The use of the Box-Behnken experimental design in the optimisation and robustness testing of a capillary electrophoresis method for the analysis of ethambutol hydrochloride in a pharmaceutical formulation.
Box-Behnken experimental designs do not appear to be extensively used in optimisation of analytical methods using capillary electrophoresis (CE). This paper describes the use of the Box-Behnken experimental design to optimise the factors affecting the separation of ethambutol hydrochloride (EB), its impurity 2-amino-1-butanol and the internal standard (phenylephrine hydrochloride) in a CE method for a pharmaceutical tablet assay. The three factors studied simultaneously were: buffer pH, buffer concentration and applied electric field, each at three levels. The method was optimised with respect to three responses: resolution between peaks, theoretical plate count and the migration time of the EB peak. A statistical programme, which applies a multiple response optimisation algorithm, was used to calculate and optimise the three responses simultaneously. The optimum conditions were established to be 58.0 mM sodium borate buffer at pH 9.50 and an applied electric field of 412 V/cm. The robustness of the method was also determined and confirmed using a second Box-Behnken design, as part of the validation exercise. System suitability values for the method were derived from the regression surface analysis. The CE method for a pharmaceutical tablet formulation was further validated according to current regulatory requirements, with respect to linearity and range, precision, specificity, accuracy and limit of quantitation. The optimised method gives a fast and efficient separation under 4 min, with complete resolution between the three peaks, and represents an improvement over the existing USP method. It can be concluded that the Box-Behnken experimental design provides a suitable means of optimising and testing the robustness of a CE pharmaceutical method.


Defensive burying following injections of cholecystokinin, bombesin, and LiCl in rats.
Past research indicates that feeding is reduced for animals injected with cholecystokinin and bombesin. One explanation for this effect suggests that these peptides act as natural satiety signals; an opposing view asserts that bombesin and cholecystokinin reduce feeding through malaise. The present experiment tested the basic assumptions associated with these positions using the defensive burying procedure. Groups of rats were given sweetened condensed milk followed by IP injections of bombesin (6, 16, and 32 micrograms/kg), cholecystokinin (0.7, 1.4, and 2.9 micrograms/kg), LiCl (6.4 mg/ml), or saline. The results showed that animals injected with cholecystokinin, bombesin, and LiCl developed learned aversions to the milk and actively buried the milk spout with their bedding. The findings provide further support for the view that bombesin and cholecystokinin induce malaise rather than satiety.


Physico-chemical characterization of asolectin-genistein liposomal system: An approach to analyze its in vitro antioxidant potential and effect in glioma cells viability.
In this study, the interaction between soy isoflavone genistein and asolectin liposomes was investigated by monitoring the effects of isoflavone on lipidic hydration, mobility, location and order. These properties were analyzed by the following techniques: horizontal attenuated total reflection Fourier transform infrared spectroscopy (HATR-FTIR), low-field (1)H nuclear magnetic resonance (NMR), high-field (31)P NMR, zeta potential, differential scanning calorimetry (DSC) and UV-vis spectroscopy. The antioxidant and antitumoral activities of the genistein liposomal system were also studied. The genistein saturation concentration in ASO liposomes corresponded to 484 μM. HATR-FTIR results indicated that genistein influences the dynamics of the lipidic phosphate, choline, carbonyl and acyl chain methylenes groups. At the lipid polar head, HATR-FTIR and (31)P NMR results showed that the isoflavone reduces the hydration degree of the phosphate group, as well as its mobility. Genistein ordered the lipid interfacial carbonyl group, as evidenced by the HATR-FTIR bandwidth analysis. This ordering effect was also observed in the lipidic hydrophobic region, by HATR-FTIR, NMR, DSC and turbidity responses. At the saturation concentration, liposome-loaded genistein inhibits the lipid peroxidation induced by hydroxyl radical in 90.9%. ASO liposome-loaded genistein at 100 μM decreased C6 glioma cell viability by 57% after 72 h of treatment. Results showed an increase of the genistein in vitro activities after its incorporation in liposomes. The data described in this work will contribute to a better understanding of the interaction between genistein and a natural-source membrane and of its influence on isoflavone biological activities. Furthermore, the antitumoral results showed that genistein-based liposomes, which contain natural-sourced lipids, may be promising as a drug delivery system to be used in the glioma therapy.


Edaravone Prevents Retinal Degeneration in Adult Mice Following Optic Nerve Injury.
To assess the therapeutic potential of edaravone, a free radical scavenger that is used for the treatment of acute brain infarction and amyotrophic lateral sclerosis, in a mouse model of optic nerve injury (ONI). Two microliters of edaravone (7.2 mM) or vehicle were injected intraocularly 3 minutes after ONI. Optical coherence tomography, retrograde labeling of retinal ganglion cells (RGCs), histopathology, and immunohistochemical analyses of phosphorylated apoptosis signal-regulating kinase-1 (ASK1) and p38 mitogen-activated protein kinase (MAPK) in the retina were performed after ONI. Reactive oxygen species (ROS) levels were assessed with a CellROX Green Reagent. Edaravone ameliorated ONI-induced ROS production, RGC death, and inner retinal degeneration. Also, activation of the ASK1-p38 MAPK pathway that induces RGC death following ONI was suppressed with edaravone treatment. The results of this study suggest that intraocular administration of edaravone may be a useful treatment for posttraumatic complications.


Autoinducer-2 and bile salts induce c-di-GMP synthesis to repress the T3SS via a T3SS chaperone.
Cyclic di-GMP (c-di-GMP) transduces extracellular stimuli into intracellular responses, coordinating a plethora of important biological processes. Low levels of c-di-GMP are often associated with highly virulent behavior that depends on the type III secretion system (T3SS) effectors encoded, whereas elevated levels of c-di-GMP lead to the repression of T3SSs. However, extracellular signals that modulate c-di-GMP metabolism to control T3SSs and c-di-GMP effectors that relay environmental stimuli to changes in T3SS activity remain largely obscure. Here, we show that the quorum sensing signal autoinducer-2 (AI-2) induces c-di-GMP synthesis via a GAPES1 domain-containing diguanylate cyclase (DGC) YeaJ to repress T3SS-1 gene expression in Salmonella enterica serovar Typhimurium. YeaJ homologs capable of sensing AI-2 are present in many other species belonging to Enterobacterales. We also reveal that taurocholate and taurodeoxycholate bind to the sensory domain of the DGC YedQ to induce intracellular accumulation of c-di-GMP, thus repressing the expression of T3SS-1 genes. Further, we find that c-di-GMP negatively controls the function of T3SSs through binding to the widely conserved CesD/SycD/LcrH family of T3SS chaperones. Our results support a model in which bacteria sense changes in population density and host-derived cues to regulate c-di-GMP synthesis, thereby modulating the activity of T3SSs via a c-di-GMP-responsive T3SS chaperone.


Characterization of transcriptional activities at a divergent promoter of the type VI secretion system in enterohemorrhagic Escherichia coli O157:H7.
The type VI secretion system (T6SS) is a novel secretion system found in many Gram-negative bacteria that plays a role in bacterial competition, virulence, and host immune evasion. The enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain EDL933 has a single functional T6SS gene cluster. In this study, we attempted to characterize the transcriptional pattern of the T6SS effector gene Z0264 in EDL933. Transcriptional analyses showed that Z0264 and other T6SS genes were transcribed in vitro in a growth-phase-dependent manner, but Z0264 was not secreted in the rich medium. Using adapter- and radioactivity-free transcription start site analysis, we identified an overlapping divergent promoter between Z0264 and Z0265. A β-galactosidase assay with truncated promoter regions showed that the divergent promoter is functional. In addition, we demonstrated the role of H-NS as a repressor in the transcription of Z0264. Notably, the cDNA PCR assay showed that the mRNA transcript from the Z0264 promoter did not include the entire main T6SS cluster, suggesting segmented gene expression by multiple promoters in the T6SS cluster. In conclusion, we identified a divergent promoter for Z0264 located in the T6SS cluster of EDL933 and characterized its in vitro transcriptional activity during growth. Our findings provide insights and a preliminary understanding of the regulatory mechanisms underlying T6SS transcription.


SIAP: an intelligent algorithm for multiple prescription pattern recognition based on weighted similarity distances.
Clinical practices have demonstrated that disease treatment can be very complex. Patients with chronic diseases often suffer from more than one disease. Complex diseases are often treated with a variety of drugs, including both primary and auxiliary treatments. This complexity and multidimensionality increase the difficulty of extracting knowledge from clinical data. In this study, we proposed a subgroup identification algorithm for complex prescriptions (SIAP). We applied the SIAP algorithm to identify the importance level of each drug in complex prescriptions. The algorithm quickly classified and determined valid prescription combinations for patients. The algorithm was validated through classification matching of classical prescriptions in traditional Chinese medicine. We collected 376 formulas and their compositions from a formulary to construct a database of standard prescriptions. We also collected 1438 herbal prescriptions from clinical data for automated prescription identification. The prescriptions were divided into training and test sets. Finally, the parameters of the two sub-algorithms of SIAP and SIAP-All, as well as those of the combination algorithm SIAP + All, were optimized on the training set. A comparison analysis was performed against the baseline intersection set rate (ISR) algorithm. The algorithm for this study was implemented with Python 3.6. The SIAP-All and SIAP + All algorithms outperformed the benchmark ISR algorithm in terms of accuracy, recall, and F1 value. The F1 values were 0.7568 for SIAP-All and 0.7799 for SIAP + All, showing improvements of 8.73% and 11.04% over the existing ISR algorithm, respectively. We developed an algorithm, SIAP, to automatically match sub-prescriptions of complex drugs with corresponding standard or classic prescriptions. The matching algorithm weights the drugs in the prescription according to their importance level. The results of this study can help to classify and analyse the drug compositions of complex prescriptions.


Histologic examination of hepatic biopsy samples as a prognostic indicator in dogs undergoing surgical correction of congenital portosystemic shunts: 64 cases (1997-2005).
To determine whether results of histologic examination of hepatic biopsy samples could be used as an indicator of survival time in dogs that underwent surgical correction of a congenital portosystemic shunt (PSS). Retrospective case series. 64 dogs that underwent exploratory laparotomy for an extrahepatic (n = 39) or intrahepatic (25) congenital PSS. All H&E-stained histologic slides of hepatic biopsy samples obtained at the time of surgery were reviewed by a single individual, and severity of histologic abnormalities (ie, arteriolar hyperplasia, biliary hyperplasia, fibrosis, cell swelling, lipidosis, lymphoplasmacytic cholangiohepatitis, suppurative cholangiohepatitis, lipid granulomas, and dilated sinusoids) was graded. A Cox proportional hazards regression model was used to determine whether each histologic feature was associated with survival time. Median follow-up time was 35.7 months, and median survival time was 50.6 months. Thirty-eight dogs were alive at the time of final follow-up; 15 had died of causes associated with the PSS, including 4 that died immediately after surgery; 3 had died of unrelated causes; and 8 were lost to follow-up. None of the histologic features examined were significantly associated with survival time. Findings suggested that results of histologic examination of hepatic biopsy samples obtained at the time of surgery cannot be used to predict long-term outcome in dogs undergoing surgical correction of a PSS.


