Characterization of the phosphoenolpyruvate carboxykinase (GTP) promoter-regulatory region. II. Identification of cAMP and glucocorticoid regulatory domains.
Hormonal regulatory elements within the phosphoenolpyruvate carboxykinase (GTP) (EC 184.108.40.206) (PEPCK) promoter region were mapped using a series of 5' deletions linked to the amino-3'-glycosyl phosphotransferase structural gene. These deletion mutants were stably transfected into the genome of FTO-2B hepatoma cells. A 47-base pair region of the PEPCK promoter was identified which was essential for stimulation by dibutyryl cAMP. A 12-base pair core sequence (CTTACGTCAGAG) within this region shows significant homology with sequences in four other cAMP-regulated genes. There are two glucocorticoid regulatory elements within the promoter, as well as an inhibitory element which depresses the level of basal gene transcription. The deletion of this inhibitory sequence prevents the induction of the chimeric gene by dexamethasone. The existence of the hormone regulatory domains within the PEPCK promoter was confirmed by attaching these elements upstream of the heterologous Herpes simplex virus thymidine kinase structural gene, containing its own promoter.


ESR1 as a recurrence-related gene in intrahepatic cholangiocarcinoma: a weighted gene coexpression network analysis.
Intrahepatic cholangiocarcinoma (iCCA) is the second most common malignant hepatic tumor and has a high postoperative recurrence rate and a poor prognosis. The key roles of most tumor recurrence-associated molecules in iCCA remain unclear. This study aimed to explore hub genes related to the postsurgical recurrence of iCCA. Differentially expressed genes (DEGs) between iCCA samples and normal liver samples were screened from The Cancer Genome Atlas (TCGA) database and used to construct a weighted gene coexpression network. Module-trait correlations were calculated to identify the key module related to recurrence in iCCA patients. Genes in the key module were subjected to functional enrichment analysis, and candidate hub genes were filtered through coexpression and protein-protein interaction (PPI) network analysis. Validation studies were conducted to detect the "real" hub gene. Furthermore, the biological functions and the underlying mechanism of the real hub gene in iCCA tumorigenesis and progression were determined via in vitro experiments. A total of 1019 DEGs were filtered and used to construct four coexpression modules. The red module, which showed the highest correlations with the recurrence status, family history, and day to death of patients, was identified as the key module. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses demonstrated that genes in the red module were enriched in genes and pathways related to tumorigenesis and tumor progression. We performed validation studies and identified estrogen receptor 1 (ESR1), which significantly impacted the prognosis of iCCA patients, as the real hub gene related to the recurrence of iCCA. The in vitro experiments demonstrated that ESR1 overexpression significantly suppressed cell proliferation, migration, and invasion, whereas ESR1 knockdown elicited opposite effects. Further investigation into the mechanism demonstrated that ESR1 acts as a tumor suppressor by inhibiting the JAK/STAT3 signaling pathway. ESR1 was identified as the real hub gene related to the recurrence of iCCA that plays a critical tumor suppressor role in iCCA progression. ESR1 significantly impacts the prognosis of iCCA patients and markedly suppresses cholangiocarcinoma cell proliferation, migration and invasion by inhibiting JAK/STAT3 signaling pathway.


Local production of kinins contributes to the endothelium dependent relaxations evoked by converting enzyme inhibitors in isolated arteries.
The angiotensin converting enzyme (ACE) inhibitor perindoprilat evokes endothelium-dependent relaxations in perfused isolated canine arteries. Kininogens, the precursors of bradykinin, elicit endothelium-dependent relaxations which are potentiated by perindoprilat, inhibited by B2-kinin antagonists and partially impaired after inhibition of NO synthase. These observations suggest that locally produced kinins may stimulate the production of NO and endothelium-derived hyperpolarizing factor, and that this action is potentiated by ACE inhibitors.


Methamphetamine neurotoxicity increases brain expression and alters behavioral functions of CB₁ cannabinoid receptors.
Cannabis is the most common secondary illicit substance in methamphetamine (METH) users, yet the outcomes of the concurrent consumption of both substances remain elusive. Capitalizing on recent findings on the implication of CB₁ cannabinoid receptors in the behavioral effects of METH, we hypothesized that METH-induced neurotoxicity may alter the brain expression of CB₁, thereby affecting its role in behavioral functions. To test this possibility, we subjected rats to a well-characterized model of METH neurotoxicity (4 mg/kg, subcutaneous × 4 injections, 2 h apart), and analyzed their CB₁ receptor brain expression three weeks later. METH exposure resulted in significant enhancements of CB₁ receptor expression across several brain regions, including prefrontal cortex, caudate-putamen, basolateral amygdala, CA1 hippocampal region and perirhinal cortex. In parallel, a different group of METH-exposed rats was used to explore the responsiveness to the potent cannabinoid agonist WIN 55,212-2 (WIN) (0.5-1 mg/kg, intraperitoneal), within several paradigms for the assessment of emotional and cognitive functions, such as open field, object exploration and recognition, and startle reflex. WIN induced anxiolytic-like effects in METH-exposed rats and anxiogenic-like effects in saline-treated controls. Furthermore, METH-exposed animals exhibited a significantly lower impact of WIN on the attenuation of exploratory behaviors and short-term (90 min) recognition memory. Conversely, METH neurotoxicity did not significantly affect WIN-induced reductions in locomotor activity, exploration time and acoustic startle. These results suggest that METH neurotoxicity may alter the vulnerability to select behavioral effects of cannabis, by inducing distinct regional variations in the expression of CB₁ receptors.


Female control of a novel form of cannibalism during copulation in a South American widow spider.
Sexual cannibalism is an extreme form of sexual conflict that may have broad evolutionary effects on mating behaviour. Latrodectus spiders (∼30 "widow" species) could enable comparative tests of the evolution of sexual cannibalism, but most species are poorly-studied. Here we describe the mating and remating behaviour of a species endemic to South America, L. mirabilis, with a focus on the occurrence and consequences of sexual cannibalism. Mating and cannibalistic behaviours were novel relative to other Latrodectus species. Cannibalism started during copulation when females grasped the male's legs with their chelicerae, and continued as females pulled the male's abdomen onto their chelicerae, without interrupting copulation. Cannibalism was initiated by females, and not facilitated by males as was observed in other Latrodectus species. Females frequently remated however, so cannibalized males might lose paternity to future rivals. We report high rates of cannibalism, with 70 % of males killed by females during their first mating, and 85 % killed by previously-mated females. We discuss our novel findings in the context of previous observations in other Latrodectus species, proposing that foundational studies in a wider range of species are necessary to support comparative tests about the evolution of sexual cannibalism within this model taxon.


Wear performance of dental ceramics after grinding and polishing treatments.
The aim of this in vitro study was to determine the two-body wear resistance of different dental ceramics after grinding and polishing treatments. Standardized specimens were prepared from three zirconia and two veneering ceramics and were subjected to different surface treatments. Zirconia ceramics were polished, ground and repolished, veneering ceramics were ground and repolished. One zirconia ceramic was investigated with a superficial glaze. Human enamel was used for reference. Surface roughness R(a) was determined using a profilometric contact surface measurement device. Two-body wear tests were performed in a chewing simulator with steatite and enamel antagonists, respectively. Specimens were loaded pneumatically in a pin-on-block design for 1.2x10(5) mastication cycles (50 N, 1.2 Hz, lateral movement: 1 mm, mouth opening: 2 mm) under simultaneous thermal cycling (600 cycles, 5/55 °C). Wear depths of specimens were determined using a 3D laser scanning device, wear areas of steatite antagonists were measured by means of light-optical micrographs. Means and standard deviations were calculated, and statistical analysis was performed using one-way analysis of variance (ANOVA) and the Bonferroni multiple comparison test for post hoc analysis (α=0.05). Scanning electron microscopy was applied for evaluating the wear performance of ceramics and antagonists. No wear was found for polished, ground and repolished zirconia. Compared to the wear depths of the enamel reference with 274.1±187.4 μm versus steatite and 123.3±131.0 μm versus enamel, relative wear depths of porcelains ranged between 0.54±0.07 and 0.62±0.09 with steatite antagonists and between 0.66±0.26 and 1.04±0.27 with enamel antagonists. Relative wear areas of steatite antagonists (enamel reference: 1.25 mm(2)) varied between 0.84±0.13 and 1.90±0.29 for zirconia and between 1.97±0.38 and 2.47±0.40 for porcelains. Enamel antagonists generally showed wear, cracks or even fractures, but revealed smooth surfaces when opposed to polished/ground/repolished zirconia and ploughed surfaces when opposed to ground/repolished porcelains or glaze. Zirconia ceramics yielded superior wear behavior and lower antagonistic wear compared to porcelains. A trend to higher ceramic and antagonistic wear was shown after grinding treatments.


Flow injection assays for NADH and ethanol using photosensitized rose bengal and luminol-copper (II) chemiluminescence system.
The online photoreaction of the rose bengal photosensitized luminol-copper (II) chemiluminescence (CL) system was used for the determination of β-nicotinamide adenine dinucleotide (NADH) and ethanol (EtOH) in pharmaceutical formulations combined with a flow injection technique. NADH can significantly enhance the CL emission of the reaction. For EtOH, alcohol dehydrogenase in soluble form was utilized in the presence of nicotinamide adenine dinucleotide resulting in NADH production. The limit of detection (3σ blank, 𝑛 = 3) of 4.0 × 10-8 and 2.17 × 10-5 M, and linear range 1.3 × 10-7 to 2.5 × 10-5 M (R2 = 0.9998, n = 6) and 0.11-2.17 × 10-3 M (R2 = 0.9996, n = 6) were obtained for NADH and EtOH respectively. The injection rate was 100 h-1 with a relative standard deviation (n = 3) of 1.5-4.8% in the range studied for both analytes. The procedure was satisfactorily applied to pharmaceutical formulations with recoveries in the range 91.6 ± 3.0% to 110 ± 2.0% for NADH and 88 ± 3.0% to 95.4 ± 4.0% for EtOH. The results obtained were very consistent and did not differ considerably from the reported approaches at a 95% confidence limit. The possible mechanism of the CL reaction is also explained briefly.


