Label-free detection of neuron-drug interactions using acoustic and Kelvin vibrational fields.
Kelvin and acoustic fields of high-frequency have been employed in the non-invasive investigation of immortalized hypothalamic neurons, in order to assess their response to different concentrations of specific drugs, toxins, a stress-reducing hormone and neurotrophic factors. In an analytical systems biology approach, this work constitutes a first study of living neuron cultures by scanning Kelvin nanoprobe (SKN) and thickness shear mode (TSM) acoustic wave techniques. N-38 hypothalamic mouse neurons were immobilized on the gold electrode of 9 MHz TSM acoustic wave devices and gold-coated slides for study by SKN. The neurons were exposed to the neurochemicals betaseron, forskolin, TCAP, and cerebrolysin. Signals were collected with the TSM in real-time mode, and with the SKN in scanning and real-time modes, as the drugs were applied at biologically significant concentrations. With the TSM, for all drugs, some frequency and resistance shifts were in the same direction, contrary to normal functioning for this type of instrument. Possible mechanisms are presented to explain this behaviour. An oscillatory signal with periodicity of approximately 2 min was observed for some neuron-coated surfaces, where the amplitude of these oscillations was altered upon application of certain neurotrophic factors. These two new techniques present novel and non-invasive electrodeless methods for detecting changes at the cellular level caused by a variety of neuroactive compounds, without killing or destroying the neurons.


Low red/far-red ratio as a signal promotes carbon assimilation of soybean seedlings by increasing the photosynthetic capacity.
Shading includes low light intensity and varying quality. However, a low red/far-red (R/Fr) ratio of light is a signal that affects plant growth in intercropping and close- planting systems. Thus, the low R/Fr ratio uncoupling from shading conditions was assessed to identify the effect of light quality on photosynthesis and CO2 assimilation. Soybean plants were grown in a growth chamber with natural solar radiation under four treatments, that is, normal (N, sunlight), N + Fr, Low (L) + Fr, and L light. Low R/Fr ratio significantly increased the total biomass, leaf area, starch and sucrose contents, chlorophyll content, net photosynthetic rate, and quantum efficiency of the photosystem II compared with normal R/Fr ratio under the same light level (P < 0.05). Proteomic analysis of soybean leaves under different treatments was performed to quantify the changes in photosynthesis and CO2 assimilation in the chloroplast. Among the 7834 proteins quantified, 12 showed a > 1.3-fold change in abundance, of which 1 was related to porphyrin and chlorophyll metabolism, 2 were involved in photosystem I (PS I), 4 were associated with PS II, 3 proteins participated in photosynthetic electron transport, and 2 were involved in starch and sucrose metabolism. The dynamic change in these proteins indicates that photosynthesis and CO2 assimilation were maintained in the L treatment by up-regulating the component protein levels compared with those in N treatment. Although low R/Fr ratio increased the photosynthetic CO2 assimilation parameters, the differences in most protein expression levels in N + Fr and L + Fr treatments compared with those in N treatment were insignificant. Similar trends were found in gene expression through quantitative reverse transcription polymerase chain reaction excluding the gene expression of sucrose synthase possible because light environment is one of the factors affecting carbon assimilation. Low R/Fr ratio (high Fr light) can increase the photosynthetic CO2 assimilation in the same light intensity by improving the photosynthetic efficiency of the photosystems.


Autophagy in Pulmonary Diseases.
The pathogenesis of pulmonary diseases is often complex and characterized by multiple cellular events, including inflammation, cell death, and cell proliferation. The mechanisms by which these events are regulated in pulmonary diseases remain poorly understood. Autophagy is an essential process for cellular homeostasis and stress adaptation in eukaryotic cells. This highly conserved cellular process involves the sequestration of cytoplasmic components in double-membrane autophagosomes, which are delivered to lysosomes for degradation. The critical roles of autophagy have been demonstrated in a wide range of pathophysiological conditions. Emerging studies have identified that autophagy plays important roles in the pathogenesis of various lung diseases. In addition, autophagy has been shown to selectively degrade subcellular targets, including proteins, organelles, and pathogens. Here, we highlight the recent advances in the molecular regulation and function of autophagy in lung diseases.


Medication-Related Experience of Deaf American Sign Language Users.
Previous studies showed that deaf and hard-of-hearing (DHH) individuals have low health literacy related to prescription labels. This study examined the DHH's experience with understanding prescription labels and how technology can impact that experience. The purpose of this qualitative study was twofold: (1) gain a more enhanced understanding of DHH experiences in understanding prescription labels with a focus on language needs, expectations, and preferences, and (2) assess the potential role of technology in addressing the communication-related accessibility issues which emerge from the data. In this study, 25 Deaf American Sign Language users who picked up a prescription from a pharmacy within the past year were interviewed. A thematic analysis, which included a systematic coding process, was used to uncover themes about their experiences picking up and using prescription medications. Thematic analyses identified that medication-related experiences centered around themes: (1) medication information seeking; (2) comfort taking medication; (3) picking up medication; and (4) communication with the pharmacy team. A large contributor to the communication experience was the perception that the pharmacist was not being respectful. Regarding comfort taking medications, 12% of participants expressed a lack of understanding medications while taking medication. This led to participants largely using online resources when seeking medication information. This study also found that technology greatly aided the participants during this experience. This study recorded the experiences within the context of limited health literacy and aversive audism found that the DHH individual repeatedly encountered communication barriers, which may contribute to their poor medication literacy. Thus, future studies should explore how to leverage the potential benefits of technology to improve the pharmacy experience of the DHH, thereby improving medication literacy. [HLRP: Health Literacy Research and Practice. 2023;7(4):e215-e224.].


Cardiomyocyte proliferation and protection against post-myocardial infarction heart failure by cyclin D1 and Skp2 ubiquitin ligase.
Cyclins and other cell-cycle regulators have been used in several studies to regenerate cardiomyocytes in ischaemic heart failure. However, proliferation of cardiomyocytes induced by nuclear-targeted cyclin D1 (D1NLS) stops after one or two rounds of cell cycles due in part to accumulation of p27Kip1, an inhibitor of cyclin-dependent kinase (CDK). Thus, expression of S-phase kinase-associated protein 2 (Skp2), a negative regulator of p27Kip1, significantly enhances the effect of D1NLS and CDK4 on cardiomyocyte proliferation in vitro. Here, we examined whether Skp2 can also improve cardiomyocyte regeneration and post-ischaemic cardiac performance in vivo. Wistar rats underwent ischaemia/reperfusion injury by ligation of the coronary artery followed by injection of adenovirus vectors for D1NLS and CDK4 with or without Skp2. Enhanced proliferation of cardiomyocytes in the presence of Skp2 was demonstrated by increased expression of Ki67, a marker of proliferating cells (1.95% vs. 4.00%), and mitotic phosphorylated histone H3 (0.24% vs. 0.58%). Compared with rats that received only D1NLS and CDK4, expression of Skp2 improved left ventricular function as measured by the maximum and minimum rates of change in left ventricular pressure, the left ventricle end-diastolic pressure, left ventricle end-diastolic volume index, and the lung/body weight ratio. Expression of Skp2 enhanced the effect of D1NLS and CDK4 on the proliferation of cardiomyocytes and further contributed to improved post-ischaemic cardiac function. Skp2 might be a versatile tool to improve the effect of cyclins on post-ischaemic regeneration of cardiomyocytes in vivo.


Documentation of the 5 as for smoking cessation by PCPs across distinct health systems.
Physicians can help patients quit smoking using the 5 As of smoking cessation. This study aimed to (1) identify the proportion of known smokers that receive smoking cessation services in the course of routine clinical practice; (2) describe demographic and comorbidity characteristics of patients receiving the 5 As in these systems; and (3) evaluate differences in performance of the 5 As across health systems, gender, and age categories. Electronic medical records of 200 current smokers from 6 unique health systems (N = 1200) were randomly selected from 2006 to 2010. Primary care encounter progress notes were hand coded for occurrences of the 5 As. Bivariate comparisons of delivery of the 3 smoking-cessation services by site, gender, and age category were analyzed using χ² tests. About 50% of smokers were advised to quit smoking, 39% were assessed for their readiness to quit, and 54% received some type of assistance to help them quit smoking. Only 2% had a documented plan for follow-up regarding their quitting efforts (arrange). Significant differences were found among sites for documentation of receiving the 5 As and between age groups receiving assistance with quitting. There was no statistically significant difference between genders in receipt of the 5 As. Documentation of adherence to the 5 As varied by site and some demographics. Adjustments to protocols for addressing cessation and readiness to quit may be warranted. Health systems could apply the methodology described in this paper to assess their own performance, and then use that as a basis to guide improvement initiatives.


Ants mediate the structure of phytotelm communities in an ant-garden bromeliad.
The main theories explaining the biological diversity of rain forests often confer a limited understanding of the contribution of interspecific interactions to the observed patterns. We show how two-species mutualisms can affect much larger segments of the invertebrate community in tropical rain forests. Aechmea mertensii (Bromeliaceae) is both a phytotelm (plant-held water) and an ant-garden epiphyte. We studied the influence of its associated ant species (Pachycondyla goeldii and Camponotus femoratus) on the physical characteristics of the plants, and, subsequently, on the diversity of the invertebrate communities that inhabit their tanks. As dispersal agents for the bromeliads, P. goeldii and C. femoratus influence the shape and size of the bromeliad by determining the location of the seedling, from exposed to partially shaded areas. By coexisting on a local scale, the two ant species generate a gradient of habitat conditions in terms of available resources (space and food) for aquatic invertebrates, the diversity of the invertebrate communities increasing with greater volumes of water and fine detritus. Two-species mutualisms are widespread in nature, but their influence on the diversity of entire communities remains largely unexplored. Because macroinvertebrates constitute an important part of animal production in all ecosystem types, further investigations should address the functional implications of such indirect effects.


Assessment of antibiotic use patterns in bovine mastitis treatment in the dairy sector in Serbia.
The dairy sector is essential for the world's food systems, playing a vital role in agricultural production. Yet, mastitis, an inflammation of the mammary gland, affects a considerable portion of the dairy cow population annually, leading to reduced productivity and sustainability. Insight into therapeutic protocols is essential for a better understanding of the situation on farms with the aim of developing new or harmonizing existing protocols in the mastitis treatment. This study conducted on dairy farm in Serbia aimed to explore the most commonly used antibiotics during bovine mastitis treatment and their association with daily milk yield. Data from 100 lactating cows, including therapeutic protocols, lactation numbers, mastitis episodes, milk yield, and antibiotic usage, were obtained from a computerized database and statistically analysed. The results demonstrated a high prevalence of narrow-spectrum antibiotics usage, with the majority of cows treated with combination therapy. The route of antibiotic application did not significantly impact daily milk yield, while treatment duration had no discernible effect. Nonetheless, specific antibiotics were associated with milk yield variations, suggesting the need for careful antibiotic selection and management in mastitis therapy. These findings emphasize the importance of prudent antibiotic use to safeguard animal health and long-term milk production sustainability.


