). This was a prospective study of pregnancies undergoing testing for maternal ZIKV infection at a center in French Guiana during the ZIKV epidemic. In ZIKV-positive women, congenital infection was defined as either a positive reverse transcription polymerase chain reaction result or identification of ZIKV-specific immunoglobulin-M in at least one placental, fetal or neonatal sample. Placental ZIKV-infection status was classified as non-exposed (placentae from non-infected women), exposed (placentae from ZIKV-infected women without congenital infection) or infected (placentae from ZIKV-infected women with proven congenital infection). Placentae were assessed by monthly prenatal ultrasound examinations, measuring placental thickness and umbilical artery Doppler parameters, and by anatomopathological examination after live birth or intrauterine death in women with ZIKV infection. The association of placental thickness during pregnancy and anatomopathological findings with the ZIKV status of the placenta was assessed. The association between placental findings and severe adverse outcome (CZS or fetal loss) in the infected group was also assessed. Among 291 fetuses/neonates/placentae from women with proven ZIKV infection, congenital infection was confirmed in 76 cases, of which 16 resulted in CZS and 11 resulted in fetal loss. The 215 remaining placentae from ZIKV-positive women without evidence of congenital ZIKV infection represented the exposed group. A total of 334 placentae from ZIKV-negative pregnant women represented the non-exposed control group. Placentomegaly (placental thickness > 40 mm) was observed more frequently in infected placentae (39.5%) than in exposed placentae (17.2%) or controls (7.2%), even when adjusting for gestational age at diagnosis and comorbidities (adjusted hazard ratio (aHR), 2.02 (95% CI, 1.22-3.36) and aHR, 3.23 (95% CI, 1.86-5.61), respectively), and appeared earlier in infected placentae. In the infected group, placentomegaly was observed more frequently in cases of CZS (62.5%) or fetal loss (45.5%) than in those with asymptomatic congenital infection (30.6%) (aHR, 5.43 (95% CI, 2.17-13.56) and aHR, 4.95 (95% CI, 1.65-14.83), respectively). Abnormal umbilical artery Doppler was observed more frequently in cases of congenital infection resulting in fetal loss than in those with asymptomatic congenital infection (30.0% vs 6.1%; adjusted relative risk (aRR), 4.83 (95% CI, 1.09-20.64)). Infected placentae also exhibited a higher risk for any pathological anomaly than did exposed placentae (62.8% vs 21.6%; aRR, 2.60 (95% CI, 1.40-4.83)). Early placentomegaly may represent the first sign of congenital infection in ZIKV-infected women, and should prompt enhanced follow-up of these pregnancies. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.


Rapid species identification and epidemiological analysis of carbapenem-resistant Acinetobacter spp. by a PCR-based open reading frame typing method.
The spread of carbapenem-resistant Acinetobacter spp. has become a global problem. In this study, 18 carbapenem-resistant Acinetobacter calcoaceticus-baumannii (ACB) complexes, identified using a conventional biochemical method at our hospital during 2004-2013, were studied for species identification and epidemiological analyses. Species identification was performed using matrix-assisted laser desorption ionization-time-of-flight MS, a partial sequence analysis of rpoB and a PCR-based ORF typing (POT) method. The POT method can not only identify the species of ACB complexes but also simultaneously determine the international epidemic clones and the genetic identities of Acinetobacterbaumannii in several hours. Carbapenem resistance gene detection by PCR, molecular epidemiological analysis by PFGE and Pasteur Institute multilocus sequence typing (MLST) analysis were performed. All three methods identified 18 isolates as A. baumannii (n=10), Acinetobacterpittii (n=4) and Acinetobacternosocomialis (n=4). A metallo-β-lactamase gene in all strains of A. pittii and A. nosocomialis and an ISAba1 gene in the upstream of the blaOXA-51-like gene in eight strains of A. baumannii were detected, respectively, as carbapenemase-related genes. Results from PFGE demonstrated that nine strains of A. baumannii were closely related genetically. Results of MLST analysis showed that A. baumannii are classifiable to sequence type 2. These results were consistent with those obtained using the POT method. This POT method can easily and rapidly identify the international epidemic clones and the identities of A. baumannii. It can be a useful tool for infection control.