Characterization of a Nitrogenase Iron Protein Substituted with a Synthetic [Fe4 Se4 ] Cluster.
The Fe protein of nitrogenase plays multiple roles in substrate reduction and cluster maturation via its redox-active [Fe4 S4 ] cluster. Here we report the synthesis and characterization of a water-soluble [Fe4 Se4 ] cluster that is used to substitute the [Fe4 S4 ] cluster of the Azotobacter vinelandii Fe protein (AvNifH). Biochemical, EPR and XAS/EXAFS analyses demonstrate the ability of the [Fe4 Se4 ] cluster to adopt the super-reduced, all-ferrous state upon its incorporation into AvNifH. Moreover, these studies reveal that the [Fe4 Se4 ] cluster in AvNifH already assumes a partial all-ferrous state ([Fe4 Se4 ]0 ) in the presence of dithionite, where its [Fe4 S4 ] counterpart in AvNifH exists solely in the reduced state ([Fe4 S4 ]1+ ). Such a discrepancy in the redox properties of the AvNifH-associated [Fe4 Se4 ] and [Fe4 S4 ] clusters can be used to distinguish the differential redox requirements for the substrate reduction and cluster maturation of nitrogenase, pointing to the utility of chalcogen-substituted FeS clusters in future mechanistic studies of nitrogenase catalysis and assembly.


On the interaction between macrophages and developmental stages of Schistosoma mansoni: effect of muramyl tripeptide phosphatidyl ethanolamine (MTP-PE) treatment on mice survival and the generation of schistosomulicidal macrophages.
Schistosomiasis is a chronic disease afflicting hundreds of millions of people throughout the world against which there is as yet no effective vaccine. In the present study we tested the effect of the immunomodulator muramyl tripeptide phosphatidyl ethanolamine (MTP-PE) on the survival of Schistosoma mansoni-infected mice and on the induction in them of schistosomulicidal macrophages. Mice exposed to 80 cercariae each and then treated with MTP-PE showed prolonged survival following either single or repeat infection. The treatment with MTP-PE, when initiated 70 days post the schistosome infection, diminished significantly the mortality of infected mice over an observed period of 110 days. In terms of treatment efficacy there was no evident difference between the intravenous and intraperitoneal mode of administration of the drug. MTP-PE treatment significantly reduced granuloma size and markedly diminished liver damaged as judged by the lower levels of alkaline phosphatase in the serum. Such treatment exerted no significant effect on the spleen or liver weight in infected mice nor on the worm burden resulting from either a single or double infection. In infected and non-treated mice, schistosomulicidal macrophages appeared after 8-10 weeks of infection. In infected mice treated with MTP-PE there was an accelerated appearance of such macrophages and these exhibited a greater cidal effect on the schistosomula. These immunostimulatory and life-prolonging effects of MTP-PE on S. mansoni-infected mice might indicate an effect of this reagent on cells involved in the granulomatous process.


Evidence for an Akt-kinase/NO/cGMP pathway in the cochlea of guinea pigs.
The protein kinase Akt (protein kinase B) can be activated by numerous growth factors via PI-3 kinase-generated phosphoinositides and is thought to have anti-apoptotic properties. Activated Akt/PKB boosts the activity of endothelial NO synthase (NOS III), which has been found in the key areas of the inner ear (e.g., hair cells and stria vascularis). In order to localize activated Akt/PKB (phospho-Akt) in the cochlea of guinea pigs, sections of ten temporal bones were observed immunohistochemically. The strongest immunoreactivity was found in and underneath inner hair cells (IHC). Within the organ of Corti, reactivity was found in supporting cells, while outer hair cells remained unstained. Spiral ganglion cells, the endothelium of the lateral wall and the vascular area of the modiolus showed moderate staining. The results give evidence that activated Akt/PKB influences the activity of the NO/cGMP pathway in the cochlea. Because of the antiapoptotic properties, activated Akt should now be examined under non-physiological conditions.


Chemistry of ecteinascidins. Part 3: preparation of 2'-N-acyl derivatives of ecteinascidin 770 and evaluation of cytotoxicity.
A three-step transformation of ecteinascidin 770 (1b) into 2'-N-indole-3-carbonyl derivative 3 via 18,6'-O-bisallyl-protected derivative 4a, which was shown to have higher cytotoxicity than 1b, is presented. In addition, a number of 2'-N amide derivatives of 1b have been prepared from 4a and their in vitro cytotoxicity were determined by measuring IC₅₀ values against human cell lines HCT116, QG56, and DU145. Benzoyl amide derivatives 7a-c showed similar in vitro cytotoxicity to 1b, whereas the nitrogen-containing heterocyclic derivatives 7d-h and cinnamoyl derivatives 9a-b showed higher cytotoxicity than 1b. In contrast, the 18,6'-O-bisallyl protected derivatives 4a-c, 6a-h, and 8a-b showed dramatic decreases in cytotoxicity relative to 1b.


Use of Busulfan to Condition Mice for Bone Marrow Transplantation.
Myeloablative gamma irradiation has traditionally been used to condition mice prior to bone marrow transplantation. However, irradiation induces high levels of inflammation that may alter patterns of reconstitution. In addition, gamma irradiators are being removed from many facilities for security reasons. Alternative conditioning regimens are thus needed. Here, we describe a protocol for the use of busulfan to condition mice for bone marrow transplantation and several of the variables to consider for effective implementation. For complete details on the use and execution of this protocol, please refer to Montecino-Rodriguez et al. (2019).


Web-based E-survey in identifying current physiotherapy practices in paediatric burns.
Physiotherapy practices in pediatric burns involves relaxation techniques and exercise training for attaining physical fitness, improving quality of life and, thereby to prevent primary and secondary complications. Physical activity and exercise act as a meaningful and purposeful tool for attaining physical fitness after burn injury. The aim of this e-survey is to identify the current practices of physiotherapy interventions in pediatric burn rehabilitation among physiotherapy students and professionals. The e-survey was created with validated questionnaire using a Google Forms to gather the information related to current physiotherapy practices in pediatric burns. After the sample size estimation, survey link were sent to identified 144 physiotherapy students and professionals using social networking sites which includes, WhatsApp, Facebook and Instagram for this cross-sectional e-survey. The frequency and percentage of survey responses were analyzed. Among 144 identified participants, 62 participants completed the survey, resulting in the response rate of (43.0%). The findings of this study revealed that> 50% of physiotherapy students, and professionals, are performing pain and scar assessment, along with their regular physiotherapy management in children with burns. In addition to these, they also practice physiotherapy for burn conditions in pediatric intensive care units (PICU), pediatric wards, and clinical outpatient department (OPD) settings. Physiotherapy students and professionals have sufficient knowledge regarding pediatric burns complications, and also, they are well aware regarding recent physiotherapy practices in pediatric burn care management.


Renal Cell Carcinoma: the Oncologist Asks, Can PSMA PET/CT Answer?
To critically review the potential clinical applications of prostate-specific membrane antigen (PSMA) radioactive ligands in renal cell carcinoma (RCC). Radioactive probes targeting PSMA hold promise in several malignancies in addition to prostate cancer, owing to the expression of PSMA by tumor neovasculature. The majority of clear cell RCCs (ccRCC), the most malignant RCC subtype, express PSMA on tumor-associated neovasculature. The endothelium of less aggressive RCC subtypes is PSMA positive in a lower, but still significant percentage of cases. PSMA might therefore represent an interesting theragnostic target in RCC. The preliminary data available suggest a potential role for PSMA-targeting radiopharmaceuticals in complementing conventional imaging for staging ccRCC patients at risk of nodal involvement and oligometastatic disease. Additional applications of PSMA imaging may be the selection and the response assessment of patients receiving anti-angiogenic treatments. The effectiveness of PSMA-targeting radionuclide therapy should also be investigated.


Efficiency of the Polymerase Chain Reaction.
The polymerase chain reaction (PCR) has found wide application in biochemistry and molecular biology such as gene expression studies, mutation detection, forensic analysis and pathogen detection. Increasingly quantitative real time PCR is used to assess copy numbers from overall yield. In this study the yield is analyzed as a function of several processes: (1) thermal damage of the template and polymerase occurs during the denaturing step, (2) competition exists between primers and templates to either anneal or form dsDNA, (3) polymerase binding to annealed products (primer/ssDNA) to form ternary complexes and (4) extension of ternary complexes. Explicit expressions are provided for the efficiency of each process, therefore reaction conditions can be directly linked to the overall yield. Examples are provided where different processes play the yield-limiting role. The analysis will give researchers a unique understanding of the factors that control the reaction and will aid in the interpretation of experimental results.


Divergent evolution of two plastid genes, rbcL and atpB, in a non-photosynthetic parasitic plant.
Plastid DNA (ptDNA) regions for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubiso) (rbcL) and the beta-subunit of ATP synthase (atpB) genes of the holoparasite Lathraea clandestina L. were sequenced. These regions were obtained by cloning either a Bam HI endonuclease generated fragment from the Lathraea ptDNA or polymerase chain reaction (PCR) amplified products. The Lathraea ptDNA contains the entire sequence for the rbcL gene which shares 94.5% homology with the Nicotiana tabacum gene, whereas atpB is maintained as a pseudogene. The intergenic region between divergently transcribed rbcL and atpB genes is shorter (758 bp) in L. clandestina plastid genome in comparison with N. tabacum (823 bp), however they have a noticeable similarity, mainly in the rbcL 5'-upstream region. A low level of the rbcL gene transcription was detected whereas no atpB transcripts were found in Latraea. The plasmid rbcL gene of the hemiparasite Melampyrum pratense and the autotroph Digitalis purpurea both from the Scrophulariaceae were cloned by PCR amplification and then sequenced. The L. clandestina rbcL gene is highly homologous to the M. pratense and D. purpurea genes. The data indicate that the evolution of the plastid atpB-rbcL region was different in parasites from the Scrophulariaceae and Orobanchaceae families.


Application of a Self-developed, Low-budget Indocyanine Green Camera in Surgical Imaging - a Single Institution's Experiences.
Indocyanine green is a fluorescent dye, the use of which is becoming more and more widespread in different areas of surgery. Several international studies deal with the dye's usefulness in intraoperative angiography, the localization of tumors, the more precise identification of anatomical structures, the detection of lymph nodes and lymph ducts, etc. The application of the dye is safe, but a suitable equipment park is required for its use, which entails relatively high costs. The aim of our research is to create a detector system on a low budget, to be used safely in everyday practice and to illustrate its operation with practical examples at our own institute. By modifying a web camera, using filter lenses and special LEDs, we created a device suitable for exciting and detecting indocyanine green fluorescence. We prove its excellent versatility during the following procedures at our institute: breast tumor surgery, kidney transplantation, bowel resection, parathyroid surgery and liver tumor resection. The finished camera has an LED light source with a peak wavelength of 780 nm, and the incoming light is filtered by a bandpass filter with a center wavelength of 832 nm. A low budget ($112), easy-to-use tool was created, which is suitable for taking advantage of the opportunities provided by indocyanine green.