TaCYP78A5 regulates seed size in wheat (Triticum aestivum).
Seed size is an important agronomic trait and a major component of seed yield in wheat. However, little is known about the genes and mechanisms that determine the final seed size in wheat. Here, we isolated TaCYP78A5, the orthologous gene of Arabidopsis CYP78A5/KLUH in wheat, from wheat cv. Shaan 512 and demonstrated that the expression of TaCYP78A5 affects seed size. TaCYP78A5 encodes the cytochrome P450 (CYP) 78A5 protein in wheat and rescued the phenotype of the Arabidopsis deletion mutant cyp78a5. By affecting the extent of integument cell proliferation in the developing ovule and seed, TaCYP78A5 influenced the growth of the seed coat, which appears to limit seed growth. TaCYP78A5 silencing caused a 10% reduction in cell numbers in the seed coat, resulting in a 10% reduction in seed size in wheat cv. Shaan 512. By contrast, the overexpression of TaCYP78A5 increased the number of cells in the seed coat, resulting in seed enlargement of ~11-35% in Arabidopsis. TaCYP78A5 activity was positively correlated with the final seed size. However, TaCYP78A5 overexpression significantly reduced seed set in Arabidopsis, possibly due to an ovule development defect. TaCYP78A5 also influenced embryo development by promoting embryo integument cell proliferation during seed development. Accordingly, a working model of the influence of TaCYP7A5 on seed size was proposed. This study provides direct evidence that TaCYP78A5 affects seed size and is a potential target for crop improvement.


Neuro-nutraceuticals: Further insights into their promise for brain health.
In this Special Issue on "Nutraceuticals: Molecular and Functional Insights into how Natural Products Nourish the Brain", the editors bring together contributions from experts in nutraceutical research to provide a contemporary overview of how select chemically identified molecules from natural products can beneficially affect brain function at the molecular level. Other contributions address key emergent issues such as bioavailability, neuronal health, inflammation and the holistic benefit of multi-targeted actions that impact upon how nutraceuticals ultimately leverage the brain to function better. In terms of the benefit of nutraceuticals it is clear that some naturally occurring molecules can be advantageous to both the young and aged brain, and that they have actions that ultimately can be directed to aid either in the improvement of cognition or in the management of debilitating neurodegenerative and neuropsychiatric conditions.


Measuring Enthalpy of Sublimation of Volatiles by Means of Piezoelectric Crystal Microbalances.
Piezoelectric Crystal Microbalances (PCM's) are widely used to study the chemical processes involving volatile compounds in any environment, such as condensation process. Since PCM's are miniaturized sensor, they are very suitable for planetary in situ missions, where can be used to detect and to measure the mass amount of astrobiologically significant compounds, such as water and organics. This work focuses on the realization and testing of a new experimental setup, able to characterize volatiles which can be found in a planetary environment. In particular the enthalpy of sublimation of some dicarboxylic acids has been measured. The importance of dicarboxylic acids in planetology and astrobiology is due to the fact that they have been detected in carbonaceous chondritic material (e.g. Murchinson), among the most pristine material present in our Solar System. In this work, a sample of acid was heated in an effusion cell up to its sublimation. For a set of temperatures (from 30 °C to 75 °C), the deposition rate on the PCM surface has been measured. From these measurements, it has been possible to infer the enthalpy of sublimation of Adipic acid, i.e. ΔH = 141.6 ± 0.8 kJ/mol and Succinic acid, i.e. ΔH = 113.3 ± 1.3 kJ/mol. This technique has so demonstrated to be a good choice to recognise a single compound or a mixture (with an analysis upstream) even if some improvements concerning the thermal stabilization of the system will be implemented in order to enhance the results' accuracy. The experiment has been performed in support of the VISTA (Volatile In Situ Thermogravimetry Analyzer) project, which is included in the scientific payload of the ESA MarcoPolo-R mission study.


Tarsal and carpal coalition and symphalangism of the Fuhrmann type. Report of a family.
We are reporting on fifteen members of a five-generation family (sixty-three members) who had an autosomal dominant osseous disorder that was characterized by tarsal and carpal coalition, symphalangism, short first metacarpals, and abnormalities of the elbow, including humeroradial fusion. This family is similar to the one reported by Fuhrmann et al.


Population genomic analyses of early-phase Atlantic Salmon (Salmo salar) domestication/captive breeding.
Domestication can have adverse genetic consequences, which may reduce the fitness of individuals once released back into the wild. Many wild Atlantic salmon (Salmo salarL.) populations are threatened by anthropogenic influences, and they are supplemented with captively bred fish. The Atlantic salmon is also widely used in selective breeding programs to increase the mean trait values for desired phenotypic traits. We analyzed a genomewide set of SNPs in three domesticated Atlantic salmon strains and their wild conspecifics to identify loci underlying domestication. The genetic differentiation between domesticated strains and wild populations was low (F ST < 0.03), and domesticated strains harbored similar levels of genetic diversity compared to their wild conspecifics. Only a few loci showed footprints of selection, and these loci were located in different linkage groups among the different wild population/hatchery strain comparisons. Simulated scenarios indicated that differentiation in quantitative trait loci exceeded that in neutral markers during the early phases of divergence only when the difference in the phenotypic optimum between populations was large. This study indicates that detecting selection using standard approaches in the early phases of domestication might be challenging unless selection is strong and the traits under selection show simple inheritance patterns.


Area-Specific Regulation of Quiescent Neural Stem Cells by Notch3 in the Adult Mouse Subependymal Zone.
In the adult mammalian brain, neural stem cells (NSCs) generate new neurons throughout the mammal's lifetime. The balance between quiescence and active cell division among NSCs is crucial in producing appropriate numbers of neurons while maintaining the stem cell pool for a long period. The Notch signaling pathway plays a central role in both maintaining quiescent NSCs (qNSCs) and promoting cell division of active NSCs (aNSCs), although no one knows how this pathway regulates these apparently opposite functions. Notch1 has been shown to promote proliferation of aNSCs without affecting qNSCs in the adult mouse subependymal zone (SEZ). In this study, we found that Notch3 is expressed to a higher extent in qNSCs than in aNSCs while Notch1 is preferentially expressed in aNSCs and transit-amplifying progenitors in the adult mouse SEZ. Furthermore, Notch3 is selectively expressed in the lateral and ventral walls of the SEZ. Knockdown of Notch3 in the lateral wall of the adult SEZ increased the division of NSCs. Moreover, deletion of the Notch3 gene resulted in significant reduction of qNSCs specifically in the lateral and ventral walls, compared with the medial and dorsal walls, of the lateral ventricles. Notch3 deletion also reduced the number of qNSCs activated after antimitotic cytosine β-D-arabinofuranoside (Ara-C) treatment. Importantly, Notch3 deletion preferentially reduced specific subtypes of newborn neurons in the olfactory bulb derived from the lateral walls of the SEZ. These results indicate that Notch isoforms differentially control the quiescent and proliferative steps of adult SEZ NSCs in a domain-specific manner.SIGNIFICANCE STATEMENT In the adult mammalian brain, the subependymal zone (SEZ) of the lateral ventricles is the largest neurogenic niche, where neural stem cells (NSCs) generate neurons. In this study, we found that Notch3 plays an important role in the maintenance of quiescent NSCs (qNSCs), while Notch1 has been reported to act as a regulator of actively cycling NSCs. Furthermore, we found that Notch3 is specifically expressed in qNSCs located in the lateral and ventral walls of the lateral ventricles and regulates neuronal production of NSCs in a region-specific manner. Our results indicate that Notch3, by maintaining the quiescence of a subpopulation of NSCs, confers a region-specific heterogeneity among NSCs in the adult SEZ.


Infusion with an oxytocin receptor antagonist delays parturition in a marsupial.
The oxytocin receptor antagonist [1-deamino-2-D-Tyr-(OEt)-4-Thr-8-Om]-oxytocin (Atosiban) is a specific antagonist of both mesotocin- and oxytocin-induced myometrial contractions in late pregnant tammars in vitro. Continuous intravenous infusion of Atosiban (1 mg kg-1 day-1) for 3 or 7 days from day 24 of the 26.5 day gestation significantly delayed births. In both the 3 day and 7 day infusion groups, all 15 control animals were pregnant and gave birth within the normal time (day 26.75 +/- 0.20, mean +/- SEM), during the infusion of saline. The neonates weighed 387 +/- 8 mg. Deliveries were observed in 15 Atosiban-treated animals significantly (P < 0.05) later than in the controls (day 27.85 +/- 0.19; neonate weight 413 +/- 9 mg). All pouch young were successfully suckled, even in the continued presence of Atosiban. Baseline plasma concentrations of the prostaglandin F metabolite (PGFM) in pregnant tammars were < 200 pg ml-1. A surge in plasma PGFM occurred at birth (811 +/- 116 pg ml-1), followed by a rapid fall to baseline concentrations within 1 h after birth. This was observed both in saline- and in Atosiban-treated animals that gave birth during the observation period, and did not differ significantly between the treatment groups. Plasma progesterone concentrations in the control and the Atosiban-treated animals showed the normal pattern of luteolysis immediately after birth. Thus, infusion of an oxytocin receptor antagonist at the end of gestation delays birth, the peripartum surge in prostaglandin release, and the fall in progesterone, suggesting that mesotocin is an important part of the hormonal cascade associated with delivery in this marsupial.


[Strain-, sex- and species-related differences of acetohexamide reductase and 20 beta-hydroxysteroid dehydrogenase activities in liver microsomes of experimental animals].
We examined physiological and genetic factors affecting acetohexamide reductase (AHR) and 20 beta-hydroxysteroid dehydrogenase (20 beta-HSD) activities in liver microsomes of experimental animals. Pronounced strain-related differences were found in both activities of AHR and 20 beta-HSD present in liver microsomes of male rats. Among rat strains tested in this study, even though a Wistar-Imamichi (WIM) rat strain was taken to lack AHR activity, it exhibited a significant 20 beta-HSD activity. These findings appeared to be in conflict with our conclusion reported so far, which AHR and 20 beta-HSD present in liver microsomes of male rats are identical enzymes. Thus the reason for this discrepancy was discussed. Furthermore, AHR and 20 beta-HSD activities were little or not observed in liver microsomes of female rats or male experimental animals other than the rat, indicating the existence of sex- and species-related differences in these two enzyme activities.


