Phenylephrine increases near-infrared spectroscopy determined muscle oxygenation in men.
Phenylephrine increases mean arterial pressure (MAP) by enhanced total peripheral resistance (TPR) but near-infrared spectroscopy (NIRS) determined muscle oxygenation (SmO2) increases. We addressed that apparent paradox during supine rest and head-up tilt (HUT). Variables were determined ± phenylephrine in males during supine rest (n = 17) and 40° HUT (n = 7). MAP, stroke volume (SV), heart rate (HR), and TPR were derived by Modelflow® and NIRS determined biceps SmO2 and (tibial) bone oxygenation (StibialO2). For ten subjects, cardiac filling and the diameter of the inferior caval vein (ICV collapsibility index: ((ICVexpiration - ICVinspiration)/ICVexpiration) × 100) were assessed by ultrasound. Pancreatic polypeptide (PP) and atrial natriuretic peptide (proANP) in plasma were determined by immunoassay. Brachial artery blood flow was assessed by ultrasound and skin oxygenation (SskinO2) monitored by white light spectroscopy. Phenylephrine increased MAP by 34% and TPR (62%; P < 0.001) during supine rest. The ICV collapsibility index decreased (24%; P < 0.001) indicating augmented cardiac preload although volume of the left atrium and ventricle did not change. SV increased (18%; P < 0.001) as HR decreased (24%; P < 0.001). ProANP increased by 9% (P = 0.002) with unaffected PP. Brachial artery blood flow tended to decrease while SskinO2 together with StibialO2 decreased by 11% (P = 0.026) and 20% (P < 0.001), respectively. Conversely, phenylephrine increased SmO2 (9%) and restored the HUT elicited decrease in SmO2 (by 19%) along with SV (P = 0.02). Phenylephrine reduces skin and bone oxygenation and tends to reduce arm blood flow, suggesting that the increase in SmO2 reflects veno-constriction with consequent centralization of the blood volume.


BubR1- and Polo-coated DNA tethers facilitate poleward segregation of acentric chromatids.
The mechanisms that safeguard cells against chromosomal instability (CIN) are of great interest, as CIN contributes to tumorigenesis. To gain insight into these mechanisms, we studied the behavior of cells entering mitosis with damaged chromosomes. We used the endonuclease I-CreI to generate acentric chromosomes in Drosophila larvae. While I-CreI expression produces acentric chromosomes in the majority of neuronal stem cells, remarkably, it has no effect on adult survival. Our live studies reveal that acentric chromatids segregate efficiently to opposite poles. The acentric chromatid poleward movement is mediated through DNA tethers decorated with BubR1, Polo, INCENP, and Aurora-B. Reduced BubR1 or Polo function results in abnormal segregation of acentric chromatids, a decrease in acentric chromosome tethering, and a great reduction in adult survival. We propose that BubR1 and Polo facilitate the accurate segregation of acentric chromatids by maintaining the integrity of the tethers that connect acentric chromosomes to their centric partners.


Shell composition has no net impact on large-scale evolutionary patterns in mollusks.
A major suspected bias in the fossil record of skeletonized groups is variation in preservability owing to differences in shell composition. However, despite extensive changes in shell composition over the 500-million-year history of marine bivalves, genus duration and shell composition show few significant relationships, and of those, virtually all are contrary to bias from preferential loss of highly reactive shell types. Distortion of large-scale temporal patterns in marine bivalves owing to preservability is thus apparently weak or randomly distributed, which increases the likelihood that observed patterns in this and other shelled groups carry a strong biological signal.


Transcription factor Tbx18 induces the differentiation of c-kit+ canine mesenchymal stem cells (cMSCs) into SAN-like pacemaker cells in a co-culture model in vitro.
Bone mesenchymal stem cells (MSCs), as well as cardiomyocytes, are derived from early mesoderm, becoming committed to their fate under the influence of different differentiation factors. We examined whether the overexpression of Tbx18 can induce the differentiation of c-kit+ cMSCs into a phenotype similar to that of native pacemaker cells and whether these transfected cells can couple to adjacent atrial cells with functional consequences. The c-kit+ cMSCs were first sorted, then transfected with different lentiviral vectors. Tbx18-c-kit+ cMSCs represented the experimental group, while EYFP-c-kit+ cMSCs and canine sinoatrial node (SAN) cells were used as controls. Within days of transfection, the hyperpolarization-activated cyclic nucleotide-gated (HCN) channel HCN4 protein and gap junction protein Connexin 45 (Cx45) expression in Tbx18-c-kit+ cMSCs were 12-fold and 5.6-fold higher, respectively, than that in EYFP-c-kit+ cMSCs. After co-culture with canine atrial cells in vitro for three days, the funny currents (If) were recorded in the Tbx18-c-kit+ cMSCs, but not in EYFP-c-kit+ cMSCs. The trend of these If currents was highly similar to that of SAN cells, although the current density was smaller. The Tbx18-EYFP-c-kit+ cMSCs showed responsiveness to β-adrenergic stimulation, and the intracellular cyclic adenosine monophosphate (cAMP) level was higher than that in EYFP-c-kit+ cMSCs. The Tbx18-EYFP-c-kit+ cMSCs delivered fluorescent dye to neighboring atrial cells via gap junctions, thus these cell pairs could communicate as a pacemaker unit. We propose that the overexpression of Tbx18 in c-kit+ cMSCs induces their differentiation to SAN-like pacemaker cells.


Modeling cell-to-cell stochastic variability in intrinsic apoptosis pathway.
Apoptosis is a cell suicide mechanism that enables metazoans to control cell number in tissues and to eliminate individual cells that threaten the animal's survival. Dependent on the type of stimulus, apoptosis can be propagated by intrinsic pathway or extrinsic pathway. Previously, we have proposed a deterministic model of intrinsic apoptosis pathway which is bistable in a robust parameter region. Cellular networks, however, are inherently stochastic and significant cell-to-cell variability in apoptosis response has been observed at single cell level. In this work, we examine the impact of intrinsic stochastic fluctuations as well as variation of protein concentrations on behavior of the intrinsic apoptosis network. First, Gillespie Stochastic Simulation Algorithm (SSA) of the model is implemented to account for intrinsic noise. Using histograms of steady-state output at varying input levels, we show that the intrinsic noise in the apoptosis network elicits a wider region of bistability. We further analyze the dependence of system stochasticity due to intrinsic fluctuations, such as steady-state noise level and random response delay time, on the input signal. We find however that the intrinsic noise is insufficient to generate significant stochastic variations at physiologically relevant level of molecular numbers. Finally, extrinsic fluctuation represented by variations of two key proteins is modeled and the resultant stochasticity of apoptosis timing is analyzed. Indeed, these protein variations can induce cell-to-cell stochastic variability at a quantitative level agreeing with experiments. Therefore, we conclude that the heterogeneity in intrinsic apoptosis responses among individual cells plausibly arises from extrinsic rather than intrinsic origin of fluctuations.


The Anthropocene: conceptual and historical perspectives.
The human imprint on the global environment has now become so large and active that it rivals some of the great forces of Nature in its impact on the functioning of the Earth system. Although global-scale human influence on the environment has been recognized since the 1800s, the term Anthropocene, introduced about a decade ago, has only recently become widely, but informally, used in the global change research community. However, the term has yet to be accepted formally as a new geological epoch or era in Earth history. In this paper, we put forward the case for formally recognizing the Anthropocene as a new epoch in Earth history, arguing that the advent of the Industrial Revolution around 1800 provides a logical start date for the new epoch. We then explore recent trends in the evolution of the Anthropocene as humanity proceeds into the twenty-first century, focusing on the profound changes to our relationship with the rest of the living world and on early attempts and proposals for managing our relationship with the large geophysical cycles that drive the Earth's climate system.


Asynchronous diversification in a specialized plant-pollinator mutualism.
Most flowering plants establish mutualistic associations with insect pollinators to facilitate sexual reproduction. However, the evolutionary processes that gave rise to these associations remain poorly understood. We reconstructed the times of divergence, diversification patterns, and interaction networks of a diverse group of specialized orchids and their bee pollinators. In contrast to a scenario of coevolution by race formation, we show that fragrance-producing orchids originated at least three times independently after their fragrance-collecting bee mutualists. Whereas orchid diversification has apparently tracked the diversification of orchids' bee pollinators, bees appear to have depended on the diverse chemical environment of neotropical forests. We corroborated this apparent asymmetrical dependency by simulating co-extinction cascades in real interaction networks that lacked reciprocal specialization. These results suggest that the diversification of insect-pollinated angiosperms may have been facilitated by the exploitation of preexisting sensory biases of insect pollinators.


Relative therapeutic efficacy of (125)I- and (131)I-labeled monoclonal antibody A33 in a human colon cancer xenograft.
A33, a monoclonal antibody that targets colon carcinomas, was labeled with (125)I or (131)I and the relative therapeutic efficacy of the 2 radiolabeled species was compared in a human colon cancer xenograft system. Nude mice bearing human SW1222 colon carcinoma xenografts were administered escalating activities of (125)I-A33 (9.25-148 MBq) or (131)I-A33 (0.925-18.5 MBq), (125)I- and (131)I-labeled control antibodies, unlabeled antibody, or no antibody. The effects of treatment were assessed using the endpoints of tumor growth delay and cure. Tumor growth delay increased with administered activity for all radiolabeled antibodies. Approximately 4.5 times more activity was required for (125)I-A33 to produce therapeutic effects that were equivalent to those of (131)I-A33. This ratio was approximately 7 for a nonspecific, noninternalizing isotype-matched, radiolabeled control antibody. Unlabeled A33 antibody had no effect on tumor growth. Approximately 10 times more activity of (125)I-A33 produced toxicity similar to that of (131)I-A33, and this ratio fell to approximately 6 for radiolabeled control antibody. Treatment with (125)I-A33 resulted in a relative therapeutic gain of approximately 2 compared with (131)I-A33 in this experimental system.