New insights into progesterone actions on prolactin secretion and prolactinoma development.
Progesterone (P4) has controversial physiological effects on the regulation of the lactotroph population. While some studies have shown a negative role for P4 in prolactin secretion and lactotroph proliferation, antagonizing estradiol effects, others demonstrated a proliferative role of P4 at the pituitary level. Usually, progesterone actions in the pituitary gland were studied through their classical, genomic pathways triggered by nuclear progesterone receptors (nPRs). However, in 2003, the scene became more complex with the discovery of another group of progesterone receptors involved in rapid, non-genomic P4 effects: the membrane progesterone receptors (mPRs), which are members of the progesterone and adipoQ receptor (PAQR) family. This review examines the historical background and current data on the study of progesterone actions on PRL secretion providing new evidence of P4 effects at the hypothalamic and at the pituitary level through non-classic P4-receptors. In addition, we explore the role of progesterone in the development of experimental prolactinomas, a controversial topic in the literature.


Adrenocorticotropin normalizes the blood levels of nitric oxide in hemorrhage-shocked rats.
Anesthetized rats were subjected to volume-controlled hemorrhagic shock by stepwise bleeding. Besides cardiovascular and respiratory functions, nitric oxide (NO)-hemoglobin formation in arterial blood was directly evaluated by means of electron spin resonance spectroscopy. During hemorrhagic shock there was a massive increase in NO-hemoglobin, associated with a fall in mean arterial pressure, pulse pressure, respiratory rate and heart rate, and there was a further increase in NO-hemoglobin 15 min after intravenous (i.v.) treatment with saline. All rats died within 30 min. The reversal of the shock condition induced by the i.v. injection of the adrenocorticotropin (ACTH) fragment 1-24 (160 microg/kg, 5 min after bleeding termination) was associated with a prompt disappearance of NO-hemoglobin. Also S-methylisothiourea (3 mg/kg i.v.), a selective inhibitor of inducible NO synthase, provoked a disappearance of NO-hemoglobin and reversal of the shock condition. The present results provide a direct demonstration that volume-controlled hemorrhagic shock is associated with highly increased blood levels of NO, as indicated by increased NO-hemoglobin, and indicate that ACTH-induced reversal of the shock condition is associated with the normalization of NO blood levels, and a parallel improvement of cardiovascular and respiratory functions. This occurs probably through the inhibition of inducible NO synthase, as suggested by the fact that S-methylisothiourea, a selective inhibitor of this NO synthase isoform, produced the same results.


PD-L1+ dendritic cells in the tumor microenvironment correlate with good prognosis and CD8+ T cell infiltration in colon cancer.
The prognostic value of tumor-associated dendritic cells (DC) in colon cancer remains poorly understood. This may be in part due to the interchangeable expression of immunostimulatory and immunoinhibitory molecules on DC. Here we investigated the prognostic impact of CD11c+ DC co-expressing the immunoinhibitory molecule PD-L1 and their spatial relationship with CD8+ T-cells in patients treated for stage III colon cancer. Tissue microarrays containing representative cores of central tumor, leading edge, and adjacent normal tissue from 221 patients with stage III colon cancer were immunostained for CD8, CD11c, PD-L1, and cytokeratin using immunofluorescent probes. Cells were quantified using StrataQuest digital image analysis software, with intratumoral and stromal regions analyzed separately. Kaplan-Meier estimates and Cox regression were used to assess survival. Intratumoral CD8+ cell density (HR = .52, 95% confidence interval [CI] .33-.83, P = .007), stromal CD11c+ cell density (HR = .52, 95% CI .33-.83, P = .006), intratumoral CD11c+ PD-L1+ cell density (HR = .57, 95% CI .35-.92, P = .021), and stromal CD11c+ PD-L1+ cell density (HR = .48, 95% CI .30-.77, P = .003) on leading-edge cores were all significantly associated with good survival. CD8+ cell density was positively correlated with both CD11c+ cell density and CD11c+ PD-L1+ cell density in tumor epithelium and stromal compartments. Here we showed that PD-L1-expressing DC in the tumor microenvironment are associated with improved survival in stage III colon cancer and likely reflect an immunologically "hot" tumor microenvironment. Further investigation into the expression of immunomodulatory molecules by tumor-associated DC may help to further elucidate their prognostic value.


Alpha-7 Nicotinic Acetylcholine Receptor Activation Inhibits Trauma Induced Pronociceptive Autoimmune Responses.
Both autonomic nervous system dysfunction and immune system activation are characteristic of chronic pain after limb injuries. Cholinergic agonists reduce immune system activation in many settings. We hypothesized, therefore, that alpha-7 nicotinic acetylcholine receptor (α7nAChR) agonist administration would reduce nociceptive and immune changes after tibia fracture and cast immobilization in mice. Fracture mice were treated with either vehicle, a low (.2 mg/kg) dose, or a high (1 mg/kg) dose of the selective α7nAChR agonist PNU-282987 for 4 weeks. We assessed hindpaw allodynia and weight bearing as behavioral outcomes. The assessment of adaptive immune responses included regional lymph node hypertrophy, germinal center formation, α7nAChR expression, and IgM deposition. Assessment of innate immune system activation focused on IL-1β and IL-6 generation in fractured hindlimb skin. We observed that mechanical allodynia and unweighting were alleviated by PNU-282987 treatment. Drug treatment also reduced popliteal lymph node hypertrophy and germinal center formation. Immunohistochemical studies localized α7nAChR to germinal center B lymphocytes, and this expression increased after fracture. Analysis of fracture limb hindpaw skin demonstrated increased inflammatory mediator (IL-1β and IL-6) levels and IgM deposition, which were abrogated by PNU-282987. Serum analyses demonstrated fracture-induced IgM reactivity against keratin 16, histone 3.2, GFAP, and NMDAR-2B. Administration of PNU-282987 reduced the enhancement of IgM reactivity. Collectively, these data suggest that the α7nAChR is involved in regulating posttraumatic innate and adaptive immune responses and the associated nociceptive sensitization. PERSPECTIVE: These studies evaluate the effects of a selective α7nAChR agonist in a tibial fracture/cast immobilization model of limb pain. Administration of the drug reduced nociceptive and functional changes 4 weeks after injury. These novel findings suggest that well-tolerated α7nAChR agonists may be viable analgesics for chronic pain after limb injuries.


KNOWLEDGE OF VESTIBULAR OCULAR DYSFUNCTION AND UTILIZATION OF VESTIBULAR OCULAR MOTOR SCREENING (VOMS) TOOL COMPONENTS AMONG PROFESSIONAL SPORTS LEAGUE ATHLETIC TRAINERS.
The Vestibular Ocular Motor Screen (VOMS) is a relatively new measurement tool intended for the non-vestibular practitioner to identify vestibular ocular dysfunction (VOD) following sport related concussion (SRC). Specific knowledge of VOD and usage of the VOMS among athletic trainers in professional sports leagues is currently unknown. The purpose of this study was to examine knowledge of VOD following SRC and utilization of the VOMS tool and other vestibular ocular tests and measures among athletic trainers in professional sports. Cross Sectional Study. A total of 117 athletic trainers from Major League Baseball, the National Basketball Association, the National Football League and Major League Soccer, with a mean of 17 ± 9 years in the athletic training profession were surveyed via Qualtrics™ with a response rate of 33%. The survey contained three primary sub-sections; demographics, knowledge of vestibular ocular deficits following SRC, and VOMS component utilization at baseline, acute and return-to-play phases of management. Total knowledge scores as well as percentage of utilization based on concussion management stages were calculated. The average percentage correct on the knowledge items was 56% (range of 30% to 65%). There was no difference in knowledge score among athletic trainers with formal post-professional concussion training (p = 0.29) compared to those with no formal training. There was no relationship found between total years practicing and total knowledge score, r = -.128, (p = 0.17). Smooth pursuit testing was the most commonly utilized (70%) in clinical practice and Visual Motion Sensitivity (VMS) was the least utilized (17%). Balance assessment measures to examine vestibular functioning remained the highest utilized examination technique at all-time points in management of SRC. The range of correct responses from 30% to 65% indicates that at the time of survey the participants had decreased knowledge of VOD following SRC. There was low overall utilization of all VOMS components despite recent evidence showing good sensitivity and low false positive rates in SRC. The results of this study identify an opportunity for future training specific to vestibular-ocular impairments and assessment following concussion injury. 2c.


Bacillus asahii sp. nov., a novel bacterium isolated from soil with the ability to deodorize the bad smell generated from short-chain fatty acids.
In a screening campaign to isolate strains with the ability to remove the bad smell associated with animal faeces, strain MA001(T) was isolated from a soil sample obtained from Shizuoka prefecture, Japan. The isolate grew at pH 6-9 but not at pH 10. Cells were Gram-positive, straight rods with peritrichous flagella and produced ellipsoidal spores. The isolate was positive for catalase and oxidase tests but negative for indole production, deamination of phenylalanine and H(2)S production. The isolate did not produce acid from any carbohydrates tested and could not grow in more than 2 % NaCl. The DNA G+C content was 39.4 mol%. The cellular fatty acids profile consisted of significant amount of C(15) branched-chain fatty acids, iso-C(15 : 0) and anteiso-C(15 : 0). Phylogenetic analysis based on 16S rRNA gene sequencing indicated that strain MA001(T) was closely related to Bacillus simplex and Bacillus psychrosaccharolyticus. DNA-DNA hybridization revealed a low relatedness of the isolate to several phylogenetically close neighbours (less than 9 %). On the basis of the phenotypic characteristics observed, phylogenetic data based on 16S rRNA gene sequencing and DNA-DNA relatedness data, it is concluded that the isolate should be classified as representing a novel species, for which the name Bacillus asahii is proposed. The type strain is MA001(T) (=JCM 12112(T)=NCIMB 13969(T)).


Synthesis, characterization, anti-inflammatory and anti-Helicobacter pylori activities of novel benzophenone tetracarboxylimide benzoyl thiourea cross-linked chitosan hydrogels.
Chitosan (Cs) was cross-linked with four various quantities of 4,4'-(5,5'‑carbonylbis(1,3-dioxoisoindoline-5,2-diyl))dibenzoyl isothiocyanate. Elemental analysis, FTIR and 1H NMR spectroscopy assured that the amino groups of chitosan reacted with the isothiocyanate groups of the cross-linker producing four new hydrogels namely as BBTU-Cs-1, BBTU-Cs-2, BBTU-Cs-3, and BBTU-Cs-4 according to the increment of their cross-linking content, respectively. SEM showed their porous structures and XRD indicated their amorphous nature. Their swell ability increased with decreasing the medium pH value and with increasing cross-linking density. In comparison with the popular COX inhibitor Celecoxib, these hydrogels showed an inhibition activity towards COX enzymes with selective inhibition towards COX-2. Their inhibition activity could be arranged as follows: Celecoxib > BBTU-Cs-4 > BBTU-Cs-3 > BBTU-Cs-2 > BBTU-Cs-1. BBTU-CS-4 hydrogel exhibited a potent inhibition against COX-2 (IC50 0.42 μg/ml) compared with that observed for the standard Celecoxib (IC50 0.26 μg/ml). BBTU-Cs-4 is more potent against H. pylori compared to the other hydrogels. BBTU-Cs-4 at a concentration of 7.81 μg/ml is able to kill 100% of the H. pylori and exhibits a preferential ability to inhibit 89.35% of COX-2 than COX-1 (0%). These findings make BBTU-Cs-4 a promising anti-H. pylori and selective anti-inflammatory agent.