Fabrication of Dielectric Elastomer Composites by Locking a Pre-Stretched Fibrous TPU Network in EVA.
Dielectric elastomer (DE) composites with high electrical breakdown strength and large voltage-induced deformation were developed by retaining pre-stretched thermoplastic polyurethane (TPU) fibers in ethylene vinyl acetate copolymer (EVA). The microstructure of the candidate E-TPU fiber membrane and EVA coated E-TPU (E-TPU/EVA) film were characterized by scanning electron microscopy (SEM). The quasi-static and dynamic mechanical property, and the electromechanical properties, including the dielectric constant, dielectric loss tangent, and electromechanical sensitivity, of the DE composites were evaluated. Initially, tensile tests demonstrated that the DE composites based on E-TPU/EVAs had a higher elongation at break of above 1000% but a low elastic modulus of approximately 1.7 MPa. Furthermore, dielectric spectroscopy showed that the E-TPU/EVA had a dielectric constant of 4.5 at the frequency of 1000 Hz, which was 1.2 times higher than that of pure EVA film. Finally, it was found from electromechanical test that the voltage induced strain of E-TPU/EVA rose to 6%, nearly 3 times higher than that of pure TPU film, indicating an excellent electromechanical property. The DE composites developed have demonstrated the potential to be good candidate materials in the fields of artificial intelligence, biomimicry and renewable energy.


Impact of advance care planning on the care of patients with heart failure: study protocol for a randomized controlled trial.
Despite the promise and popularity of advance care planning, there is insufficient evidence that advance care planning helps patients to meet their end-of-life care preferences, especially in Asian settings. Thus, the proposed study aims to assess whether patients with advanced heart failure who are receiving advance care planning have a greater likelihood of receiving end-of-life care consistent with their preferences compared to patients receiving usual care. Secondary objectives are to compare differences in health care expenditures, quality of life, anxiety and depression, understanding of own illness, participation in decision-making and concordance with their caregiver's preferences for end-of-life care, between patients with advanced heart failure receiving advance care planning and usual care. This is a two-arm randomized controlled trial of advance care planning versus usual care (control) conducted at two institutions in Singapore. Two hundred and eighty-two patients with advanced heart failure (n = 94 in the advance care planning arm; n = 188 in the control arm receiving usual care) will be recruited from these centers and followed for 1 year or until they die, whichever is earlier. Additionally, the study will include up to one caregiver per patient enrolled. If advance care planning is proven to be effective, the results will help to promote its uptake among health care providers and patients both within Singapore and in other countries. NCT02299180 . Registered on 18 November 2014.


[Cost-effectiveness evaluation of drugs: multidisciplinary collaboration and transparency are key].
A new German law introduced the cost-effectiveness evaluation of drugs. In Germany, the Institute for Quality and Efficiency in Health Care (IQWiG) is responsible for such evaluations. Currently, however, there is a heavy and controversial debate about the correct method to be applied. A recent proposal of IQWiG on the method to be used for cost-effectiveness evaluations has been dismissed by an expert panel. Moreover, previous IQWiG assessments are criticized and the institute is accused of providing insufficient transparency of its evaluation procedures. The head organizations of the compulsory health insurances focus on their own method to analyze and judge new drugs (EVITA), which is claimed to provide the desired results faster than IQWiG. In Germany, the current situation appears obscure and there is the threat of confusing and contradictory methods for cost-effectiveness evaluations yielding inconsistent assessments. This paper aims at outlining the current situation in Germany and providing potential solutions. It requires an early and multidisciplinary collaboration to achieve the goal of a valid and fully transparent cost-effectiveness evaluation.


Correlation of bond strength with surface roughness using a new roughness measurement technique.
The correlation between shear bond strength and surface roughness was investigated using new surface measurement methods. Bonding agents and associated resin composites were applied to set amalgam after mechanically roughening its surface. Surface treatments were noe (as set against glass), 80 grit, and 600 grit abrasive paper. Surface roughness (R(a) as measured parallel and perpendicular (+) to the direction of the polishing scratches and true profile length were measured. A knife-edge was applied (rate = 2.54 mm/min) at the bonding agent/amalgam interface of each sample until failure. Coefficients of determination for mean bond strength vs either roughness (R(a), of profile length were significantly higher for measurements in parallel directions than for those measurements in (+) directions. The shear bond strength to set amalgam for a PENTA-containing adhesives system (L.D. Caulk Division) was not significantly different from that of a PENTA-free adhesive (3M Dental Products Division), even though PENTA has been reported to increase bond strength to nonprecious metals. The shear bond strength of resin composite to amalgam is correlated to surface roughness when it is measured parallel to the polishing scratches. This correlation is significantly lower when surface roughness is measured in the typical manner, perpendicular to the polishing scratches.


The zebrafish stress axis: molecular fallout from the teleost-specific genome duplication event.
The teleost-specific whole genome duplication event 350 million years ago resulted in a variety of duplicated genes that exist in fish today. In this review, we examine whether molecular components involved in the functioning of the hypothalamus-pituitary-interrenal (HPI) axis are present as single or duplicate genes. Specifically, we looked at corticotropin releasing hormone (CRH), adrenocorticotropic hormone (ACTH) and the glucocorticoid receptor (GR). The focus is on zebrafish but a variety of species are covered whenever data is available through literature or genomic database searches. Duplicate CRH genes are retained in the salmoniformes and cypriniformes, and the peptide sequences are very similar or identical. Zebrafish, along with the Acanthopterygii, are the exceptions as they have a single CRH gene. Also, two copies of the proopiomelanocortin (POMC) gene, which encodes for ACTH and other peptides, have been observed in all teleosts except tilapia and sea bass. In zebrafish, ACTH is derived from only one POMC gene, since the cleavage site is mutated in the other gene. All teleosts examined to date have two GRs, including the recent discoveries of duplicate GRs in two species of cyprinids (carp and fathead minnow). Zebrafish are the only known exception with one GR gene. The loss of duplicate genes is not a general feature of the zebrafish genome, but zebrafish have lost the duplicate CRH, ACTH and GR genes in the past 33 million years, after possessing two of each for the previous 300 million years. The evolutionary pressures underlying the rapid loss of these HPI axis genes, and the implications on the development and the functioning of the evolutionarily conserved cortisol stress response in zebrafish are currently unknown.


High-risk human papillomavirus oncogenes disrupt the Fanconi anemia DNA repair pathway by impairing localization and de-ubiquitination of FancD2.
Persistent expression of high-risk HPV oncogenes is necessary for the development of anogenital and oropharyngeal cancers. Here, we show that E6/E7 expressing cells are hypersensitive to DNA crosslinking agent cisplatin and have defects in repairing DNA interstrand crosslinks (ICL). Importantly, we elucidate how E6/E7 attenuate the Fanconi anemia (FA) DNA crosslink repair pathway. Though E6/E7 activated the pathway by increasing FancD2 monoubiquitination and foci formation, they inhibited the completion of the repair by multiple mechanisms. E6/E7 impaired FancD2 colocalization with double-strand breaks (DSB), which subsequently hindered the recruitment of the downstream protein Rad51 to DSB in E6 cells. Further, E6 expression caused delayed FancD2 de-ubiquitination, an important process for effective ICL repair. Delayed FancD2 de-ubiquitination was associated with the increased chromatin retention of FancD2 hindering USP1 de-ubiquitinating activity, and persistently activated ATR/CHK-1/pS565 FancI signaling. E6 mediated p53 degradation did not hamper the cell cycle specific process of FancD2 modifications but abrogated repair by disrupting FancD2 de-ubiquitination. Further, E6 reduced the expression and foci formation of Palb2, which is a repair protein downstream of FancD2. These findings uncover unique mechanisms by which HPV oncogenes contribute to genomic instability and the response to cisplatin therapies.


Personal values in adolescence and psychological distress in adults: A cross-sectional study based on a retrospective recall.
The aim of this study is to examine the relationship between personal values in adolescence retrospectively assessed and psychological distress in adults in a representative sample of community adults in Japan. We used the J-SHINE data collected in 2010 and 2017. Personal values in adolescence were retrospectively measured in the 2017 survey in two ways: (1) value priorities developed from Schwartz's theory of basic values; and (2) commitment to values measured by Personal Values Questionnaire II (PVQ-II). Psychological distress was measured by using K6 in 2010. Multiple regression analysis was conducted to clarify the association between personal values in adolescence and psychological distress in adults, adjusting for sociodemographic variables, smoking, alcohol drinking, and economic status at age 15. Enduring active challenging, cherishing family and friends, and the commitment to values were significantly and negatively associated with psychological distress in adults. Pursuing one's interest was significantly and positively associated with psychological distress. Having a value priority of enduring active challenging, cherishing family and friends, and the commitment to values in adolescence may reduce psychological distress in adults.


Some immunohistochemical features of argyrophilic grain dementia with normal cortical choline acetyltransferase levels but extensive subcortical pathology and markedly reduced dopamine.
Detailed immunohistochemical and biochemical studies are reported on two cases of progressive dementia showing no Alzheimer-type pathology but extensive argyrophilic grains as described previously by Braak and Braak. These cases had no specific clinical features, and the pathology of these brains showed subcortical gliosis (proliferation of astrocytes and microglia) without significant neuronal losses. Interesting novel immunohistochemical findings were the profuse appearance of complement-activated oligodendrocytes and oligodendroglial microtubular masses. Their appearance seems to indicate oligodendroglial reactions to widespread damage of myelinated axons. Cortical levels of choline acetyltransferase were normal, but striatal levels of dopamine and its metabolites were markedly reduced. This disease may be consistent with the criteria for progressive subcortical gliosis.


Long-term analysis of the IFM 99 trials for myeloma: cytogenetic abnormalities [t(4;14), del(17p), 1q gains] play a major role in defining long-term survival.
In multiple myeloma, many prognostic parameters have been proposed. However, all of these predict shorter survival. To identify patients with a longer life expectancy, we updated the data of patients treated in the IFM (Intergroupe Francophone du Myelome) 99-02 and 99-04 trials. A series of 520 patients was analyzed. Median follow-up was 90.5 months. To perform a comprehensive analysis of the major prognostic factors, we reanalyzed all patients for 1q gains [in addition to updating del(13), t(4;14), and del(17p) analyses]. It was possible to identify a subgroup of patients (representing 20% of total patients) with an 8-year survival of 75%. These patients were defined by the absence of t(4;14), del(17p), and 1q gain and β(2)-microglobulin less than 5.5 mg/L. We propose that all patients with newly diagnosed multiple myeloma be evaluated for these three chromosomal changes, not only to define high-risk patients but also to identify those with a longer life expectancy.