Acceptability and feasibility of a behavioral and mobile health intervention (COMBIND) shown to increase uptake of prevention of mother to child transmission (PMTCT) care in India.
A cluster-randomized trial recently demonstrated that an integrated behavioral and mobile technology intervention improved uptake of key components of a Prevention of Mother to Child Transmission (PMTCT) Option B+ program, among HIV- infected pregnant/breastfeeding women in India. To guide scale-up and optimize programmatic implementation, we conducted a mixed-methods evaluation of the feasibility and acceptability of this intervention. The COMmunity Home Based INDia (COMBIND) study, was conducted in four districts of Maharashtra, India and randomized 119 integrated counseling and testing centers (ICTC) and their outreach workers (ORWs) to the COMBIND intervention, an integrated mHealth application that allowed digital data capture, PMTCT educational videos, SMS alerts for missed visits and reminder for visits, combined with personal empowerment and motivational interviewing training for ORWs. This qualitative evaluation was done through 15 in-depth interviews (IDIs) with ORWs and 15 IDIs with HIV-infected pregnant/breastfeeding women from the intervention arm. Utilizing a concurrent nested mixed-method evaluation approach, we assess the feasibility and acceptability of the study intervention. All 30 participants reported that the PMTCT videos were essential in providing easy to understand information on critical aspects of HIV and necessary care related to PMTCT practices. A majority of the ORWs reported that the personal empowerment training with motivational interviewing skills training increased their confidence, motivation and gave them the tools for effectively supporting their clients. The mHealth application improved their working style as it facilitated targeted PMTCT information support, systemized data capture, streamlined their health education delivery practice and provided a sense of work satisfaction. The SMS appointment alerts improved retention in HIV care for mother and baby to the smaller proportion that had access to their phones. Despite reported improvements in knowledge and communication, few ORWs reported that structural challenges such as limited drug stocks, lack of HIV kits or unavailability of trained staff at ICTC, may hamper the uptake of PMTCT services, thus resulting in limited significant impacts of COMBIND on PMTCT outcomes. This study found that COMBIND intervention is scalable, feasible, beneficial and very well accepted by ORWs and patients, however structural challenges in goods and services remain.


Causes and pattern of unilateral hand injuries.
To assess the causes and pattern of hand injuries in patients with isolated unilateral acute hand injuries managed at the Kenyatta National Hospital (KNH). A prospective cross-sectional descriptive study. Kenyatta National Hospital, Nairobi, Kenya, between May and August 2006. All patients with isolated unilateral acute hand injuries who presented at the casualty department. A total of 99 patients were recruited. The mean age was 28.2 years with the modal age being 21-30 years. More males were injured than females. Occupational injuries and assaults were the most prevalent causes of hand injury. Majority of the injuries included lacerations, fractures and tendon injuries. The distal phalanges of the ring and long fingers were the most common sites of injury on the digits. Occupational hand injuries, a largely preventable problem, are the most prevalent cause of hand injury. This offers opportunity for strategies in preventing a large number of hand injuries by initiating safety measures at the work place.


Tunable blue-green-emitting wurtzite ZnS:Mg nanosheet-assembled hierarchical spheres for near-UV white LEDs.
Mg-doped ZnS hierarchical spheres have been synthesized via hydrothermal method using mixed solvents of ethylenediamine and DI water without any surface-active agent. The surface morphology and microstructure studies revealed that the hierarchical spheres were consisted of many well-aligned nanosheets with width 10 nm and length about 50 ~ 100 nm. X-Ray diffraction results show that the ZnS:Mg hierarchical spheres have wurtzite structure with high crystallinity. The absorption edge in the diffuse reflection spectra shifts towards lower wavelength with increasing Mg concentration, indicating an expansion in the bandgap energy that is estimated to be in the range of 3.28 to 3.47 eV. Blue-green photoluminescence with tunable intensity and peak position was observed depending on the Mg content. The Mg2+-activated ZnS phosphor can be good candidates for blue-green components in near-UV white light-emitting diodes.


Molecular frame photoemission by a comb of elliptical high-order harmonics: a sensitive probe of both photodynamics and harmonic complete polarization state.
Due to the intimate anisotropic interaction between an XUV light field and a molecule resulting in photoionization (PI), molecular frame photoelectron angular distributions (MFPADs) are most sensitive probes of both electronic/nuclear dynamics and the polarization state of the ionizing light field. Consequently, they encode the complex dipole matrix elements describing the dynamics of the PI transition, as well as the three normalized Stokes parameters s1, s2, s3 characterizing the complete polarization state of the light, operating as molecular polarimetry. The remarkable development of advanced light sources delivering attosecond XUV pulses opens the perspective to visualize the primary steps of photochemical dynamics in time-resolved studies, at the natural attosecond to few femtosecond time-scales of electron dynamics and fast nuclear motion. It is thus timely to investigate the feasibility of measurement of MFPADs when PI is induced e.g., by an attosecond pulse train (APT) corresponding to a comb of discrete high-order harmonics. In the work presented here, we report MFPAD studies based on coincident electron-ion 3D momentum imaging in the context of ultrafast molecular dynamics investigated at the PLFA facility (CEA-SLIC), with two perspectives: (i) using APTs generated in atoms/molecules as a source for MFPAD-resolved PI studies, and (ii) taking advantage of molecular polarimetry to perform a complete polarization analysis of the harmonic emission of molecules, a major challenge of high harmonic spectroscopy. Recent results illustrating both aspects are reported for APTs generated in unaligned SF6 molecules by an elliptically polarized infrared driving field. The observed fingerprints of the elliptically polarized harmonics include the first direct determination of the complete s1, s2, s3 Stokes vector, equivalent to (ψ, ε, P), the orientation and the signed ellipticity of the polarization ellipse, and the degree of polarization P. They are compared to so far incomplete results of XUV optical polarimetry. We finally discuss the comparison between the outcomes of photoionization and high harmonic spectroscopy for the description of molecular photodynamics.


Respiratory controversies in the critical care setting. Does high-frequency ventilation offer benefits over conventional ventilation in adult patients with acute respiratory distress syndrome?
High-frequency ventilation is the application of mechanical ventilation with a respiratory rate > 100 breaths/min. High-frequency oscillatory ventilation (HFOV) is the form of high-frequency ventilation most widely used in adult critical care. The principles of lung-protective ventilation have matured in parallel with the technology for HFOV. The 2 basic principles of lung-protective ventilation are the use of small tidal volume and maintenance of adequate alveolar recruitment. Research in animal models and humans demonstrate that HFOV can support gas exchange with much smaller tidal volume than can be achieved with conventional ventilation. HFOV also provides more effective lung recruitment than conventional mechanical ventilation. However, at present, evidence is lacking that survival in adults with acute respiratory distress syndrome is improved by HFOV. Although HFOV may improve P(aO(2)) in some patients, this improvement is often transitory. Available evidence does not support that pulmonary inflammation is reduced with HFOV in adult acute respiratory distress syndrome. Heavy sedation and often paralysis are necessary. The promise of HFOV as a lung-protective ventilation strategy remains attractive, but additional clinical trials are needed to determine whether this approach is superior to lung-protective ventilation with conventional mechanical ventilation.


Community nursing services for postdischarge chronically ill patients.
To examine community nursing services for patients with cardiovascular diseases, chronic respiratory diseases and other general medical conditions, making the transition from hospital to home. The original study design was a randomised controlled trial. This study is a secondary analysis of the hospital records documented by community nurses for the study-group patients. The sample consisted of 46 subjects, randomly drawn from the main study group of the study. The community nursing records were analysed using the Omaha System. Self-reported health status and readmission data were retrieved from the data base of the original study. The three groups of patients experienced problems across the four domains in the Omaha System. Community nursing interventions did not differ greatly by disease groups. The primary purpose of home visits was observation, followed by treatment and procedures and health teaching. The community nurses in the study spent more effort providing health teaching to the respiratory group than to their counterparts. The outcome measures are self-reported health status and hospital readmission rates. For self-reported health status, significant differences were observed in the respiratory and cardiovascular group before and after community nursing services. For hospital readmission rate, no significant difference was found. To improve the well being of chronically ill patients, a comprehensive home intervention programme, emphasising continuous needs of monitoring and case management, is fundamental to producing desired, measurable effects. This paper adds the understanding of home-care services provided by community nurses to chronically ill patients. The scope of nursing services emphasises the significance of a positive, patient-centred, caring and appropriate client-practitioner relationship to improve the self-reported health of patients.


Compilation and analysis of eukaryotic POL II promoter sequences.
A representative set of 168 eukaryotic POL II promoters has been compiled from the EMBL library and subjected to computer signal search analysis. Application of this technique to E. coli promoters as a control ensemble revealed the well known consensus sequences at -35 and -10 which indicates that the methods are adequate to approach problems of this kind. The results obtained from the eukaryotic promoter set can be summarized as follows: Common sequence features are confined to a region between -50 and +10 relative to the transcriptional initiation site. The only well conserved consensus sequence is TATAAA, centered at -28. A weak motif, CA followed preferentially by pyrimidines, surrounds the cap-site. Two pentanucleotides which have been shown by experiments to stimulate transcription of certain genes, GGGCG and CCAAT, are moderately over-represented in the upstream region (between -129 and -50). However, they occur at highly variable distances from the initiation site.


Effects of the plant growth-promoting bacterium Burkholderia phytofirmans PsJN throughout the life cycle of Arabidopsis thaliana.
Plant growth-promoting rhizobacteria (PGPR) induce positive effects in plants, such as increased growth or reduced stress susceptibility. The mechanisms behind PGPR/plant interaction are poorly understood, as most studies have described short-term responses on plants and only a few studies have analyzed plant molecular responses under PGPR colonization. Here, we studied the effects of the PGPR bacterial model Burkholderiaphytofirmans PsJN on the whole life cycle of Arabidopsis thaliana plants. We reported that at different plant developmental points, strain PsJN can be found in the rhizosphere and also colonizing their internal tissues. In early ontogeny, strain PsJN increased several growth parameters and accelerated growth rate of the plants. Also, an Arabidopsis transcriptome analysis revealed that 408 genes showed differential expression in PsJN-inoculated plants; some of these genes are involved in stress response and hormone pathways. Specifically, genes implicated in auxin and gibberellin pathways were induced. Quantitative transcriptional analyses of selected genes in different developmental stages revealed that the beginning of these changes could be evidenced early in development, especially among the down-regulated genes. The inoculation with heat-killed bacteria provoked a more severe transcriptional response in plants, but was not able to induce plant growth-promotion. Later in ontogeny, the growth rates of inoculated plants decreased with respect to the non-inoculated group and, interestingly, the inoculation accelerated the flowering time and the appearance of senescence signs in plants; these modifications correlate with the early up-regulation of flowering control genes. Then, we show that a single inoculation with a PGPR could affect the whole life cycle of a plant, accelerating its growth rate and shortening its vegetative period, both effects relevant for most crops. Thus, these findings provide novel and interesting aspects of these relevant biological interactions.