High-quality graphene transfer via directional etching of metal substrates.
Large-scale applications of graphene require its high-efficiency transfer from growth metal substrates to any other substrates of interest. The wrinkles and folds generated during the transfer process of graphene by the well-known poly(methyl methacrylate) (PMMA)-assisted technique is a critical issue. Here, we report an improvement of this method by applying a directional etching process for the removal of the growth Cu substrates, using a pair of electrodes inserted into the etchant with a constant current to form an electrochemical system. The controlled redox reactions between the Cu and the solution environment result in the etching of Cu in a part-by-part manner strictly from one end to the other. The consistency of the Cu etching direction can avoid the formation of an easily destroyable Cu structure and release the stress concentration that is usually generated during the random etching process, and finally yield significantly improved quality of the transferred graphene film with a lowered density of wrinkles, cracks/folds, adlayers, reduced root-mean-square of surface roughness, and increased performance in sheet resistance and carrier mobility.


Pharmacokinetic analysis of 5-[18F]fluorouracil tissue concentrations measured with positron emission tomography in patients with liver metastases from colorectal adenocarcinoma.
The purpose of our study was to develop a pharmacokinetic model to quantify the intracellular 5-fluorouracil (5-FU) concentration in liver metastases, which is expected to be closely correlated to therapy response. In addition, the influence of the biomodulator folinic acid on the action of 5-FU in the metastases was investigated. After i.v. application of 5-FU labeled with the positron emitter fluorine-18 (5-[18F]FU), the kinetics of the regional 5-[18F]FU/uptake was measured dynamically with positron emission tomography over 120 min in 14 patients with a total of 27 liver metastases from colorectal adenocarcinoma. Activity-time curves were evaluated in the metastases, the normal liver tissue, as well as in the aorta and analyzed by a six-compartment model. The catabolic breakdown of 5-FU to alpha-fluoro-beta-alanine (FBAL) in the normal liver tissue was modeled to separate the catabolites from the cytostatic agent 5-[18F]FU and the active 5-[18F]fluorodeoxyuridine nucleotides. With our model, all measured activity-time courses could be described adequately with only small interindividual variations in parameters connected with liver and blood. Extrahepatic clearance of 5-FU was estimated as 0.66 +/- 0.33 liters/min, whereas the hepatic clearance was 0.52 +/- 0.25 liters/min. The Michaelis-Menten parameters describing the nonlinear conversion of 5-FU to FBAL were Km = 11.3 +/- 6.4 micromol and Vmax = 147.1 +/- 130.7 micromol/min. The maximum FBAL concentration in the liver was reached between 35 and 65 min after i.v. 5-FU infusion. The most sensitive parameters for therapy monitoring were k(in) and k(out), which characterize the transport in and out of the intracellular volume of the metastases, respectively. Tumor response can only be expected if k(in) is high and k(out) is low ("trapping"). These criteria were met by 6 of the 27 metastases, which were identical to those with high values for the area under the intracellular 5-FU concentration curve (AUC[meta,IC]5-FU). The parameters k(in) and k(out) were also used to investigate the influence of the biomodulating agent folinic acid on drug effect. Five of the six metastases that showed trapping belonged to patients who received folinic acid. With the exception of one patient, however, all patients who received folinic acid had multiple metastases, of which only one was able to trap 5-FU. Because patient response can only be expected when all metastases trap 5-FU, folinic acid showed no effect on the overall clinical response. With the quantitative modeling approach used, trapping of 5-FU can be assessed noninvasively and on an individual basis. This makes it possible to adjust the dose for each individual patient to optimize the treatment schedule.


Identification of two laccase genes in the cultivated mushroom Agaricus bisporus.
A cDNA library was constructed in lambda gt11 using mRNA from 11-d-old mycelium of Agaricus bisporus. Three clones containing laccase sequence were identified using an affinity-purified anti-laccase antibody. From one of these clones, a 333 bp sequence was used to identify further cDNA clones (including one which is close to full length) and a genomic clone. The coding sequences found were of two similar but not identical versions with differences at 36 out of 520 residues of deduced amino acid sequence. The laccase genes each encode a sequence expressed as a 2.3 kb mRNA, specifying a 520 residue polypeptide including a 19 amino acid residue signal peptide that is absent from the N terminus of the mature (extracellular) protein. The coding sequence of lcc1 is interrupted by 14 short introns. The lcc1 and lcc2 genes are not allelic as they do not segregate in uninucleate spores derived from a four-spored basidium. Comparison of the deduced amino acid sequences with that of the other fungal laccases that have been cloned, and with the very similar ascorbate oxidases from higher plants shows that whilst some sequence is absolutely conserved at and around the amino acid residues involved in copper binding, the overall sequence similarities are low.


The metabolites of the cardioprotective drug dexrazoxane do not protect myocytes from doxorubicin-induced cytotoxicity.
The clinically approved cardioprotective agent dexrazoxane (ICRF-187) and two of its hydrolyzed metabolites (a one-ring open form of dexrazoxane and ADR-925) were examined for their ability to protect neonatal rat cardiac myocytes from doxorubicin-induced damage. Dexrazoxane may protect against doxorubicin-induced damage to myocytes through its strongly metal-chelating hydrolysis product ADR-925, which could act by displacing iron bound to doxorubicin or chelating free or loosely bound iron, thus preventing site-specific iron-based oxygen radical damage. The results of this study showed that whereas dexrazoxane was able to protect myocytes from doxorubicin-induced lactate dehydrogenase release, neither of the metabolites displayed any protective ability. Dexrazoxane also reduced apoptosis in doxorubicin-treated myocytes. The ability of dexrazoxane and its three metabolites to displace iron from a fluorescence-quenched trapped intracellular iron-calcein complex was also determined to see whether the metabolites were taken up by myocytes. Although ADR-925 was taken up in the absence of calcium in the medium, in the presence of calcium, its uptake was greatly slowed, presumably because it formed a complex with calcium. Both of the one-ring open metabolites were taken up by myocytes and displaced iron from its complex with calcein. These results suggest either that the anionic metabolites do not have the same access to iron pools in critical cellular compartments, that their uptake is slowed in the presence of calcium, or, less likely, that dexrazoxane protects by some other mechanism.


Pilot program integrating outpatient opioid treatment within a rural primary care setting.
Canada is experiencing an opioid crisis. In rural areas, limited access to specialty addictions services, public transportation, and many of the social determinants of health create a unique set of challenges for people who use substances. The Rural Outpatient Opioid Treatment (ROOT) program was created to bring some of the structure of an inpatient treatment program into a rural primary care setting in Ontario, Canada. The program uses a harm reduction approach to provide group recovery work, primary care, peer support, smoking cessation, opioid agonist therapy, screening and treatment for hepatitis C and HIV, and longitudinal follow-up. Sixteen participants have enrolled in three rounds of the ROOT program to date. A program evaluation shows that opioid use decreased while use of other substances remained high, in particular methamphetamine use, which is increasing more broadly in the local area. Participants described feeling cared for and appreciated the 'seamless' nature of the multidisciplinary program, the peer support provided, and their new and expanded social networks. The rural context created both benefits and challenges for their substance use, recovery, and for community programming. In conclusion, the evaluation of this pilot program demonstrates that it is possible to successfully integrate an outpatient substance-use treatment program into rural primary care.


Microtubules are needed for the perinuclear positioning of aquaporin-2 after its endocytic retrieval in renal principal cells.
Water reabsorption in the renal collecting duct is regulated by arginine vasopressin (AVP). AVP induces the insertion of the water channel aquaporin-2 (AQP2) into the plasma membrane of principal cells, thereby increasing the osmotic water permeability. The redistribution of AQP2 to the plasma membrane is a cAMP-dependent process and thus a paradigm for cAMP-controlled exocytic processes. Using primary cultured rat inner medullary collecting duct cells, we show that the redistribution of AQP2 to the plasma membrane is accompanied by the reorganization of microtubules and the redistribution of the small GTPase Rab11. In resting cells, AQP2 is colocalized with Rab11 perinuclearly. AVP induced the redistribution of AQP2 to the plasma membrane and of Rab11 to the cell periphery. The redistribution of both proteins was increased when microtubules were depolymerized by nocodazole. In addition, the depolymerization of microtubules prevented the perinuclear positioning of AQP2 and Rab11 in resting cells, which was restored if nocodazole was washed out and microtubules repolymerized. After internalization of AQP2, induced by removal of AVP, forskolin triggered the AQP2 redistribution to the plasma membrane even if microtubules were depolymerized and without the previous positioning of AQP2 in the perinuclear recycling compartment. Collectively, the data indicate that microtubule-dependent transport of AQP2 is predominantly responsible for trafficking and localization of AQP2 inside the cell after its internalization but not for the exocytic transport of the water channel. We also demonstrate that cAMP-signaling regulates the localization of Rab11-positive recycling endosomes in renal principal cells.


Synthesis and 5-lipoxygenase inhibitory activities of eicosanoid compounds.
Ten eicosanoid compounds (3, 6, 9, 11, 12, 15, 18, 21, 23, and 25), methyl (6E,8Z,11Z,14Z)-5-hydroxy-6,8,11,14-eicosatetraenoate (5-HETE, 10), leukotriene A4 (26), and (5S,6E,8E,10E,12RS,14E)-5,12-dihydroxy-6,8,10,14-eicosatetraenoic acid (5,12-diHETE, 27) were prepared and their inhibitory activities against the 5-lipoxygenase from guinea pig polymorphonuclear leukocytes (PMNL) were tested. 5,6-Methanoleukotriene A4 (18) was especially a potent and specific inhibitor of the 5-lipoxygenase without inhibiting the cyclooxygenase and the 12-lipoxygenase. Leukotriene A4, 5-HETE, and 5,12-diHETE also have inhibitory activities against the 5-lipoxygenase at micromolar concentrations, which can regulate the formation of slow-reacting substance of anaphylaxis intracellulary.


Characterization of N protein self-association in coronavirus ribonucleoprotein complexes.
Mouse hepatitis virus (MHV) nucleocapsid (N) protein binds to the large, single-stranded, positive-sense viral genomic RNA to form a helical nucleocapsid structure in mature virions. In addition N protein binds the intracellular form of the genomic RNA, all of the MHV subgenomic mRNAs, and expressed non-MHV RNA transcripts to form ribonucleoprotein (RNP) complexes in infected cells. Among the intracellular viral RNP complexes, only the genomic RNP complex is packaged into virus particles. The present study demonstrated that N protein in the MHV virion nucleocapsid and in the intracellular genome-length RNP complex that bound to viral envelope M protein was tightly self-associated such that its association was retained even after extensive RNase A-treatment of the RNP complexes. The RNase A-resistant tight N protein association in the virion nucleocapsid was not mediated by an intermolecular disulfide bridge between N proteins. In contrast, N protein association in the majority of the intracellular RNP complexes was susceptible to RNase A-treatment. Because the RNP complexes that specifically interact with the M protein are selectively packaged into MHV particles, the present data suggested that there was a distinct difference between N protein association in viral genomic RNP complexes that undergo packaging into virus particles and the subgenomic RNP complexes that are not packaged into MHV particles.