What you don't know can hurt you: adverse psychologic effects in members of BRCA1-linked and BRCA2-linked families who decline genetic testing.
To identify members of hereditary breast and ovarian cancer families who are at risk for adverse psychologic effects of genetic testing. A prospective cohort study with baseline (preeducation) assessments of predictor variables (ie, sociodemographic factors, cancer history, and cancer-related stress symptoms) was performed. The primary outcome variable (presence of depressive symptoms) was assessed at baseline and at 1- and 6-month follow-up evaluations. Participants were 327 adult male and female members of BRCA1- and BRCA2-linked hereditary breast and ovarian cancer families, who were identified as carriers, noncarriers, or decliners of genetic testing. The presence of cancer-related stress symptoms at baseline was strongly predictive of the onset of depressive symptoms in family members who were invited but declined testing. Among persons who reported high baseline levels of stress, depression rates in decliners increased from 26% at baseline to 47% at 1-month follow-up; depression rates in noncarriers decreased and in carriers showed no change (odds ratio [OR] for decliners v noncarriers=8.0; 95% confidence interval [CI], 1.9 to 33.5; P=.0004). These significant differences in depression rates were still evident at the 6-month follow-up evaluation (P=.04). In BRCA1/2-linked families, persons with high levels of cancer-related stress who decline genetic testing may be at risk for depression. These family members may benefit from education and counseling, even if they ultimately elect not to be tested, and should be monitored for potential adverse effects.


Prognostic factors of all-cause mortalities in continuous ambulatory peritoneal dialysis: a cohort study.
The role of small solute clearance on mortalities in patients with CAPD has been controversial. We therefore conducted a study with 3 years' follow up in adult patients who participated in the CAPD-first policy. There were 11,523 patients with end-stage renal disease who participated in the CAPD-first policy between 2008 and 2011. Among them, 1,177 patients were included in the retrospective cohort study. A receiver operating characteristic curve was applied to calibrate the cutoffs of tKt/V, rKt/V and tCrcl. Kaplan-Meier and Cox-regression models with time varying covariates were applied to estimate overall death rate, probability of death and prognosis, respectively. The cutoffs of rKt/V and tKt/V were 0.25 and 1.75, respectively. The Cox regression suggested that the higher these clearance parameters, the lower the risks of death after adjusting for covariables. The risks of death for those above these cutoffs were 57% (HR = 0.43, 95% CI: 0.31, 0.60) and 29% (HR = 0.71, 95% CI: 0.52, 0.98) lower for rKt/V and tKt/V, respectively. Age, serum albumin, hemoglobin, systolic blood pressure, and ultra-filtration volume significantly affected the mortality outcome. Our study suggested that the cutoffs of 0.25 and 1.75 for rKt/V and tKt/V might be associated with mortality in CAPD patients. A minimum tKt/V of 1.75 should be targeted, but increased dialysis dosage to achieve tKt/V > 2.19 adds no further benefit. Serum albumin, hemoglobin, SBP, and UF volume are also associated with mortality. However, our study may face with selection and other unobserved confounders, so further randomized controlled trials are required to confirm these cutoffs.


Laminar alterations in gamma-aminobutyric acidA, muscarinic, and beta adrenoceptors and neuron degeneration in cingulate cortex in Alzheimer's disease.
The laminar distribution of binding to a number of postsynaptic neurotransmitter receptors was assessed autoradiographically in postmortem samples of area 23a in posterior cingulate cortex from 13 Alzheimer and nine age-matched control cases. Specific binding in all Alzheimer cases was compared to that in control cases, and the following alterations were observed: reduced muscimol binding in most layers; no changes in pirenzepine binding; and elevated cyanopindolol binding in layers Ic, IIIc, and IV. The Alzheimer cases were classified further on the basis of neuronal degeneration: class 1, no neuron loss; class 2, greatest losses in layer II or III; class 3, greatest losses in layer IV; and class 4, greatest losses in layer V or VI. This classification uncovered further alterations in ligand binding patterns. First, muscimol binding was reduced in layers II and III only in class 2 cases and in layers V and VI only in class 4 cases. Second, pirenzepine binding was reduced in layers Ic, IIIa-b, and VI of class 1 cases and layers Va and VI of class 4 cases. In spite of neuron degeneration in classes 2 and 3, there was no change in pirenzepine binding in these classes. Third, elevated cyanopindolol binding occurred in classes 3 and 4, whereas classes 1 and 2 had normal levels of binding. These results suggest that cases of Alzheimer's disease express heterogeneities in neocortical pathology which are reflected in the laminar patterns of binding to postsynaptic receptors. Reductions in muscimol binding to the gamma-aminobutyric acidA receptor had the closest relationship with neuron degeneration, whereas pirenzepine binding appeared to reflect a compensation in muscarinic receptors for changes in neuron densities.