A randomised controlled feasibility trial and qualitative evaluation of an early years language development intervention: study protocol of the 'outcomes of Talking Together evaluation and results' (oTTer) project.
Problems with oral language skills in childhood have been linked with poor educational, employment, and mental health outcomes. In the UK, there is increasing concern about the oral language skills of children, particularly children from areas of social disadvantage. Research emphasises the importance of the home language environment as a fundamental bedrock for the development of oral language skills. It is vital, therefore, that support is available to help families in need to provide the optimal language environment for their child. Talking Together is a 6-week home visiting programme recently commissioned by Better Start Bradford to develop parents' knowledge of the importance of a good language environment and help to improve parent-child interactions. This study represents the initial steps in developing a definitive trial of the Talking Together programme. This study is a two-arm randomised controlled feasibility study in which families referred into the Talking Together programme and consent to participate in the trial will be randomly allocated to either an intervention group or a waiting control group. We will assess the recruitment and retention rates, the representativeness of our sample, the appropriateness of our measures, and the sample size needed for a definitive trial. We will also carry out a qualitative evaluation to explore the acceptability of trial procedures for families and service providers, fidelity of delivery, time and resources for training, and barriers and facilitators to engagement with the programme. Clear progression criteria will be used to assess suitability for a definitive trial. This feasibility study will inform the development of a definitive trial of this home-based visiting programme, which will add to the sparse evidence base on which practitioners can draw when supporting families in need. The lessons learnt from this feasibility study will also inform the wider evaluation work of the Better Start Bradford Innovation Hub. The trial is registered with the ISRCTN registry: study ID ISRCTN13251954. Date of registration: 21 February 2019 (the trial was retrospectively registered).


Using Facebook and participant information clips to recruit emergency nurses for research.
To examine the use of social networking sites in recruiting research participants. Workplace violence is an important issue for staff and patients. One workplace that reports the highest levels of violence is the emergency department. The ability to research issues such as workplace violence in real time is important in addressing them expeditiously, and social media can be used to advertise and recruit research subjects, implement studies and disseminate information. The experience of recruiting subjects through social networks, specifically Facebook, and the use of participant information clips (PICs) for advertising. A brief discussion of the history of advertising and communication using the internet is presented to provide an understanding of the trajectory of social media and implications for recruitment in general. The paper then focuses on the lead author's experience of recruiting subjects using Facebook, including its limitations and advantages, and her experience of using participant information clips. The low cost of advertising and recruiting participants this way, as well as the convenience provided to participants, resulted in almost half the study's total participants being obtained within 72 hours. Using Facebook to target a younger age range of nurses to participate in a study was successful and yielded a large number of completed responses in a short time period at little cost to the researcher. Recording the PIC was cheap, and posting it and a link to the site on pre-existing group pages was free, providing valuable viral marketing and snowball recruiting. Future researchers should not overlook using social network sites for recruitment if the demographics of the desired study population and subject matter permit it.


Local and sex-specific biases in crossover vs. noncrossover outcomes at meiotic recombination hot spots in mice.
Meiotic recombination initiated by programmed double-strand breaks (DSBs) yields two types of interhomolog recombination products, crossovers and noncrossovers, but what determines whether a DSB will yield a crossover or noncrossover is not understood. In this study, we analyzed the influence of sex and chromosomal location on mammalian recombination outcomes by constructing fine-scale recombination maps in both males and females at two mouse hot spots located in different regions of the same chromosome. These include the most comprehensive maps of recombination hot spots in oocytes to date. One hot spot, located centrally on chromosome 1, behaved similarly in male and female meiosis: Crossovers and noncrossovers formed at comparable levels and ratios in both sexes. In contrast, at a distal hot spot, crossovers were recovered only in males even though noncrossovers were obtained at similar frequencies in both sexes. These findings reveal an example of extreme sex-specific bias in recombination outcome. We further found that estimates of relative DSB levels are surprisingly poor predictors of relative crossover frequencies between hot spots in males. Our results demonstrate that the outcome of mammalian meiotic recombination can be biased, that this bias can vary depending on location and cellular context, and that DSB frequency is not the only determinant of crossover frequency.


The role of the cerebellum in reconstructing social action sequences: a pilot study.
Recent research has revealed that the cerebellum plays a critical role in social reasoning and in particular in understanding false beliefs and making trait attributions. One hypothesis is that the cerebellum is responsible for the understanding of sequences of motions and actions, which may be a prerequisite for social understanding. To investigate the role of action sequencing in mentalizing, we tested patients with generalized cerebellar degenerative lesions on tests of social understanding and compared their performance with matched healthy volunteers. The tests involved understanding violations of social norms making trait and causal attributions on the basis of short behavioral sentences and generating the correct chronological order of social actions depicted in cartoons (picture sequencing task). Cerebellar patients showed clear deficits only on the picture sequencing task when generating the correct order of cartoons depicting false belief stories and showed at or close to normal performance for mechanical stories and overlearned social scripts. In addition, they performed marginally worse on trait attributions inferred from verbal behavioral descriptions. We conclude that inferring the mental state of others through understanding the correct sequences of their actions requires the support of the cerebellum.


Complete genome sequence of Kocuria rhizophila BT304, isolated from the small intestine of castrated beef cattle.
Members of the species Kocuria rhizophila, belonging to the family Micrococcaceae in the phylum Actinobacteria, have been isolated from a wide variety of natural sources, such as soil, freshwater, fish gut, and clinical specimens. K. rhizophila is important from an industrial viewpoint, because the bacterium grows rapidly with high cell density and exhibits robustness at various growth conditions. However, the bacterium is an opportunistic pathogen involved in human infections. Here, we sequenced and analyzed the genome of the K. rhizophila strain BT304, isolated from the small intestine of adult castrated beef cattle. The genome of K. rhizophila BT304 consisted of a single circular chromosome of 2,763,150 bp with a GC content of 71.2%. The genome contained 2359 coding sequences, 51 tRNA genes, and 9 rRNA genes. Sequence annotations with the RAST server revealed many genes related to amino acid, carbohydrate, and protein metabolism. Moreover, the genome contained genes related to branched chain amino acid biosynthesis and degradation. Analysis of the OrthoANI values revealed that the genome has high similarity (> 97.8%) with other K. rhizophila strains, such as DC2201, FDAARGOS 302, and G2. Comparative genomic analysis further revealed that the antibiotic properties of K. rhizophila vary among the strains. The relatively small number of virulence-related genes and the great potential in production of host available nutrients suggest potential application of the BT304 strain as a probiotic in breeding beef cattle.


Food-derived bioactives as potential regulators of the IL-12/IL-23 pathway implicated in inflammatory bowel diseases.
The gene-specific modulation of inflammatory cytokines by food bioactives represents a possible approach to the nutritional or pharmaceutical prevention and treatment of inflammatory bowel disease (IBD). There is evidence for a key role of the interleukin-12beta1/23 receptor (IL-12 Rbeta1/23 R) pathway in IBD, and that reduction of the normal expression of the IL-23 R gene may provide a therapeutic target for this disease. The binding of interleukin-23 (IL-23) to its receptor IL-23 R regulates a newly defined effector T-cell subset, Th17 cells, characterised by the production of interleukin-17 (IL-17) and other cytokines, including tumour necrosis factor-alpha (TNF-alpha). In this study we developed an assay that measured IL-17 and TNF-alpha expression after incubation with specific dietary bioactives in the human T-cell Kit 225. It is anticipated that these changes will reflect differences in IL-23 R production, albeit indirectly. The cell line Kit 225 has similarities to Th17 cells, a subset of T cells producing IL-17 and TNF-alpha, and in initial experiments we demonstrated that the cells express both IL-23 receptor subunits, as well as IL-17 and TNF-alpha genes. Upon verification that stimulation of Kit 225 cells with 1ng/mL IL-23 significantly upregulated IL-17 and TNF-alpha gene expression, and IL-17 production, we supplemented cells with selected food bioactives, caffeic acid phenethyl ester (CAPE), epigallocatechin gallate (EGCG), docosahexaenoic acid (DHA), and linoleic acid (LA), and with phorbol myristate acetate (PMA) and sodium salicylate, used as pro-inflammatory and anti-inflammatory controls, respectively. In both unstimulated cells and after IL-23 stimulation, bioactives modulated the pro-inflammatory cytokines involved in IBD, underlining the possible role of foods in this disease. EGCG and DHA, which significantly inhibited both IL-17 and TNF-alpha expression, appeared particularly interesting.


No mutations of FGFR3 in normal urothelium in the vicinity of urothelial carcinoma of the bladder harbouring activating FGFR3 mutations in patients with bladder cancer.
Mutations in the fibroblast growth factor receptor 3 (FGFR3) gene causing constitutive oncogenic protein activation have been shown to be frequent in papillary noninvasive bladder tumours and are associated with a low risk of progression and a favourable outcome. FGFR3 alterations have also been found in benign urothelial papilloma and flat urothelial hyperplasia suggesting FGFR3 alterations as an early event in bladder tumorigenesis. To date there is no data available on FGFR3 mutations in normal urothelium from patients with bladder cancer. We therefore analysed 64 samples of histopathological unsuspicious normal urothelium from 38 patients with FGFR3 mutated bladder tumours and 15 samples of urothelium from patients (n = 15) without any urothelial malignancy as a control group. Urothelial cells were microdissected from formalin-fixed, paraffin-embedded material. After DNA isolation whole genome amplification was done by I-PEP-PCR. FGFR3 mutations were detected using SNaPshot analysis. All samples could successfully be investigated. FGFR3 analyses did not reveal any mutation in the urothelium from neither the control group nor the bladder cancer group. All urothelial samples showed a wildtype sequence for FGFR3. These data suggest that mutations in the FGFR3 gene are not the earliest genetic alterations in bladder carcinogenesis and are associated with a hyperproliferative (hyperplastic) phenotype in the urothelium. Chromosomal alterations like deletions on chromosome 9q or aberrant promoter hypermethylation could play more important roles in early urothelial transformation than mutational FGFR3 activation.