Maternal valproate dosage and foetal malformations.
OBJECTIVE- To study the possible dose dependence of the foetal malformation rate after exposure to sodium valproate in pregnancy METHODS- Analysis of records of all foetuses in the Australian Registry of Antiepileptic Drugs in Pregnancy exposed to valproate, to carbamazepine, lamotrigine or phenytoin in the absence of valproate, and to no antiepileptic drugs. RESULTS- The foetal malformation rate was higher (P<0.05) in the 110 foetuses exposed to valproate alone (17.1%), and in the 165 exposed to valproate, whether alone or together with the other antiepileptic drugs (15.2%), than in the 297 exposed to the other drugs without valproate (2.4%). It was also higher (P<0.10) than in the 40 not exposed to antiepileptic drugs (2.5%). Unlike the situation for the other drugs, the malformation rate in those exposed to valproate increased with increasing maternal drug dosage (P<0.05). The rate was not altered by simultaneous exposure to the other drugs. Valproate doses exceeding 1400 mg per day seemed to be associated with a more steeply increasing malformation rate than at lower doses and with a different pattern of foetal malformations. CONCLUSIONS- Foetal exposure to valproate during pregnancy is associated with particularly high, and dose-dependent risks of malformation compared with other antiepileptic drugs, and may possibly involve different teratogenetic mechanisms.


Ligation of single-stranded oligodeoxyribonucleotides by T4 RNA ligase.
Despite its unique ability to ligate single-stranded DNA molecules, T4 RNA ligase has so far seen little use in molecular biology due to long reaction times, modest yields, and apparent inability to promote ligation of long oligodeoxyribonucleotides. We describe here a set of reaction conditions which dramatically shorten the reaction time and give reproducible 40 to 60% ligation of DNA fragments of up to 40 bases in length. These improvements open promising new fields of application to T4 RNA ligase.


Methyl 6-dimethyl-amino-4-hy-droxy-2-naphtho-ate.
In the title compound, C(14)H(15)NO(3), the ester group is oriented so that the carbonyl group points in the opposite direction to the hy-droxy group. The mol-ecule as a whole is almost planar (the r.m.s. deviation of the non-H atoms is 0.0268 Å). In the crystal, mol-ecules are linked by inter-molecular O-H⋯O hydrogen bonds into infinite chains that propagate parallel to the c axis.


Chronic central neuropeptide Y infusion in normal rats: status of the hypothalamo-pituitary-adrenal axis, and vagal mediation of hyperinsulinaemia.
Neuropeptide Y in the hypothalamus is a potent physiological stimulator of feeding, and may contribute to the characteristic metabolic defects of obesity when hypothalamic levels remain chronically elevated. Since corticosterone and insulin are important regulators of fuel metabolism, the longitudinal effects of chronic (6 days) intracerebroventricular infusion of neuropeptide Y in normal rats on the hypothalamo-pituitary-adrenal axis and on insulin secretion were studied. Neuropeptide Y-infused rats were either allowed to eat ad libitum, or were pair-fed with normophagic control rats. Neuropeptide Y increased the basal plasma concentrations of adrenocorticotropic hormone and corticosterone during the first 2 days of its intracerebroventricular infusion and increased cold stress-induced plasma adrenocorticotropic hormone concentrations. After 4-6 days of central neuropeptide Y infusion, however, basal plasma adrenocorticotropic hormone and corticosterone concentrations were no different from control values (except in ad libitum-fed rats in which corticosteronaemia remained elevated), they were unaffected by the stress of cold exposure, and the hypothalamic content of corticotropin-releasing factor immunoreactivity was significantly decreased. A state of hyperinsulinaemia was present throughout the 6 days of intracerebroventricular neuropeptide Y infusion, being more marked in the ad libitum-fed than in the pair-fed group. The proportions of insulin, proinsulin, and conversion intermediates in plasma and pancreas were unchanged. Hyperinsulinaemia of the pair-fed neuropeptide Y-infused rats was accompanied by muscle insulin resistance and white adipose tissue insulin hyperresponsiveness, as assessed by the in vivo uptake of 2-deoxyglucose. Finally, bilateral subdiaphragmatic vagotomy prevented both the basal and the marked glucose-induced hyperinsulinaemia of animals chronically infused with neuropeptide Y, demonstrating that central neuropeptide Y-induced hyperinsulinaemia is mediated by the parasympathetic nervous system.


Determination of the relative amounts of three crystal forms of a benzimidazole drug in complex finished formulations by FT-Raman spectroscopy.
A 5% (m/m) premix for animal use was quantitatively characterized for the polymorph composition of its benzimidazole drug substance. Raman spectra of reference samples (pure polymorphs A, B and C in lactose at a concentration of 5%, m/m) were compared with the spectra of benzimidazole samples with a known polymorph composition and with the spectra of uncharacterized premixes. The raw intensities of 78 selected wavenumbers were vector-normalized and application of stepwise linear regression models estimated the relative quantities of the benzimidazole-drug polymorphs A, B and C in the different samples. Modelling results of the samples with known polymorph composition were in compliance with the expected concentrations, validating the proposed methodology. The benzimidazole drug substance in the premixes was predominantly polymorph B. Although statistically not significant, some traces of polymorph A could not be ruled out. Similar analyses were performed to evaluate the solid-state stability of the benzimidazole drug substance in another drug formulation, i.e. a suspension-emulsion. Suspension-emulsions originally determined as containing polymorph B benzimidazole drug substance were stored for 12 months at 25 degrees C/60%RH. FT-Raman spectroscopy revealed that no polymorph transformations occurred during this storage.


On the structure of immune-stimulating saponin-lipid complexes (iscoms).
Immune-stimulating complexes (iscoms) are stable complexes of cholesterol, phospholipid and Quil A, a triterpene saponin mixture in the size range from 40 to 100 nm. They can be used as antigen carriers in subunit vaccines. In this paper it is demonstrated that iscoms are rigid, negatively charged vesicles in which small water soluble molecules like carboxyfluorescein cannot be retained. The negative zeta-potential prevents iscoms from aggregation. The chemical composition of iscoms in one dispersion varied considerably. A typical example of the composition of iscoms is cholesterol/phospholipid/Quil A = 1.0:1.2:6.2 by weight for the iscom matrix, that is iscoms without antigen, and 1.0:1.3:5.1 for antigen-containing iscoms. A hypothetical model for the structure of the iscom matrix and related structures is presented, based on analytical chemical, physico-chemical and electronmicroscopic data. In this model iscoms are considered to be multi-micellar structures, shaped and stabilized by hydrophobic interactions, electrostatic repulsion, steric factors and possibly hydrogen bonds. The individual micelles are relatively flat, ring-shaped structures, the center offering space for one of the two bulky sugar chains of the saponins.


Cytochrome P450 mediated metabolic activation of chrysophanol.
Chrysophanol, a major anthraquinone component occurring in many traditional Chinese herbs, is accepted as important active component with various pharmacological actions such as antibacterial and anticancer activity. Previous studies demonstrated that exposure to chrysophanol induced cytotoxicity, but the mechanisms of the toxic effects remain unknown. In the present metabolism study, three oxidative metabolites (M1-M3, aloe-emodine, 7-hydroxychrysophanol, and 2-hydroxychrysophanol) and five GSH conjugates (M4-M8) were detected in rat and human liver microsomal incubations of chrysophanol supplemented with GSH, and the formation of the metabolites was NADPH dependent except M4 and M5. M4 and M5 were directly derived from parent compound chrysophanol, M6 arose from M2, and M7 and M8 resulted from the oxidation of M4 and M5. Metabolites M5 and M6 were also observed in bile of rats after exposure to chrysophanol, M1-M3 and one NAC conjugate (M9) were detected in urine of rats administrated chrysophanol, and urinary metabolite M9 originated from the degradation of biliary GSH conjugation M6. Recombinant P450 enzyme incubation and microsome inhibition studies demonstrated that P450 1A2 was the primary enzyme responsible for the metabolic activation of chrysophanol and that P450 2B6 and P450 3A4 also participated in the generation of the oxidative metabolites. These findings helped us to understand the mechanisms of chrysophanol-induced cytotoxicity.


Concurrent conventional & high-definition transcranial direct current stimulation for treatment of schizophrenia with co-morbid obsessive-compulsive disorder: A case report.
Non-invasive brain stimulation techniques such as conventional transcranial direct current stimulation (tDCS) and high definition tDCS (HD-tDCS) are increasingly being used as add-on treatment options in schizophrenia and obsessive-compulsive disorder (OCD). This is reporting of the use of a novel accelerated, symptom-specific, add-on tDCS (combining conventional and high definition) protocol in a patient with both schizophrenia and OCD. The intervention showed clinical utility by reducing both schizophrenia and OCD symptoms.


The First 40 Years of AIDS: Promising Programs, Limited Success.
HIV-prevention program planning, implementation, and evaluation began in the United States shortly after reports of a mysterious, apparently acquired, immune deficiency syndrome appeared in summer 1981. In San Francisco, New York City, and elsewhere, members of LGBT communities responded by providing accurate information, giving support, and raising money. During the first decade of the AIDS pandemic (1981-1990), social and behavioral scientists contributed by designing theory-based and practical interventions, combining interventions into programs, and measuring impact on behavior change and HIV incidence. In the second decade (1991-2000), federal, state, and local agencies and organizations played a more prominent role in establishing policies and procedures, funding research and programs, and determining the direction of intervention efforts. In the third decade (2001-2010), biomedical interventions were prioritized over behavioral interventions and have dominated attempts in the fourth decade (2011-2020) to integrate biomedical, behavioral, and structural interventions into coherent, efficient, and cost-effective programs to end AIDS.


[Manifestations and prognosis of thyrotropin-secreting pituitary adenomas: a case series of three patients].
Among pituitary adenomas, thyrotropinomas were previously considered as extremely rare and resistant to therapy. However, the common use of the sensitive TSH measurement and the improvement of pituitary imaging have modified their clinical and hormonal presentation. We here report three cases of TSH secreting pituitary adenoma that highlight the great diversity of the clinical, hormonal and morphological presentation, and their better prognosis. In the presence of an inappropriate TSH secretion, the main differential diagnosis is the syndrome of thyroid hormone resistance. The role of somatostatin analogue prior to the surgical treatment of TSH secreting pituitary adenoma is also underlined in this report. The increasing frequency and early diagnosis of TSH secreting pituitary adenoma may be explained by ultrasensitive methods now used for TSH measurement and progress in pituitary imaging, mainly with MRI. This changing spectrum in the presentation and the excellent response to somatostatin analogues improved in the prognosis of the disease.