How do patients use antimalarial drugs? A review of the evidence.
Patient adherence is a major determinant of the therapeutic response to antimalarial drugs, as most treatments are taken at home without medical supervision. With the introduction of new, effective, but more expensive antimalarials, there is concern that the high levels of efficacy observed in clinical trials may not be translated into effectiveness in the normal context of use. We reviewed available published evidence on adherence to antimalarial drugs and community drug usage; 24 studies were identified of which nine were 'intervention' studies, seven were classified as 'outcome studies', and the remainder were purely descriptive studies of antimalarial adherence. Definitions, methods, and results varied widely. Adherence was generally better when treatments were effective, and was improved by interventions focusing on provider knowledge and behaviour, packaging, and provision of correct dosages. There is insufficient information on this important subject, and current data certainly do not justify extrapolation from results with ineffective drugs to new effective treatments. Research in this area would benefit from of standardization of methodologies and the application of pharmacokinetic modelling.


Unraveling the complexity of protein backbone dynamics with combined (13)C and (15)N solid-state NMR relaxation measurements.
Typically, protein dynamics involve a complex hierarchy of motions occurring on different time scales between conformations separated by a range of different energy barriers. NMR relaxation can in principle provide a site-specific picture of both the time scales and amplitudes of these motions, but independent relaxation rates sensitive to fluctuations in different time scale ranges are required to obtain a faithful representation of the underlying dynamic complexity. This is especially pertinent for relaxation measurements in the solid state, which report on dynamics in a broader window of time scales by more than 3 orders of magnitudes compared to solution NMR relaxation. To aid in unraveling the intricacies of biomolecular dynamics we introduce (13)C spin-lattice relaxation in the rotating frame (R1ρ) as a probe of backbone nanosecond-microsecond motions in proteins in the solid state. We present measurements of (13)C'R1ρ rates in fully protonated crystalline protein GB1 at 600 and 850 MHz (1)H Larmor frequencies and compare them to (13)C'R1, (15)N R1 and R1ρ measured under the same conditions. The addition of carbon relaxation data to the model free analysis of nitrogen relaxation data leads to greatly improved characterization of time scales of protein backbone motions, minimizing the occurrence of fitting artifacts that may be present when (15)N data is used alone. We also discuss how internal motions characterized by different time scales contribute to (15)N and (13)C relaxation rates in the solid state and solution state, leading to fundamental differences between them, as well as phenomena such as underestimation of picosecond-range motions in the solid state and nanosecond-range motions in solution.


Adherence to Antiretroviral Therapy and Its Effect on Survival of HIV-Infected Individuals in Jharkhand, India.
Research in India has extensively examined the factors associated with non-adherence to antiretroviral therapy (ART) with limited focus on examining the relationship between adherence to ART regimen and survival status of HIV infected patients. This study examines the effect of optimal adherence to ART on survival status of HIV infected patients attending ART centers in Jharkhand, India. Data from a cohort of 239 HIV infected individuals who were initiated ART in 2007 were compiled from medical records retrospectively for 36 months. Socio-demographic characteristics, CD4 T cell count, presence of opportunistic infections at the time of ART initiation and ART regimen intake and survival status was collected periodically. Optimal adherence was assessed using pill count methods; patients who took <95% of the specified regimens were identified as non-adherent. Cox-proportional hazard model was used to determine the relative hazards of mortality. More than three-fourths of the patients were male, on an average 34 year old and median CD4 T cell count was 118 cells/cmm at the time of ART registration. About 57% of the patients registered for ART were found to be adherent to ART. A total of 104 patients died in 358.5 patient-years of observation resulting in a mortality rate of 29 per 100 patient-years (95% confidence interval (CI): 23.9-35.2) and median survival time of 6.5 months (CI: 2.7-10.9). The mortality rate was higher among patients who were non-adherent to ART (64.5, CI: 50.5-82.4) than who were adherent (15.4, CI: 11.3-21.0). The risk of mortality was fourfold higher among individuals who were non-adherent to ART than who were adherent (Adjusted hazard ratio: 3.9, CI: 2.6-6.0). Adherence to ART is associated with a higher chance of survival of HIV infected patients, ascertaining the need for interventions to improve the ART adherence and early initiation of ART.