The role of propagating modes in silver nanowire arrays for transparent electrodes.
Silver nanowires have been shown to demonstrate enhanced transmission and promising potential for next-generation transparent electrodes. In this paper, we systematically investigated the electrical and optical properties of 1D and 2D silver nanowire arrays as a function of diameter and pitch and compared their performance to that of silver thin films. Silver nanowires were found to exhibit enhanced transmission over thin films due to propagating resonance modes between nanowires. We evaluated the angular dependence and dispersion relation of these propagating modes and demonstrate that larger nanowire diameters and pitches are favored for achieving higher solar transmission at a particular sheet resistance. Silver nanowires may achieve achieve solar transmission > 90% with sheet resistances of a few Ω/sq and figure of merit σdc/σop > 1000.


The "continuum" of a unified theory of diseases.
This essay's theme was inspired by a question asked by a child: 'Why do I get ill?' The question is very interesting, but has no easy answer. This paper discusses a few possible answers to this difficult question. Through the life of a person, from birth to death, there is a "continuum" in the pathological conditions a person may experience. The body, as a whole, suffers deeply any time there is an acute or a chronic condition that is either maltreated or neglected. Chronic and acute diseases in the medical history of a person constitute a rigidly related chain of immune responses in the form of a real "continuum" that at every point in time indicates the end result of this continuum. The idea promoted here is that suppression of diseases, through excess of chemical drugs or other means, many times overwhelms the body's natural defenses and forces the immune system to compromise and start a deeper line of defense, which then constitute the beginning of a new chronic condition. Thus, the original inflammation of an acute condition may continue as a sub-acute inflammatory process on a deeper level. Acute inflammatory conditions must therefore be treated very carefully from their beginnings in childhood in order not to force the immune system to compromise. It is also suggested here that all chronic degenerative conditions have a sub-acute inflammatory character, and that "inflammation" constitutes the main common parameter of all diseases.


Biogenesis of Hydrogen Sulfide and Thioethers by Cystathionine Beta-Synthase.
The transsulfuration pathway enzymes cystathionine beta-synthase (CBS) and cystathionine gamma-lyase are thought to be the major source of hydrogen sulfide (H2S). In this study, we assessed the role of CBS in H2S biogenesis. We show that despite discouraging enzyme kinetics of alternative H2S-producing reactions utilizing cysteine compared with the canonical condensation of serine and homocysteine, our simulations of substrate competitions at biologically relevant conditions suggest that cysteine is able to partially compete with serine on CBS, thus leading to generation of appreciable amounts of H2S. The leading H2S-producing reaction is condensation of cysteine with homocysteine, while cysteine desulfuration plays a dominant role when cysteine is more abundant than serine and homocysteine is limited. We found that the serine-to-cysteine ratio is the main determinant of CBS H2S productivity. Abundance of cysteine over serine, for example, in plasma, allowed for up to 43% of CBS activity being responsible for H2S production, while excess of serine typical for intracellular levels effectively limited such activity to less than 1.5%. CBS also produced lanthionine from serine and cysteine and a third of lanthionine coming from condensation of two cysteines contributed to the H2S pool. Our study characterizes the H2S-producing potential of CBS under biologically relevant conditions and highlights the serine-to-cysteine ratio as the main determinant of H2S production by CBS in vivo. Our data clarify the function of CBS in H2S biogenesis and the role of thioethers as surrogate H2S markers. Antioxid. Redox Signal. 28, 311-323.


Clonal change of blaSIM-1-carrying Acinetobacter spp. from 2003 to 2008 in the hospital where it was initially discovered.
SIM-1 metallo-β-lactamase was first discovered from carbapenem-resistant Acinetobacter spp. isolated in a Korean University Hospital in 2003, and was recently reported to have been discovered in A. baylyi isolated in nearby China. The aims of this study were to reveal clonal changes in bla(SIM-1)-harboring Acinetobacter isolates collected from 2003 to 2008 in the same Korean hospital, where they were first discovered to gain further insight into the relation between bla(SIM-1)-harboring plasmids and Acinetobacter spp. Among 1,761 nonduplicated imipenem-resistant Acinetobacter spp. isolates, 29 isolates were identified as bla(SIM-1) carriers. They were categorized into nine types according to pulsed-field gel electrophoresis findings. While most bla(SIM-1)-carrying isolates from 2003 to 2005 belonged to A. pittii, those from 2006 to 2007 were mostly isolates of A. nosocomialis. Most of the bla(SIM-1) genes were carried on ca. 280-kb plasmids and were only discovered in non-baumannii Acinetobacter spp. Integrons carrying the bla(SIM-1) gene were identical in structure in all species. These findings suggest that the plasmids were transferable, but not promiscuous. Further surveillance should be continued to detect and control further spread of the bla(SIM-1) gene, as the appearance of the bla(SIM-1) gene in different Acinetobacter spp. in different countries has already begun.


The C terminus of the metabotropic glutamate receptor subtypes 2 and 7 specifies the receptor signaling pathways.
There is accumulating evidence that the specificity of the transduction cascades activated by G protein-coupled receptors cannot solely depend on the nature of the coupled G protein. To identify additional structural determinants, we studied two metabotropic glutamate (mGlu) receptors, the mGlu2 and mGlu7 receptors, that are both coupled to G(o) proteins but are known to affect different effectors in neurons. Thus, the mGlu2 receptor selectively blocks N- and L-type Ca(2+) channels via a protein kinase C-independent pathway, whereas the mGlu7 receptor selectively blocks P/Q-type Ca(2+) channels via a protein kinase C-dependent pathway, and both effects are pertussis toxin-sensitive. We examined the role of the C-terminal domain of these receptors in this coupling. Chimeras were constructed by exchanging the C terminus of these receptors and transfected into neurons. Different chimeric receptors bearing the C terminus of mGlu7 receptor blocked selectively P/Q-type Ca(2+) channels, whereas chimeras bearing the C terminus of mGlu2 receptor selectively blocked N- and L-type Ca(2+) channels. These results show that the C terminus of mGlu2 and mGlu7 receptors is a key structural determinant that allows these receptors to select a specific signaling pathway in neurons.


Pattern-sensitive epilepsy: electroclinical characteristics, natural history, and delineation of the epileptic syndrome.
To elucidate the electroclinical features and long-term outcome of patients with pattern-sensitive epilepsy. We reviewed the clinical and electroencephalographic (EEG) findings of 73 (43 female and 30 male) patients in whom pattern-sensitive epilepsy was diagnosed at Mayo Clinic (Rochester, Minnesota, U.S.A.) from 1950 through 1999. We contacted patients and their relatives by letter or telephone to obtain the latest seizure and quality-of-life outcomes. The median age at onset of seizures was 12.8 years (range, 0.6-32.9 years). Most patients had absence, myoclonic, or generalized tonic-clonic seizures. Interictal epileptiform discharges in the EEG were detected in 61 (83.6%) patients and were generalized in 54 (74%). Paroxysmal epileptiform discharges in the EEG elicited with standard patterns were all generalized in two thirds of patients but were restricted to the posterior head region in one-third. Eight (11%) patients did not exhibit photosensitivity. Television was the most common precipitant [30 patients (41%)]. Twenty-nine patients gave a clear history of one or more seizures precipitated while viewing environmental patterns such as window screens, garments, tablecloths, and ceiling tiles; the rest of the patients admitted that they preferred to avoid looking at patterned objects because these objects made them uncomfortable. The electroclinical features suggested juvenile myoclonic epilepsy in 14 patients, progressive myoclonus epilepsy in three, progressive familial cerebellar ataxia with myoclonus in two, and severe myoclonic epilepsy of infancy in one. During a median follow-up period of 15.7 years, 25 (45.5%) of 55 patients who were followed up for > or =5 years achieved complete seizure remission. The median age at remission was 24.4 years. The absence of progressive neurologic disease was correlated significantly with remission; a family history of seizures showed a trend in favor of remission. More than two thirds of the patients did not consider the seizures an impediment to their family life or to educational and occupational achievements. Although pattern sensitivity as a trait occurs in various epileptic syndromes, pattern-sensitive epilepsy is a readily distinguishable subtype of the visually provoked reflex epilepsies. In our opinion, the location and extent of the excitable region or regions within the visual cortex concerned with different attributes of visual function dictate susceptibility to a specific trigger (intermittent light, pattern, or color) or closely related multiple triggers and the resultant electroclinical phenomenon.


Geodemographic insights on the COVID-19 pandemic in the State of Wisconsin and the role of risky facilities.
The COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to impact the United States. While age and comorbid health conditions remain primary concerns in the community-based transmission of the virus, empirical evidence continues to suggest that substantial variability exists in the geographic and geodemographic distribution of COVID-19 infection rates. The purpose of this paper is to provide an alternative, spatiotemporal perspective on the pandemic using the state of Wisconsin as a case study. Specifically, in this paper, we explore the geographic nuances of COVID-19 and its spread in Wisconsin using a suite of spatial statistical approaches. We link detected hot spots of COVID-19 to local geodemographic profiles and the presence of high-risk facilities, including federal and state correctional facilities. The results suggest that the virus disproportionately impacts several communities and geodemographic groups and that proximity to risky facilities correlates to increased community infection rates.


Effect of nimodipine on intracellular brain pH, cortical blood flow, and EEG in experimental focal cerebral ischemia.
Intracellular brain pH, cortical blood flow, and electroencephalograms (EEG's) were recorded in severely and moderately ischemic regions in 10 control and 10 nimodipine-treated rabbits prior to and following major branch occlusion of the middle cerebral artery (MCA). Preocclusion cortical blood flow was 51 ml/100 gm/min and intracellular brain pH was 7.01 in both the control and the treated animals. After MCA occlusion, the severely ischemic regions in the control group showed initial and 4-hour postocclusion flows of 12.7 and 5.2 ml/100 gm/min with a brain pH of 6.64 and 6.08, respectively. In animals given nimodipine after MCA occlusion, blood flow increased from 10.5 to 18.8 ml/100 gm/min, with an associated elevation in intracellular brain pH from 6.57 to 6.91. Comparable findings were observed in areas of moderate ischemia. Improvements in cortical blood flow, intracellular brain pH, and EEG attenuations produced by nimodipine were all statistically significant. Inspection of the cortex revealed reversal of cortical pallor and small-vessel spasm following treatment with nimodipine. It is hypothesized that nimodipine exerts its effects through reversal of ischemia-induced secondary vasoconstriction, and that this drug may be an important adjunctive treatment for patients with focal cerebral ischemia.