Study of preoperative diagnostic modalities in Chinese patients with superficial esophageal squamous cell carcinoma.
Endoscopic ultrasonography (EUS) and magnifying endoscopy (ME) reliably determine indications for endoscopic resection in patients with superficial esophageal squamous cell carcinoma (SESCC). ME is widely accepted for predicting the invasion depth of superficial esophageal cancer with satisfying accuracy. However, the addition of EUS is controversial. To evaluate the diagnostic efficiency of ME vs EUS for invasion depth prediction and investigate the influencing factors in patients with SESCC to determine the best diagnostic model in China. We retrospectively analyzed patients with suspected SESCC who completed both ME and EUS and then underwent endoscopic or surgical resection at Sun Yat-Sen University Cancer Center between January 2018 and December 2021. We evaluated and compared the diagnostic efficiency of EUS and ME according to histological results, and investigated the influencing factors. We included 152 lesions from 144 patients in this study. The diagnostic accuracies of ME and EUS in differentiating invasion depth were not significantly different (73.0% and 66.4%, P = 0.24); both demonstrated moderate consistency with the pathological results (ME: kappa = 0.58, 95% confidence interval [CI]: 0.48-0.68, P < 0.01; EUS: kappa = 0.46, 95%CI: 0.34-0.57, P < 0.01). ME was significantly more accurate in the diagnosis of high-grade intraepithelial (HGIN) or carcinoma in situ (odds ratio [OR] = 3.62, 95%CI: 1.43-9.16, P = 0.007) subgroups. Using a miniature probe rather than conventional EUS can improve the accuracy of lesion depth determination (82.3% vs 49.3%, P < 0.01). Less than a quarter of circumferential occupation and application of a miniature probe were independent risk factors for the accuracy of tumor invasion depth as assessed by EUS (< 1/4 circumferential occupation: OR = 3.07, 95%CI: 1.04-9.10; application of a miniature probe: OR = 5.28, 95%CI: 2.41-11.59, P < 0.01). Of the 41 lesions (41/152, 27.0%) that were misdiagnosed by ME, 24 were corrected by EUS (24/41, 58.5%). Preoperative diagnosis of SESCC should be conducted endoscopically using white light and magnification. In China, EUS can be added after obtaining patient consent. Use of a high-frequency miniature probe or miniature probe combined with conventional EUS is preferable.


Afghanistan's national strategy for improving quality in health care.
When the Ministry of Public Health (MoPH) of the Islamic Republic of Afghanistan began reconstructing the health system in 2003, it faced serious challenges. Decades of war had severely damaged the health infrastructure and the country's ability to deliver health services. A national health resources assessment in 2002 revealed huge structural and resource disparities fundamental to improving health care. For example, only 9% of the population was able to access basic health services, and about 40% of health facilities had no female health providers, severely constraining access of women to health care. Multiple donor programs and the MoPH had some success in improving quality, but questions about sustainability, as well as fragmentation and poor coordination, existed. In 2009, MoPH resolved to align and accelerate quality improvement efforts as well as build structural and skill capacity. The MoPH established a new quality unit within the ministry and undertook a year-long consultative process that drew on international evidence and inputs from all levels of the health system to developed a National Strategy for Improving Quality in Health Care consisting of a strategy implementation framework and a five-year operational plan. Even in resource-restrained countries, under the most adverse circumstances, quality of health care can be improved at the front-lines and a consensual and coherent national quality strategy developed and implemented.


Neurosurgical Implant Safety in 7 T MRI: A Scoping Review.
The use of 7 Tesla (T) magnetic resonance imaging (MRI) is expanding across neurosurgical and neurologic specialties. However, few neurosurgical-related implants have been tested for safety at 7 T, limiting its use in patients with cranial fixation, shunt placements, and other implants. Implant safety can be determined via the American Society for Testing Materials International (ASTM) guidelines. To assess the current state of neurosurgical implant safety at 7 T, a systematic search was performed using PubMed, MEDLINE, Web of Knowledge, and citation matching. Studies written in English that included at least one neurosurgical implant and at least one safety outcome were included. Data were extracted for implant studied, implant composition, deflection angle, torque, temperature change, and ASTM guidelines followed. PRISMA reporting guidelines for scoping reviews were followed. Overall, 18 studies consisting of 45 unique implants were included. Implants included cranial fixation devices, aneurysm clips, spinal rods, pedicle screws, ventriculoperitoneal (VP) shunts, deep brain stimulation devices, and electroencephalogram (EEG) caps and electrodes. Cranial fixation devices, deep brain stimulation devices, spinal rods, and pedicle screws are likely 7 T MRI compatible based on outcomes reported. Aneurysm clips and EEG devices had variable safety outcomes. The VP shunts studied lost functionality after 7 T MRI exposure. We identified several implants that are likely compatible with 7 T MRI. Given the growth in 7 T imaging and expansion of the technology, neurosurgical implants should be constructed with the aforementioned considerations. Caution must be taken with all implants, especially aneurysm clips, programmable VP shunts, and EEG recording devices. It is also noteworthy that several implant testing reports did not report following ASTM standards. This scoping review seeks to concisely summarize all neurosurgical-related implants that have been tested for safety in 7 T MRI. EVIDENCE LEVEL: 2 TECHNICAL EFFICACY: Stage 2.


Mutations in the COL5A1 coding sequence are not common in patients with spontaneous cervical artery dissections.
The dermal connective tissue of most patients with spontaneous cervical artery dissections (sCAD) contains abnormal collagen fibers. This suggests a predisposing connective tissue defect. The ultrastructural abnormalities in the skin of patients with sCAD have similarity with the morphological alterations in patients with Ehlers-Danlos syndrome type II, a dominant hereditary disorder that has been correlated in some patients to mutations within the genes encoding type V collagen. The aim of this study was to assess the alpha 1 chain of type V collagen (COL5A1) as a candidate gene for sCAD. We searched for mutations in the COL5A1 gene in cDNA from cultured fibroblasts of 19 patients with sCAD using single-strand conformational polymorphism analysis and nucleotide sequence analysis of polymerase chain reaction-amplified fragments of the whole COL5A1 coding sequence. We detected 1 missense mutation leading to a predicted amino acid (192D/N) substitution within the N-terminal propeptide in 2 siblings. All other patients showed regular COL5A1 sequences with some silent polymorphisms. Mutations in the COL5A1 gene do not appear to be a major factor in the etiology of sCAD.


Pharmacokinetic properties of enrofloxacin in rabbits.
The pharmacokinetic properties of the fluoroquinolone antimicrobial enrofloxacin were studied in New Zealand White rabbits. Four rabbits were each given enrofloxacin as a single 5 mg/kg of body weight dosage by IV, SC, and oral routes over 4 weeks. Serum antimicrobial concentrations were determined for 24 hours after dosing. Compartmental modeling of the IV administration indicated that a 2-compartment open model best described the disposition of enrofloxacin in rabbits. Serum enrofloxacin concentrations after SC and oral dosing were best described by a 1- and 2-compartment model, respectively. Overall elimination half-lives for IV, SC, and oral routes of administration were 2.5, 1.71, and 2.41 hours, respectively. The half-life of absorption for oral dosing was 26 times the half-life of absorption after SC dosing (7.73 hours vs 0.3 hour). The observed time to maximal serum concentration was 0.9 hour after SC dosing and 2.3 hours after oral administration. The observed serum concentrations at these times were 2.07 and 0.452 micrograms/ml, respectively. Mean residence times were 1.55 hours for IV injections, 1.46 hours for SC dosing, and 8.46 hours for oral administration. Enrofloxacin was widely distributed in the rabbit as suggested by the volume of distribution value of 2.12 L/kg calculated from the IV study. The volume of distribution at steady-state was estimated at 0.93 L/kg. Compared with IV administration, bioavailability was 77% after SC dosing and 61% for gastrointestinal absorption. Estimates of predicted average steady-state serum concentrations were 0.359, 0.254, and 0.226 micrograms/ml for IV, SC, and oral administration, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)


Serine phosphoacceptor sites within the core protein of hepatitis B virus contribute to genome replication pleiotropically.
The core protein of hepatitis B virus can be phosphorylated at serines 155, 162, and 170. The contribution of these serine residues to DNA synthesis was investigated. Core protein mutants were generated in which each serine was replaced with either alanine or aspartate. Aspartates can mimic constitutively phosphorylated serines while alanines can mimic constitutively dephosphorylated serines. The ability of these mutants to carry out each step of DNA synthesis was determined. Alanine substitutions decreased the efficiency of minus-strand DNA elongation, primer translocation, circularization, and plus-strand DNA elongation. Aspartate substitutions also reduced the efficiency of these steps, but the magnitude of the reduction was less. Our findings suggest that phosphorylated serines are required for multiple steps during DNA synthesis. It has been proposed that generation of mature DNA requires serine dephosphorylation. Our results suggest that completion of rcDNA synthesis requires phosphorylated serines.


[Carbapenems resistance in Pseudomonas aeruginosa during a 10-year period at Rawson Hospital].
Pae is a non-fermenting, Gram-negative bacillus that is associated with nosocomial infections and can easily be transmitted through the hands of health staff from patient to patient. Pae has a wide variety of natural resistance mechanisms and a great capacity to acquire new resistance mechanisms or to increase the expression of their natural resistance. Carbapenems resistance may be offered by impermeability mechanisms, increase in the efflux or enzymatic pump (production of carbapenemases). To determine prevailing carbapenems resistance mechanisms in Pae and their evolution. 372 Pae strains were collected during a ten-year period. The strains were identified through biochemical tests and performed a sensitivity test through diffusion and microbiological method. carbapenems resistance mechanisms for efflux in the year 2000 were , 50%; 2001 16.6%, 2002 6.3%, 2003 23.9%, 2004 18%, 2005 8.8%, 2006 17.8%, 2007 31.8%, 2008 2.5%, 2009 5.2%. As regards impermeability: 2000 33%, 2001 21.4%, 2002 8.5%, 2003 30.4%, 2004 14.7%, 2005 28.8%, 2006 23.5%, 2007 13.6%, 2008 2.5%, 2009 5.2%. The microbiological test was negative for all studied strains. carbapenems resistance during this period was offered by impermeability and an increase in the expression of the efflux mechanism, not by enzymatic mechanism in our hospital. This is a very important issue from an epidemiological point of view because of the rapid horizontal transmission of the strains with this resistance mechanism.