Lameness in dairy heifers; impacts of hoof lesions present around first calving on future lameness, milk yield and culling risk.
The importance of lameness in primiparous dairy heifers is increasingly recognised. Although it is accepted that clinical lameness in any lactation increases the risk of future lameness, the impact of foot lesions during the first lactation on long-term lameness risk is less clear. This retrospective cohort study aimed to investigate the impacts of foot lesions occurring around the time of first calving in heifers on future lameness risk, daily milk yield and survival within a dairy herd. Records were obtained for 158 heifers from one UK dairy herd. Heifers were examined in 2 month blocks from 2 months pre-calving through to 4 months post-calving. Sole lesions and white line lesions were scored on a zero to 10 scale and digital dermatitis on a zero to 3 scale. Outcomes investigated were; lameness risk based on weekly locomotion scores, average daily milk yield and culling risk. Mixed effect models were used to investigate associations between maximum lesion scores and outcomes. Lesion scores in the highest score categories for claw horn lesions (sole lesions and white line lesions) in the 2 to 4 month post-calving period were associated with an increased risk of future lameness; heifers with white line lesion scores ≥3 compared with scores zero to 1 and heifers with sole lesion scores ≥4 compared with score 2, at this time point, had a predicted increased risk of future lameness of 1.6 and 2.6 respectively. Sole lesions ≥4 were also associated with a reduction in average daily milk yield of 2.68kg. Managing heifers to reduce claw horn lesions during this time period post-calving may provide health, welfare and production benefits for the long-term future of those animals. A novel finding from the study was that mild lesion scores compared with scores zero to 1, were associated with a reduced risk of future lameness for white line lesions and sole lesions occurring in the pre-calving or 2 to 4 months post-calving periods respectively. Mild sole lesions in the pre-calving period were also associated with a reduced risk of premature culling. One hypothesis for this result is that a mild insult may result in adaptive changes to the foot leading to greater biomechanical resilience and so increased longevity.


Colorectal cancer susceptibility loci on chromosome 8q23.3 and 11q23.1 as modifiers for disease expression in Lynch syndrome.
Recently, six colorectal cancer (CRC) susceptibility loci have been identified, and two single-nucleotide polymorphisms (SNPs)--rs16892766 (8q23.3) and rs3802842 (11q23.1)--from two of these regions have been found to be significantly associated with an increased CRC risk in patients with Lynch syndrome. The objective of this study was to genotype nine SNPs within these six loci to confirm previous findings and investigate whether they act as modifiers of disease risk in patients with Lynch syndrome. The patient cohort consisted of 684 mutation-positive patients with Lynch syndrome from 298 Australian and Polish families. Nine SNPs were genotyped: rs16892766 (8q23.3), rs7014346 and rs6983267 (8q24.21), rs10795668 (10p14), rs3802842 (11q23.1), rs10318 and rs4779584 (15q13.3), and rs4939827 and rs4464148 (18q21.1). The data were analysed to investigate possible associations between the presence of variant alleles and the risk of developing disease. An association between SNP rs3802842 on chromosome 11q23.1 and rs16892766 on chromosome 8q23.3 and the risk of developing CRC and age of diagnosis was found in MLH1 mutation carriers. Female MLH1 mutation carriers harbouring the homozygous variant genotype for SNP rs3802842 have the highest risk of developing CRC. When the number of risk alleles for the two SNPs combined was analysed, a difference of 24 years was detected between individuals carrying three risk alleles and those carrying no risk alleles. The authors were able to replicate the association between the CRC susceptibility loci on chromosomes 8q23.3 and 11q23 and the risk of developing CRC in patients with Lynch syndrome, but the association could only be detected in MLH1 mutation carriers in this study.


Impact of renin-angiotensin system inhibition on microalbuminuria in type 2 diabetes: a post hoc analysis of the Shiga Microalbuminuria Reduction Trial (SMART).
The Shiga Microalbuminuria Reduction Trial (SMART) showed the advantage of ARB over CCB beyond the blood pressure (BP)-lowering effect in reducing microalbuminuria. To further assess the impact of BP control or renin-angiotensin system inhibition on microalbuminuria, the SMART patients were re-analyzed. Hypertensive patients with type 2 diabetes and microalbuminuria were randomly assigned to valsartan or amlodipine treatment groups for 24 weeks. Target blood pressure was set at <130/80 mmHg. Changes in the urinary albumin creatinine ratio (ACR) from baseline were assessed in the valsartan monotherapy (VM) group (n=33), the amlodipine monotherapy (AM) group (n=36), the concomitant valsartan and angiotensin-converting enzyme inhibitor group (VA) (n=33), and the concomitant amlodipine and angiotensin-converting enzyme inhibitor (AA) group (n=38). At the end of the study, mean BP was not different among the four treatment groups. The changes in ACR from baseline to the end of the treatment period in VM, AM, VA, and AA were -36%, +30%, -26%, and +8%, respectively. The dissociation between the anti-albuminuric and antihypertensive effects of valsartan or amlodipine was observed in the respective monotherapy groups. In the AA group, however, a significant positive relationship was found between the changes in ACR and those in systolic BP. In conclusion, RAS inhibitors may be necessary in order for calcium channel blockers to have an effect on microalbuminuria. Therefore, RAS inhibitors are first-line drugs for hypertensive patients with type 2 diabetes and microalbuminuria.