Comparison of regional brain deficit patterns in common psychiatric and neurological disorders as revealed by big data.
Neurological and psychiatric illnesses are associated with regional brain deficit patterns that bear unique signatures and capture illness-specific characteristics. The Regional Vulnerability Index (RVI) was developed toquantify brain similarity by comparing individual white matter microstructure, cortical gray matter thickness and subcortical gray matter structural volume measures with neuroanatomical deficit patterns derived from large-scale meta-analytic studies. We tested the specificity of the RVI approach for major depressive disorder (MDD) and Alzheimer's disease (AD) in a large epidemiological sample of UK Biobank (UKBB) participants (N = 19,393; 9138 M/10,255F; age = 64.8 ± 7.4 years). Compared to controls free of neuropsychiatric disorders, participants with MDD (N = 2,248; 805 M/1443F; age = 63.4 ± 7.4) had significantly higher RVI-MDD values (t = 5.6, p = 1·10-8), but showed no detectable difference in RVI-AD (t = 2.0, p = 0.10). Subjects with dementia (N = 7; 4 M/3F; age = 68.6 ± 8.6 years) showed significant elevation in RVI-AD (t = 4.2, p = 3·10-5) but not RVI-MDD (t = 2.1, p = 0.10) compared to controls. Even within affective illnesses, participants with bipolar disorder (N = 54) and anxiety disorder (N = 773) showed no significant elevation in whole-brain RVI-MDD. Participants with Parkinson's disease (N = 37) showed elevation in RVI-AD (t = 2.4, p = 0.01) while subjects with stroke (N = 247) showed no such elevation (t = 1.1, p = 0.3). In summary, we demonstrated elevation in RVI-MDD and RVI-AD measures in the respective illnesses with strong replicability that is relatively specific to the respective diagnoses. These neuroanatomic deviation patterns offer a useful biomarker for population-wide assessments of similarity to neuropsychiatric illnesses.


Axial elongation associated with biomechanical factors during near work.
To investigate the changes occurring in the axial length, choroidal thickness, and anterior biometrics of the eye during a 10-minute near task performed in downward gaze. Twenty young adult subjects (10 emmetropes and 10 myopes) participated in this study. To measure ocular biometrics in downward gaze, an optical biometer was inclined on a custom-built height- and tilt-adjustable table. Baseline measures were collected after each subject performed a distance primary gaze control task for 10 minutes to provide washout period for previous visual tasks before each of three different accommodation/gaze conditions. These other three conditions included a near task (2.5 diopters [D]) in primary gaze and a near (2.5 D) and a far (0 D) accommodative task in downward gaze (25 degrees), all for 10 minutes' duration. Immediately after and then 5 and 10 minutes from the commencement of each trial, measurements of ocular biometrics (e.g., anterior biometrics, axial length, choroidal thickness, and retinal thickness) were obtained. Axial length increased with accommodation and was significantly greater for downward gaze with accommodation (mean ± SD change, 23 ± 13 μm at 10 minutes) compared with primary gaze with accommodation (8 ± 15 μm at 10 minutes) (p < 0.05). A small amount of choroidal thinning was also found during accommodation that was statistically significant in downward gaze (13 ± 14 μm at 10 minutes; p < 0.05). Accommodation in downward gaze also caused greater changes in anterior chamber depth and lens thickness compared with accommodation in primary gaze. Axial length, choroidal thickness, and anterior eye biometrics change significantly during accommodation in downward gaze as a function of time. These changes seem to be caused by the combined influence of biomechanical factors (i.e., extraocular muscle forces, ciliary muscle contraction) associated with near tasks in downward gaze.


Higher risk of revision for infection using systemic clindamycin prophylaxis than with cloxacillin.
Background and purpose - Clindamycin has not been compared with other antibiotics for prophylaxis in arthroplasty. Since 2009, the Swedish Knee Arthroplasty Register (SKAR) has been collecting information on the prophylactic antibiotic regime used at every individual operation. In Sweden, when there is allergy to penicillin, clindamycin has been the recommended alternative. We examined whether there were differences in the rate of revision due to infection depending on which antibiotic was used as systemic prophylaxis. Patients and methods - Patients who had a total knee arthroplasty (TKA) performed due to osteoarthritis (OA) during the years 2009-2015 were included in the study. Information on which antibiotic was used was available for 80,018 operations (55,530 patients). Survival statistics were used to calculate the rate of revision due to infection until the end of 2015, comparing the group of patients who received cloxacillin with those who received clindamycin as systemic prophylaxis. Results - Cloxacillin was used in 90% of the cases, clindamycin in 7%, and cephalosporins in 2%. The risk of being revised due to infection was higher when clindamycin was used than when cloxacillin was used (RR =1.5, 95% CI: 1.2-2.0; p = 0.001). There was no significant difference in the revision rate for other causes (p = 0.2). Interpretation - We advise that patients reporting allergic reaction to penicillin should have their allergic history explored. In the absence of a clear history of type-I allergic reaction (e.g. urticaria, anaphylaxis, or bronchospasm), we suggest the use of a third-generation cephalosporin instead of clindamycin as perioperative prophylaxis when undergoing a TKR. No recommendation can be given regarding patients with type-1 allergy.


Influence of air flow rate and backwashing on the hydraulic behaviour of a submerged filter.
The aim of this study was to evaluate backwashing effects on the apparent porosity of the filter media and on the hydraulic behaviour of a pilot scale submerged filter, prior to biofilm colonization, under different hydraulic retention times, and different air flow rates. Tracer curves were analysed with two mathematical models for ideal and non-ideal flow (axial dispersion and Wolf and Resnick models). The filter media was lava stones sieved to 4.5 mm. Backwashing causes attrition of media particles, decreasing the void volume of the filter media and, consequently, the tracer flow is more uniform. The eroded media presented lower dead volumes (79% for the filter with aeration and 8% for the filter without aeration) compared with the new media (83% for the filter with aeration and 22% for the filter without aeration). The flow patterns of eroded and new media were different because the more regular shape of the particles decreases the void volume of the filter media. The dead volume is attributed, in the case of the filter with aeration, to the turbulence caused by the air bubbles that generate preferential channelling of the bulk liquid along the filter media, creating large zones of stagnant liquid and, for the filter without aeration, to the channels formed due to the irregular shaped media.


Structural and functional connectivity in healthy aging: Associations for cognition and motor behavior.
Age-related behavioral declines may be the result of deterioration of white matter tracts, affecting brain structural (SC) and functional connectivity (FC) during resting state. To date, it is not clear if the combination of SC and FC data could better predict cognitive/motor performance than each measure separately. We probed these relationships in the cingulum bundle, a major white matter pathway of the default mode network. We aimed to attain deeper knowledge about: (a) the relationship between age and the cingulum's SC and FC strength, (b) the association between SC and FC, and particularly (c) how the cingulum's SC and FC are related to cognitive/motor performance separately and combined. We examined these associations in a healthy and well-educated sample of 165 older participants (aged 64-85). SC and FC were acquired using probabilistic tractography to derive measures to capture white matter integrity within the cingulum bundle (fractional anisotropy, mean, axial and radial diffusivity) and a seed-based resting-state functional MRI correlation approach, respectively. Participants performed cognitive tests measuring processing speed, memory and executive functions, and motor tests measuring motor speed and grip force. Our data revealed that only SC but not resting state FC was significantly associated with age. Further, the cingulum's SC and FC showed no relation. Different relationships between cognitive/motor performance and SC/FC separately were found, but no additive effect of the combined analysis of cingulum's SC and FC for predicting cognitive/motor performance was apparent.


Improved haplotype-based detection of ongoing selective sweeps towards an application in Arabidopsis thaliana.
The increasing amount of genome information allows us to address various questions regarding the molecular evolution and population genetics of different species. Such genome-wide data sets including thousands of individuals genotyped at hundreds of thousands of markers require time-efficient and powerful analysis methods. Demography and sampling introduce a bias into present population genetic tests of natural selection, which may confound results. Thus, a modification of test statistics is necessary to introduce time-efficient and unbiased analysis methods. We present an improved haplotype-based test of selective sweeps in samples of unequally related individuals. For this purpose, we modified existing tests by weighting the contribution of each individual based on its uniqueness in the entire sample. In contrast to previous tests, this modified test is feasible even for large genome-wide data sets of multiple individuals. We utilize coalescent simulations to estimate the sensitivity of such haplotype-based test statistics to complex demographic scenarios, such as population structure, population growth and bottlenecks. The analysis of empirical data from humans reveals different results compared to previous tests. Additionally, we show that our statistic is applicable to empirical data from Arabidopsis thaliana. Overall, the modified test leads to a slight but significant increase of power to detect selective sweeps among all demographic scenarios. The concept of this modification might be applied to other statistics in population genetics to reduce the intrinsic bias of demography and sampling. Additionally, the combination of different test statistics may further improve the performance of tests for natural selection.


Evaluation of oxygen transfer rates in stirred-tank bioreactors for clinical manufacturing.
Several methods are available for determining the volumetric oxygen transfer coefficient in bioreactors, though their application in industrial bioprocess has been limited. To be practically useful, mass transfer measurements made in nonfermenting systems must be consistent with observed microbial respiration rates. This report details a procedure for quantifying the relationship between agitation frequency and oxygen transfer rate that was applied in stirred-tank bioreactors used for clinical biologics manufacturing. The intrinsic delay in dissolved oxygen (DO) measurement was evaluated by shifting the bioreactor pressure and fitting a first-order mathematical model to the DO response. The dynamic method was coupled with the DO lag results to determine the oxygen transfer rate in Water for Injection (WFI) and a complete culture medium. A range of agitation frequencies was investigated at a fixed air sparge flow rate, replicating operating conditions used in Pichia pastoris fermentation. Oxygen transfer rates determined by this method were in excellent agreement with off-gas calculations from cultivation of the organism (P = 0.1). Fermentation of Escherichia coli at different operating parameters also produced respiration rates that agreed with the corresponding dynamic method results in WFI (P = 0.02). The consistency of the dynamic method results with the off-gas data suggests that compensation for the delay in DO measurement can be combined with dynamic gassing to provide a practical, viable model of bioreactor oxygen transfer under conditions of microbial fermentation.