[Prenatal cytogenetic diagnosis using chromosomal preparations of the native biopsy samples of chorionic villi, cultured chorionic cells and amniocytes].
Analysis of the results of prenatal cytogenetic diagnosis carried out in the first and second pregnancy trimesters in more than 300 women permitted comparing the efficacies of two methodologic approaches, diagnostic amniocentesis and chorion sampling , with due consideration for the methodologic errors typical of these methods and of the tested biologic material. Up to 5% of the diagnoses are erroneous if the diagnosis is based on chorion sampling data, whereas in amniocentesis the share of diagnostic errors is lower by an order. The authors have given a theoretical rationale for and tried a methodologic approach, involving the employment of the 'direct' chromosomal preparations from villous chorion biopsy specimens and the so-called 'maintained' cell culture technique, that permits obtaining chromosomal preparations of higher quality and, consequently, helps improve the accuracy of chromosomal diagnosis.


Epidemiologic study by DNA typing of a Candida albicans outbreak in heroin addicts.
The epidemiology of an outbreak of Candida endophthalmitis in heroin addicts was studied by DNA typing. Although one biotype was prevalent, the 13 isolates of Candida albicans from seven of the patients were placed into six separate groups by DNA type. Thus, the outbreak of candidiasis was not, as had been concluded from biotyping, due to a C. albicans strain of common origin.


Inter- and intrafractional dose uncertainty in hypofractionated Gamma Knife radiosurgery.
The purpose of this study is to evaluate inter- and intrafractional dose variations resulting from head position deviations for patients treated with the Extend relocatable frame system utilized in hypofractionated Gamma Knife radiosurgery (GKRS). While previous reports characterized the residual setup and intrafraction uncertainties of the system, the dosimetric consequences have not been investigated. A digital gauge was used to measure the head position of 16 consecutive Extend patients (62 fractions) at the time of simulation, before each fraction, and immediately following each fraction. Vector interfraction (difference between simulation and prefraction positions) and intrafraction (difference between postfraction and prefraction positions) shifts in patient position were calculated. Planned dose distributions were shifted by the offset to determine the time-of-treatment dose. Variations in mean and maximum target and organ at risk (OAR) doses as a function of positional shift were evaluated. The mean vector interfraction shift was 0.64 mm (Standard Deviation (SD): 0.25 mm, maximum: 1.17 mm). The mean intrafraction shift was 0.39 mm (SD: 0.25 mm, maximum: 1.44 mm). The mean variation in mean target dose was 0.66% (SD: 1.15%, maximum: 5.77%) for inter-fraction shifts and 0.26% (SD: 0.34%, maximum: 1.85%) for intrafraction shifts. The mean variation in maximum dose to OARs was 7.15% (SD: 5.73%, maximum: 30.59%) for interfraction shifts and 4.07% (SD: 4.22%, maximum: 17.04%) for intrafraction shifts. Linear fitting of the mean variation in maximum dose to OARs as a function of position yielded dose deviations of 10.58%/mm for interfractional shifts and 7.69%/mm for intrafractional shifts. Positional uncertainties when per-forming hypofractionated Gamma Knife radiosurgery with the Extend system are small and comparable to frame-based uncertainties (< 1 mm). However, the steep dose gradient characteristics of GKRS mean that the dosimetric consequences of positional uncertainties should be considered as part of treatment planning. These dose uncertainties should be evaluated in the context of tumor response and OAR tolerance for hypofractionated treatment scenarios where any increase in dose may be tempered by the increased protection hypofractionation provides to normal tissue.


Recursive SVM biomarker selection for early detection of breast cancer in peripheral blood.
Breast cancer is worldwide the second most common type of cancer after lung cancer. Traditional mammography and Tissue Microarray has been studied for early cancer detection and cancer prediction. However, there is a need for more reliable diagnostic tools for early detection of breast cancer. This can be a challenge due to a number of factors and logistics. First, obtaining tissue biopsies can be difficult. Second, mammography may not detect small tumors, and is often unsatisfactory for younger women who typically have dense breast tissue. Lastly, breast cancer is not a single homogeneous disease but consists of multiple disease states, each arising from a distinct molecular mechanism and having a distinct clinical progression path which makes the disease difficult to detect and predict in early stages. In the paper, we present a Support Vector Machine based on Recursive Feature Elimination and Cross Validation (SVM-RFE-CV) algorithm for early detection of breast cancer in peripheral blood and show how to use SVM-RFE-CV to model the classification and prediction problem of early detection of breast cancer in peripheral blood.The training set which consists of 32 health and 33 cancer samples and the testing set consisting of 31 health and 34 cancer samples were randomly separated from a dataset of peripheral blood of breast cancer that is downloaded from Gene Express Omnibus. First, we identified the 42 differentially expressed biomarkers between "normal" and "cancer". Then, with the SVM-RFE-CV we extracted 15 biomarkers that yield zero cross validation score. Lastly, we compared the classification and prediction performance of SVM-RFE-CV with that of SVM and SVM Recursive Feature Elimination (SVM-RFE). We found that 1) the SVM-RFE-CV is suitable for analyzing noisy high-throughput microarray data, 2) it outperforms SVM-RFE in the robustness to noise and in the ability to recover informative features, and 3) it can improve the prediction performance (Area Under Curve) in the testing data set from 0.5826 to 0.7879. Further pathway analysis showed that the biomarkers are associated with Signaling, Hemostasis, Hormones, and Immune System, which are consistent with previous findings. Our prediction model can serve as a general model for biomarker discovery in early detection of other cancers. In the future, Polymerase Chain Reaction (PCR) is planned for validation of the ability of these potential biomarkers for early detection of breast cancer.


Effect of hexavalent chromium on histone biotinylation in human bronchial epithelial cells.
Chromium is a potent human mutagen and carcinogen. The capability of chromium to cause cancers has been known for more than a century, and numerous epidemiological studies have been performed to determine its carcinogenicity. In the post-genome era, cancer has been found to relate to epigenetic mutations. However, very few researches have focused on hexavalent chromium (Cr(VI))-induced epigenetic alterations. The present study was designed to investigate whether Cr(VI) would affect the level of a newfound epigenetic modification: histone biotinylation. Histone acetylation and histone biotinylation were studied in detail using human bronchial epithelial (16HBE) cells as an in vitro model after Cr(VI) treatment. Our study showed that Cr(VI) treatment decreased histone acetylation level in 16HBE cells. In addition, low doses of Cr(VI) (≤0.6μM) elevated the level of histone biotinylation. Furthermore, immunoblot analysis of biotinidase (BTD), a major protein which maintains homeostasis of histone biotinylation, showed that the distribution of BTD became less even and more concentrated at the nuclear periphery in cells exposed to Cr(VI). Moreover, Cr(VI)-induced histone deacetylation may take part in the regulation of histone biotinylation. Together, our study provides new insight into the mechanisms of Cr(VI)-induced epigenetic regulation that may contribute to the chemoprevention of Cr(VI)-induced cancers and may have important implications for epigenetic therapy.


Organization of terminal reiterations in the virion DNA of herpesvirus saimiri.
The population of herpesvirus saimiri (HVS) genomes extracted from extracellular virions are double-stranded, linear DNA molecules of about 160 kilobase pairs (kbp) each composed of a central segment of 110 to 112 kbp and 36% (G + C) (i.e. 'light' or L-DNA) linked to direct reiterations of a 1.44 kbp repeat unit of 71% (G + C) (i.e. 'heavy' or H-DNA) at each terminus. In this paper, we show that the population of HVS DNA molecules contains approximately equal concentrations of genomes with all possible integral numbers of complete repeat units (i.e. from greater than 30 to 1) at either 'left' or 'right' ends but that all molecular ends are derived by a unique cleavage at a site close to the single ApaI restriction endonuclease site of the H-DNA repeat unit. Junctions of proximal H-DNA repeat units with L-DNA occur at, or very close to, the sequence present at the molecular ends. The transition from L- to H-DNA occurs abruptly at this site at the 'right' end of the L-DNA component but some rearranged restriction enzyme cleavage sites typical of H-DNA are found within the first 0.8 kbp of the L-DNA sequences at the 'left' H-L DNA junction. HVS appears to provide an extreme example of the general process whereby herpesvirus DNAs are matured from concatemeric intermediates by a site-specific cleavage/recombination process involving random choice between equivalent sites for the initiation of the process and with choices between alternative termination sites being limited by a headful packaging mechanism.


Consistency of muscle synergies during pedaling across different mechanical constraints.
The purpose of the present study was to determine whether muscle synergies are constrained by changes in the mechanics of pedaling. The decomposition algorithm used to identify muscle synergies was based on two components: "muscle synergy vectors," which represent the relative weighting of each muscle within each synergy, and "synergy activation coefficients," which represent the relative contribution of muscle synergy to the overall muscle activity pattern. We hypothesized that muscle synergy vectors would remain fixed but that synergy activation coefficients could vary, resulting in observed variations in individual electromyographic (EMG) patterns. Eleven cyclists were tested during a submaximal pedaling exercise and five all-out sprints. The effects of torque, maximal torque-velocity combination, and posture were studied. First, muscle synergies were extracted from each pedaling exercise independently using non-negative matrix factorization. Then, to cross-validate the results, muscle synergies were extracted from the entire data pooled across all conditions, and muscle synergy vectors extracted from the submaximal exercise were used to reconstruct EMG patterns of the five all-out sprints. Whatever the mechanical constraints, three muscle synergies accounted for the majority of variability [mean variance accounted for (VAF) = 93.3 ± 1.6%, VAF (muscle) > 82.5%] in the EMG signals of 11 lower limb muscles. In addition, there was a robust consistency in the muscle synergy vectors. This high similarity in the composition of the three extracted synergies was accompanied by slight adaptations in their activation coefficients in response to extreme changes in torque and posture. Thus, our results support the hypothesis that these muscle synergies reflect a neural control strategy, with only a few timing adjustments in their activation regarding the mechanical constraints.


Initial plans for a large-scale investigation into the chronic health effects of earthquakes in Italy: building on Barbara Pacelli's legacy.
Barbara Pacelli, a young Italian epidemiologist, passed away unexpectedly in September 2019. During her prolific professional life, she gave several scientific contributions to natural disaster epidemiology, particularly in relation to the medium and long-term health effects of earthquakes. In this opinion paper, we reflect on Barbara's legacy and outline potential actions that could arise from her work. Particularly, availability of electronic health records would enable a systematic and large-scale investigation into the long-term health effects of earthquakes in Italy, a country with high seismic risk. This effort would have high societal value as it would likely enable mitigation of substantial morbidity and mortality in areas affected by earthquakes. In this paper, we define scope, objectives, potential data sources, and analysis methods that could be used to systematically assess the chronic health effects of recent earthquakes in Italy. Keywords: earthquakes; chronic diseases; electronic health records; retrospective cohort; case crossover study.