Changes in aniseikonia of an axial anisometrope at various stages of orthokeratology lens wear.
To report a case of reduced aniseikonia in a myopic axial anisometrope during orthokeratology (OK) lens wear. A 19-year-old female university student with myopic anisometropia presented for an OK lens fitting consultation. At baseline, perceptual interocular image size difference or aniseikonia of 1.45% was found, with a smaller image seen by the right eye compared to the left eye. The patient was fitted with a pair of OK lenses and interocular image size differences, subjective refraction and corneal topography were measured after 7, 14 and 47 days of overnight lens wear. Hyperopic shifts in central refraction and corresponding flattening of the central cornea was measured during OK treatment. Aniseikonia reduced after OK wear with the most significant change measured from baseline to day 7. Aniseikonia recorded after 7, 14 and 47 nights of lens wear was 0.05%, 0.35% and 0.85%, respectively. Although minimum differences in refractive error between eyes was reached after 47 days of OK, aniseikonia was greater than that measured after 7 and 14 days of OK. This case report demonstrates reduction in aniseikonia with OK lens wear in a myopic patient with axial anisometropia, although this effect was not sustained beyond 1 week of lens wear. As the impact of corneal curvature changes on aniseikonia is not well understood, future studies on the impact of OK on aniseikonia and associated asthenopia is required.


Nutrition therapy for overweight children and adolescents with type 2 diabetes.
The increased number of children and adolescents diagnosed with type 2 diabetes presents new challenges to pediatricians, nurses, and dietitians. Treatment with medications and lifestyle changes is similar to that for adults with type 2 diabetes, but must be developmentally appropriate for children and adolescents. Registered dieticians must apply knowledge from treatment for overweight children and adolescents without diabetes with treatment for adults with type 2 diabetes to provide effective nutrition therapy for this population.


An extra-adrenal abdominal pheochromocytoma causing ectopic ACTH syndrome.
We report a 55-year-old woman with ectopic adrenocorticotropin (ACTH) secretion caused by extra-adrenal pheochromocytoma. The patient presented with a 6-month history of hypertension and diabetes mellitus. Her serum and urinary cortisol levels were extremely high and dexamethasone failed to suppress the cortisol secretion. Her plasma ACTH levels were also elevated (>300 pg/mL) and irresponsive to corticotropin-releasing hormone (CRH) or metyrapone administration. Gel filtration analysis of the patient's plasma detected the existence of large molecular weight ACTH being eluted with a major peak of authentic 1-39 ACTH. Abdominal computed tomographic scan and magnetic resonance imaging revealed a 5-cm paraganglioma located underneath the left kidney, in which (123)I-MIBG tracer specifically accumulated. Bilateral adrenal glands were diffusely enlarged. After surgical removal of the paraganglioma, the patient's clinical symptoms improved and biochemistry normalized including plasma ACTH, urinary free cortisol, and urinary catecholamines. Subsequent histologic evaluation of the transected paranglioma tissue revealed ACTH, synaptin, and chromogranin-A histologically immunostaining. Culture of primary cells collected from the resected paraganglioma demonstrated of in vitro production of ACTH, noradrenaline, and adrenaline. This is the first report of ectopic ACTH syndrome induced by an extra-adrenal abdominal paraganglioma.


The neurology of ambiguity.
One of the primordial functions of the brain is the acquisition of knowledge. The apparatus that it has evolved to do so is flexible enough to allow it to acquire knowledge about unambiguous conditions on the one hand (colour vision being a good example), and about situations that are capable of two or more interpretations, each one of which has equal validity with the others. However, in the latter instance, we can only be conscious of one interpretation at any given moment. The study of ambiguity thus gives us some insights into how activity at different stations of the brain can result in a micro-consciousness for an attribute, and also tell us something about interactions between different cerebral areas that result in several potential micro-conscious correlates, though only one predominates at any given time. Finally, the study of ambiguity also gives us insights into the neurological machinery that artists have tapped to create the ambiguity that is commonly a hallmark of great works of art.


3D Printed Models of Trochlear Dysplasia and Trochleoplasty Simulation for Trainee Education.
Trochlear dysplasia is a major contributor to patellofemoral instability and subsequent failure of isolated soft tissue reconstruction procedures in the treatment of recurrent patellar dislocation and/or subluxation. Trochleoplasty procedures aim to address abnormal osseous trochlear morphologic factors that contribute to patellar maltracking. However, teaching these techniques is limited by the lack of reliable training models for trochlear dysplasia and trochleoplasty simulation. Although a cadaveric knee model of trochlear dysplasia for trochleoplasty simulation has been recently described, cadaveric knees are less amenable for use in trochleoplasty planning and surgeon training because of the absence of reliable, natural dysplastic anatomic relationships, such as suprapatellar spurs due to the rarity of dysplastic cadavers and the high cost of cadaveric specimens. Furthermore, readily available sawbone models represent "normal" osseous trochlear morphology and are difficult to modify and bend due to their material composition. Given this, we have developed a cost-effective, reliable, and anatomically accurate three-dimensional (3D) knee model of trochlear dysplasia for trochleoplasty simulation and trainee education.


Loss of ZBED6 Protects Against Sepsis-Induced Muscle Atrophy by Upregulating DOCK3-Mediated RAC1/PI3K/AKT Signaling Pathway in Pigs.
Sepsis-induced muscle atrophy often increases morbidity and mortality in intensive care unit (ICU) patients, yet neither therapeutic target nor optimal animal model is available for this disease. Here, by modifying the surgical strategy of cecal ligation and puncture (CLP), a novel sepsis pig model is created that for the first time recapitulates the whole course of sepsis in humans. With this model and sepsis patients, increased levels of the transcription factor zinc finger BED-type containing 6 (ZBED6) in skeletal muscle are shown. Protection against sepsis-induced muscle wasting in ZBED6-deficient pigs is further demonstrated. Mechanistically, integrated analysis of RNA-seq and ChIP-seq reveals dedicator of cytokinesis 3 (DOCK3) as the direct target of ZBED6. In septic ZBED6-deficient pigs, DOCK3 expression is increased in skeletal muscle and myocytes, activating the RAC1/PI3K/AKT pathway and protecting against sepsis-induced muscle wasting. Conversely, opposite gene expression patterns and exacerbated muscle wasting are observed in septic ZBED6-overexpressing myotubes. Notably, sepsis patients show increased ZBED6 expression along with reduced DOCK3 and downregulated RAC1/PI3K/AKT pathway. These findings suggest that ZBED6 is a potential therapeutic target for sepsis-induced muscle atrophy, and the established sepsis pig model is a valuable tool for understanding sepsis pathogenesis and developing its therapeutics.


Prevalence of ectoparasites in dogs of Shimoga, Karnataka.
A study was carried out to ascertain the status of ecto-parasites infesting dogs of Shimoga region. A total of 120 dogs with the history of skin problems presented to the local hospitals and local pet clinics of Shimoga region were screened for different ecto-parasites. The ticks, fleas and lice were collected, processed and identified. The skin scrapings were also collected from the affected dogs and processed for identification of mites. Out of 120 dogs examined, 59 (49.1 %) had harboured ecto-parasites. Among 59 infested pet dogs, 22 (37.28 %) positive for Fleas, 18 (30.5 %) for ticks, 09 (15.2 %) for Lice, 07 (11.8 %) for Sarcoptic mange and 03 (5.0 %) were for Demodectic mange conditions. The two species of fleas were identified as Ctenocephalides canis 13 (59 %) and Ctenocephalides felis 9 (41 %). The ticks and lice species were identified as Riphicephalus sanguineus and Trichodectus canis respectively. The mite species infecting dogs were identified as Sarcoptes scabeii and Demodex canis based on the morphological character. The Prevalence of ectoparasites was more in stray and adults dogs compared to pet dogs and puppies respectively.


Bit error rate of focused Gaussian beams in weak oceanic turbulence: comment.
Recent calculations [J. Opt. Soc. Am. A31, 1963 (2014)JOAOD60740-323210.1364/JOSAA.31.001963] of the scintillation index of focused beams did not take into account the second order of the perturbation theory, and therefore led the author to erroneous conclusions regarding dependence of the scintillation index for the focused beams on the aperture size. The log-normal intensity probability distribution used for the Bit Error Rate calculations in [J. Opt. Soc. Am. A31, 1963 (2014)JOAOD60740-323210.1364/JOSAA.31.001963] is also invalid for the wide focused beams. As a result, most of the conclusions of [J. Opt. Soc. Am. A31, 1963 (2014)JOAOD60740-323210.1364/JOSAA.31.001963] are incorrect.


Importance of surface roughness on perfluorooctanoic acid (PFOA) transport in unsaturated porous media.
Understanding the subsurface transport of perfluorooctanoic acid (PFOA) is of considerable interest for evaluating its potential risks to humans and ecosystems. In this study, packed-column experiments were conducted to examine the influence of surface roughness on PFOA transport in unsaturated glass beads, quartz sand and limestone porous media. Results showed decreasing moisture content significantly increased the air-water interfacial adsorption of PFOA and led to greater retardation in all three types of porous media. Particularly, rougher surface (limestone > quartz sand > glass beads) and smaller grain size (i.e. a larger solid specific surface area, SSSA) significantly enhanced PFOA retardation under unsaturated conditions. These results were further supported by bubble column experiments and SSSA analysis of porous media, which demonstrate that except for the factors affecting PFOA transport in solid-water interface (e.g. surface charge and chemical heterogeneity), the greater retardation of PFOA during transport is attributed to the larger air-water interfacial areas associated with rougher surface and smaller grain size and hence greater interfacial adsorption of PFOA. Our results indicated the importance of surface roughness on the retention and transport of PFOA in the unsaturated zone.