Video-based motion-resilient reconstruction of three-dimensional position for functional near-infrared spectroscopy and electroencephalography head mounted probes.
Significance: We propose a video-based, motion-resilient, and fast method for estimating the position of optodes on the scalp. Aim: Measuring the exact placement of probes (e.g., electrodes and optodes) on a participant's head is a notoriously difficult step in acquiring neuroimaging data from methods that rely on scalp recordings (e.g., electroencephalography and functional near-infrared spectroscopy) and is particularly difficult for any clinical or developmental population. Existing methods of head measurements require the participant to remain still for a lengthy period of time, are laborious, and require extensive training. Therefore, a fast and motion-resilient method is required for estimating the scalp location of probes. Approach: We propose an innovative video-based method for estimating the probes' positions relative to the participant's head, which is fast, motion-resilient, and automatic. Our method builds on capitalizing the advantages and understanding the limitations of cutting-edge computer vision and machine learning tools. We validate our method on 10 adult subjects and provide proof of feasibility with infant subjects. Results: We show that our method is both reliable and valid compared to existing state-of-the-art methods by estimating probe positions in a single measurement and by tracking their translation and consistency across sessions. Finally, we show that our automatic method is able to estimate the position of probes on an infant head without lengthy offline procedures, a task that has been considered challenging until now. Conclusions: Our proposed method allows, for the first time, the use of automated spatial co-registration methods on developmental and clinical populations, where lengthy, motion-sensitive measurement methods routinely fail.


A fear-inducing odor alters PER2 and c-Fos expression in brain regions involved in fear memory.
Evidence demonstrates that rodents learn to associate a foot shock with time of day, indicating the formation of a fear related time-stamp memory, even in the absence of a functioning SCN. In addition, mice acquire and retain fear memory better during the early day compared to the early night. This type of memory may be regulated by circadian pacemakers outside of the SCN. As a first step in testing the hypothesis that clock genes are involved in the formation of a time-stamp fear memory, we exposed one group of mice to fox feces derived odor (TMT) at ZT 0 and one group at ZT 12 for 4 successive days. A separate group with no exposure to TMT was also included as a control. Animals were sacrificed one day after the last exposure to TMT, and PER2 and c-Fos protein were quantified in the SCN, amygdala, hippocampus, and piriform cortex. Exposure to TMT had a strong effect at ZT 0, decreasing PER2 expression at this time point in most regions except the SCN, and reversing the normal rhythm of PER2 expression in the amygdala and piriform cortex. These changes were accompanied by increased c-Fos expression at ZT0. In contrast, exposure to TMT at ZT 12 abolished the rhythm of PER2 expression in the amygdala. In addition, increased c-Fos expression at ZT 12 was only detected in the central nucleus of the amygdala in the TMT12 group. TMT exposure at either time point did not affect PER2 or c-Fos in the SCN, indicating that under a light-dark cycle, the SCN rhythm is stable in the presence of repeated exposure to a fear-inducing stimulus. Taken together, these results indicate that entrainment to a fear-inducing stimulus leads to changes in PER2 and c-Fos expression that are detected 24 hours following the last exposure to TMT, indicating entrainment of endogenous oscillators in these regions. The observed effects on PER2 expression and c-Fos were stronger during the early day than during the early night, possibly to prepare appropriate systems at ZT 0 to respond to a fear-inducing stimulus.


Recurrent neural networks in computer-based clinical decision support for laryngopathies: an experimental study.
The main goal of this paper is to give the basis for creating a computer-based clinical decision support (CDS) system for laryngopathies. One of approaches which can be used in the proposed CDS is based on the speech signal analysis using recurrent neural networks (RNNs). RNNs can be used for pattern recognition in time series data due to their ability of memorizing some information from the past. The Elman networks (ENs) are a classical representative of RNNs. To improve learning ability of ENs, we may modify and combine them with another kind of RNNs, namely, with the Jordan networks. The modified Elman-Jordan networks (EJNs) manifest a faster and more exact achievement of the target pattern. Validation experiments were carried out on speech signals of patients from the control group and with two kinds of laryngopathies.


Involvement of CYP 2C9 in mediating the proinflammatory effects of linoleic acid in vascular endothelial cells.
Polyunsaturated fatty acids such as linoleic acid are well known dietary lipids that may be atherogenic by activating vascular endothelial cells. In the liver, fatty acids can be metabolized by cytochrome P450 (CYP) enzymes, but little is known about the role of these enzymes in the vascular endothelium. CYP 2C9 is involved in linoleic acid epoxygenation, and the major product of this reaction is leukotoxin (LTX). We investigated the role of CYP-mediated mechanisms of linoleic acid metabolism in endothelial cell activation by examining the effects of linoleic acid or its oxidized metabolites such as LTX and leukotoxin diol (LTD). The effect of linoleic acid on CYP 2C9 gene expression was studied by RT-PCR. Oxidative stress was monitored by measuring DCF fluorescence and intracellular glutathione levels, and electrophoretic mobility shift assay was carried out to study the activation of oxidative stress sensitive transcription factors. Analysis of oxidized lipids was carried out by liquid chromatography/mass spectrometry. Linoleic acid treatment for six hours increased the expression of CYP 2C9 in endothelial cells. Linoleic acid-mediated increase in oxidative stress and activation of AP-1 were blocked by sulfaphenazole, a specific inhibitor of CYP 2C9. The linoleic acid metabolites LTX and LTD increased oxidative stress and activation of transcription factors only at high concentrations. Our data show that CYP 2C9 plays a key role in linoleic acid-induced oxidative stress and subsequent proinflammatory events in vascular endothelial cells by possibly causing superoxide generation through uncoupling processes.


Heterogeneous nuclear ribonucleoprotein L interacts with the 3' border of the internal ribosomal entry site of hepatitis C virus.
Translation initiation of hepatitis C virus (HCV) RNA occurs by internal entry of a ribosome into the 5' nontranslated region in a cap-independent manner. The HCV RNA sequence from about nucleotide 40 up to the N terminus of the coding sequence of the core protein is required for efficient internal initiation of translation, though the precise border of the HCV internal ribosomal entry site (IRES) has yet to be determined. Several cellular proteins have been proposed to direct HCV IRES-dependent translation by binding to the HCV IRES. Here we report on a novel cellular protein that specifically interacts with the 3' border of the HCV IRES in the core-coding sequence. This protein with an apparent molecular mass of 68 kDa turned out to be heterogeneous nuclear ribonucleoprotein L (hnRNP L). The binding of hnRNP L to the HCV IRES correlates with the translational efficiencies of corresponding mRNAs. This finding suggests that hnRNP L may play an important role in the translation of HCV mRNA through the IRES element.


Beyond dose escalation: clinical options for relapse or resistance in chronic myelogenous leukemia.
The development of imatinib has changed the management of chronic myeloid leukemia (CML), producing high response rates in most patients. However, most individuals treated with imatinib, 400 mg, have residual molecular disease, and both intrinsic and acquired resistance can occur. The newer tyrosine kinase inhibitors, dasatinib and nilotinib, are effective in patients with imatinib-resistant CML, In patients with relapse or resistance, current guidelines recommend escalating the dose of imatinib or switching to new tyrosine kinase inhibitors. Dasatinib has been investigated in patients who were resistant or intolerant to imatinib. Switching to dasatinib, 70 mg, twice daily has been shown to be more effective than high-dose imatinib. Another study found that dasatinib, 100 mg, once daily was just as effective as the twice-daily regimen but was better tolerated. Nilotinib is also effective in most patients with resistance or intolerance to imatinib and is associated with minimal toxicity. Other inhibitors, such as bosutinib and INNO-406, are being developed with favorable early results. New drugs are still needed, particularly for individuals who are resistant to tyrosine kinase inhibitors or those with the T3151 mutation. Emerging CML therapies, some of which have different mechanisms of action from those of tyrosine kinase inhibitors, have shown promising results and could offer an alternative to these patients as monotherapy or in combination.


Immunology taught by human genetics.
Human genetic studies are rarely conducted for immunological purposes. Instead, they are typically driven by medical and evolutionary goals, such as understanding the predisposition or resistance to infectious or inflammatory diseases, the pathogenesis of such diseases, and human evolution in the context of the long-standing relationships between humans and their commensal and environmental microbes. However, the dissection of these experiments of Nature has also led to major immunological advances. In this review, we draw on some of the immunological lessons learned in the three branches of human molecular genetics most relevant to immunology: clinical genetics, epidemiological genetics, and evolutionary genetics. We argue that human genetics has become a new frontier not only for timely studies of specific features of human immunity, but also for defining general principles of immunity. These studies teach us about immunity as it occurs under "natural" conditions, through the transition from the almost complete wilderness that existed worldwide until about a century ago to the current unevenly distributed medically shaped environment. Hygiene, vaccines, antibiotics, and surgery have considerably decreased the burden of infection, but these interventions have been available only recently, so have yet to have a major impact on patterns of genomic diversity, making it possible to carry out unbiased evolutionary studies at the population level. Clinical genetic studies of childhood phenotypes have not been blurred by modern medicine either. Instead, medical advances have actually facilitated such studies, by making it possible for children with life-threatening infections to survive. In addition, the prevention and treatment of infectious diseases have increased life expectancy at birth from ∼20 yr to ∼80 yr, providing unique opportunities to study the genetic basis of immunological phenomena against which there is no natural counterselection, such as reactivation and secondary infectious diseases and breakdown of self-tolerance manifesting as autoimmunity, in populations of adult and aging patients. Recently developed deep sequencing and stem cell technologies are of unprecedented power, and their application to human genetics is opening up exciting and timely possibilities for young immunologists seeking uncharted waters to explore.


