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This dissertation examines the fundamental yet disavowed role of anti-Black enslavist legacies in shaping contemporary forms of gender-based violence (GBV) perpetration in the United States. Through contrapuntal analysis of landmark scholarship on domestic abuse perpetration, I argue that abusive men's tactics of surveillance, coercion, and entrapment both mirror and exploit the infrastructure of racialized subjugation inaugurated through chattel slavery. Applying Black feminist frameworks to seminal perpetration theories, I elucidate how Lundy Bancroft's formulation of an "ownership mentality" and Evan Stark's "coercive control" model invoke yet disavow the afterlife of mastery, an abjection reflective of white gender theory's broader refusal to reckon with its imbrication in racial violence. My conceptualization of a "geography of perpetration" illuminates the spatial strategies whereby abusive men harness the enduring prerogatives and tools of masters to ensnare women in personal life. I situate this "master's discourse" within a continuum that links plantation domination to present-day carceral and policing paradigms across disparate arenas of life. Disturbing continuities manifest in police officers' vastly disproportionate rates of intimate partner battering, epitomizing the impunity society grants to masculinity marked as legitimate, which is itself deeply embedded in enslavist notions of mastery. Ultimately, I contend that ending violence against women, first, requires abolition, and, second, that abolitionist efforts to dismantle GBV must confront its roots in the ongoing cultural and material effects of enslavism, as failing to do so risks inadvertently perpetuating the very power structures abolitionists seek to transform. Towards this end, I argue the reparations movement holds potential to catalyze an urgent reckoning with the afterlife of mastery in America as a fundamental force structuring violence against women in America, amongst a host of other ty

We introduce TQCompressor, a novel method for neural network model compression with improved tensor decompositions. We explore the challenges posed by the computational and storage demands of pre-trained language models in NLP tasks and propose a permutation-based enhancement to Kronecker decomposition. This enhancement makes it possible to reduce loss in model expressivity which is usually associated with factorization. We demonstrate this method applied to the GPT-2$_{small}$. The result of the compression is TQCompressedGPT-2 model, featuring 81 mln. parameters compared to 124 mln. in the GPT-2$_{small}$. We make TQCompressedGPT-2 publicly available. We further enhance the performance of the TQCompressedGPT-2 through a training strategy involving multi-step knowledge distillation, using only a 3.1% of the OpenWebText. TQCompressedGPT-2 surpasses DistilGPT-2 and KnGPT-2 in comparative evaluations, marking an advancement in the efficient and effective deployment of models in resource-constrained environments.

BACKGROUND: The goal of the present study was to investigate intraoperative factors associated with major neurological complications at 1 year following surgery for necrotizing enterocolitis. MATERIAL AND METHODS: The study consisted of a retrospective review of medical charts of patients operated for over one calendar year in one institution. Data collected included demographic data, cardiac resuscitation at birth, Bell classification, antibiotics usage, time of day of surgery, surgical technique, surgical duration, type of ventilation, intraoperative vasoactive agents, and albumin use, nadir cerebral saturation, the decrease in cerebral saturation from baseline, the time period when cerebral saturation was at least 20% below baseline, and the mean arterial pressure at nadir cerebral saturation. Reported follow-up complications were assessed during formal neonatologist consultation and additional imaging exploration as needed. Analyses included descriptive statistics, and univariable and multivariable statistics. RESULTS: The study included 32 patients with no prior clinical neurological complications, of which 25 had normal cerebral imaging. Severe neurological complications occurred in nine patients at 1 year: Intraventricular hemorrhage (N = 2) and Periventricular leukomalacia (N = 7). However, preoperative cerebral imaging was lacking in seven patients. Consequently, the observed neurological complications at 1 year might be present before the surgery. Multivariable analysis found the decrease in cerebral saturation ≥36% from baseline as the only factor associated with the occurrence of those complications. CONCLUSION: Intraoperative decrease of cerebral oxygen saturation below ≥36% from baseline is associated with severe neurological complications in neonates undergoing surgery for necrotizing enterocolitis.