Normal reference ranges of inferior vena cava doppler indices from 14 to 40 weeks of gestation.
To establish normal reference ranges of inferior vena cava (IVC) Doppler velocities in normal singleton pregnancies from 14 to 40 weeks. A prospective descriptive study was conducted on normal singleton pregnancies with confirmed gestational age. The sonographic measurements of IVC Doppler indices, including preload index, peak velocity index for vein, and pulsatility index for vein, were determined by experienced sonographers. A total of 645 measurements were performed, ranging from 12 to 30 measurements available for each gestational week. The relationship between gestational age and IVC Doppler indices was not linear. The best-fitted regression equations for the indices are as follows: [Formula: see text] [Formula: see text] [Formula: see text] (GA = gestational age in week) All IVC Doppler indices rapidly declined from 14 to 20 weeks of gestation and then they were relatively constant or slightly decreased throughout the rest of pregnancy. The tables of normal reference ranges were constructed. Normal reference ranges of IVC Doppler indices for each gestational week from 14 to 40 weeks were constructed. This normative data may be a useful noninvasive tool in evaluation of fetal cardiac function.


Solitary cutaneous metastasis of superficial bladder cancer.
Cutaneous metastases from bladder cancer are an uncommon finding. We present a patient with superficial bladder cancer whose first manifestation of disseminated disease was a solitary cutaneous metastasis of the scrotum.


Synthesis of monodisperse organosilica nanoparticles with hollow interiors and porous shells using silica nanospheres as templates.
A versatile method for the formation of monodisperse, bridged silsesquioxane nanoparticles with hollow interiors and porous shells has been developed using silica nanospheres as templates. Tunable size and shell thickness, as well as high surface areas and large pore volumes of the hollow particles, allow for practical application of these nanoparticles in many fields.


Microbiological, clinical and molecular findings of non-typhoidal Salmonella bloodstream infections associated with malaria, Oriental Province, Democratic Republic of the Congo.
In sub-Saharan Africa, non-typhoidal Salmonella (NTS) can cause bloodstream infections, referred to as invasive non-typhoidal Salmonella disease (iNTS disease); it can occur in outbreaks and is often preceded by malaria. Data from Central Africa is limited. Clinical, microbiological and molecular findings of NTS recovered in a blood culture surveillance project (2009-2014) were analyzed. In March-July 2012 there was an epidemic increase in malaria infections in the Oriental Province of the Democratic Republic of the Congo (DRC). In one referral hospital, overall hospital admissions in June 2012 were 2.6 times higher as compared to the same period in the years before and after (336 versus an average of 128 respectively); numbers of malaria cases and blood transfusions were nearly three- and five-fold higher respectively (317 versus 112 and 250 versus 55). Case fatality rates (in-hospital deaths versus all admissions) peaked at 14.6 %. Salmonella Typhimurium and Salmonella Enteritidis together accounted for 88.9 % of pathogens isolated from blood cultures collected during an outreach visit to the affected districts in June 2012. Children infected with Salmonella Enteritidis (33 patient files available) tended to be co-infected with Plasmodium falciparum more often than children infected with Salmonella Typhimurium (40 patients files available) (81.8 % versus 62.5 %). Through the microbiological surveillance project (May 2009-May 2014) 113 unique NTS isolates were collected (28.5 % (113/396) of pathogens); most (95.3 %) were recovered from children < 15 years. Salmonella Typhimurium (n = 54) and Salmonella Enteritidis (n = 56) accounted for 47.8 % and of 49.6 % NTS isolates respectively. Multilocus variable-number tandem-repeat analysis (MLVA) revealed more heterogeneity for Salmonella Typhimurium than for Salmonella Enteritidis. Most (82/96, 85.4 %) NTS isolates that were available for antibiotic susceptibility testing were multidrug resistant. All isolates were susceptible to ceftriaxone and azithromycin. During the peak of an epidemic increase in malaria in the DRC in 2012, a high proportion of multidrug resistant Salmonella Typhimurium and Salmonella Enteritidis were isolated from blood cultures. Overall, the two serovars showed subtle differences in clinical presentation and genetic diversity.


The experimental study of fluorine-leaching ability of granite with different solutions: A new insight into the dynamic of groundwater fluorine levels along coastal zones.
Drinking-water fluorosis is universal along coastal zones, and the seawater or brine water intrusion is occasionally supposed to enrich groundwater fluorine levels. However, there is no conclusive proof, and the laws and mechanisms remain ambiguous. Granite, the common fluorine-bearing rock, is selected and experimented upon to reveal the characteristics and laws of fluorine's leaching ability during the intrusion of seawater. The fluorine-leaching ability increases with the increasing ratios of seawater or brine water, the increasing levels of NaCl or NaHCO3, and the decreasing levels of CaCl2. Such results directly confirm that seawater or brine water intrusion, as well as the conditions of higher Na+, HCO3- and lower Ca2+, promotes fluorine-leaching ability from granite. The intensities of SiOSi, SiOFe, SiOAl bonds decrease but those of OH bonds increase with a higher ratio of seawater or brine water, the higher levels of NaCl or NaHCO3, and the lower levels of CaCl2. This indicates the more silicate dissolution and stronger OH-F exchange evoked by seawater or brine water intrusion are responsible for the higher fluorine-leaching from granite. Therefore, the process of seawater or brine water intrusion should be important for the groundwater enrichment dynamics along coastal zones.


Transcriptome and methylome sequencing reveals altered long non-coding RNA genes expression and their aberrant DNA methylation in equine sarcoids.
Recent publications confirmed that long non-coding RNAs (lncRNAs) perform an essential function in gene-specific transcription regulation. Nevertheless, despite its important role, lncRNA has not yet been described in equine sarcoids, the skin neoplasia of horses. Therefore, the aim of this study is to deepen the knowledge about lncRNA expression in the pathogenesis of equine sarcoids and provide new insight into the regulatory function of lncRNA in the bovine papillomavirus-dependent neoplasia of horse dermal tissues. RNA sequencing (RNA-seq) data from 12 equine sarcoid samples and the corresponding controls were reanalyzed in this study. A total of 3396 differentially expressed (DE) lncRNAs and 128 DElncRNA-DE genes (DEGs) pairs were identified. Differentially expressed lncRNAs predicted target genes were enriched in pathways associated with inter alia the extracellular matrix disassembly and cancer pathways. Furthermore, methylation data from the same samples were integrated into the analysis, and 12 DElncRNAs were described as potentially disturbed by aberrant methylation. In conclusion, this study presents novel data about lncRNA's role in the pathogenesis of equine sarcoids.


Efficacy of Immune Checkpoint Inhibitors against Advanced or Metastatic Neuroendocrine Neoplasms: A Systematic Review and Meta-Analysis.
We performed a systematic review and meta-analysis of the treatment efficacy of immune checkpoint inhibitors (ICIs) in advanced/metastatic neuroendocrine neoplasms (NENs). MEDLINE and EMBASE were searched to identify studies that provide data on treatment response and/or survival outcomes of advanced/metastatic NEN patients treated with ICIs. The overall response rate (ORR) was pooled using a random-effects model. Meta-regression was performed to explore factors influencing the ORR. Individual patient data (IPD) meta-analysis of survival was performed using stratified Cox regression. Ten studies (464 patients) were included. The overall pooled ORR was 15.5% (95% confidence interval (CI), 9.5-24.3%), and it varied according to the primary site (thoracic, 24.7%; gastro-entero-pancreatic, 9.5%), tumor differentiation (poorly differentiated, 22.7%; well-differentiated, 10.4%), and drug regimen (combination, 25.3%; monotherapy, 10.1%). All these variables significantly influenced the ORR. Tumor differentiation was associated with both overall survival and progression-free survival (hazard ratio of poorly differentiated tumors, 4.2 (95% CI, 2.0-8.7) and 2.6 (95% CI, 1.6-4.4), respectively). Thus, the treatment efficacy of ICIs for advanced/metastatic NENs varied according to primary site, tumor differentiation, and drug regimen. Poorly differentiated NENs showed a better ORR than well-differentiated NENs but had a negative impact on survival.


Perioperative Management and Outcomes of Hyperthyroid Patients Unable to Tolerate Antithyroid Drugs.
Prior to thyroidectomy for hyperthyroidism, it is recommended that patients are managed with antithyroid drugs (ATDs) and rendered euthyroid to decrease the risk of thyroid storm. However, not all patients tolerate ATD and the risk of thyroid storm during thyroidectomy in these patients is unclear. Therefore, the aim of this study was to compare the management and outcomes of hyperthyroid patients that were on ATDs prior to surgery to those who were not. A prospectively maintained, single-institution database was queried for all hyperthyroid patients who were initially treated with ATDs and underwent thyroidectomy from January 1, 2012, to June 18, 2018. Patients were divided into two groups: (1) those on ATDs at surgery (ATD group) and (2) those who could not tolerate and stopped ATDs prior to surgery (no-ATD group). Demographic and clinical data were collected. Primary outcomes were readmissions/emergency department visits and postoperative complications within 30 days of thyroidectomy. Of the 248 patients, 231 were in the ATD group and 17 (7%) were in the no-ATD group. There were no mortalities or thyroid storm events in either group. There was no difference in Clavien-Dindo Grade 2 or 3 complications between the two groups. There were no ED visits or 30-day readmissions in the no-ATD group compared to 17 (7%) events in the ATD group (p = 1.0). While it is preferable to render patients euthyroid prior to thyroidectomy for hyperthyroidism, results of this study suggest that when patients cannot tolerate ATDs, it is possible to perform thyroidectomy without increased risk of thyroid storm or intra- and postoperative complications.


High Metabolic Dependence on Oxidative Phosphorylation Drives Sensitivity to Metformin Treatment in MLL/AF9 Acute Myeloid Leukemia.
Acute myeloid leukemia (AML) is a group of hematological cancers with metabolic heterogeneity. Oxidative phosphorylation (OXPHOS) has been reported to play an important role in the function of leukemic stem cells and chemotherapy-resistant cells and are associated with inferior prognosis in AML patients. However, the relationship between metabolic phenotype and genetic mutations are yet to be explored. In the present study, we demonstrate that AML cell lines have high metabolic heterogeneity, and AML cells with MLL/AF9 have upregulated mitochondrial activity and mainly depend on OXPHOS for energy production. Furthermore, we show that metformin repressed the proliferation of MLL/AF9 AML cells by inhibiting mitochondrial respiration. Together, this study demonstrates that AML cells with an MLL/AF9 genotype have a high dependency on OXPHOS and could be therapeutically targeted by metformin.