Regulatory T cells prevent Th2 immune responses and pulmonary eosinophilia during respiratory syncytial virus infection in mice.
During viral infection, inflammation and recovery are tightly controlled by competing proinflammatory and regulatory immune pathways. Respiratory syncytial virus (RSV) is the leading global cause of infantile bronchiolitis, which is associated with recurrent wheeze and asthma diagnosis in later life. Th2-driven disease has been well described under some conditions for RSV-infected mice. In the present studies, we used the Foxp3(DTR) mice (which allow specific conditional depletion of Foxp3(+) T cells) to investigate the functional effects of regulatory T cells (Tregs) during A2-strain RSV infection. Infected Treg-depleted mice lost significantly more weight than wild-type mice, indicating enhanced disease. This enhancement was characterized by increased cellularity in the bronchoalveolar lavage (BAL) fluid and notable lung eosinophilia not seen in control mice. This was accompanied by abundant CD4(+) and CD8(+) T cells exhibiting an activated phenotype and induction of interleukin 13 (IL-13)- and GATA3-expressing Th2-type CD4(+) T cells that remained present in the airways even 14 days after infection. Therefore, Treg cells perform vital anti-inflammatory functions during RSV infection, suppressing pathogenic T cell responses and inhibiting lung eosinophilia. These findings provide additional evidence that dysregulation of normal immune responses to viral infection may contribute to severe RSV disease.


Locomotor adaptation to resistance during treadmill training transfers to overground walking in human SCI.
Treadmill training has been used as a promising technique to improve overground walking in patients with spinal cord injury (SCI). Previous findings showed that a gait pattern may adapt to a force perturbation during treadmill training and show aftereffects following removal of the force perturbation. We hypothesized that aftereffects would transfer to overground walking to a greater extent when the force perturbation was resisting rather than assisting leg swing during treadmill training. Ten subjects with incomplete SCI were recruited into this study for two treadmill training sessions: one using swing resistance and the other using swing assistance during treadmill stepping. A controlled resistance/assistance was provided to the subjects' right knee using a customized cable-driven robot. The subjects' spatial and temporal parameters were recorded during the training. The same parameters during overground walking were also recorded before and after the training session using an instrumented walkway. Results indicated that stride length during treadmill stepping increased following the release of resistance load and the aftereffect transferred to overground walking. In contrast, stride length during treadmill stepping decreased following the release of assistance load, but the aftereffect did not transfer to overground walking. Providing swing resistance during treadmill training could enhance the active involvement of the subjects in the gait motor task, thereby aiding in the transfer to overground walking. Such a paradigm may be useful as an adjunct approach to improve the locomotor function in patients with incomplete SCI.


Recurrent neural networks with backtrack-points and negative reinforcement applied to cost-based abduction.
Abduction is the process of proceeding from data describing a set of observations or events, to a set of hypotheses which best explains or accounts for the data. Cost-based abduction (CBA) is an AI formalism in which evidence to be explained is treated as a goal to be proven, proofs have costs based on how much needs to be assumed to complete the proof, and the set of assumptions needed to complete the least-cost proof are taken as the best explanation for the given evidence. In this paper, we present two techniques for improving the performance of high order recurrent networks (HORN) applied to cost-based abduction. In the backtrack-points technique, we use heuristics to recognize early that the network trajectory is moving in the wrong direction; we then restore the network state to a previously stored point, and apply heuristic perturbations to nudge the network trajectory in a different direction. In the negative reinforcement technique, we add hyperedges to the network to reduce the attractiveness of local minima. We apply these techniques to a suite of six large CBA instances, systematically generated to be difficult.


Synthesis of sub-micrometer biphasic Au-AuGa2/liquid metal frameworks.
A novel biphasic sub-micrometer Au-AuGa2/liquid metal framework, consisting of solid nanoparticles encapsulating liquid metal (LM) droplets, is introduced. By utilizing oxide-free galvanic replacement of a Ga-alloy LM with alkaline KAuBr4, the controllable process of gold-based encapsulation of individual sub-micrometer LM droplets capped with polyvinylpyrrolidone, lysozyme or sodium alginate is demonstrated. The morphology, structure and composition of the encapsulated droplets are characterized in-depth via scanning electron microscopy, energy-dispersive X-ray spectroscopy, and X-ray diffraction. For the first time, the existence of two kinetic regimes leading to the controllable synthesis of either intermetallic AuGa2 or Au nanoparticles to encapsulate LM droplets is highlighted via the study of the evolution of the pH of the reaction medium. The as-formed biphasic sub-micrometer Au-LM frameworks demonstrate moderate stability in an aqueous environment and formidable structural and chemical stability in ambient air at room temperature. The encapsulation process can be adapted to the micro-patterning of LM droplet surfaces to produce conductive Au-encapsulated LM droplets then sintered to recover LM lines, while the sub-micrometer Au-LM droplets can be reintegrated onto larger LM surfaces and volumes to create multi-metallic and multi-scale biphasic structures with controllable electrical properties. The adaptability of the encapsulation process to the macro-, micro- and nanoscale of liquid metal features with a wide range of surfactants could in principle allow its use in various systems and applications.


Skin Globotriaosylceramide 3 Load Is Increased in Men with Advanced Fabry Disease.
The X-chromosomally linked life-limiting Fabry disease (FD) is associated with deposits of the sphingolipid globotriaosylceramide 3 (Gb3) in various tissues. Skin is easily accessible and may be used as an additional diagnostic and follow-up medium. Our aims were to visualize skin Gb3 deposits in FD patients applying immunofluorescence and to determine if cutaneous Gb3 load correlates with disease severity. At our Fabry Center for Interdisciplinary Therapy we enrolled 84 patients with FD and 27 healthy controls. All subjects underwent 5-mm skin punch biopsy at the lateral lower leg and the back. Skin samples were processed for immunohistochemistry using antibodies against CD77 (i.e. Gb3). Cutaneous Gb3 deposition was quantified in a blinded manner and correlated to clinical data. We found that Gb3 load was higher in distal skin of male FD patients compared to healthy controls (p<0.05). Men (p<0.01) and women (p<0.05) with a classic FD phenotype had higher distal skin Gb3 load than healthy controls. Men with advanced disease as reflected by impaired renal function, and men and women with small fiber neuropathy had more Gb3 deposits in distal skin samples than males with normal renal function (p<0.05) and without small fiber neuropathy. Gb3 deposits were not different between patients with and without enzyme replacement therapy. Immunofluorescence on minimally invasive skin punch biopsies may be useful as a tool for assessment and follow-up in FD patients.


Alternatives to the Swan-Ganz catheter.
While the pulmonary artery catheter (PAC) is still interesting in specific situations, there are many alternatives. A group of experts from different backgrounds discusses their respective interests and limitations of the various techniques and related measured variables. The goal of this review is to highlight the conditions in which the alternative devices will suffice and when they will not or when these alternative techniques can provide information not available with PAC. The panel concluded that it is useful to combine different techniques instead of relying on a single one and to adapt the "package" of interventions to the condition of the patient. As a first step, the clinical and biologic signs should be used to identify patients with impaired tissue perfusion. Whenever available, echocardiography should be performed as it provides a rapid and comprehensive hemodynamic evaluation. If the patient responds rapidly to therapy, either no additional monitoring or pulse wave analysis (allowing continuous monitoring in case potential degradation is anticipated) can be applied. If the patient does not rapidly respond to therapy or complex hemodynamic alterations are observed, pulse wave analysis coupled with TPTD is suggested.


Development of the pattern of cell renewal in the crypt-villus unit of chimaeric mouse small intestine.
We have previously shown that the epithelium of each adult intestinal crypt in chimaeric mice is derived from a single progenitor cell. Whether the crypts are monoclonal from the outset-that is, are formed by the proliferation of a single cell-or whether their formation is initiated by several cells was not known. Here we report that many crypts contain cells of both chimaeric genotypes in the neonatal period indicating a polyclonal origin at this stage of morphogenesis. The cellular organization of the early neonatal crypt is therefore different from that of the adult crypt, which includes a zone of 'anchored' stem cells above the crypt base. Within 2 weeks, however, the crypt progenitor cell and its descendants displace all other cells from the crypt and the crypt attains monoclonality. The distribution of enterocytes on chimaeric villi in the neonate shows a mottled pattern of mosaicism which is progressively replaced by coherent sheets of cells from the crypts, and within two weeks the orderly adult clonal pattern is established.


Machine Learning-Based Ensemble Recursive Feature Selection of Circulating miRNAs for Cancer Tumor Classification.
Circulating microRNAs (miRNA) are small noncoding RNA molecules that can be detected in bodily fluids without the need for major invasive procedures on patients. miRNAs have shown great promise as biomarkers for tumors to both assess their presence and to predict their type and subtype. Recently, thanks to the availability of miRNAs datasets, machine learning techniques have been successfully applied to tumor classification. The results, however, are difficult to assess and interpret by medical experts because the algorithms exploit information from thousands of miRNAs. In this work, we propose a novel technique that aims at reducing the necessary information to the smallest possible set of circulating miRNAs. The dimensionality reduction achieved reflects a very important first step in a potential, clinically actionable, circulating miRNA-based precision medicine pipeline. While it is currently under discussion whether this first step can be taken, we demonstrate here that it is possible to perform classification tasks by exploiting a recursive feature elimination procedure that integrates a heterogeneous ensemble of high-quality, state-of-the-art classifiers on circulating miRNAs. Heterogeneous ensembles can compensate inherent biases of classifiers by using different classification algorithms. Selecting features then further eliminates biases emerging from using data from different studies or batches, yielding more robust and reliable outcomes. The proposed approach is first tested on a tumor classification problem in order to separate 10 different types of cancer, with samples collected over 10 different clinical trials, and later is assessed on a cancer subtype classification task, with the aim to distinguish triple negative breast cancer from other subtypes of breast cancer. Overall, the presented methodology proves to be effective and compares favorably to other state-of-the-art feature selection methods.