The solute carrier transporters and the brain: Physiological and pharmacological implications.
Solute carriers (SLCs) are the largest family of transmembrane transporters that determine the exchange of various substances, including nutrients, ions, metabolites, and drugs across biological membranes. To date, the presence of about 287 SLC genes have been identified in the brain, among which mutations or the resultant dysfunctions of 71 SLC genes have been reported to be correlated with human brain disorders. Although increasing interest in SLCs have focused on drug development, SLCs are currently still under-explored as drug targets, especially in the brain. We summarize the main substrates and functions of SLCs that are expressed in the brain, with an emphasis on selected SLCs that are important physiologically, pathologically, and pharmacologically in the blood-brain barrier, astrocytes, and neurons. Evidence suggests that a fraction of SLCs are regulated along with the occurrences of brain disorders, among which epilepsy, neurodegenerative diseases, and autism are representative. Given the review of SLCs involved in the onset and procession of brain disorders, we hope these SLCs will be screened as promising drug targets to improve drug delivery to the brain.


Acute Liver Failure from Herpes Simplex Virus in an Immunocompetent Patient Due to Direct Inoculation of the Peritoneum.
Herpes simplex virus (HSV) hepatitis is a rare cause of acute liver failure (ALF). It carries a mortality rate of 80% if untreated, thus early identification and treatment are critical. Without high clinical suspicion, HSV hepatitis is difficult to diagnose. A 48-year-old Hispanic female presented with a 4-day history of abdominal pain and a vaginal cuff tear requiring laparoscopic repair. She subsequently developed postsurgical disseminated HSV, resulting in ALF. Acyclovir was initiated, but she was resistant to treatment. She was given additional foscarnet and responded without requiring a liver transplant.


Variability in mRNA expression of ABC- and SLC-transporters in human intestinal cells: comparison between human segments and Caco-2 cells.
The Caco-2 cell monolayer model is widely used as a tool for evaluating human intestinal permeability and interaction with transporters. Therefore, we determined mRNA levels for 15 of the most frequently studied uptake and efflux transporters (MDR1, MRP2-3, BCRP, OCTN2, PepT1, OATP-B, OATP8, OCT1-3, OAT1-3, MCT1) using real-time PCR in Caco-2 cells and in human jejunum and colon. The expression levels in the Caco-2 cells did not significantly vary between different passages (p29-43) and batches for any of the genes measured. However, levels increased with culture time (1-5 weeks) for PepT1, MDR1, MRP2, OATP-B and BCRP. The general rank order of the gene expression levels in Caco-2 cells was established as follows: MRP2>OATP-B>PepT1>>MDR1>MCT1 approximately MRP3 approximately BCRP approximately OCTN2>>OCT3>OCT1>OAT2. Four genes were absent: OATP8, OCT2, OAT1, and OAT3. Ranking of 11 expressed genes showed a significant correlation between human jejunum and 2-5-week-old Caco-2 cells. The expression profile in colon was, however, very different compared to both Caco-2 cells and jejunum. We conclude that the Caco-2 cells in our hands express similar transporters as the human jejunum, but are different from colon, indicating their usefulness for obtaining small intestinal transport data. In addition, we also suggest that cells with a well-defined range of culture ages should be used to minimize variability in data from experiments and even erroneous conclusions.


Silica Nanoparticles Induce Hepatotoxicity by Triggering Oxidative Damage, Apoptosis, and Bax-Bcl2 Signaling Pathway.
The increase in the usage of silica nanoparticles (SiNPs) in the industrial and medical fields has raised concerns about their possible adverse effects on human health. The present study aimed to investigate the potential adverse effects of SiNPs at daily doses of 25 and 100 mg/kg body weight intraperitoneally (i.p.) for 28 consecutive days on markers of liver damage in adult male rats. Results revealed that SiNPs induced a marked increase in serum markers of liver damage, including lactate dehydrogenase (LDH), alanine aminotransferase (ALAT), and aspartate aminotransferase (ASAT). SiNPs also induced an elevation of reactive oxygen species (ROS) production in liver, along with an increase in oxidative stress markers (NO, MDA, PCO, and H2O2), and a decrease in antioxidant enzyme activities (CAT, SOD, and GPx). Quantitative real-time PCR showed that SiNPs also induced upregulation of pro-apoptotic gene expression (including Bax, p53, Caspase-9/3) and downregulation of anti-apoptotic factors Bcl-2. Moreover, histopathological analysis revealed that SiNPs induced hepatocyte alterations, which was accompanied by sinusoidal dilatation, Kupffer cell hyperplasia, and the presence of inflammatory cells in the liver. Taken together, these data showed that SiNPs trigger hepatic damage through ROS-activated caspase signaling pathway, which plays a fundamental role in SiNP-induced apoptosis in the liver.


Evaluating the burden of endogenous Cushing's syndrome using a web-based questionnaire and validated patient-reported outcome measures.
Endogenous Cushing's syndrome (CS) is a rare endocrine condition caused by chronic oversecretion of cortisol, resulting in a diverse constellation of symptoms. This study examined the ongoing burden of illness (BOI), from the first appearance of symptoms through treatment, which is currently not well evaluated. A quantitative, cross-sectional, web-enabled survey including 5 validated patient reported outcomes (PRO) measures was conducted in patients with CS who had been diagnosed ≥ 6 months prior and who had received ≥ 1 treatment for their endogenous CS at the time of the survey. Fifty-five patients participated in this study; 85% were women. The mean age was 43.4 ± 12.3 years (± standard deviation, SD). On average, respondents reported a 10-year gap between the first occurrence of symptoms and diagnosis; 80% underwent surgical treatment for CS. Respondents experienced symptoms on 16 days in a typical month, and their health-related quality of life was moderately impacted based on the CushingQoL score. Weight gain, muscle fatigue, and weakness were the most common symptoms and 69% percent of patients reported moderate or severe fatigue using the Brief Fatigue Inventory. Following treatment, the occurrence of most symptoms declined over time, although anxiety and pain did not significantly decrease. Overall, 38% of participants reported an annual average of 25 missed workdays due to CS symptoms. These results demonstrate a BOI in CS despite ongoing treatment and illustrate the need for interventions to address persistent symptoms, particularly weight gain, pain, and anxiety.


Mastoid obliteration and external auditory canal reconstruction with silicone block in canal wall down mastoidectomy.
To prevent cavity problems in canal wall down mastoidectomy, silicone block for mastoid obliteration was used. In this retrospective cohort study, 39 patients (21 males and 18 females) underwent canal wall down mastoidectomy and mastoid obliteration using silicone block. We evaluated the postoperative outcome, the time until epithelialization of the cavity, graft success rate, and the hearing outcome. The time until complete epithelialization of the mastoid cavity was 35.5 ± 5.4 days. We had a graft success rate of 100% during the follow-ups. The postoperative evaluation revealed 36 dry ears (92.3%) patients without any cavity problems. However, one ear developed granulation tissue, and two ears had partially exposed silicone block, which required revision mastoidectomy. Regarding hearing outcomes, a complication such as deaf ear was not reported. Silicone block is safe and suitable for mastoid obliteration and external auditory canal reconstruction in canal wall down mastoidectomy. 4.


Aging-Associated Alterations in Mammary Epithelia and Stroma Revealed by Single-Cell RNA Sequencing.
Aging is closely associated with increased susceptibility to breast cancer, yet there have been limited systematic studies of aging-induced alterations in the mammary gland. Here, we leverage high-throughput single-cell RNA sequencing to generate a detailed transcriptomic atlas of young and aged murine mammary tissues. By analyzing epithelial, stromal, and immune cells, we identify age-dependent alterations in cell proportions and gene expression, providing evidence that suggests alveolar maturation and physiological decline. The analysis also uncovers potential pro-tumorigenic mechanisms coupled to the age-associated loss of tumor suppressor function and change in microenvironment. In addition, we identify a rare, age-dependent luminal population co-expressing hormone-sensing and secretory-alveolar lineage markers, as well as two macrophage populations expressing distinct gene signatures, underscoring the complex heterogeneity of the mammary epithelia and stroma. Collectively, this rich single-cell atlas reveals the effects of aging on mammary physiology and can serve as a useful resource for understanding aging-associated cancer risk.


Anisotropic 1 H STD-NMR Spectroscopy: Exploration of Enantiomer-Polypeptide Interactions in Chiral Oriented Environments.
The front cover artwork is provided by Dr. Philippe Lesot's group (NMR in Oriented Media, ICMMO, UMR CNRS 8182) at Université Paris-Saclay, France. The image shows four pieces of a puzzle: the magnet of an NMR spectrometer, the principle of the 1 H STD-NMR experiment and the 3D helical structure of the poly-γ-benzyl-L-glutamate polymer leading to a chiral liquid-crystalline phase that discriminates the enantiomers of a model chiral solute (1-phenethyl alcohol). Putting these pieces of the puzzle together allows us to identify the hydrogen sites of each enantiomer interacting with the polypeptide side chain. These new outcomes are a further step towards a global understanding of the chiral recognition that occurs in such media. Read the full text of the Research Article at 10.1002/cphc.202200508.


Diagnostic yield of repeat evaluation for asymptomatic microscopic hematuria after negative initial workup.
The American Urological Association guideline for asymptomatic microhematuria recommends in patients with a negative initial workup, repeat workup should be considered for those with persistent/recurrent microhematuria. However, there is little data on the yield of repeat evaluation. Our hypothesis was that repeat workup yields a low detection rate of urologic malignancy. We retrospectively reviewed all patients at our institution who underwent microhematuria workup with cystoscopy and upper tract imaging from May 2010 to June 2016. Microhematuria was defined as ≥3 RBCs/HPF on a properly collected specimen in the absence of a benign cause. Demographics, age, smoking history, history of radiation, and findings on repeat cystoscopy and imaging were collected. Our primary endpoint was a new diagnosis of urologic malignancy. Our initial cohort included 1,332 patients, of whom 21 were diagnosed with urothelial carcinoma and 7 with suspicious renal masses on initial workup. A total of 637 patients with negative initial workup had persistent/recurrent microhematuria. Repeat cystoscopy was performed in 161 (25%) patients at a median of 39 months, and repeat upper tract imaging was performed in 317 (50%) patients at a median of 39 months. Overall, repeat cystoscopy revealed new bladder cancer in 2 (1.2%) patients and repeat imaging revealed new suspicious renal mass in 4 (1.3%) patients. We observed a low number of newly diagnosed malignancies among patients with persistent/recurrent asymptomatic microhematuria who had a prior negative workup. Additional research is required to determine the utility of a repeat AMH workup.