The neuronal extracellular matrix restricts distribution and internalization of aggregated Tau-protein.
Alzheimer's disease (AD) is a chronic degenerative disorder characterized by fibrillary aggregates of Aß and Tau-protein. Formation and progression of these pathological hallmarks throughout the brain follow a specific spatio-temporal pattern which provides the basis for neuropathological staging. Previously, we could demonstrate that cortical and subcortical neurons are less frequently affected by neurofibrillary degeneration if they are enwrapped by a specialized form of the hyaluronan-based extracellular matrix (ECM), the so called 'perineuronal net' (PN). PNs are composed of large aggregating chondroitin sulfate proteoglycans connected to a hyaluronan backbone, stabilized by link proteins and cross-linked via tenascin-R. Recently, PN-associated neurons were shown to be better protected against iron-induced neurodegeneration compared to neurons without PN, indicating a neuroprotective function. Here, we investigated the role of PNs in distribution and internalization of exogenous Tau-protein by using organotypic slice cultures of wildtype mice as well as mice lacking the ECM-components aggrecan, HAPLN1 or tenascin-R. We could demonstrate that PNs restrict both distribution and internalization of Tau. Accordingly, PN-ensheathed neurons were less frequently affected by Tau-internalization, than neurons without PN. Finally, the PNs as well as their three investigated components were shown to modulate the processes of distribution as well as internalization of Tau.


Hypotony and choroidal detachment as late complications of trabeculectomy.
Seven eyes of six patients with glaucoma developed a flat anterior chamber, hypotony, and choroidal detachment two to 26 months after technically uncomplicated trabeculectomy. No signs warranting immediate surgical intervention, such as wound or bleb leakage, contact between cornea and lens, or massive choroidal detachments touching in the midvitreous cavity were found on initial examination. After treatment with topical corticosteroids and cycloplegic eyedrops, five of seven eyes showed marked clinical improvement within one month. The two remaining eyes eventually returned to normal anterior chamber depth, but required repeat filtration surgery for intraocular pressure control.


Superporous agarose--reticulated vitreous carbon electrodes for electrochemical sandwich bioassays.
We report on the use of flow-through electrodes fabricated from a composite of superporous agarose (SPA) and reticulated vitreous carbon (RVC) for carrying out sandwich bioassays via a model sandwich assay scheme. The flow-through design of the SPA-RVC electrodes allows for ease in solution handling with the use of micropipettors while allowing sandwich assays to be performed on the SPA matrix inside the RVC. A sandwich bioassay was devised for detecting biotinylated bovine serum albumin (b-BSA) as a proof-of-concept scheme to demonstrate applicability of SPA-RVC electrodes to carry out sandwich assays. In this bioassay scheme, SPA-RVC electrodes with avidin molecules immobilized on the SPA matrix were incubated with low quantities of b-BSA followed by incubation with avidinylated alkaline phosphatase (av-ALP). This construct creates a sandwich bioassay whereby b-BSA is sandwiched between the two avidin complexes. Av-ALP labels captured on the bound b-BSA catalytically hydrolyze conversion of 4-aminophenylphosphate (PAPP) to electrochemically active 4-aminophenol (PAP) which is then voltammetrically detected inside the RVC. The lower concentration detection limit for b-BSA was 0.32+/-0.1 ng mL(-1) and the absolute detection limit was 32+/-10 pg. Non-specific binding of av-ALP enzyme labels onto the avidin-activated SPA-RVC electrodes was low. Catalytic generation of PAP by non-specifically bound av-ALP occurs at a rate less than 2% of that for PAP generation by av-ALP in [(SPA-av)-(b-BSA-b)-(av-ALP)] sandwich constructs.


Ornithine decarboxylase in Phycomyces: in vitro and in vivo properties.
The properties of ornithine decarboxylase from Phycomyces blakesleeanus were examined. Enzyme from mycelial cultures was extracted and purified approximately 70-fold. The apparent molecular weight is 96K. The Michaelis constants with respect to ornithine and pyridoxal 5'-phosphate are 90 and 0.37 microM, respectively. Putrescine is a potent competitive inhibitor with a Ki of 75 microM. Exposure of ornithine decarboxylase to sulfhydryl-modifying reagents resulted in a rapid inhibition of activity. In vivo addition of putrescine produced characteristic decreases in cellular ornithine decarboxylase activity. Light stimulation of dark-adapted mycelial cultures also decreased cellular ornithine decarboxylase activity.


Analysis of the role of RsbV, RsbW, and RsbY in regulating {sigma}B activity in Bacillus cereus.
The alternative sigma factor sigma(B) is an important regulator of the stress response of Bacillus cereus. Here, the role of the regulatory proteins RsbV, RsbW, and RsbY in regulating sigma(B) activity in B. cereus is analyzed. Functional characterization of RsbV and RsbW showed that they act as an anti-sigma factor antagonist and an anti-sigma factor, respectively. RsbW can also act as a kinase on RsbV. These data are in line with earlier functional characterizations of RsbV and RsbW homologs in B. subtilis. The rsbY gene is unique to B. cereus and its closest relatives and is predicted to encode a protein with an N-terminal CheY domain and a C-terminal PP2C domain. In an rsbY deletion mutant, the sigma(B) response upon stress exposure was almost completely abolished, but the response could be restored by complementation with full-length rsbY. Expression analysis showed that rsbY is transcribed from both a sigma(A)-dependent promoter and a sigma(B)-dependent promoter. The central role of RsbY in regulating the activity of sigma(B) indicates that in B. cereus, the sigma(B) activation pathway is markedly different from that in other gram-positive bacteria.


Coronary stenting or balloon angioplasty for chronic total coronary occlusions: the Taiwan experience (a single-center report).
The authors conducted this study to compare the restenosis and reocclusion rates of primary balloon angioplasty alone versus angioplasty followed by stenting in Taiwanese patients with chronic total occlusions. They also evaluated whether stenting reduced the incidence of restenosis and improved left ventricular function in these patients. From October 1998 to April 2000, a total of 294 patients with chronic total occlusion (Thrombolysis in Myocardial Infarction grade 0 flow) underwent recanalization using balloon angioplasty alone or followed by stent implantation. Of these, only 129 patients were included after procedural failure and patients lost to follow-up; 62 patients were placed in the stent group, while 67 patients were assigned to the percutaneous transluminal coronary angioplasty (PTCA) group. Coronary angiography was performed at baseline and at 6 months follow-up or earlier if angina or objective evidence of ischemia involving the target vessel or other vessels was present. Procedural success was 60%. Minimal lumen diameter increased significantly after stenting: 2.97 +/-0.41 vs 2.24 +/-0.41 (p < 0.001); 60% of patients in the stent group were free of restenosis, whereas only 33% in the PTCA group were free of restenosis at follow-up. Only 1 patient in the stent group had reocclusion, as opposed to 17 (25%) patients in the PTCA group (p < 0.001). The follow-up minimal lumen diameter (MLD) at 6 months was significantly larger in the stent group: 1.80 +/-0.85 mm vs 1.08 +/-0.82 mm (p < 0.001). Left ventricular function improved in the stent group, but not in the PTCA group (58.44 +/-16.58% to 63.60 +/-14.59% [p < 0.001] vs 54.13 +/-15.66% to 54.31 +/-15.60% [p = 0.885]). More patients had angina in the PTCA group than in the stented group 43 vs 29 (p = 0.053). The postprocedural MLD and reference vessel diameter (RVD) were the strong predictors of restenosis and follow-up MLD (p < 0.001). Stenting of chronically occluded arteries significantly reduced the incidence of reocclusion and restenosis, at the same time improving left ventricular function in these patients. This should be the procedure of choice after successful angioplasty of chronically occluded vessels.


The stimulatory and inhibitory role of the hypothalamus in the regulation of ovulation in grass frog, Rana temporaria L.
In our previous experiments it was found that lesions placed in the infundibular hypothalamus induced an advanced ovulation in hibernating frogs, Rana temporaria. It was suggested that this premature ovulation was the effect of gonadotropin-releasing hormone (GnRH) due to the injury of some hypothalamic area inhibiting its release or its action on the pituitary gonadotrophs. To investigate this hypothesis, the following experiments were undertaken: (1) an attempt to induce ovulation with exogenous GnRH in hibernating frogs; (2) an attempt to inhibit ovulation with anti-GnRH serum in preovulatory hibernating animals nonlesioned and with lesions of the infundibular hypothalamus; and (3) administration of bromocriptine in hibernating animals to assess whether this substance, as an agonist of dopamine, possesses an inhibitory action on the ovulation. The following results were obtained: (1) lesions of the infundibular hypothalamus in hibernating preovulatory females caused an advanced ovulation during hibernation; (2) the exogenous GnRH administered to preovulatory females induced ovulation during hibernation; (3) the anti-GnRH serum injected into hibernating preovulatory lesioned females inhibited preterm ovulation during, but not after, hibernation; (4) the immunoneutralization of endogenous GnRH in nonlesioned females resulted in an inhibition of the posthibernatory ovulation; (5) bromocriptine inhibited posthibernatory ovulation in nonlesioned hibernating animals. Thus, the results of these experiments support the suggestion that induction of accelerated ovulation in lesioned hibernating animals involved the releasing action of GnRH. This action of GnRH seemed to be facilitated by the ablation of inhibitory dopaminergic function of hypothalamus in lesioned animals.


Use of computers and the Internet for health information by patients with epilepsy.
The purpose of this study was to describe computer and Internet use among an online group and a clinic-based group of people with epilepsy. Greater than 95% of the online group and 60% of the clinic group have access to computers and the Internet. More than 99% of the online group and 57% of the clinic group used the Internet to find health information. A majority of people reported being likely to employ an Internet-based self-management program to control their epilepsy. About 43% reported searching for general information on epilepsy, 30% for medication, 23% for specific types of epilepsy, and 20% for treatment. This study found that people with epilepsy have access to computers and the Internet, desire epilepsy-specific information, and are receptive to online health information on how to manage their epilepsy.


Use of ICD-10-CM T codes in hospital claims data to identify adverse drug events in Taiwan.
Adverse drug events (ADEs) are a major public health concern worldwide and may prolong hospital stays, causing a burden on the healthcare system and increasing the associated costs. Therefore, optimizing medication use and reducing ADEs are priorities for public health. Medication safety can be monitored and improved by identifying ADEs. The utilization of diagnoses coded according to the International Statistical Classification of Diseases and Related Health Problems (ICD) system for the identification of ADEs has been firmly established. In Taiwan, however, the validity of recording ADEs on the basis of inpatient ICD-10-CM T codes has not been evaluated. Therefore, this study investigated the potential usefulness of ICD-10-CM T codes in routine hospital data for the identification of ADEs and for increasing the rate of reporting. We use hospital claims data of hospitalized patients from one medical centre in northern Taiwan between 1 July 2016 and 30 June 2018. We defined an ADE to have taken place if an ICD-10-CM T code was present among the primary or secondary diagnosis codes. The inpatients who were discharged with T codes in a primary or secondary diagnosis were identified by the computerized T code information platform, and the retrospective review of the medical charts was performed by pharmacists to confirm the ADEs. 1384 inpatients who were discharged with the relevant T codes in a primary or secondary diagnosis were identified during the study period. Code T36 (poisoning by, adverse effect of or underdosing of systemic antibiotics) was the most common code, accounting for 56.6%, followed by T42 (17.7%; poisoning by, adverse effect of or underdosing of antiepileptic, sedative-hypnotic or antiparkinsonism drug). Overall, 789 clinically significant ADEs were identified after medical chart review. The dermatologic system was the most commonly involved. The overall positive predictive value for a flagged code representing an ADE was 57%. Furthermore, the use of T codes to confirm the number of ADE cases increased the ADE reporting rate by 9.17%. The PPV of ICD-10-CM T codes analysed in our study was insufficient for identifying ADEs during hospitalization. The sensitivity and specificity of this were inadequate. However, the T code system can be used as an auxiliary resource for pharmacists to identify potential ADEs and report the information as prompts on the physician order entry system. When a physician prescribes a drug that may cause an ADE in a patient, an alert is issued to ensure medication safety. In conclusion, the T codes did not perform well in our study and caution should be exercised in their use to identify ADEs on their own.