GATA1 insufficiencies in primary myelofibrosis and other hematopoietic disorders: consequences for therapy.
GATA1, the founding member of a family of transcription factors, plays important roles in the development of hematopoietic cells of several lineages. Although loss of GATA1 has been known to impair hematopoiesis in animal models for nearly 25 years, the link between GATA1 defects and human blood diseases has only recently been realized. Areas covered: Here the current understanding of the functions of GATA1 in normal hematopoiesis and how it is altered in disease is reviewed. GATA1 is indispensable mainly for erythroid and megakaryocyte differentiation. In erythroid cells, GATA1 regulates early stages of differentiation, and its deficiency results in apoptosis. In megakaryocytes, GATA1 controls terminal maturation and its deficiency induces proliferation. GATA1 alterations are often found in diseases involving these two lineages, such as congenital erythroid and/or megakaryocyte deficiencies, including Diamond Blackfan Anemia (DBA), and acquired neoplasms, such as acute megakaryocytic leukemia (AMKL) and the myeloproliferative neoplasms (MPNs). Expert commentary: Since the first discovery of GATA1 mutations in AMKL, the number of diseases that are associated with impaired GATA1 function has increased to include DBA and MPNs. With respect to the latter, we are only just now appreciating the link between enhanced JAK/STAT signaling, GATA1 deficiency and disease pathogenesis.


Effect of interprofessional collaboration among nursing home professionals on end-of-life care in nursing homes.
As end-of-life (EOL) care in nursing homes is gradually increasing, interprofessional collaboration in EOL care in nursing homes is becoming important. However, a method for measuring interprofessional collaboration has not been established. Therefore, this study aimed to clarify the effect of interprofessional collaboration on EOL care in nursing homes. Questionnaires were mailed to the facility directors of 378 nursing homes in Kanagawa Prefecture, Japan, and distributed to nurses, care managers, and professional caregivers. Three professionals from each nursing home completed the same questionnaire, which included 9 items on EOL care: shared facility policy, residents' wishes, each professional's roles, person in charge of the facility, residents' conditions, mental status of residents' families, emergency codes, residents' key people, and sufficient discussion among professionals. Based on the professionals' responses, interprofessional collaboration was assessed. We used multivariable analysis, with interprofessional collaboration as an independent factor. The outcome was the amount of EOL care in the nursing home. A total of 180 (47.6%) nursing homes participated. Multivariable analysis showed that interprofessional collaboration (beta [β] coefficient 2.5, 95% confidence interval [CI] 0.45-4.48; p = 0.017), availability of EOL care bonuses (β coefficient 4.4, 95% CI 1.41-7.38; p = 0.004), physician support for emergency care during off time (β coefficient 5.4, 95% CI 1.86-8.94; p = 0.003), and EOL care conferences (β coefficient 4.1, 95% CI 1.19-6.99; p = 0.006) were significant factors associated with the amount of EOL care in the nursing homes. We found evidence in the adjusted model that interprofessional collaboration among facility professionals is effective for EOL care in nursing homes.


Arthroscopic Bankart repair and capsular shift for recurrent anterior shoulder instability: functional outcomes and identification of risk factors for recurrence.
Arthroscopic Bankart repair and capsular shift is a well-established technique for the treatment of anterior shoulder instability. The purpose of this study was to evaluate the outcomes following arthroscopic Bankart repair and capsular shift and to identify risk factors that are predictive of recurrence of glenohumeral instability. We performed a retrospective review of a prospectively collected database consisting of 302 patients who had undergone arthroscopic Bankart repair and capsular shift for the treatment of recurrent anterior glenohumeral instability. The prevalence of patient and injury-related risk factors for recurrence was assessed. Cox proportional hazards models were used to estimate the predicted probability of recurrence within two years. The chief outcome measures were the risk of recurrence and the two-year functional outcomes assessed with the Western Ontario shoulder instability index (WOSI) and disabilities of the arm, shoulder and hand (DASH) scores. The rate of recurrent glenohumeral instability after arthroscopic Bankart repair and capsular shift was 13.2%. The median time to recurrence was twelve months, and this complication developed within one year in 55% of these patients. The risk of recurrence was independently predicted by the patient's age at surgery, the severity of glenoid bone loss, and the presence of an engaging Hill-Sachs lesion (all p < 0.001). These variables were incorporated into a model to provide an estimate of the risk of recurrence after surgery. Varying the cutoff level for the predicted probability of recurrence in the model from 50% to lower values increased the sensitivity of the model to detect recurrences but decreased the positive predictive value of the model to correctly predict failed repairs. There was a significant improvement in the mean WOSI and DASH scores at two years postoperatively (both p < 0.001), but the mean scores in the group with recurrence were significantly lower than those in the group without recurrence (both p < 0.001). Our study identified factors that are independently associated with a higher risk of recurrence following arthroscopic Bankart repair and capsular shift. These data can be useful for counseling patients undergoing this procedure for the treatment of recurrent glenohumeral instability and individualizing treatment options for particular groups of patients. Prognostic level I. See Instructions for authors for a complete description of levels of evidence.


DNA and RNA analysis by flow cytometry of the leukemia in AKR mice.
The development of the leukemia-lymphoma complex was studied in AKR/J (H-2k) mice using flow cytometry and staining with acridine orange. Investigation of cytokinetics and of cellular RNA content showed that during the neonatal period all mice had a significant increase of S phase cells in the thymus, bone marrow, lymph nodes and spleen reflecting extramedullary hematopoiesis. Concomitantly, G0/G1 cells were significantly reduced in the thymus, lymph nodes and spleen when compared to 6-week old mice of the same strain. No changes in the cell cycle or in RNA content were observed until 10 months of age in congeneic AKR (H-2b) mice, which do not develop leukemia during the first year of life. In leukemia-prone AKR/J (H-2k) mice, however, it was shown that the first appearance of a leukemic process may be recognized in the thymus by a significant increase of cells in G1 phase of the cell cycle which have a high RNA content. These changes were first seen at 5 months of age before the increased expression of MuLV antigen signals preleukemic alterations at 6 months of age and long before morphological changes appear (8-10 months). Furthermore, at 6 months these mice showed a significant elevation of cells in S phase which always appeared initially in the thymus. By 10 months of age, when the mice were overtly leukemic, these changes had progressed in all lymphoid organs and in the peripheral blood. At the same time a unique population of cells was observed that was characterized by cells in S and G2M with very low RNA content. The method used is applicable to further analysis of the precise locus of development of leukemia in the thymus of AKR/J (H-2k) mice, analysis of the nature of the earliest malignant cells, and investigation of the influence of viruses in the pathogenesis of AKR leukemia.


A putative role for microRNA-205 in mammary epithelial cell progenitors.
In an effort to understand the potential role of microRNAs (miRNAs) in mammary-gland stem or progenitor cells, miRNA microarrays were performed on subpopulations of the mouse mammary epithelial cell (MEC) line COMMA-DbetaGeo. This cell line contains a heterogeneous subpopulation of progenitors characterized by the expression of stem cell antigen 1 (Sca-1; encoded by Ly6a). Microarray analysis indicated that the Sca-1 subpopulations have distinct miRNA expression profiles. Functional studies were performed on miR-205, which was highly expressed in the Sca-1-positive (Sca-1(+)) cells. When miR-205 was overexpressed in vitro, the COMMA-DbetaGeo cells underwent several significant morphological and molecular changes. miR-205 overexpression led to an expansion of the progenitor-cell population, decreased cell size and increased cellular proliferation. In addition, the colony-forming potential of the two Sca-1 subpopulations was increased. Target prediction for miR-205 indicated that it might regulate the expression of the tumor-suppressor protein PTEN. Overexpression studies using reporter constructs confirmed that PTEN expression is regulated by miR-205. In addition to PTEN, several other putative and previously validated miR-205 targets were identified by microarray analysis, including the previously reported miR-205 targets ZEB1 and ZEB2. Additionally, in normal mouse MECs, high expression of miR-205 was observed in stem-cell-enriched cell populations isolated by FACS using established cell-surface markers.


Matching of the postmortem hypothalamus from patients and controls.
The quality of postmortem hypothalamus research depends strongly on a thorough clinical investigation and documentation of the patient's disorder and therapies. In addition, a systematic and professional neuropathological investigation of the entire brain of both the cases and the controls is absolutely crucial. In the experience of the Netherlands Brain Bank (NBB), about 20% of the clinical neurological diagnoses, despite being made in first rate clinics, have to be revised or require extra diagnoses after a complete and thorough neuropathologic review by the NBB. The neuropathology examination may reveal for instance that the elderly "controls" already have preclinical neurodegenerative alterations. In postmortem studies, the patient and control groups must be matched for as many as possible of the known confounding factors. This is necessary to make the groups as similar as possible, except for the topic being investigated. Confounding factors are present (i) before, (ii) during, and (iii) after death. They are, respectively: (i) genetic background, systemic diseases, duration and gravity of illness, medicines and addictive compounds used, age, sex, gender identity, sexual orientation, clock- and seasonal time of death, and lateralization; (ii) agonal state, stress of dying; and (iii) postmortem delay, freezing procedures, fixation, and storage time. Agonal state is generally estimated by measuring the pH of the brain. However, there are disorders in which pH is lower as a part of the disease process. Because of the large number of potentially confounding factors that differ according to, for instance, brain area and disease, a brain bank should have a large number of controls at its disposal for appropriate matching. If matching fails for some confounders, the influence of the confounders may be determined by statistical methods, such as analysis of variance or the regression models.


Tumor masses support naive T cell infiltration, activation, and differentiation into effectors.
Studies of T cell responses to tumors have focused on the draining lymph node (LN) as the site of activation. We examined the tumor mass as a potential site of activation after adoptive transfer of naive tumor-specific CD8 T cells. Activated CD8 T cells were present in tumors within 24 h of adoptive transfer and proliferation of these cells was also evident 4-5 d later in mice treated with FTY720 to prevent infiltration of cells activated in LNs. To confirm that activation of these T cells occurred in the tumor and not the tumor-draining LNs, we used mice lacking LNs. Activated and proliferating tumor-infiltrating lymphocytes were evident in these mice 24 h and 4 d after naive cell transfer. T cells activated within tumors acquired effector function that was evident both ex vivo and in vivo. Both cross-presenting antigen presenting cells within the tumor and tumor cells directly presenting antigen activated these functional CD8 effectors. We conclude that tumors support the infiltration, activation, and effector differentiation of naive CD8 T cells, despite the presence of immunosuppressive mechanisms. Thus, targeting of T cell activation to tumors may present a tool in the development of cancer immunotherapy.