Purification, crystallization and crystallographic analysis of Dictyostelium discoideum phenylalanine hydroxylase in complex with dihydrobiopterin and FeIII.
Dictyostelium discoideum phenylalanine hydroxylase (DicPAH; residues 1-415) was expressed in Escherichia coli and purified for structural analysis. Apo DicPAH and DicPAH complexed with dihydrobiopterin (BH(2)) and Fe(III) were crystallized using 0.06 M PIPES pH 7.0, 26%(w/v) PEG 2000 by the hanging-drop vapour-diffusion method. Crystals of apo DicPAH and the DicPAH-BH(2)-Fe(III) complex diffracted to 2.6 and 2.07 A resolution, respectively, and belonged to space group P2(1), with unit-cell parameters a = 70.02, b = 85.43, c = 74.86 A, beta = 110.12 degrees and a = 70.97, b = 85.33, c = 74.89 A, beta = 110.23 degrees , respectively. There were two molecules in the asymmetric unit. The structure of DicPAH has been solved by molecular replacement.


Induction of cell death in renal cell carcinoma with combination of D-fraction and vitamin C.
Although several conventional therapeutic options for advanced renal cell carcinoma (RCC) are currently available, the unsatisfactory outcomes demand establishing more effective interventions. D-fraction (PDF), a bioactive proteoglucan of Maitake mushroom, demonstrates anticancer and immunomodulatory activities, which are also shown to be potentiated by vitamin C (VC). We thus hypothesized that a combination of PDF and VC (PDF + VC) could be an alternative approach to more effectively inhibit the growth of RCC. We examined the dose-dependent effects of PDF + VC on RCC cell viability and also performed biochemical assays to explore the growth regulatory mechanism. Human RCC, ACHN cell line, was employed and exposed to varying concentrations of PDF or VC and their combinations. Cell viability at specified times was determined by MTT assay. Lipid peroxidation assay, cell cycle analysis, and Western blot analysis were also performed. PDF or VC alone led to the significant reduction in cell viability at 72 hours with PDF >500 µg/mL and VC ≥300 µM. When various combinations of PDF and VC were tested, the combination of the ineffective concentrations of PDF (300 µg/mL) and VC (200 µM) resulted in ~90% cell death in 24 hours. Lipid peroxidation assay then indicated significantly (~2.5 fold) elevated oxidative stress with this PDF + VC. Cell cycle analysis also indicated a G1 cell cycle arrest following a 6-hour PDF + VC treatment. Western blots further revealed a downregulation of Bcl2, an upregulation of Bax, and proteolytic activation of PARP (poly[ADP-ribose] polymerase) in PDF + VC-treated cells, indicating induction of apoptosis. The present study demonstrates that the combination of PDF and VC can become highly cytotoxic, inducing severe cell death in ACHN cells. This cytotoxic mechanism appears to be primarily attributed to oxidative stress, accompanied by a G1 cell cycle arrest. Such cell death induced by PDF + VC could be more likely linked to apoptosis, as indicated by the modulation of apoptosis regulators (Bcl2, Bax, and PARP). Therefore, as PDF and VC may work synergistically to induce apoptotic cell death, they may have clinical implications in an alternative, improved therapeutic modality for advanced RCC.


Conservative treatment of upper gastrointestinal bleeding in portal hypertension.
The main aim of conservative treatment of upper gastrointestinal bleeding in portal hypertension is aim to treat and prevent esophageal variceal hemorrhage. Controlled trials show that the hemostasis rate following vaso-active therapy (vasopressin and analogues, somatostatin) is only slightly superior to the spontaneous hemostasis rate. Complications caused by vasopressin treatment can be avoided by concomitant application of nitroglycerin or by alternative treatment with somatostatin. Balloon tamponade is slightly superior to vasopressin for arresting variceal hemorrhage. Injection sclerotherapy influences acute bleeding most positively. Analysis of controlled trials favors sclerotherapy for prophylaxis of rebleeding, but beta-adrenoceptor blockers appear to be almost equally good.


The role of folic acid in deficiency states and prevention of disease.
Folic acid, a water-soluble vitamin, has been used since the 1940s to treat some cases of macrocytic anemia without neurologic disease. Folate deficiency is best diagnosed with red blood cell folate levels along with macrocytosis and/or megaloblastic anemia. In addition to reversing overt deficiency, the vitamin may reduce the incidence of neural tube defects by 45% in women who receive 400 micrograms per day. It is recommended that all women of childbearing age take 400 micrograms of folate per day. Elevations in homocysteine levels, a metabolite intimately associated with folate, are also being found with increasing regularity in those with cardiovascular diseases. Homocysteine levels are reduced by folic acid administration. Therefore, there is some biologic plausibility, but not currently direct proof, for the assumption that folate supplements may prevent heart disease, stroke, and peripheral arterial disease. Controlled trials should take place before widespread food supplementation with folate is carried out on a large scale because of the possibility of outbreaks of permanent B12-related neurologic damage in those with undiagnosed pernicious anemia. However, if a patient has a premature cardiovascular event and has minimal risk factors, ordering a test to determine homocysteine level may be advisable, and if elevated, treating with folic acid supplement as long as B12 deficiency does not coexist.


Diagnosis of lung cancer by the analysis of exhaled breath with a colorimetric sensor array.
The pattern of volatile organic compounds (VOCs) in the exhaled breath of patients with lung cancer may be unique. New sensor systems that detect patterns of VOCs have been developed. One of these sensor systems, a colorimetric sensor array, has 36 spots composed of different chemically sensitive compounds impregnated on a disposable cartridge. The colours of these spots change based on the chemicals with which they come into contact. In this proof of principle study, the ability of this sensor system to detect a pattern of VOCs unique to lung cancer is assessed. Individuals with lung cancer, those with other lung diseases and healthy controls performed tidal breathing of room air for 12 min while exhaling into a device designed to draw their breath across a colorimetric sensor array. The colour changes that occurred for each individual were converted into a numerical vector. The vectors were analysed statistically, using a random forests technique, to determine whether lung cancer could be predicted from the responses of the sensor. 143 individuals participated in the study: 49 with non-small cell lung cancer, 18 with chronic obstructive pulmonary disease 15 with idiopathic pulmonary fibrosis 20 with pulmonary arterial hypertension 20 with sarcoidosis and 21 controls. A prediction model was developed using observations from 70% of the subjects. This model was able to predict the presence of lung cancer in the remaining 30% of subjects with a sensitivity of 73.3% and a specificity of 72.4% (p = 0.01). The unique chemical signature of the breath of patients with lung cancer can be detected with moderate accuracy by a colorimetric sensor array.


The ascent of man: deductions based on a multivariate analysis of the brain.
This paper attempts to investigate taxonomic and phylogenetic relationships through an analysis of the distribution of mass within the brain. A multivariate analysis was performed on encephalization ratios for various divisions of the brain, employing volume measurements reported for 63 species by Stephan et al. [1970]. The ratios for each brain region were considered to lie along one of the dimensions of eight dimensional 'brain space', and distances between all species were calculated and employed in the determination of 'family trees'. It was assumed that modern species have, in many cases, brains representative of different stages of an evolutionary progression towards higher encephalization, and it was assumed that no 'backwards evolution' occurred. The family trees based on these data and assumptions were rigidly determined and do not represent mere opinion but, rather, inescapable conclusions if one accepts the premises. Most of the findings were in very good agreement with traditional or popular ideas, and this includes conclusions that tree shrews were ancestral to prosimians and that simians are derived from a tarsioid ancestor. Other findings, however, were just as strikingly deviant from current popular and expert thought. Present methods demanded, for example, that man must have a platyrrhine ancestry. While one may reject this particular conclusion it remains true that by present measures the human brain is much more like that of an American wooley or spider monkey than like that of either the chimpanzee or the gorilla.


Beyond glucose lowering: glucagon-like peptide-1 receptor agonists, body weight and the cardiovascular system.
Glucagon-like peptide-1 (GLP-1) belongs to the incretin hormone family: in the presence of elevated blood glucose, it stimulates insulin secretion and inhibits glucagon production. In addition, GLP-1 slows gastric emptying. GLP-1 secretion has also been reported to potentially affect patients with type 2 diabetes (T2DM) compared with non-diabetics and, as enzymatic inactivation by dipeptidyl peptidase-4 (DPP-4) shortens the GLP-1 half-life to a few minutes, GLP-1 receptor agonists such as exenatide twice daily (BID) and liraglutide have been developed, and have become part of the management of patients with T2DM. This review focuses on the potential beneficial effects of these compounds beyond those associated with improvements in blood glucose control and weight loss, including changes in the cardiovascular and central nervous systems. This was a state-of-the-art review of the literature to evaluate the relationships between GLP-1, GLP-1 receptor agonists, weight and the cardiovascular system. GLP-1 receptor agonists improve glucose control and do not significantly increase the risk of hypoglycaemia. Also, this new class of antidiabetic drugs was shown to favour weight loss. Mechanisms may involve central action, direct action by reduction of food intake and probably indirect action through slowing of gastric emptying. The relative importance of each activity remains unclear. Weight loss may improve cardiovascular outcomes in patients with T2DM, although GLP-1 receptor agonists may have other direct and indirect effects on the cardiovascular system. Reductions in myocardial infarct size and improvements in cardiac function have been seen in animal models. Beneficial changes in cardiac function were also demonstrated in patients with myocardial infarcts or heart failure. Indirect effects could involve a reduction in blood pressure and potential effects on oxidation. However, the mechanisms involved in the pleiotropic effects of GLP-1 receptor agonists have yet to be completely elucidated and require further study. These compounds may play an important role in the treatment of patients with T2DM as their potential effects go beyond glucose-lowering (weight loss, potential improvement of cardiovascular risk factors). However, to better understand their place in the management of T2DM, further experimental and clinical prospective studies are required.