Possible pitfalls in the study of IgG receptors produced by herpesvirus-infected cells. Brief report.
The presence of specific antibodies to human herpesviruses in certain purified human IgG preparations from commercial sources may produce misleading results when such preparations are used in studied on the IgG receptors produced by virus-infected cells.


Eosinophilic Esophagitis: Incidence, Diagnosis, Management, and Future Directions.
Eosinophilic esophagitis (EoE) is an antigen-mediated esophageal disease defined by the presence of esophageal eosinophilia and symptoms of esophageal dysfunction. The pathophysiology involves an allergen-driven Th2 T cell response that triggers infiltration of eosinophils into the esophagus leading to inflammation, remodeling, and fibrosis. This results in disruption of esophageal function and accompanying symptoms - most notably dysphagia. Effective therapies target inflammation or fibrostenotic complications and include proton pump inhibitors, swallowed topical steroids, dietary exclusion, and dilation. Clinical trials testing promising biologic therapies are ongoing.


Multiplexed single-cell proteomics using SCoPE2.
Many biological systems are composed of diverse single cells. This diversity necessitates functional and molecular single-cell analysis. Single-cell protein analysis has long relied on affinity reagents, but emerging mass-spectrometry methods (either label-free or multiplexed) have enabled quantifying >1,000 proteins per cell while simultaneously increasing the specificity of protein quantification. Here we describe the Single Cell ProtEomics (SCoPE2) protocol, which uses an isobaric carrier to enhance peptide sequence identification. Single cells are isolated by FACS or CellenONE into multiwell plates and lysed by Minimal ProteOmic sample Preparation (mPOP), and their peptides labeled by isobaric mass tags (TMT or TMTpro) for multiplexed analysis. SCoPE2 affords a cost-effective single-cell protein quantification that can be fully automated using widely available equipment and scaled to thousands of single cells. SCoPE2 uses inexpensive reagents and is applicable to any sample that can be processed to a single-cell suspension. The SCoPE2 workflow allows analyzing ~200 single cells per 24 h using only standard commercial equipment. We emphasize experimental steps and benchmarks required for achieving quantitative protein analysis.


Analyzing physical activity impact based on ubiquitous nonlinear dynamics and electroencephalography data.
Understanding complex systems is made easier with the tools provided by the theory of nonlinear dynamic systems. It provides novel ideas, algorithms, and techniques for signal processing, analysis, and classification. Presently, these ideas are being applied to the investigation of how physiological signals evolve. The study applies nonlinear dynamics theory to electroencephalogram (EEG) signals to better comprehend the range of alcoholic mental states. One of the main contributions of this paper is an algorithm for automatically distinguishing between sober and drunken EEG signals based on their salient features. The study utilized various entropy-based features, including ApEn, SampEn, Shannon and Renyi entropies, PE, TS, FE, WE, and KSE, to extract information from EEG signals. To identify the most relevant features, the study employed ranking methods like T-test, Wilcoxon, and Bhattacharyya, and trained SVM classifiers with the selected features. The Bhattacharyya ranking method was found to be the most effective in achieving high classification accuracy, sensitivity, and specificity. Classification accuracy of 95.89%, the sensitivity of 94.43%, and specificity of 96.67% are achieved by the SVM classifier with radial basis function (RBF) for polynomial Kernel using the Bhattacharyya ranking method. From the result, it is clear that the model serves as a cost-effective and accurate decision-support tool for doctors in diagnosing alcoholism and for rehabilitation centres to monitor the effectiveness of interventions aimed at mitigating or reversing brain damage caused by alcoholism.


Expression of EphrinB2 and EphB4 in glioma tissues correlated to the progression of glioma and the prognosis of glioblastoma patients.
The ligand EphrinB2 and the corresponding receptor EphB4 are up-regulated and involved in tumour growth in various human cancers. However, little is known about how this receptor-ligand complex contributes to the progression of glioma. This prompted us to study the association between the expressions of EphrinB2 and EphB4, clinicopathological variables, and glioma patient outcome. Immunohistochemical staining was performed to detect the expression patterns of EphrinB2 and EphB4 in the biopsies from 96 patients with primary gliomas. Kaplan-Meier survival and Cox regression analyses were performed to evaluate the prognosis of patients. Immunohistochemical analysis revealed that the expression of EphrinB2 was significantly correlated with that of EphB4 (r=0.86, p=0.002). EphrinB2 and EphB4 were significantly associated with the Karnofsky performance scale (KPS) score and World Health Organization grades of patients with gliomas, respectively. Especially, the positive expression rates of EphrinB2 and EphB4 were significantly higher in patients with higher grade (both p=0.001) and lower KPS score (p=0.002 and 0.003, respectively). Multivariate Cox regression analysis revealed that EphrinB2 and EphB4 expressions were both independent prognostic factors for progress-free survival of glioblastoma patients (both p=0.02). Our data indicated for the first time that EphrinB2 and EphB4 expressions increase according to the histopathological grade and KPS score of glioma, and their expression levels are related to the progression-free survival of glioblastoma patients.


Lung damage from exposure to pulsed ultrasound.
Motivated by a recent finding that threshold pressures for hemorrhage in mouse lung exposed to the fields of an electrohydraulic lithotripter were less than 2 MPa, we extended the exposures to pulsed ultrasound. Sharply defined thresholds of the order of 1 MPa were found with 10 microseconds length pulses and roughly twice that value for 1 microsecond pulses. The thresholds at 4 MHz are greater than at 1 MHz. The thresholds are comparable for focused and unfocused fields. As would be expected for a cavitation-like phenomenon, temporal average intensity is a very poor predictor of this effect. In the extreme case, lesions were found at temporal average intensities on the order of 1 mW/cm2.


Middle small-bowel segment Lewis score may predict long-term outcomes among patients with quiescent Crohn's disease.
Video capsule endoscopy (VCE) has been proven to accurately diagnose small-bowel inflammation and predict flares among patients with quiescent Crohn's disease (CD). However, data regarding its predictive role in this population over an extended follow-up are scarce. To predict clinical exacerbation and to assess the yield of Lewis score in identifying CD patients with future clinical exacerbation during an extended follow-up (>24 months). A post hoc analysis study. Adult patients with quiescent small-bowel CD who were followed with VCE, inflammatory biomarkers and magnetic resonance enterography in a prospective study (between 2013 and 2018). We extracted extended clinical data (up to April 2022). The primary composite outcome (i.e. clinical exacerbation) was defined as intestinal surgery, endoscopic dilation, CD-related admission, corticosteroid administration, or biological/immunomodulator treatment change during follow-up. Of the 61 patients in the study [median age 29 (24-37) years, male 57.4%, biologic treatment 46.7%], 18 patients met the primary outcome during an extended follow-up [median 58.0 (34.5-93.0) months]. On univariable analysis, complicated [hazard ratio (HR) 7.348, p = 0.002] and stricturing disease phenotype (HR 5.305, p = 0.001) were associated with higher risk for clinical exacerbation during follow-up. A baseline VCE middle small-bowel segment Lewis score (midLS) ⩾ 135 identified patients with future exacerbation [AUC (area under the curve) 0.767, 95% confidence interval (CI) 0.633-0.902, p = 0.001, HR 6.317, 93% negative predictive value], whereas the AUC of the conventional Lewis score was 0.734 (95% CI: 0.589-0.879, p = 0.004). Sensitivity analysis restricted to patients with either complicated (n = 34) or stricturing (n = 26) disease phenotype revealed that midLS still predicted clinical exacerbation during follow-up (AUC 0.747/0.753, respectively), in these patients. MidLS predicts treatment failure in quiescent CD patients (median follow-up of 5 years) independently of disease phenotype.


Long-term evaluation of the central vault after phakic Collamer® lens (ICL) implantation using OCT.
The aim of this work is to evaluate the changes in the central vault after using implantable Collamer® lenses (ICL) to correct myopia. This cohort study included 133 eyes; mean spherical equivalent -9.47 ± 3.71 diopters (D) (range: -2.75 to -22.25 D) who underwent ICL V4 implantation. Besides a complete ophthalmologic examination, the central vaulting was measured with Visante OCT and evaluated between different periods of follow-up (1 week, 1, 3, and 6 months, and yearly postoperatively). Post-operative time ranged from 6 to 73 months. Within the first 6 months, the mean central vaulting dropped from 510 ± 238 μm (range 100 to 940 μm) to 439 ± 231 μm (range 90 to 910) a significant decrease by 71 ± 58 μm (range -170 to 30 μm) (p = 0.028) with a trend to a lower reduction after this period. After 36 month from ICL implantation, this variation tends to be smaller with the majority of the eyes having a mean decrease smaller than 2 μm per month. Initial vault (Spearman Rho = -0.237, p = 0.006) was the factor more significantly associated with decrease in vault; eyes presenting a reduction in vault >100 μm had an average initial vault that was 141 μm and 184 μm higher than eyes with no change or with increase in vault over time, respectively. This study shows a continuous reduction of central vault over time. The reduction is highest during the first 6 months and tends to be lower and slow down over time. The decrease of vaulting was more pronounced in eyes with larger initial vaulting.


Electrical anal stimulation and the prediction of bladder stability after surgical relief of male outflow obstruction.
In an attempt to predict which unstable bladders in males with outflow obstruction would convert to stability following surgery, we performed functional electrical stimulation (FES) of the anus during pre-operative rapid fill cystometry in 13 patients with benign prostatic hypertrophy (BPH) and 5 patients with bladder neck obstruction (BNO), all of whom had associated instability. Instability was suppressed in only 2 patients with BPH. Although suppression was achieved in 4 patients with BNO, no correlation was found with either post-operative stability or symptomatology in this group or the patients with BPH and we conclude that FES has no predictive value in this context.