Diagnosing 'male' depression in men diagnosed with prostate cancer: the next step in effective translational psycho-oncology interventions?
Depression in men diagnosed with prostate cancer is associated with several adverse outcomes. However, some data suggest that standard methods of assessing depression in males via the criteria for Major Depressive Disorder (MDD) may omit several extra key symptoms of male depression. Therefore, this study tested the comparative effects of standard MDD-based diagnostic criteria for depression and criteria for 'male depression' in a sample of men diagnosed with prostate cancer. 191 men diagnosed with prostate cancer completed a postal survey questionnaire containing questions about background variables, the Patient Health Questionnaire-9 for depression (PHQ9) and the Gotland Male Depression Scale (GMDS). Comparisons were made of the relative prevalence of depression according to these scales, plus a scale that combined the PHQ9 and GMDS extra items for male depression Although there were significant correlations between total PHQ9 and GMDS scores, over one-third of variance in the GMDS was not accounted for by the PHQ9, and sensitivity of the PHQ9 against the GMDS showed that about 24% of those patients identified as depressed on the GMDS would not be similarly identified on the PHQ9. Different prevalence rates from the two scales suggested that they were assessing different sets of symptoms of depression. A combined PHQ9-GMDS scale of 15 items was used to produce a profile of male depression in these patients. Adequate and reliable assessment of depression in men diagnosed with prostate cancer may require use of additional symptoms to those listed for MDD, and treatment planning and delivery could be more precise and effective using this methodology.


Nucleotide sequence of Moloney leukemia virus: 3' end reveals details of replications, analogy to bacterial transposons, and an unexpected gene.
We have determined the sequence of a cloned DNA fragment 1108 base pairs long which corresponds to the 3' end of the Moloney murine leukemia provirus. The clone was obtained as the primary product of reverse transcription and begins with the Moloney "strong stop" sequence, then extends towards the 5' end of the provirus. Our sequence: (i) proves that reverse transcriptase switches templates during minus strand synthesis; (ii) defines the limits of the 515-base-pair repeats which occupy both ends of the integrated provirus; (iii) shows that the structure of the proviral repeats has strong analogy to bacterial insertion sequences, indicating that the Moloney provirus is a transposon; (iv) identifies the putative promotor for genomic transcription within these repeats; (v) shows that the presumed origin of second strand synthesis, which lies just outside the 3' repeat, has tertiary structure analogous to single-stranded bacteriophage origins of replication; (vi) solves the amino acid sequence of most of pI5E, the carboxy-terminal product of the env gene; (vii) allows detailed mapping of the mink cell focus-forming virus substitution locus in a central location within the gp70 region of the env gene; and (viii) identifies a long open translation frame to the right of the env gene (R gene) which could be involved in leukemogenesis.


Mitogenic activity and signaling mechanism of 2-(14,15- epoxyeicosatrienoyl)glycerol, a novel cytochrome p450 arachidonate metabolite.
Arachidonic acid is an essential constituent of cell membranes that is esterified to the sn-2 position of glycerophospholipids and is released from selected phospholipid pools by tightly regulated phospholipase cleavage. Metabolism of the released arachidonic acid by the cytochrome P450 enzyme system (cP450) generates biologically active compounds, including epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetraenoic acids. Here we report that 2-(14,15-epoxyeicosatrienoyl)glycerol (2-14,15-EG), a novel cP450 arachidonate metabolite produced in the kidney, is a potent mitogen for renal proximal tubule cells. This effect is mediated by activation of tumor necrosis factor alpha-converting enzyme (ADAM17), which cleaves membrane-bound transforming growth factor alpha (proTGF-alpha) and releases soluble TGF-alpha as a ligand that binds and activates epidermal growth factor receptor (EGFR). The present studies additionally demonstrate that the structurally related 14,15-EET stimulates release of soluble heparin-binding EGF-like growth factor as an EGFR ligand by activation of ADAM9, another member of the ADAM family. Thus, in addition to the characterization of 2-14,15-EG's mitogenic activity and signaling mechanism, our study provides the first example that two structurally related biologically active lipid mediators can activate different metalloproteinases and release different EGFR ligands in the same cell type to activate EGFR and stimulate cell proliferation.