[The heart in dermatomyositis and polymyositis ].
We studied eleven consecutive patients: eight with Dermatomyositis (DM) and three with Polymyositis (PM) from the cardiological point of view through non invasive methods. Nine patients (82%) had some kind of cardiopulmonary complications as shown by any of the used methods. eight (73%) referred some kind of cardiopulmonary symptoms, mainly dyspnea; Physical examination; in seven (64%) was abnormal, detecting increased second pulmonary sound in four (36%), findings of mitral valve prolapse (MVP) in two (18%) and in two (18%) S3 gallop; Electrocardiogram: in seven (64%) was abnormal; six (55%) had some kind of heart enlargement corresponding four (36%) to right atrial or ventricular hypertrophy (RAH & RVH) and two (18%) to left ventricular hypertrophy (LVH), three (27%) had incomplete or complete right bundle branch block, one (9%) had bifascicular block and one (9%) left anterior hemiblock. Two (18%) had sinus tachycardia and two (18%) atrial premature contractions; d) chest ray: six (55%) were abnormal, among them, three (27%) had pulmonary fibrosis, three (27%) had RAH and/or RVH, two (18%) had LVH and one (9%) pericardial effusion; e) Echocardiogram: was abnormal in eight (73%), corresponding three (27%) to RVH, three to MVP which has been considered rare, in two (18%) congestive cardiomyopathy, in two (18%) pericardial effusion and in one (9%) type "A" paradoxical septal movement.


Susceptibility and gene interaction study of the angiotensin II type 1 receptor (AGTR1) gene polymorphisms with non-alcoholic fatty liver disease in a multi-ethnic population.
Angiotensin II type 1 receptor (AGTR1) has been reported to play a fibrogenic role in non-alcoholic fatty liver disease (NAFLD). In this study, five variants of the AGTR1 gene (rs3772622, rs3772627, rs3772630, rs3772633, and rs2276736) were examined for their association with susceptibility to NAFLD. Subjects made up of 144 biopsy-proven NAFLD patients and 198 controls were genotyped using TaqMan assays. The liver biopsy specimens were histologically graded and scored according to the method of Brunt. Single locus analysis in pooled subjects revealed no association between each of the five variants with susceptibility to NAFLD. In the Indian ethnic group, the rs2276736, rs3772630 and rs3772627 appear to be protective against NAFLD (p = 0.010, p = 0.016 and p = 0.026, respectively). Haplotype ACGCA is shown to be protective against NAFLD for the Indian ethnic subgroup (p = 0.03). Gene-gene interaction between the AGTR1 gene and the patatin-like phospholipase domain-containing 3 (PNPLA3) gene, which we previously reported as associated with NAFLD in this sample, showed a strong interaction between AGTR1 (rs3772627), AGTRI (rs3772630) and PNPLA3 (rs738409) polymorphisms on NAFLD susceptibility (p = 0.007). Further analysis of the NAFLD patients revealed that the G allele of the AGTR1 rs3772622 is associated with increased fibrosis score (p = 0.003). This is the first study that replicates an association between AGTR1 polymorphism and NAFLD, with further details in histological features of NAFLD. There is lack of evidence to suggest an association between any of the five variants of the AGTR1 gene and NAFLD in the Malays and Chinese. In the Indians, the rs2276736, rs3772630 and rs3772627 appear to protect against NAFLD. We report novel findings of an association between the G allele of the rs3772622 with occurrence of fibrosis and of the gene-gene interaction between AGTR1gene and the much-studied PNPLA3 gene.


Grassland systems of red meat production: integration between biodiversity, plant nutrient utilisation, greenhouse gas emissions and meat nutritional quality.
Government policies relating to red meat production take account of the carbon footprint, environmental impact, and contributions to human health and nutrition, biodiversity and food security. This paper reviews the impact of grazing on these parameters and their interactions, identifying those practices that best meet governments' strategic goals. The recent focus of research on livestock grazing and biodiversity has been on reducing grazing intensity on hill and upland areas. Although this produces rapid increases in sward height and herbage mass, changes in structural diversity and plant species are slower, with no appreciable short-term increases in biodiversity so that environmental policies that simply involve reductions in numbers of livestock may not result in increased biodiversity. Furthermore, upland areas rely heavily on nutrient inputs to pastures so that withdrawal of these inputs can threaten food security. Differences in grazing patterns among breeds increase our ability to manage biodiversity if they are matched appropriately to different conservation grazing goals. Lowland grassland systems differ from upland pastures in that additional nutrients in the form of organic and inorganic fertilisers are more frequently applied to lowland pastures. Appropriate management of these nutrient applications is required, to reduce the associated environmental impact. New slurry-spreading techniques and technologies (e.g. the trailing shoe) help reduce nutrient losses but high nitrogen losses from urine deposition remain a key issue for lowland grassland systems. Nitrification inhibitors have the greatest potential to successfully tackle this problem. Greenhouse gas (GHG) emissions are lower from indoor-based systems that use concentrates to shorten finishing periods. The challenge is to achieve the same level of performance from grass-based systems. Research has shown potential solutions through the use of forages containing condensed tannins or establishing swards with a high proportion of clover and high-sugar grasses. Relative to feeding conserved forage or concentrates, grazing fresh grass not only reduces GHG emissions but also enhances the fatty acid composition of meat in terms of consumer health. It is possible to influence biodiversity, nutrient utilisation, GHG emissions and the nutritional quality of meat in grass-based systems, but each of these parameters is intrinsically linked and should not be considered in isolation. Interactions between these parameters must be considered carefully when policies are being developed, in order to ensure that strategies designed to achieve positive gains in one category do not lead to a negative impact in another. Some win-win outcomes are identified.


Mtl1 is required to activate general stress response through Tor1 and Ras2 inhibition under conditions of glucose starvation and oxidative stress.
Mtl1 is a member of the cell wall integrity (CWI) pathway of Saccharomyces cerevisiae, which functions as a cell wall sensor for oxidative stress. Genome-wide transcriptional analysis revealed a cluster of genes that were down-regulated in the absence of Mtl1. Many of these genes were potentially regulated by the general stress response factor Msn2/Msn4. In response to rapamycin, caffeine, glucose starvation and oxidative stress provoked by H(2)O(2), mtl1 presents a significant loss of viability as well as a deficiency in the transcriptional response mediated by Msn2/Msn4. The Mtl1 function was required (i) to induce ribosomal gene repression, (ii) to induce the general stress response driven by the transcription factor Msn2/Msn4, and (iii) to activate the CWI pathway in response to both glucose starvation and oxidative stress. We also detected higher cAMP levels in the mtl1 mutant than in wild type cells indicative of up-regulated RAS2-PKA activity. Disruption of TOR1, disruption of RAS2, or hyperactivation of Rho1 restored both the viability and the transcriptional function (both ribosomal and Msn2/Msn4-dependent gene expression) in the mtl1 mutant to almost wild type levels when cells were starved of glucose or stressed with H(2)O(2). Taking our results together, we propose an essential role for Mtl1 in signaling oxidative stress and quiescence to the CWI pathway and to the general stress response through Rho1 and the inhibition of either the TOR1 or RAS2 functions. These mechanisms would be required to allow cells to adapt to both oxidative and nutritional stresses.


Aerolysin--a paradigm for membrane insertion of beta-sheet protein toxins?
The determination of the crystal structure of the bacterial protein proaerolysin provided the first view of a pore-forming toxin constructed mainly from beta-sheet. The structure that was obtained and subsequent crystallographic and biochemical studies have together allowed us to explain how the toxin is transformed from a water-soluble dimer to a heptameric transmembrane pore. Recent discoveries of structural similarities between aerolysin and other toxins suggest that the structure/function studies we have made may prove useful in understanding the actions of a number of pore-forming proteins.


ATRX loss refines the classification of anaplastic gliomas and identifies a subgroup of IDH mutant astrocytic tumors with better prognosis.
Mutation/loss of alpha-thalassemia/mental retardation syndrome X-linked (ATRX) expression has been described in anaplastic gliomas. The present study explored the role of ATRX status in the molecular classification of anaplastic gliomas and its impact on survival in the biomarker cohort of the NOA-04 anaplastic glioma trial. Patients (n = 133) of the NOA-04 trial were analyzed for ATRX expression using immunohistochemistry. ATRX status was correlated with age, histology, isocitrate dehydrogenase (IDH), 1p/19q, alternative lengthening of telomeres (ALT) and O6-methylguanine-DNA methyltransferase (MGMT) status, and the trial efficacy endpoints. Loss of ATRX expression was detected in 45 % of anaplastic astrocytomas (AA), 27 % of anaplastic oligoastrocytomas (AOA) and 10 % of anaplastic oligodendrogliomas (AO). It was mostly restricted to IDH mutant tumors and almost mutually exclusive with 1p/19q co-deletion. The ALT phenotype was significantly correlated with ATRX loss. ATRX and 1p/19q status were used to re-classify AOA: AOA harboring ATRX loss shared a similar clinical course with AA, whereas AOA carrying 1p/19q co-deletion shared a similar course with AO. Accordingly, in a Cox regression model including ATRX and 1p/19q status, histology was no longer significantly associated with time to treatment failure. Survival analysis showed a marked separation of IDH mutant astrocytic tumors into two groups based on ATRX status: tumors with ATRX loss had a significantly better prognosis (median time to treatment failure 55.6 vs. 31.8 months, p = 0.0168, log rank test). ATRX status helps better define the clinically and morphologically mixed group of AOA, since ATRX loss is a hallmark of astrocytic tumors. Furthermore, ATRX loss defines a subgroup of astrocytic tumors with a favorable prognosis.


Plant ionome diagnosis using sound balances: case study with mango (Mangifera Indica).
Plant ionomes and soil nutrients are commonly diagnosed in agronomy using concentration and nutrient ratio ranges. However, both diagnoses are biased by redundancy of information, subcompositional incoherence and non-normal distribution inherent to compositional data, potentially leading to conflicting results and wrong inferences. Our objective was to present an unbiased statistical approach of plant nutrient diagnosis using a balance concept and mango (Mangifera indica) as test crop. We collected foliar samples at flowering stage in 175 mango orchards. The ionomes comprised 11 nutrients (S, N, P, K, Ca, Mg, B, Cu, Zn, Mn, Fe). Traditional multivariate methods were found to be biased. Ionomes were thus represented by unbiased balances computed as isometric log ratios (ilr). Soil fertility attributes (pH and bioavailable nutrients) were transformed into balances to conduct discriminant analysis. The orchards differed more from genotype than soil nutrient signatures. A customized receiver operating characteristic (ROC) iterative procedure was developed to classify tissue ionomes between balanced/misbalanced and high/low-yielders. The ROC partitioning procedure showed that the critical Mahalanobis distance of 4.08 separating balanced from imbalanced specimens about yield cut-off of 128.5 kg fruit tree(-1) proved to be a fairly informative test (area under curve = 0.84-0.92). The [P | N,S] and [Mn | Cu,Zn] balances were found to be potential sources of misbalance in the less productive orchards, and should thus be further investigated in field experiments. We propose using a coherent pan balance diagnostic method with median ilr values of top yielders centered at fulcrums of a mobile and the critical Mahalanobis distance as a guide for global nutrient balance. Nutrient concentrations in weighing pans assisted appreciating nutrients as relative shortage, adequacy or excess in balances.