Loss of CBP causes T cell lymphomagenesis in synergy with p27Kip1 insufficiency.
CBP can function as a tumor suppressor, but the mechanisms that govern oncogenesis in its absence are unknown. Here we show that CBP inactivation in mouse thymocytes leads to lymphoma. Although CBP has been implicated in the transactivation functions of p53, development of these tumors does not seem to involve loss of p53 activity. CBP-null tumors show reduced levels of p27Kip1 and increased levels of cyclin E and Skp2, two oncoproteins that can promote p27Kip1 proteolysis. Reduction of p27Kip1 by introduction of a p27Kip1-null allele into CBP knockout mice accelerates lymphomagenesis and seems to obviate the requirement for Skp2 and cyclin E upregulation. These data suggest that CBP loss mediates lymphomagenesis in cooperation with a mechanism that reduces p27Kip1 abundance.


Organ culture of chicken cecum: morphologic and physiologic observations after 24 and 48 h of culture.
Cecum from chickens, 4 wk old, can best be maintained for 24 h in a serum-free organ culture system using Trowell T8 medium agar sheet at 25 degrees C. As determined by light microscopy as well as scanning and transmission electron microscopy, mucosal architecture involving classic ultrastructure of chicken cecal mucosa was preserved. Protein content of cecal explants did not change up to 48 h of culture. DNA content of cecal explants did not change up to 24 h of culture but decreased significantly to two-thirds of control in 48 h of culture. Based on the morphologic and physiologic findings, it became evident that this organ culture system using Trowell T8 medium at 25 degrees C can be successfully used as an in vitro experimental model for as long as 24 h. The organ culture system could be a useful tool, from the structural integrity of ceca observed in this study, in investigating mucosal function and mucosal response to drugs, carcinogens, trophic factors, and pathogens.


Evolutionary potential of transcription factors for gene regulatory rewiring.
Gene regulatory networks evolve through rewiring of individual components-that is, through changes in regulatory connections. However, the mechanistic basis of regulatory rewiring is poorly understood. Using a canonical gene regulatory system, we quantify the properties of transcription factors that determine the evolutionary potential for rewiring of regulatory connections: robustness, tunability and evolvability. In vivo repression measurements of two repressors at mutated operator sites reveal their contrasting evolutionary potential: while robustness and evolvability were positively correlated, both were in trade-off with tunability. Epistatic interactions between adjacent operators alleviated this trade-off. A thermodynamic model explains how the differences in robustness, tunability and evolvability arise from biophysical characteristics of repressor-DNA binding. The model also uncovers that the energy matrix, which describes how mutations affect repressor-DNA binding, encodes crucial information about the evolutionary potential of a repressor. The biophysical determinants of evolutionary potential for regulatory rewiring constitute a mechanistic framework for understanding network evolution.


10-nanosecond dead time and low afterpulsing with a free-running reach-through single-photon avalanche diode.
The reduction of detector dead time represents an enabling factor in several photon counting applications. In this work, we investigate the free-running operation of reach-through single-photon avalanche diodes (SPADs) at ultra-low dead times. By employing a fast active quenching circuit with direct bonding to the detector, we are able to achieve a 10 ns dead time with a thick SPAD by Excelitas, still maintaining extremely low afterpulsing probabilities (below 1.5%).


Effect of Trigonella foenum graceum on blood glucose levels in normal and alloxan-diabetic mice.
The hypoglycemic effects of a decoction and an ethanol extract of Trigenolla foenum graceum seeds on the serum glucose levels of normal and alloxan diabetic mice were studied. A single 0.5 ml oral dose of 40-80% decoctions to normal as well as alloxanized mice was followed by hypoglycemia developed over a 6-h period. Reduction in blood glucose concentration was highly significant, was maximum at 6 h and was dose-dependent. The hypoglycemia caused by the ethanol extract (200-400 mg/kg) in alloxanized mice was also dose-dependent and 200 mg/kg was comparable in effect to 200 mg/kg tolbutamide.


Fate and transport of elemental copper (Cu0) nanoparticles through saturated porous media in the presence of organic materials.
Column experiments were performed to assess the fate and transport of nanoscale elemental copper (Cu(0)) particles in saturated quartz sands. Both effluent concentrations and retention profiles were measured over a broad range of physicochemical conditions, which included pH, ionic strength, the presence of natural organic matter (humic and fulvic acids) and an organic buffer (Trizma). At neutral pHs, Cu(0) nanoparticles were positively charged and essentially immobile in porous media. The presence of natural organic matter, trizma buffer, and high pH decreased the attachment efficiency facilitating elemental copper transport through sand columns. Experimental results suggested the presence of both favourable and unfavourable nanoparticle interactions causes significant deviation from classical colloid filtration theory.


Lorentz-force-induced motion in conductive media.
This project was designed to assess whether MRI imaging could detect Lorentz-force-induced motion in conductive samples. Experiments were performed by applying alternating voltages across 2% agar and 18% bovine gels placed in the field of a 1.5-T MRI scanner. Motion-sensitized time-gated MRI images that were obtained and analyzed with custom-developed software used in previous studies revealed the production of movement in both agar and gel samples. Motion was most pronounced in the plane vertical to the sample and had the greatest amplitude when the current path was perpendicular to the scanner's magnetic field. These findings are compatible with the vector cross product nature of the Lorentz force and suggest that the imaging of Lorentz-force-induced motion in conductive samples is feasible. Whether this approach can be extended to study electrically active tissues such as the peripheral nerves, brain and heart remains to be seen.


Dietary Curcumin Supplementation Could Improve Muscle Quality, Antioxidant Enzyme Activities and the Gut Microbiota Structure of Pelodiscus sinensis.
This experiment aimed to assess the impact of different dietary curcumin (CM) levels on growth, muscle quality, serum-biochemical parameters, antioxidant-enzyme activities, gut microbiome, and liver transcriptome in Chinese soft-shelled turtles (Pelodiscus sinensis). Five experimental diets were formulated to include graded levels of curcumin at 0 (control, CM0), 0.5 (CM0.5), 1 (CM1), 2 (CM2) and 4 g/kg (CM4). Each diet was randomly distributed to quadruplicate groups of turtles (164.33 ± 5.5 g) for 6 weeks. Our findings indicated that dietary curcumin supplementation did not have a significant influence on growth performance (p > 0.05); however, it significantly improved the muscular texture profiles (p < 0.05). Serum total superoxide dismutase (SOD), liver catalase (CAT), and total antioxidant capacity (T-AOC) activities increased significantly as dietary curcumin levels rose from 0.5 to 4 g/kg (p < 0.05). Dietary curcumin supplementation improved gut microbiota composition, as evidenced by an increase in the proportion of dominant bacteria such as Lactobacillus and Flavobacterium. Liver transcriptome analysis revealed that curcumin altered metabolic pathways in the liver. In conclusion, based on the evaluation of the activities of SOD in serum and CAT in liver under current experimental design, it was determined that the appropriate dietary curcumin supplementation for Chinese soft-shelled turtles is approximately 3.9 g/kg.


Single-cell RNA sequencing uncovers heterogeneous transcriptional signatures in tumor-infiltrated dendritic cells in prostate cancer.
Prostate cancer (PCa) is one of the two solid malignancies in which a higher T cell infiltration in the tumor microenvironment (TME) corresponds with a worse prognosis for the tumor. The inability of T cells to eliminate tumor cells despite an increase in their number reinforces the possibility of impaired antigen presentation. In this study, we investigated the TME at single-cell resolution to understand the molecular function and communication of dendritic cells (DCs) (as professional antigen-presenting cells). According to our data, tumor cells stimulate the migration of immature DCs to the tumor site by inducing inflammatory chemokines. Many signaling pathways such as TNF-α/NF-κB, IL2/STAT5, and E2F up-regulated after DCs enter the tumor location. In addition, some molecules such as GPR34 and SLCO2B1 decreased on the surface of DCs. The analysis of molecular and signaling alterations in DCs revealed some suppression mechanisms of tumors, such as removing mature DCs, reducing the DC's survival, inducing anergy or exhaustion in the effector T cells, and enhancing the differentiation of T cells to Th2 and Tregs. In addition, we investigated the cellular and molecular communication between DCs and macrophages in the tumor site and found three molecular pairs including CCR5/CCL5, CD52/SIGLEC10, and HLA-DPB1/TNFSF13B. These molecular pairs are involved in the migration of immature DCs to the TME and disrupt the antigen-presenting function of DCs. Furthermore, we presented new therapeutic targets by the construction of a gene co-expression network. These data increase our knowledge of the heterogeneity and the role of DCs in PCa TME.


Magnetic Resonance-Based Characterization of Myocardial Architecture.
Advances in technology have made it possible to image the microstructure of the heart with diffusion-weighted magnetic resonance. The technique provides unique insights into the cellular architecture of the myocardium and how this is perturbed in a range of disease contexts. In this review, the physical basis of diffusion MRI and the challenges of implementing it in the beating heart are discussed. Cutting edge acquisition and analysis techniques, as well as the results of initial clinical studies, are reported.


The tolerance of Jatropha curcas seedlings to NaCl: an ecophysiological analysis.
Jatropha curcas L. is a biodiesel crop that is resistant to drought stress. However, the salt tolerance of this plant has not yet been studied. To address this question, J. curcas seedlings were grown in a fertilised substrate to evaluate the effects of salinity stress on growth, leaf water relation and organic solutes, leaf and root mineral concentrations, chlorophyll fluorescence parameters, and carbohydrate concentration. The experiment consisted of six treatments with different concentrations of NaCl in the irrigation water: 0 (control), 30, 60, 90, 120 and 150 mM. The total biomass exhibited a salt-induced decrease in the 60 mM or higher NaCl concentrations. The Cl(r) concentration was higher than the Na(+) concentration in all of the plant tissues. The water potential and relative water content of the leaves were not affected by any of the salt treatments. However, salinity induced a decline in the leaf K(+) concentration, together with a significant enhancement in the leaf P, S, Fe, Zn, Mn and Cu levels. The net assimilation of CO₂ also decreased with the salt treatment, due in part to non-stomatal limitation from the increase in C(a)/C(i) and a decrease in the maximum quantum efficiency (F(v)/F(m)) of photosystem II and soil plant analysis development (SPAD) units. This work suggests that J. curcas seedlings exhibit a moderate tolerance to salinity, as the plants were able to tolerate up to 4 dS m(-1) (EC water irrigation; 30 mM NaCl). The negative influences of salinity in this crop are mainly due to Cl(r) and/or Na(+) toxicity and to a nutritional imbalance caused by an increase in the Na(+)/K(+) ratio. The osmotic effect of salinity in this species is negligible, perhaps due to its strong control of leaf transpiration, which reduces water loss.