A Systematic Review on Intensity Modulated Radiation Therapy for Mediastinal Hodgkin's Lymphoma.
Secondary malignant neoplasms (SMNs) and cardiovascular diseases induced by chemotherapy and radiotherapy represent the main cause of excess mortality for early-stage Hodgkin lymphoma patients, especially when the mediastinum is involved. Conformal radiotherapy techniques such as Intensity-Modulated Radiation Therapy (IMRT) could allow a reduction of the dose to the organs-at-risk (OARs) and therefore limit long-term toxicity. We performed a systematic review of the current literature regarding comparisons between IMRT and conventional photon beam radiotherapy, or between different IMRT techniques, for the treatment of mediastinal lymphoma. IMRT allows a substantial reduction of the volumes of OARs exposed to high doses, reducing the risk of long-term toxicity. This benefit is conterbalanced by the increase of volumes receiving low doses, that could potentially increase the risk of SMNs. Treatment planning should be personalized on patient and disease characteristics. Dedicated techniques such as "butterfly" VMAT often provide the best trade-off.


Evidence of residual micellar structures in a lipid nanocapsule dispersion. A multi-technique approach.
Nanoemulsions are metastable emulsions in the nanometric range which can be obtained using low-energy processes. A decade ago, it was demonstrated that a non-negligible amount of residual surfactant micelles may coexist with the oil nanodroplets in a model oil/surfactant system. Those micelles were called "wasted" micelles as they did not participate in the formation of the nanodroplets. Little attention has been focused on the potential presence or effect of such secondary structures in nanoemulsions used as drug delivery systems. Here, we present an extensive characterization of lipid nanocapsules, a nanoemulsion obtained from a medium-chain triglyceride mixed with a pegylated surfactant by a process comprising a temperature-dependent phase inversion followed by a cold-water quench. Lipid nanocapsules demonstrate a very good shelf stability. First, for clarity and academic purposes, we briefly present the pros and the cons of the various diffusion-based characterization techniques used i.e., multi-angle and single-angle dynamic light scattering, nanoparticle tracking analysis, fluorescence recovery after photobleaching, and diffusometry nuclear magnetic resonance. Then, combining all these techniques, we show that up to 40 wt% of the surfactant is not involved in the lipid nanocapsule construction but forms residual micellar structures. Those micelles also contain a small quantity of medium-chain triglyceride (2 wt% of the initial amount) and encapsulate around 40 wt% of a fluorescent dye originally dispersed in the oily phase.


Evidence and consequences of academic drift in the field of dental research: A bibliometric analysis 2000-2015.
The mission of academic excellence has resulted in a science system that incentivises publications within high impact, often basic science journals, and less in application-oriented journals. For the dental research field this so-called academic drift can result in a research portfolio that moves away from research that serves dental healthcare. Therefore, we examined if and how academic drift has changed the dental research field. Web of Science data were used to develop a network map for dental research containing journal clusters that show similar citation behavior. From the year 2000 up to 2015, we explored the intensity of knowledge exchange between the different clusters through citation relations. Next, we analyzed changes in research focus of dental research institutes in seven countries, in dental research, clinical medicine research, basic science, public health research and other fields. Within the citation network, 85.5% of all references in dental journals concern references to other dental journals. The knowledge contribution of non-dental research fields to dental research was limited during the studied period. At the same time, the share of output of dental research institutes in dental research has declined. The research activity of the dental research institutes increased mainly in basic science while the knowledge input from basic science into dental research did not increase. Our findings suggest that the dental research portfolio is influenced by academic drift. This academic drift has increased the disbalance towards basic science, and presents a challenge for the scientific progress in dental healthcare services.


Research Note: Texture characteristics of wooden breast fillets deboned at different postmortem times.
The study aimed to understand whether deboning time contributes to the altered texture attributes of wooden breast (WB) fillets. To this purpose, 30 unaffected (NORM) and 30 severely affected (WB) carcasses were selected at 15 min postmortem (PM) and allotted into treatments with different deboning times: A (5 NORM + 5 WB; right fillets deboned at 15 min PM), B (5 NORM + 5 WB; right fillets deboned at 3 h PM), and C (5 NORM + 5 WB; right fillets deboned at 6 h PM). Left fillets from each carcass were deboned at 24 h PM. Multiple instrumental texture analyses were performed on the cranial-middle portion of the Pectoralis major muscles. Irrespective of deboning time, all fillets were subjected to a single 30% compression in the raw state at 24 h PM. All fillets were cooked at 24 h PM and subjected to shear force assessments at 48 h PM using blunt Meullenet-Owens razor shear, Meullenet-Owens razor shear, and Warner-Bratzler shear force. Using an ANOVA mixed model, deboning time was evaluated as a fixed effect within muscle condition (sampling session and carcass ID as random effects). Compression force assessment of raw meat at 24 h PM showed that WB fillets exhibited the greatest hardness when they were deboned at 6 h PM (15 min = 35.4 N; 3 h = 30.9 N; 6 h = 48.0 N; 24 h = 30.6 N; P < 0.05). Differently, deboning time had no effect (P > 0.05) on raw compression force values in NORM fillets. In cooked NORM fillets, shear force values were the greatest in fillets deboned at 15 min, and shear force gradually declined with deboning time through 24 h PM. On the contrary, no changes (P > 0.05) in shear values due to different deboning times were observed in cooked WB meat regardless of shear method. These results suggest that early PM changes in breast muscles and their influence on meat texture are different between normal and WB fillets.


Measurement of enzymatic activation energy by continuous heating with microwaves.
A microwave heating method and an associated numerical method for computing the reaction velocities in a single experiment are proposed for rapid determination of the activation energy of enzymatic reactions. Application of this new technique to different enzymatic reactions is reported.


The interaction of opsonins with human polymorphonuclear leucocytes (PMN). I. The influence of human complement (C) and IgG on ingestion and digestion of C-resistant E. coli.
The influence of opsonization on ingestion and digestion of C-resistant E. coli was tested in a phagocytic system consisting of a monolayer of human PMN to which was added E. coli opsonized in different ways. At the end of the phagocytosis, noningested bacteria were separated and the monolayer was removed. The cells were then fractionated. The number of bacteria at different time intervals and the amount and distribution of lysosomal enzymes in the cell fractions were determined. It became apparent that the ingestion of cell-attached bacteria was independent of opsonins. However, the intracellular destruction was enhanced following opsonization with C only, although this was not the result of a greater discharge of lysosomal enzymes into the phagosome. We postulate that sublethal damage of E. coli by C renders the bacteria more sensitive to the bactericidal activity of the phagocytes.


[Yersinia enterocolitica tonsillitis].
We present five cases of tonsillitis secondary to Yersinia enterocolitica. No response to commonly-used antibiotics and continuous professional contact with pigs were common. The definitive diagnosis was established by bacteriological isolation on tonsil tissue samples.


[The Importance of De- /Vascularisation of the Ureter during Reconstruction after Trauma].
Serious errors may occur in the reconstruction of injured ureters if the degree of -devascularisation is not taken in account. The problems involved in ureter reconstruction after traumatic reconstruction are presented by means of two case -studies reviewed by the Expert Committee for Re-viewing Errors in Medical Treatment of the Ärztekammer Nordrhein. In a 55-year-old man there was a persistent drainage disorder of the left kidney following implantation of a Y-prosthesis with "lateralisation of the ureter". Following renewed splint-ing of the ureter and percutaneous nephrostomy, operative ureterolysis with partial resection of the ureter and end-to-end anastomosis resulted in anastomostic necrosis with a ureter-skin fistula. Fol-lowing a renewed nephrostomy, the left kidney had to be removed because of considerable loss in function. In a 66-year-old woman with right-sided salpingo-oophorectomy and right-sided -colonectomy there was extensive devascularisation of the right ureter with complete -severance in the lower third. An emergency uretero-uretero anastomosis led to scar-induced stenosis of the anastomosed segment of the ureter. Following unsuccessful attemps at splinting, the ureter was reimplanted using the psoas hitch procedure. Because of a fistula to the small intestine, relaparotomy with segmental resection of the small intestine and opening of an abscess -because of wound infection were necessary. A satisfactory final result was obtained after about nine months. When reconstructing damaged -ureters, numerous factors must be taken into account, e. g., the type, extent and location of the -damage to the ureter(s), the time of detection, the duration and severity of the sequelae for the kidney(s), the patient's concomitant condition and the suitability of the reconstruction procedure. Further-more, the degree of de- / vascularisation is of decisive impor-tance.


Prevention of Unplanned Hospital Admissions in Multimorbid Patients Using Computational Modeling: Observational Retrospective Cohort Study.
Enhanced management of multimorbidity constitutes a major clinical challenge. Multimorbidity shows well-established causal relationships with the high use of health care resources and, specifically, with unplanned hospital admissions. Enhanced patient stratification is vital for achieving effectiveness through personalized postdischarge service selection. The study has a 2-fold aim: (1) generation and assessment of predictive models of mortality and readmission at 90 days after discharge; and (2) characterization of patients' profiles for personalized service selection purposes. Gradient boosting techniques were used to generate predictive models based on multisource data (registries, clinical/functional and social support) from 761 nonsurgical patients admitted in a tertiary hospital over 12 months (October 2017 to November 2018). K-means clustering was used to characterize patient profiles. Performance (area under the receiver operating characteristic curve, sensitivity, and specificity) of the predictive models was 0.82, 0.78, and 0.70 and 0.72, 0.70, and 0.63 for mortality and readmissions, respectively. A total of 4 patients' profiles were identified. In brief, the reference patients (cluster 1; 281/761, 36.9%), 53.7% (151/281) men and mean age of 71 (SD 16) years, showed 3.6% (10/281) mortality and 15.7% (44/281) readmissions at 90 days following discharge. The unhealthy lifestyle habit profile (cluster 2; 179/761, 23.5%) predominantly comprised males (137/179, 76.5%) with similar age, mean 70 (SD 13) years, but showed slightly higher mortality (10/179, 5.6%) and markedly higher readmission rate (49/179, 27.4%). Patients in the frailty profile (cluster 3; 152/761, 19.9%) were older (mean 81 years, SD 13 years) and predominantly female (63/152, 41.4%, males). They showed medical complexity with a high level of social vulnerability and the highest mortality rate (23/152, 15.1%), but with a similar hospitalization rate (39/152, 25.7%) compared with cluster 2. Finally, the medical complexity profile (cluster 4; 149/761, 19.6%), mean age 83 (SD 9) years, 55.7% (83/149) males, showed the highest clinical complexity resulting in 12.8% (19/149) mortality and the highest readmission rate (56/149, 37.6%). The results indicated the potential to predict mortality and morbidity-related adverse events leading to unplanned hospital readmissions. The resulting patient profiles fostered recommendations for personalized service selection with the capacity for value generation.