Modelling survival and mortality risk to 15 years of age for a national cohort of children with serious congenital heart defects diagnosed in infancy.
Congenital heart defects (CHDs) are a significant cause of death in infancy. Although contemporary management ensures that 80% of affected children reach adulthood, post-infant mortality and factors associated with death during childhood are not well-characterised. Using data from a UK-wide multicentre birth cohort of children with serious CHDs, we observed survival and investigated independent predictors of mortality up to age 15 years. Data were extracted retrospectively from hospital records and death certificates of 3,897 children (57% boys) in a prospectively identified cohort, born 1992-1995 with CHDs requiring intervention or resulting in death before age one year. A discrete-time survival model accounted for time-varying predictors; hazards ratios were estimated for mortality. Incomplete data were addressed through multilevel multiple imputation. By age 15 years, 932 children had died; 144 died without any procedure. Survival to one year was 79.8% (95% confidence intervals [CI] 78.5, 81.1%) and to 15 years was 71.7% (63.9, 73.4%), with variation by cardiac diagnosis. Importantly, 20% of cohort deaths occurred after age one year. Models using imputed data (including all children from birth) demonstrated higher mortality risk as independently associated with cardiac diagnosis, female sex, preterm birth, having additional cardiac defects or non-cardiac malformations. In models excluding children who had no procedure, additional predictors of higher mortality were younger age at first procedure, lower weight or height, longer cardiopulmonary bypass or circulatory arrest duration, and peri-procedural complications; non-cardiac malformations were no longer significant. We confirm the high mortality risk associated with CHDs in the first year of life and demonstrate an important persisting risk of death throughout childhood. Late mortality may be underestimated by procedure-based audit focusing on shorter-term surgical outcomes. National monitoring systems should emphasise the importance of routinely capturing longer-term survival and exploring the mechanisms of mortality risk in children with serious CHDs.


Divergent syntheses of iodinated isobenzofuranones and isochromenones by iodolactonization of 2-alkynylbenzoic acids in ionic liquids.
The regiochemical outcome of the iodolactonization of 2-alkynylbenzoic acids, carried out at 100 °C in ionic liquids (ILs) as unconventional solvents and with molecular iodine as the iodine source, in the absence of external bases, was found to be strongly dependent on the nature of the IL medium. In particular, while the use of N-ethyl-N-methylmorpholinium dicyanamide (Mor1,2N(CN)2) promoted the stereoselective formation of (E)-3-(iodomethylene)isobenzofuran-1(3H)-ones, through an anti-5-exo-dig cyclization route, the use of 1-ethyl-3-methylimidazolium ethyl sulfate (EmimEtSO4) tended to favor the 6-endo-dig cyclization mode, with preferential or selective formation of 4-iodo-1H-isochromen-1-ones. In any case, the IL solvent could be easily recycled after extraction of the product from the reaction mixture with diethyl ether. DFT calculations have been carried out to clarify the role of the IL's nature in favoring either the anti-5-exo-dig cyclization route or the 6-endo-dig mode. In the case of iodocyclization of 2-ethynylbenzoic acid, only the 5-exo-dig mode was observed in both EmimEtSO4 and Mor1,2N(CN)2 solvents. The structures of two representative products have been confirmed by X-ray diffraction analysis.


Charging C60 islands with the AFM tip.
We show that electrons can be transferred on demand from an AFM tip into single bulk-like C60 islands, which are supported on the insulating NaCl(001) surface. We exemplify this by controlled charge-manipulation experiments conducted in ultrahigh vacuum by noncontact AFM (nc-AFM), electrostatic force microscopy (EFM) and Kelvin probe force microscopy (KPFM). KPFM shows a homogeneous contrast at the islands, which is a signature for an equal distribution of the electrons in the T1u band. The charge dissipates during half a day due to an interaction of the charged C60 islands with defects in the near surface region of NaCl. Our results open the perspective in photo-voltaics to study charge attachment, stability and charge exchange with the environment of any C60 bulk-like system.


Smartphone-based colorimetric detection via machine learning.
We report the application of machine learning to smartphone-based colorimetric detection of pH values. The strip images were used as the training set for Least Squares-Support Vector Machine (LS-SVM) classifier algorithms that were able to successfully classify the distinct pH values. The difference in the obtained image formats was found not to significantly affect the performance of the proposed machine learning approach. Moreover, the influence of the illumination conditions on the perceived color of pH strips was investigated and further experiments were conducted to study the effect of color change on the learning model. Non-integer pH levels are identified as their nearest integer pH values, whereas the test results for integer pH levels using JPEG, RAW and RAW-corrected image formats captured under different lighting conditions lead to perfect classification accuracy, sensitivity and specificity, which proves that colorimetric detection using machine learning based systems is able to adapt to various experimental conditions and is a great candidate for smartphone-based sensing in paper-based colorimetric assays.