The p38 MAPK pathway is critical for 5,5'-dibromodiindolylmethane-induced apoptosis to prevent oral squamous carcinoma cells.
Cruciferous vegetables contain isothiocyanates including diindolylmethane (DIM) that exhibit cancer chemopreventive effects. We developed a series of synthetic ring-substituted DIM analogs including 5,5'-dibromoDIM that exhibited better inhibitory activity in breast and colon cancer cells than DIM. In this study, we investigated whether 5,5'-dibromoDIM inhibits the proliferation of KB and YD-10B oral squamous carcinoma cell lines. 5,5'-dibromoDIM decreased the cell survival and inhibited the growth of oral cancer cells. Exposure of KB and YD-10B cells to 5,5'-dibromoDIM induced caspase-dependent apoptosis evidenced by poly-ADP ribose polymerase cleavage, accumulation of sub-G1 population, and nuclear condensation and fragmentation. In addition, apoptotic cell death was correlated with damage to the mitochondrial membrane potential through a decrease in the level of Bcl-2 protein expression. Mechanistic studies showed that mitochondria-dependent apoptosis induced by 5,5'-dibromoDIM was mediated by the p38 mitogen-activated protein kinase pathway but not the ERK1/2 and JNK pathway. These results highlight 5,5'-dibromoDIM as an important chemopreventive agent for the clinical treatment of oral cancer through the p38 mitogen-activated protein kinase pathway.


Lipid and cholesterols modulate the dynamics of SARS-CoV-2 viral ion channel ORF3a and its pathogenic variants.
SARS-CoV-2 accessory protein, ORF3a is a putative ion channel which immensely contributes to viral pathogenicity by modulating host immune responses and virus-host interactions. Relatively high expression of ORF3a in diseased individuals and implication with inflammasome activation, apoptosis and autophagy inhibition, ratifies as an effective target for developing vaccines and therapeutics. Herein, we present the elusive dynamics of ORF3a-dimeric state using all-atoms molecular dynamics (MD) simulations at μ-seconds scale in a heterogeneous lipid-mimetic system in multiple replicates. Additionally, we also explore the effect of non-synonymous pathogenic mutations on ORF3a ion channel activity and viral pathogenicity in different SARS-CoV-2 variants using various structure-based protein stability (ΔΔG) tools and computational saturation mutagenesis. Our study ascertains the role of phosphatidylcholines and cholesterol in modulating the structure of ORF3a, which perturbs the size and flexibility of the polar cavity that allows permeation of large cations. Discrete trend in ion channel pore radius and area per lipid arises the premise that presence of lipids might also affect the overall conformation of ORF3a. MD structural-ensembles, in some replicates rationalize the crucial role of TM2 in maintaining the native structure of ORF3a. We also infer that loss of structural stability primarily grounds for pathogenicity in more than half of the pathogenic variants of ORF3a. Overall, the effect of mutation on alteration of ion permeability of ORF3a, proposed in this study brings mechanistic insights into variant consequences on viral membrane proteins of SARS-CoV-2, which can be utilized for the development of novel therapeutics to treat COVID-19 and other coronavirus diseases.


First report of epizootic trichomoniasis in wild finches (family Fringillidae) in southern Fennoscandia.
Forty-one outbreaks of mortality in wild finches were reported in southern Norway, Sweden, and Finland in the second half of 2008 (n = 40) and in February 2009 (n = 1). Greenfinches (Carduelis chloris) and occasional chaffinches (Fringilla coelebs) primarily were affected. Forty-eight greenfinches, eight chaffinches, one hawfinch (Coccothraustes coccothraustes), and one blue tit (Parus caeruleus) from 22 incidents were examined postmortem. Birds were in poor nutritional condition and had necrotizing ingluvitis, esophagitis, and/or oropharyngitis. Viable trichomonads with morphology consistent with Trichomonas gallinae were demonstrated successfully in 65% and 71% of fresh carcasses examined by culture and wet mount, respectively. No primary bacterial pathogens were detected. To our knowledge, this is the first report of epizootic trichomoniasis in wild finches in Europe outside of the UK.


A comparative analysis of open surgery vs endoscopic balloon dilation for pediatric subglottic stenosis.
Minimally invasive endoscopic techniques are an appealing alternative to open surgical management of pediatric subglottic stenosis (SGS), but more information is needed to understand the comparative risks, benefits, and limitations of such interventions. To compare the effectiveness of endoscopic balloon dilation (EBD) and laryngotracheoplasty (LTP) in pediatric patients with SGS and to identify patient and disease factors that are associated with successful EBD. A retrospective medical record review of children undergoing EBD and LTP for SGS in a tertiary care children's hospital from 2006 through 2012. Success was defined as decannulation or tracheotomy avoidance. Additional outcomes were total number of procedures and number of unplanned procedures. Univariate χ2 analyses and multivariate regression analyses were performed to identify patient and disease factors statistically associated with success within treatment groups. Overall, 86 of 90 patients (96%) successfully avoided tracheotomy or were decannulated. Fourteen patients were successfully treated with EBD, but for 13 patients, EBD failed, and they underwent LTP. A total of 76 patients underwent LTP. In univariate analyses, patients for whom EBD was successful were more likely to have mild (grade 1 or 2; n = 10) than severe (grade 3 or 4; n = 4) SGS compared with patients for whom EBD failed (grade 1 or 2, n = 0 vs grade 3 or 4, n = 13) (P < .001). Three patients who underwent initial EBD had worsening stenosis. Patients initially treated with EBD were more likely to require unplanned surgical intervention during their treatment (6 of 27; 22%) than patients initially treated with LTP (3 of 63; 5%) (P = .01). Patients initially treated with EBD had a lower number of airway interventions and/or evaluations under anesthesia (mean, 6.7) during their course of treatment than patients initially treated with LTP (mean, 9.2) (P = .003). In multivariate analyses, only severe SGS was significantly associated with failure of initial EBD (13 of 13 [100%] with type 3 or 4 vs 4 of 14 with type 1 or 2 [29%]) (P = .002). For severe SGS, EBD has limited application compared with LTP, and in some cases failed EBD is even detrimental, increasing the risk of unplanned urgent interventions compared with LTP.


Shr of group A streptococcus is a new type of composite NEAT protein involved in sequestering haem from methaemoglobin.
A growing body of evidence suggests that surface or secreted proteins with NEAr Transporter (NEAT) domains play a central role in haem acquisition and trafficking across the cell envelope of Gram-positive bacteria. Group A streptococcus (GAS), a β-haemolytic human pathogen, expresses a NEAT protein, Shr, which binds several haemoproteins and extracellular matrix (ECM) components. Shr is a complex, membrane-anchored protein, with a unique N-terminal domain (NTD) and two NEAT domains separated by a central leucine-rich repeat region. In this study we have carried out an analysis of the functional domains in Shr. We show that Shr obtains haem in solution and furthermore reduces the haem iron; this is the first report of haem reduction by a NEAT protein. More specifically, we demonstrate that both of the constituent NEAT domains of Shr are responsible for binding haem, although they are missing a critical tyrosine residue found in the ligand-binding pocket of other haem-binding NEAT domains. Further investigations show that a previously undescribed region within the Shr NTD interacts with methaemoglobin. Shr NEAT domains, however, do not contribute significantly to the binding of methaemoglobin but mediate binding to the ECM components fibronectin and laminin. A protein fragment containing the NTD plus the first NEAT domain was found to be sufficient to sequester haem directly from methaemoglobin. Correlating these in vitro findings to in vivo biological function, mutants analysis establishes the role of Shr in GAS growth with methaemoglobin as a sole source of iron, and indicates that at least one NEAT domain is necessary for the utilization of methaemoglobin. We suggest that Shr is the prototype of a new group of NEAT composite proteins involved in haem uptake found in pyogenic streptococci and Clostridium novyi.


Effect of systemic candidiasis on blastogenesis of lymphocytes from germfree and conventional rats.
Germfree and conventional rats were challenged (intravenously) with Candida albicans and sacrificed at various times after infection, and their spleen cells were harvested to examine the effect of disseminated candidiasis on in vitro lymphocyte hypersensitivity to Candida antigens (CA). Results showed that conventional rat splenocytes, initially responsive in vitro to stimulation by CA, manifested a depression in CA-specific responsiveness after challenge with viable C. albicans (days 3 to 6 postchallenge). In addition, the latter splenocyte response to phytohemagglutinin (PHA) and concanavalin A (ConA) was suppressed by 3 to 6 days after challenge with Candida. In contrast to conventional rats, the response of germfree rat splenocytes to CA was insignificant before challenge with C. albicans, and it was increased at 9 days after infection. The response of uninfected germfree rat splenocytes to PHA and ConA was significantly lower than that of unchallenged conventional rats. Challenge with viable C. albicans did not result in a suppression of gnotobiotic rat splenocyte responses to PHA and ConA, but rather, the disseminated infection resulted in as much as fivefold increases in PHA or ConA-induced blastogenesis. These findings suggest that disseminated candidiasis is capable of suppressing blastogenesis in immunologically mature conventional rats and of improving lymphocyte blastogenesis from immunologically immature germfree rats.


Religion: a mediator of Anglo-American and Mexican attributional differences toward symptoms of schizophrenia?
This study examined the relationship of religiosity to attributions toward schizophrenia, within a cultural context. Previous research suggests that on self-report measures, Mexicans endorse holding greater moral-religious values than do their Anglo-American counterparts. Research also indicates that Mexicans, relative to Anglo-Americans, tend to hold fewer blameworthy attributions and are less likely to view patients with schizophrenia as responsible for the symptoms of the disorder. In an analog study of 88 Mexican and 88 Anglo-American college students asked to imagine that they have a brother with schizophrenia, this study assessed two competing hypotheses regarding the role of religion in shaping reactions to schizophrenia in a family member. For hypothesis 1, Baron and Kenny's mediational model (1986) was used to assess whether moral religious values may play a direct mediating role between ethnicity and controllability attributions for schizophrenia. In other words, based on observations of previous researchers, this set of analyses assessed whether ethnic differences in controllability attributions might be explained by a religious or spiritual tendency in Mexicans to view negative events, such as mental disability, as rooted in divine factors beyond the patient's personal control. In contrast, a second, competing, hypothesis was also assessed in this study: namely, that greater religiosity would be positively correlated with increasing perceptions of control over the symptoms of schizophrenia. This hypothesis stems from the premise of several investigators that religious individuals may be more likely to perceive another's adversity (such as having schizophrenia) as a punishment for prior wrongdoings or for failure to try to help oneself. Results indicate support for the latter hypothesis. Implications of the paradoxical finding, that Mexicans were both more religious and more external in their attributions, are discussed.