"Fast track surgery" in the north-west of Italy: influence on the orientation of surgical practice.
Fast track surgery is a peri-operative management model, including different strategies to improve patients' convalescence, avoid metabolic alterations, reduce complications, and shorten hospital stay. Prerequisite is coordination between different practitioners (surgeon, anaesthetist, nurse, nutritionist, physiotherapist). The purpose of our investigation is to understand the level of fast track surgery application in Piedmont and to evidence analogies and differences among departments. We projected an investigation proposing, to every surgery department in Piedmont, a multiple-choice questionnaire evaluating the level of fast track surgery peri-operative interventions' application. Data analysis was conducted in two points of view: the transversal one with an overview of answer's percentages, the longitudinal one correlating data through Pearson's index (r). We collected answers by 78 % of balloted departments (38 on 49). Transversal analysis, including the evaluation of percentages of each question, shows that intra-operative period is the most influenced by fast track principles, and that only 12 departments of 38 apply complete protocols. Longitudinal analysis, estimating the whole of each department's answers, demonstrates the absence of statistical significance in the correlation between fast track surgery application and territorial (r = 0.18), economic (r = 0.31), or age (r = 0.06) variables. Influence of fast track surgery is significantly present in our territory, even though it is not fully concretized in protocols. The choice of fast track depends on the instruction, the environment and the sensibility of each surgeon. Knowledge of geographic distribution of departments applying this model can be useful to organize common protocols, starting from more experienced hospitals.


Clinical course of venous thromboembolism following abdominally based microsurgical breast reconstruction: A case series.
Venous thromboembolism (VTE) is a potentially devastating complication following abdominally based microsurgical breast reconstruction, with a reported incidence of 0.08-4%. The authors aim to describe disease presentation and clinical course following VTE diagnosis in patients within their practice. A retrospective chart review identified patients who underwent microsurgical breast reconstruction from January 2007 through December 2018. Patients with VTE diagnosed within 90 days of surgery were included. Demographics, co-morbidities, signs and symptoms, and characteristics of oncologic, surgical, and post-operative care were analyzed. Seven hundred one patients underwent microsurgical breast reconstruction. Eleven patients with pulmonary embolism (PE) and four with deep vein thrombosis (DVT) were identified, resulting in VTE incidence of 2.1% (0.57% DVT, 1.6% PE). Patients were on average 51 years old and had an average body mass index (BMI) of 31.7 kg/m2. Two had a history of VTE, and none had a known hypercoagulable disorder. Using the 2005 Caprini model, all were high risk and seven were highest risk. Among those with PE, the most common symptom was shortness of breath, and the most common signs were desaturation or supplemental oxygen requirements. VTE was diagnosed on average 14.2 days post-operatively (range 2-52 days). VTE is an infrequent complication following abdominally based microsurgical breast reconstruction. We recommend a high index of suspicion in women reporting shortness of breath or having desaturation, especially in those with high BMI, high Caprini scores, post-operative complications, or early return to the operating room.


Biomaterial-associated infection: locating the finish line in the race for the surface.
Biomaterial-associated infections occur on both permanent implants and temporary devices for restoration or support of human functions. Despite increasing use of biomaterials in an aging society, comparatively few biomaterials have been designed that effectively reduce the incidence of biomaterial-associated infections. This review provides design guidelines for infection-reducing strategies based on the concept that the fate of biomaterial implants or devices is a competition between host tissue cell integration and bacterial colonization at their surfaces.


Acid-base characterization, coordination properties towards copper(II) ions and DNA interaction studies of ribavirin, an antiviral drug.
We have studied processes of copper(II) ion binding by ribavirin, an antiviral agent used in treating hepatitis C, which is accompanied usually by an increased copper level in the serum and liver tissue. Protonation equilibria and Cu(II) binding were investigated using the UV-visible, EPR and NMR spectroscopic techniques as well as the DFT (density functional theory) calculations. The spectroscopic data suggest that the first complex is formed in the water solution at pH as low as 0.5. In this compound Cu(II) ion is bound to one of the nitrogen atoms from the triazole ring. Above pH6.0, the metal ion is surrounded by two nitrogen and two oxygen atoms from two ligand molecules. The DFT calculations allowed to determine the exact structure of this complex. We found that in the lowest energy isomer two molecules of the ligand coordinate via O and N4 atoms in trans positions. The hypothetical oxidative properties of the investigated system were also examined. It proved not to generate plasmid DNA scission products. However, the calf thymus (CT)-DNA binding studies showed that it reacts with ribavirin and its cupric complex. Moreover, the interaction with the complex is much more efficient.


Sarcopenic obesity is associated with a faster decline in renal function in people with type 2 diabetes.
To evaluate the association between sarcopenic obesity and the decline in estimated GFR in people with type 2 diabetes. We enrolled 745 people with type 2 diabetes (mean age 64.6 years, 53.6% men). Body composition was evaluated using dual-energy X-ray absorptiometry. Skeletal muscle index, calculated as appendicular non-fat mass (kg) divided by height squared (m2 ), was used to determine sarcopenia. Sarcopenic obesity was defined as the coexistence of sarcopenia and a ratio of android to gynoid fat mass greater than the median values in each gender. The association of sarcopenic obesity both with the annual rate of decline in estimated GFR and a >30% decline in estimated GFR was evaluated using multivariate linear regression models and Cox proportional hazard models, respectively. Participants with sarcopenic obesity were at an increased risk of a high annual rate of decline in estimated GFR, even after adjustment for the confounding variables (standardized β = -0.228, P <0.001). Sarcopenic obesity was also significantly associated with risk of a >30% decline in estimated GFR (hazard ratio 4.52, 95% CI 2.16-9.47; P < 0.01) in multivariate model. Sarcopenic obesity evaluated by dual energy X-ray absorptiometry is associated with a faster decline in renal function in people with type 2 diabetes.


Efficacy and selectivity of tumor-treating field therapy for triple-negative breast cancer cells via in-house delivery device.
Triple-negative breast cancer continues to be one of the leading causes of death in women, making up 7% of all cancer deaths. Tumor-treating electric fields are low-energy, low-frequency oscillating electric fields that induce an anti-proliferative effect on mitotic cells in glioblastoma multiforme, non-small cell lung cancer, and ovarian cancer. Little is known about effects of tumor-treating fields on triple-negative breast cancer and known research for tumor-treating fields only utilizes low (< 3 V/cm) electric field intensities. We have developed an in-house field delivery device capable of high levels of customization to explore a much wider variety of electric field and treatment parameters. Furthermore, we investigated the selectivity of tumor-treating field treatment between triple-negative breast cancer and human breast epithelial cells. Tumor-treating fields show greatest efficacy against triple-negative breast cancer cell lines between 1 and 3 V/cm electric field intensities while having little effect on epithelial cells. These results provide a clear therapeutic window for tumor-treating field delivery to triple-negative breast cancer.


The conducting tissue in the adult chicken atria. A histological and immunohistochemical analysis.
A three-dimensional reconstruction from serial sections of adult chicken heart was made to verify whether Purkinje cells, that can be recognized by a number of well-known histological criteria, form specialized tracts in the adult chicken atria. This reconstruction revealed a loosely arranged network of Purkinje cells connecting the two atria. This network has not been described before. No tracts could be detected between the sinoatrial and the atrioventricular nodes. These atrial Purkinje cells express the atrial and ventricular myosin isoform, as determined by the use of monoclonal antibodies that were prepared against atrial and ventricular myosin isoform, respectively. Some atrial myocytes that are topographically closely related to the Purkinje cells and that cannot be distinguished from the surrounding myocytes with conventional histological criteria, express, apart from the atrial myosin isoform, also the ventricular myosin isoform. The similar expression pattern of these two cell types and their close topographical relationship suggest the presence of a more elaborate system specialized in conduction than the well-known conductive system found with conventional histological techniques.


[Pharmacological influences on the brain level and transport of GABA. I) Effect of various antipileptic drugs on brain levels of GABA].
The effects of some antipileptic drugs on the level of GABA in the mouse brain was studied using standardized chromatographic methods. Diphenylhydantoin and carbamazepine determined a marked increase of the cerebral GABA level, whereas phenobarbital, diazepam a,d clonazepam were without effect. The results seem to indicate that diphenyihydantoin and carbamazepine, by increasing the GABA level, may act through an enchancement of the GABA-ergic transmission.


Cyclin-dependent kinase 9 (CDK9) predicts recurrence in Middle Eastern epithelial ovarian cancer.
Cyclin-dependent kinase 9 (CDK9) has been shown to play an important role in tumorigenesis of several malignancies. However, the expression of CDK9 in ovarian cancer from Middle Eastern ethnicity remains unknown. A tissue microarray of 441 epithelial ovarian cancer (EOC) samples was used to study the expression of CDK9 immunohistochemically and their clinico-pathological associations were determined. Cox proportional hazards regression model was used for univariate and multivariate analysis of recurrence-free survival. CDK9 over-expression was noted in 56.2 % (248/441) of EOCs and was associated with adverse clinico-pathological parameters such as distant metastasis (p < 0.0001), stage IV tumors (p < 0.0001), tumor recurrence (p = 0.0105) and high Ki-67 index (p < 0.0001). Importantly, CDK9 over-expression was an independent predictor of poor recurrence-free survival (Hazard ratio = 1.51; 95 % confidence interval = 1.15-1.98; p = 0.0030). We also found that CDK9 outperforms Ki-67 as a predictor of tumor recurrence in EOC. Our results show that CDK9 expression correlates with markers of advanced disease in Middle Eastern EOC and is also a prognostic marker. CDK9 overexpression also identifies a subset of patients with highest likelihood of recurrence across the patient cohort. These patients may benefit from additional alternative therapies targeting CKD9.


Physician counseling of pregnant women about active and secondhand smoking in Argentina.
Describe physicians' practices of smoking cessation and secondhand smoke (SHS) exposure counseling during prenatal visits. Cross-sectional survey. A total of 13 public and private hospitals from three cities in Argentina. A total of 300 obstetrician/gynecologists. Self-administered survey included knowledge and attitudes about tobacco use during pregnancy, frequency, type and duration of smoking cessation counseling, barriers to counseling, communication skills, level of understanding and personal smoking history. Composite outcomes of four items, each representative of counseling on smoking cessation and SHS exposure. A total of 235 (78.3%) questionnaires were completed; 54.5% men, mean age 45, 35% current smokers. Only 22% had received training in smoking cessation counseling and 48.5% reported insufficient knowledge to provide smoking cessation advice. Although 88.9% always or almost always advised women to stop smoking, 75% believed it was acceptable for pregnant women to smoke up to 6 cigarettes per day. The risk of SHS exposure was 'always or almost always discussed' by only 34.5% of physicians. Multivariate logistic regression showed that lack of training was associated with less counseling about smoking cessation (OR 0.18; 95% CI 0.04-0.82) and SHS exposure (OR 0.27; 95% CI 0.12-0.59). Current compared to never smokers had lower odds of smoking cessation counseling (OR 0.39; 95% CI 0.05-0.82). Current smokers were less likely than former smokers to counsel about SHS (OR 0.25; 95% CI 0.11-0.62). Smoking cessation counseling during pregnancy in Argentina occurs infrequently, interventions are needed to assist physicians motivate and counsel women to quit smoking and avoid SHS exposure. Physicians taking care of pregnant women also need to quit smoking.