Ascites Increases Expression/Function of Multidrug Resistance Proteins in Ovarian Cancer Cells.
Chemotherapy resistance is the major reason for the failure of ovarian cancer treatment. One mechanism behind chemo-resistance involves the upregulation of multidrug resistance (MDR) genes (ABC transporters) that effectively transport (efflux) drugs out of the tumor cells. As a common symptom in stage III/IV ovarian cancer patients, ascites is associated with cancer progression. However, whether ascites drives multidrug resistance in ovarian cancer cells awaits elucidation. Here, we demonstrate that when cultured with ascites derived from ovarian cancer-bearing mice, a murine ovarian cancer cell line became less sensitive to paclitaxel, a first line chemotherapeutic agent for ovarian cancer patients. Moreover, incubation of murine ovarian cancer cells in vitro with ascites drives efflux function in these cells. Functional studies show ascites-driven efflux is suppressible by specific inhibitors of either of two ABC transporters [Multidrug Related Protein (MRP1); Breast Cancer Related Protein (BCRP)]. To demonstrate relevance of our findings to ovarian cancer patients, we studied relative efflux in human ovarian cancer cells obtained from either patient ascites or from primary tumor. Immortalized cell lines developed from human ascites show increased susceptibility to efflux inhibitors (MRP1, BCRP) compared to a cell line derived from a primary ovarian cancer, suggesting an association between ascites and efflux function in human ovarian cancer. Efflux in ascites-derived human ovarian cancer cells is associated with increased expression of ABC transporters compared to that in primary tumor-derived human ovarian cancer cells. Collectively, our findings identify a novel activity for ascites in promoting ovarian cancer multidrug resistance.


Treatment of proteins with dietary polyphenols lowers the formation of AGEs and AGE-induced toxicity.
Advanced glycation endproducts (AGEs) are a group of harmful compounds produced either endogenously or during thermal food processing. Once absorbed by humans via food intake, AGEs can cause oxidative cell damage and contribute to pathological development of various diseases. The AGE-inhibitory activity of dietary polyphenols in vitro has been extensively reported before, but the current study is pioneering in examining the antiglycation activity of five selected dietary polyphenols (phloretin, naringenin, epicatechin, chlorogenic acid, and rosmarinic acid) during the thermal protein glycation process. When added into the glucose-casein glycation model heated at 120 °C for 2 h, these polyphenols were capable of inhibiting the formation of both total fluorescent AGEs and nonfluorescent carboxymethyllysine (CML). The thermal stability and transformation of polyphenols are likely important factors affecting their antioxidant activity and inhibitory efficacy of reactive carbonyl species formation. Treatment with epicatechin would lower not only AGE formation but also AGE-induced cytotoxicity and oxidative stress to human retinal pigment epithelial (ARPE-19) cells.


Interferon gamma-activated human monocytes downregulate transferrin receptors and inhibit the intracellular multiplication of Legionella pneumophila by limiting the availability of iron.
We have investigated the role of iron in the intracellular biology of Legionella pneumophila in human monocytes and in the effector arm of cell-mediated immune defense against this intracellular bacterial pathogen. To determine if L. pneumophila intracellular multiplication is iron dependent, we studied the effect of the iron chelator deferoxamine on L. pneumophila infection of monocytes. Deferoxamine at 15 microM completely inhibited L. pneumophila intracellular multiplication. The inhibitory effect of deferoxamine was reversed with equimolar iron-saturated transferrin but not apotransferrin. To examine the potential role of iron in monocyte activation, we investigated the influence of iron-saturated transferrin on L. pneumophila multiplication in IFN gamma-activated monocytes. Iron transferrin, but not apotransferrin, neutralized the capacity of activated monocytes to inhibit L. pneumophila multiplication. To explore a potential mechanism by which activated monocytes might limit the availability of intracellular iron, we examined transferrin receptor expression on nonactivated and activated monocytes cultured in vitro for 5 d. By fluorescence-activated flow cytometry, activated monocytes exhibited markedly fewer transferrin receptors than nonactivated monocytes. By Scatchard analysis of 125I-transferrin binding to monocytes, nonactivated monocytes had 38,300 +/- 12,700 (mean +/- SE) transferrin binding sites, whereas activated monocytes had 10,300 +/- 1,600, a reduction of 73%. Activated and nonactivated monocytes had a similar mean Kd (1.8 +/- 0.2 nM). This study demonstrates that (a) L. pneumophila intracellular multiplication is iron dependent; (b) activated monocytes inhibit L. pneumophila multiplication by limiting the availability of intracellular iron; and (c) transferrin receptors are downregulated on IFN gamma-activated monocytes.


Interaction between smoking and asbestos in human lung adenocarcinoma: role of K-ras mutations.
To investigate the role of tobacco smoking and asbestos fibers in the etiology of human lung cancer, we examined the activating point mutations in the K-ras oncogene in DNA samples from 49 patients. Mutations were found more often in tissue from adenocarcinomas (12/21) than in tissue from tumors other than nonadenocarcinomas of the lung (3/28). Among the adenocarcinoma patients, asbestos exposure was predictive of K-ras mutation (odds ratio, 4.9; 95% confidence interval, 0.7-34.3); in patients with other types of lung cancer, the relation appeared to be an inverse one, but the numbers were small. The proportion of heavy smokers (over 50 pack-years) was 60% among people with K-ras mutations and 35% among the K-ras-negative subjects, suggesting that smoking causes K-ras mutations. If mutations in K-ras genes are caused by smoking, asbestos would act as a promoting agent by conferring selective growth conditions for clonal expansion on these mutated cells. Asbestos may favor recruitment of (initiated) K-ras mutation-positive cells in the multistage process of carcinogenesis by stimulating cellular growth.