Genitourinary Prosthetics: A Primer for the Non-urologic Surgeon.
Genitourinary prosthetics are used for correction of functional deficits and to improve the quality of lives of affected patients. General surgeons must evaluate patients scheduled for nonurologic surgery with urologic devices that can impact their perioperative management. Lack of recognition of these prosthetics preoperatively can lead to unnecessary morbidity for the patient and have legal implications for the surgeon. Close consultation with a urologist may avoid common complications associated with these devices and allows for surgical assistance when operative misadventures do occur. This article reviews 3 common urologic prosthetics: testicular prosthesis, artificial urinary sphincter, and penile prosthesis.


Cloning of malarial genes coding for high molecular weight antigens: isolation of fragments of Plasmodium falciparum genes coding for proteins of 145 000 molecular weight.
RNA preparations from Plasmodium falciparum were shown to direct the cell free synthesis of many high molecular weight proteins, some of which could be immunoprecipitated by sera from humans immune to P. falciparum. Among these was an abundant protein with a molecular weight of 145 000. Messenger RNAs coding for high molecular weight antigens were enriched by size fractionation and used after reverse transcription to identify genomic restriction fragments in Southern blots. Genomic restriction fragments were cloned into plasmid and phage lambda vectors. Two recombinants which were recognized by the high molecular weight probe were characterized by hybrid-select-translation. In both cases the predominant in vitro translation products were proteins with a molecular weight of 145 000.


A behavioral screen for mediators of age-dependent TDP-43 neurodegeneration identifies SF2/SRSF1 among a group of potent suppressors in both neurons and glia.
Cytoplasmic aggregation of Tar-DNA/RNA binding protein 43 (TDP-43) occurs in 97 percent of amyotrophic lateral sclerosis (ALS), ~40% of frontotemporal dementia (FTD) and in many cases of Alzheimer's disease (AD). Cytoplasmic TDP-43 inclusions are seen in both sporadic and familial forms of these disorders, including those cases that are caused by repeat expansion mutations in the C9orf72 gene. To identify downstream mediators of TDP-43 toxicity, we expressed human TDP-43 in a subset of Drosophila motor neurons. Such expression causes age-dependent deficits in negative geotaxis behavior. Using this behavioral readout of locomotion, we conducted an shRNA suppressor screen and identified 32 transcripts whose knockdown was sufficient to ameliorate the neurological phenotype. The majority of these suppressors also substantially suppressed the negative effects on lifespan seen with glial TDP-43 expression. In addition to identification of a number of genes whose roles in neurodegeneration were not previously known, our screen also yielded genes involved in chromatin regulation and nuclear/import export- pathways that were previously identified in the context of cell based or neurodevelopmental suppressor screens. A notable example is SF2, a conserved orthologue of mammalian SRSF1, an RNA binding protein with roles in splicing and nuclear export. Our identification SF2/SRSF1 as a potent suppressor of both neuronal and glial TDP-43 toxicity also provides a convergence with C9orf72 expansion repeat mediated neurodegeneration, where this gene also acts as a downstream mediator.


Determination of acetylcholine and dopamine content in thalamus and striatum after excitotoxic lesions of the pedunculopontine tegmental nucleus in rats.
The pedunculopontine tegmental nucleus (PPTg) contains cholinergic neurons whose principal ascending connections are with thalamic nuclei and structures associated with the striatum. It has been hypothesized that PPTg neurons are more closely associated with the substantia nigra (and therefore striatal motor systems) than with the ventral tegmental area (and therefore limbic striatal functions). In the present experiments we have examined the hypothesis that the PPTg is similarly associated with motor nuclei in the thalamus. Rats received unilateral ibotenate lesions of PPTg and were sacrificed 1, 2, 4 or 7 days later. Discrete thalamic nuclei, and samples of caudate-putamen and nucleus accumbens, were punched out and thalamic acetylcholine (ACh) and striatal ACh and dopamine (DA) content examined. Anteroventral nucleus had decreased ACh content after PPTg lesion, but a time dependent increase was found in mediodorsal nucleus; ACh concentration was unchanged in thalamic reticular nucleus or medial geniculate. No long-term lesion-dependent changes in striatal ACh or DA content were found. The effects of PPTg lesion on thalamic ACh content are consistent with the hypothesis that it has effects on motor nuclei, but also indicate that PPTg lesions have complex and dynamic effects on thalamic ACh content.


Wrapped Up: The Motility of Polarly Flagellated Bacteria.
A huge number of bacterial species are motile by flagella, which allow them to actively move toward favorable environments and away from hazardous areas and to conquer new habitats. The general perception of flagellum-mediated movement and chemotaxis is dominated by the Escherichia coli paradigm, with its peritrichous flagellation and its famous run-and-tumble navigation pattern, which has shaped the view on how bacteria swim and navigate in chemical gradients. However, a significant amount-more likely the majority-of bacterial species exhibit a (bi)polar flagellar localization pattern instead of lateral flagella. Accordingly, these species have evolved very different mechanisms for navigation and chemotaxis. Here, we review the earlier and recent findings on the various modes of motility mediated by polar flagella.


The stereoselective recognition of substrates by phosphoinositide kinases. Studies using synthetic stereoisomers of dipalmitoyl phosphatidylinositol.
Soluble phosphatidylinositol (PtdIns) 4- and 3-kinase activities were partially purified and characterized from human placental extracts. The placental PtdIns 4-kinase (type 3) has a Km for ATP of 460 microM and is kinetically different to a partially purified human erythrocyte, membrane-bound, PtdIns 4-kinase (type 2). These three inositol lipid kinases were then used to compare their substrate specificities against the four synthetic stereoisomers of dipalmitoyl PtdIns. Only the placental 4-kinase was influenced by the chirality of the glycerol moiety of PtdIns. However, neither of the 4-kinases was able to phosphorylate L-PtdIns and, therefore, these kinases have an absolute requirement for the inositol ring to be linked to the glyceryl backbone of the lipid through the D-1 position. Phosphoinositide 3-kinase, on the other hand, was found to phosphorylate both D- and L-PtdIns. While the 3-kinase phosphorylated exclusively the D-3 position of D-PtdIns, further analyses demonstrated that the same enzyme phosphorylated two sites on L-PtdIns, namely the D-6 and D-5 positions of the inositol ring. Some implications of these findings are discussed.


Identification of 130 kDa cell surface LDL-binding protein from smooth muscle cells as a partially processed T-cadherin precursor.
Atypical cell surface lipoprotein-binding proteins of 105 kDa and 130 kDa are present in membranes of vascular smooth muscle cells. We recently identified the 105 kDa protein from human aortic media as T-cadherin, an unusual glycosylphosphatidylinositol (GPI)-anchored member of the cadherin family of cell adhesion proteins. The goal of the present study was to determine the identity of 130 kDa lipoprotein-binding protein of smooth muscle cells. We applied different approaches that included protein sequencing of purified protein from human aortic media, the use of human T-cadherin peptide-specific antisera, and enzymatic treatment of cultured cells with trypsin and GPI-specific phospholipase C. Our results indicate that the 130 kDa protein is a partially processed form of T-cadherin which is attached to the membrane surface of smooth muscle cells via a GPI anchor and contains uncleaved N-terminal propeptide sequence. Our data disclose that, in contrast to classical cadherins, T-cadherin is expressed on the cell surface in both its precursor (130 kDa) and mature (105 kDa) forms.


Improved Survival of IPF patients Treated With Antifibrotic Drugs Compared With Untreated Patients.
Pirfenidone and nintedanib unequivocally inhibit FVC decline, but have been inconsistently linked to reduced mortality in phase III studies. On the contrary, real-world data show a survival benefit of antifibrotic drugs. However, it is unknown what this benefit is across different Gender, Age, and Physiology (GAP) stages. Is there a difference in transplant-free (TPF) survival of IPF patients receiving antifibrotic drugs (IPFAF) compared with an untreated cohort (IPFnon-AF)? Is this different for patients with GAP stage I, II, or III. This is a single-center observational cohort study using prospectively included patients diagnosed with IPF between 2008-2018. Primary outcomes were TPF survival difference and 1-, 2-, and 3-year cumulative mortality for IPFAF and IPFnon-AF. This was repeated after stratification for GAP stage. In total, 457 patients were included. The median transplant-free survival was 3.4 years in IPFAF (n = 313) and 2.2 years in IPFnon-AF (n = 144, p = 0.005). For GAP stage II, a median survival of 3.1 and 1.7 years was noted for IPFAF (n = 143) and IPFnon-AF (n = 59, p < 0.001), respectively. A significantly lower 1-, 2-, and 3- year cumulative mortality was found for IPFAF with GAP stage II (1 yr: 7.0% vs 35.6%, 2 yr: 26.6% vs 55.9%, and 3 yr: 46.9% vs 69.5%). The 1-year cumulative mortality of IPFAF with GAP III was also significantly lower (19.0% vs 65.0%). This large real-world study showed a survival benefit in IPFAF compared with IPFnon-AF. This especially holds true for patients with GAP stage II and III.


Cannabidiol attenuates fear memory expression in female rats via hippocampal 5-HT1A but not CB1 or CB2 receptors.
Growing evidence from male rodent and human studies suggests that cannabidiol (CBD) modulates the expression of aversive memories and anxiety-related responses. The limited data on whether and how CBD influences these aspects in females could have therapeutic implications given the increased susceptibility of women to anxiety- and stress-related disorders relative to men. Female studies are also essential to examine inherent aspects that potentially contribute to differences in responsiveness to CBD. Here we addressed these questions in adult female rats. Contextually fear-conditioned animals acutely treated with CBD (1.0-10 mg/kg) were tested 45 min later. In subsequent experiments, we investigated the estrous cycle effects and the contribution of dorsal hippocampus (DH) serotonin 1A (5-HT1A) and cannabinoid types 1 (CB1) and 2 (CB2) receptors to CBD-induced effects on memory retrieval/expression. The effects of pre-retrieval systemic or intra-DH CBD administration on subsequent fear extinction were also assessed. Lastly, we evaluated the open arms avoidance and stretched-attend postures in females exposed to the elevated plus-maze after systemic CBD treatment. CBD 3.0 and 10 mg/kg administered before conditioned context exposure reduced females' freezing. This action remained unchanged across the estrous cycle and involved DH 5-HT1A receptors activation. Pre-retrieval CBD impaired memory reconsolidation and lowered fear during early extinction. CBD applied directly to the DH was sufficient to reproduce the effects of systemic CBD treatment. CBD 3.0 and 10 mg/kg reduced anxiety-related responses scored in the elevated plus-maze. Our findings demonstrate that CBD attenuates the behavioral manifestation of learned fear and anxiety in female rats.