Experience with outlet strut fracture of the Björk-Shiley convexoconcave mitral valve prosthesis.
Within a period of 15 months, we encountered five patients with outlet strut fracture of the Björk-Shiley convexoconcave mitral valve prosthesis. All of the patients had acute failure of the left side of the heart and pulmonary edema. The diagnosis was made by review of the chest roentgenogram which showed features of pulmonary edema; both the outlet strut and the tilting disc were missing from the mitral prosthesis. The "missing" components might be found in the left side of the heart or in the course of the thoracic aorta. Emergency reoperation was carried out on three patients, and there were two survivors. It is essential to recognize this potentially fatal complication and undertake immediate surgical intervention. We performed the original valvular replacement operations in four of the five patients. The probability of outlet strut fracture occurring in our series of 237 patients with a Björk-Shiley convexoconcave mitral valve prosthesis is 1.69%, or 0.076 per 100 patient months.


Depression and anxiety disorders in children and adolescents with velo-cardio-facial syndrome (VCFS).
Velo-cardio-facial syndrome (VCFS) is characterized by a high prevalence of depression and anxiety disorders in childhood and adolescence. These disorders are a source of great impairment in everyday functioning, as well as important risk factors for the emergence of later psychotic disorders. Impairment in daily and social functioning as well as loss of IQ throughout growth are also are well-established correlates of the VCFS. This study aimed to confirm the high prevalence of depression and anxiety disorders. The second objective was to ascertain the correlation between anxious and depressive symptoms and the decline in adaptive and cognitive functioning. A total of 73 children and adolescents with VCFS (mean age 11.9 years) underwent psychiatric evaluation. Subjects were further divided into four age groups: ages 6-9, 9-12, 12-15 and 15-18 years. Assessments measuring intelligence, anxious and depressive symptoms, and adaptation skills reported by parents were submitted to a subsample of 62 children (mean age 12.2 years); 62.2 % of the sample showed an anxiety disorder, specific phobia being the most represented at all ages. Lifetime depression concerned 27 % of the sample, peaking at age 12-15 years. Anxious and depressive symptoms and low IQ were significantly associated with low adaptive functioning. Anxiety and depression are common disorders in children and adolescents with VCFS and have a great impact on adaptive functioning. Clinicians should pay great attention to diagnosis and treatment.


[The current jurisdiction of the Federal Administrative Court concerning the public service employer's obligation within the scope of the Public Support Fund for Civil Servants].
Civil servants are entitled to a subsidy for their health insurance expenses provided the medical interventions are basically necessary and the amount of related expenses adequate. The definition of the borderline between accepted and unaccepted expenses for medical treatment is factually based on the criteria of evidence-based medicine. Accordingly, expenses are not refundable if incurred for treatments and medical products whose efficiency is doubtful or which generally lack scientific acceptance, though under narrowly defined conditions, even "outsider methods" might be approved of. Thus, the law governing these subsidies is more closely related to private insurance than to social security law.


21-(4-Methyl-phenyl-sulfon-yl)-4,7,13,16-tetra-oxa-1,10,21-triaza-bicyclo-[8.8.5]tricosane-19,23-dione: an N-tosyl-ated macrobicyclic dilactam.
The macrobicyclic title compound, C(23)H(35)N(3)O(8)S, contains two tertiary amide bridgehead N atoms and a toluene-sulfonamide N atom in the center of the five-atom bridging strand. The mol-ecule has a central cavity that is defined by the 18-membered ring identified by the N(2)O(4) donor atom set and two 15-membered rings with N(3)O(2) donor atom sets. The toluene-sulfonamide N atom adopts an exo orientation with respect to the central cavity, and the tosyl group is oriented on one side of the aza-bridging strand that connects the bridgehead N atoms.


Association of germline or somatic TP53 missense mutation with oncogene amplification in tumors developed in patients with Li-Fraumeni or Li-Fraumeni-like syndrome.
Germline TP53 mutations are found in Li-Fraumeni syndrome (LFS) patients, predisposed to soft tissue sarcoma and other malignancies. The mutations and succeeding genetic events are thought to cause LFS-associated cancer, whose genetic alterations have rarely been investigated. Here, we study two LFS or Li-Fraumeni-like syndrome (LFLS) patients whose cancers showed aggressive phenotypes. Patient 1 with LFS and TP53(R273H) developed a rhabdomyosarcoma twice at the ages of 18 months and 21 years. A single-nucleotide polymorphism array-based analysis revealed two amplicons in the second tumor; one at 5q11.2 containing MAP3K1 and the other at 11q22.2 containing BIRC2/3 and YAP1. Increase of kinase signaling of MAP3K1 along with anti-apoptosis function of BIRC2/3 may have facilitated progression of this tumor. Patient 2 with LFLS and wild-typeTP53 suffered from acute myeloid leukemia. The leukemic cells had TP53(I195T) and two amplicons; one at 8q24.1 containing DEPDC6 and the other at 8q24.2 containing TRIB1, MYC, and PVT1. Quantitative PCR confirmed amplification of the genes and FISH revealed co-amplification of DEPDC6 and PVT1 in the same double minutes. Quantitative RT-PCR revealed increased expression levels of TRIB1, but no or little expression of DEPDC6, MYC, and PVT1. The results indicate that TRIB1 may be the target gene in the amplicon in the leukemia cells. Mutant TP53 can be engaged in pathways triggering gene amplification through impairment of DNA double-stranded break repair. The amplified candidate oncogenes identified in this study may have played a part in cancer development and lead to the poor outcome of LFS or LFLS-associated tumors.


Xenobiotic triacylglycerol formation in isolated hepatocytes.
The formation of neutral lipophilic metabolites from five xenobiotic carboxylic acids was studied in isolated rat hepatocytes. Oleic acid was used as a positive control. Rates of formation of lipids lay in the order: oleic acid greater than phytanic acid greater than ibuprofen greater than 3-phenoxybenzoic acid greater than indomethacin and 3-phenylbutanoic acid (rates were undetectable with the last two substrates). The process was saturable with the maximum rates at about 0.5 mM substrate concentration. Supplementation of the hepatocyte system with glycerol enhanced the yields of lipid products. The hepatocytes also effectively modelled the in vivo metabolism of ibuprofen, 3-phenoxybenzoic acid and 3-phenylbutanoic acid with oxidations and classical conjugation reactions predominating over xenobiotic lipid formation.


Microstructure and Superior Corrosion Resistance of an In-Situ Synthesized NiTi-Based Intermetallic Coating via Laser Melting Deposition.
A nickel-titanium (NiTi)-based intermetallic coating was in-situ synthesized on a Ti-6Al-4V (TC4) substrate via laser melting deposition (LMD) using Ni-20Cr and TC4 powders. Scanning electron microscopy, X-ray diffraction, a digital microhardness tester and an electrochemical analyzer were used to evaluate the microstructure, Vicker's microhardness and electrochemical corrosion resistance of the intermetallic coating. Results indicate that the microstructure of the intermetallic coating is composed of NiTi2, NiTi and Ni3Ti. The measured microhardness achieved is as high as ~850 HV0.2, ~2.5 times larger than that of the TC4 alloy, which can be attributed to the solid solution strengthening of Al and Cr, dispersion strengthening of the intermetallic compounds, and grain refinement strengthening from the rapid cooling of LMD. During the electrochemical corrosion of 3.5% NaCl solution, a large amount of Ti ions were released from the intermetallic coating surface and reacted with Cl- ions to form [TiCl6]2 with an increase in corrosion voltage. In further hydrolysis reactions, TiO2 formation occurred when the ratio of [TiCl6]2- reached a critical value. The in-situ synthesized intermetallic coating can achieve a superior corrosion resistance compared to that of the TC4 alloy.


Improvements in sleep indices during exam stress due to consumption of a Bifidobacterium longum.
Targeting the gut microbiome as an effective therapeutic strategy for psychological disorders has shown promise in recent years. Variation in the composition of the microbiota and restoration of a stable microbiome using targeted interventions (psychobiotics) including Bifidobacteria have shown promise in pre-clinical studies, but more human data is required on the potential health benefits of these live microorganisms. Bifidobacterium including Bif. longum 1714 has been shown to dampen the effects of acute stress in humans. However, its effects over a period of prolonged stress have not been examined. A randomised, placebo-controlled, repeated measures, cross-over intervention study was conducted to examine the effects of a probiotic intervention on measures of stress, cognitive performance, and mood in healthy human volunteers. Twenty male students participated in this crossover study. Post-intervention assessments took place during the university exam period, which was used as a naturalistic chronic stressor. Self-reported measures of stress, depression, sleep quality, physical activity, gastrointestinal symptoms, cognition, and mood were assessed by questionnaire. In addition, tests from the Cambridge Neuropsychological Test Automated Battery (CANTAB) were administered to all participants. Stress and depression scores increased in both placebo and probiotic treated groups during the exam period. While overall sleep quality and duration of sleep improved significantly in the probiotic treated group during exam stress compared with the placebo treated group, B. longum 1714, similar to placebo treatment, showed no efficacy in improving measures of working memory, visual memory, sustained attention or perception. Overall, while B. longum 1714 shows promise in improving sleep quality and duration, it did not alleviate symptoms of chronic stress, depression, or any measure of cognitive assessment. Thus, further mechanistic studies into the ability of B. longum 1714 to modulate sleep during prolonged periods of stress are now warranted.


Contributions from lateral interaction mechanisms to the human ERG can be studied with a two-frequency method.
We present a two-frequency method to investigate lateral interaction components (liERGs) in the human electroretinogram. Adjacent half cycles of sinusoidal gratings were modulated sinusoidally with different temporal frequencies f1 and f2. The liERGs were defined by the Fourier components at the intermodulation frequencies /f1 - f2/ and f1 + f2 which indicate nonlinear interactions between the half cycles. Significant liERGs were found in all subjects with a monotonic increase of the liERG magnitude in the spatial frequency range from f(s)=0.07 to 2.4 cpd. When /f1 - f2/ was below 5 Hz, liERGs were masked by noise intrusions. In a control experiment we demonstrated that the liERGs were not evoked by stray light artifacts. The liERGs may help to further differentiate the responses that are evoked by patterned stimuli within the retina.