Labour Market Attachment, Workplace Infection Control Procedures and Mental Health: A Cross-Sectional Survey of Canadian Non-healthcare Workers during the COVID-19 Pandemic.
The COVID-19 pandemic has led to large proportions of the labour market moving to remote work, while others have become unemployed. Those still at their physical workplace likely face increased risk of infection, compared to other workers. The objective of this paper is to understand the relationship between working arrangements, infection control programs (ICP), and symptoms of anxiety and depression among Canadian workers, not specifically working in healthcare. A convenience-based internet survey of Canadian non-healthcare workers was facilitated through various labour organizations between April 26 and June 6, 2020. A total of 5180 respondents started the survey, of which 3779 were assessed as employed in a full-time or part-time capacity on 2 March 2020 (prior to large-scale COVID-19 pandemic responses in Canada). Of this sample, 3305 (87.5%) had complete information on main exposures and outcomes. Anxiety symptoms were measured using the Generalised Anxiety Disorder screener (GAD-2), and depressive symptoms using the Patient Health Questionnaire screener (PHQ-2). For workers at their physical workplace (site-based workers) we asked questions about the adequacy and implementation of 11 different types of ICP, and the adequacy and supply of eight different types of personal protective equipment (PPE). Respondents were classified as either: working remotely; site-based workers with 100% of their ICP/PPE needs met; site-based workers with 50-99% of ICP/PPE needs met; site-based workers with 1-49% of ICP/PPE needs met; site-based workers with none of ICP/PPE needs met; or no longer employed. Regression analyses examined the association between working arrangements and ICP/PPE adequacy and having GAD-2 and PHQ-2 scores of three and higher (a common screening point in both scales). Models were adjusted for a range of demographic, occupation, workplace, and COVID-19-specific factors. A total of 42.3% (95% CI: 40.6-44.0%) of the sample had GAD-2 scores of 3 and higher, and 34.6% (95% CI: 32.-36.2%) had PHQ-2 scores of 3 and higher. In initial analyses, symptoms of anxiety and depression were lowest among those working remotely (35.4 and 27.5%), compared to site-based workers (43.5 and 34.7%) and those who had lost their jobs (44.1 and 35.9%). When adequacy of ICP and PPE was taken into account, the lowest prevalence of anxiety and depressive symptoms was observed among site-based workers with all of their ICP needs being met (29.8% prevalence for GAD-2 scores of 3 and higher, and 23.0% prevalence for PHQ-2 scores of 3 and higher), while the highest prevalence was observed among site-based workers with none of their ICP needs being met (52.3% for GAD-2 scores of 3 and higher, and 45.8% for PHQ-2 scores of 3 and higher). Our results suggest that the adequate design and implementation of employer-based ICP have implications for the mental health of site-based workers. As economies re-open the ongoing assessment of ICP and associated mental health outcomes among the workforce is warranted.


Intravenous indocyanine green dye is insufficient for robust immune cell labelling in the human retina.
It is not currently possible to reliably visualise and track immune cells in the human central nervous system or eye. Previous work demonstrated that indocyanine green (ICG) dye could label immune cells and be imaged after a delay during disease in the mouse retina. We report a pilot study investigating if ICG can similarly label immune cells within the human retina. Twelve adult participants receiving ICG angiography as part of routine standard of care were recruited. Baseline retinal images were obtained prior to ICG administration then repeated over a period ranging from 2 hours to 9 days. Matched peripheral blood samples were obtained to examine systemic immune cell labelling and activation from ICG by flow cytometry with human macrophage cultures as positive controls. Differences between the delayed near infrared ICG imaging and 488 nm autofluorescence was observed across pathologies, likely arising from the retinal pigment epithelium (RPE). Only one subject demonstrated ICG signal on peripheral blood myeloid cells and only three distinct cell-sized signals appeared over time within the retina of three participants. No significant increase in immune cell activation markers were detected after ICG administration. ICG accumulated in the endosomes of macrophage cultures and was detectable above a minimum concentration, suggesting cell labelling is possible. ICG can label RPE and may be used as an additional biomarker for RPE health across a range of retinal disorders. Standard clinical doses of intravenous ICG do not lead to robust immune cell labelling in human blood or retina and further optimisation in dose and route are required.