Despite the advent of effective, curative treatments for hepatitis C virus (HCV), a preventative vaccine remains essential for the global elimination of HCV. It is now clear that the induction of broadly neutralising antibodies (bNAbs) is essential for the rational design of such a vaccine. This review details the current understanding of epitopes on the HCV envelope, characterising the potency, breadth and immunodominance of antibodies induced against these epitopes, as well as describing the interactions between B-cell receptors and HCV infection, with a particular focus on bNAb heavy and light chain variable gene usage. Additionally, we consider the importance of a public repertoire for antibodies against HCV, compiling current knowledge and suggesting that further research in this area may be critical to the rational design of an effective HCV vaccine.

Considerable concerns relating to the duration of protective immunity against severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) exist, with evidence of antibody titers declining rapidly after infection and reports of reinfection. Here, we monitor the antibody responses against SARS-CoV-2 receptor-binding domain (RBD) for up to 6 months after infection. While antibody titers are maintained, ∼13% of the cohort's neutralizing responses return to background. However, encouragingly, in a selected subset of 13 participants, 12 have detectable RBD-specific memory B cells and these generally are increasing out to 6 months. Furthermore, we are able to generate monoclonal antibodies with SARS-CoV-2 neutralizing capacity from these memory B cells. Overall, our study suggests that the loss of neutralizing antibodies in plasma may be countered by the maintenance of neutralizing capacity in the memory B cell repertoire.

Background & Aims: Neutralising antibodies (NAbs) play a key role in clearance of HCV. NAbs have been isolated and mapped to several domains on the HCV envelope proteins. However, the immunodominance of these epitopes in HCV infection remains unknown, hindering efforts to elicit optimal epitope-specific responses. Furthermore, it remains unclear which epitope-specific responses are associated with broad NAb (bNAb) activity in primary HCV infection. The aim of this study was to define B cell immunodominance in primary HCV, and its implications on neutralisation breadth and clearance. Methods: Using samples from 168 patients with primary HCV infection, the antibody responses targeted 2 immunodominant domains, termed domains B and C. Genotype 1 and 3 infections were associated with responses targeted towards different bNAb domains. Results: No epitopes were uniquely targeted by clearers compared to those who developed chronic infection. Samples with bNAb activity were enriched for multi-specific responses directed towards the epitopes antigenic region 3, antigenic region 4, and domain D, and did not target non-neutralising domains. Conclusions: This study outlines for the first time a clear NAb immunodominance profile in primary HCV infection, and indicates that it is influenced by the infecting virus. It also highlights the need for a vaccination strategy to induce multi-specific responses that do not target non-neutralising domains. Lay summary: Neutralising antibodies will likely form a key component of a protective hepatitis C virus vaccine. In this work we characterise the predominant neutralising and non-neutralising antibody (epitope) targets in acute hepatitis C virus infection. We have defined the natural hierarchy of epitope immunodominance, and demonstrated that viral genotype can impact on this hierarchy. Our findings highlight key epitopes that are associated with broadly neutralising antibodies, and the deleterious impact of mounting a response towards so

The Age-Friendly Cities and Communities Guide was released by the World Health Organization over a decade ago with the aim of creating environments that support healthy ageing. The comprehensive framework includes the domains of outdoor spaces and buildings, transportation, housing, social participation, respect and inclusion, civic participation and employment, communication and information, and community and health services. A major critique of the age-friendly community movement has argued for a more clearly defined scope of actions, the need to measure or quantify results and increase the connections to policy and funding levers. This paper provides a quantifiable spatial indicators framework to assess local lived environments according to each Age-Friendly Cities and Communities (AFC) domain. The selection of these AFC spatial indicators can be applied within local neighbourhoods, census tracts, suburbs, municipalities, or cities with minimal resource requirements other than applied spatial analysis, which addresses past critiques of the Age-Friendly Community movement. The framework has great potential for applications within local, national, and international policy and planning contexts in the future.

BACKGROUND: Few publications in the literature examine enhanced recovery after scoliosis surgery (ERAS) in children, despite significant scientific interest in adults. The objective of the current study was to describe an ERAS protocol for surgical correction of adolescent idiopathic scoliosis (AIS) and its results. METHODS: ERAS outcomes were measured in two patient cohorts. Historical controls and ERAS groups were selected from patients managed for scoliosis surgery in 2015 and 2018, respectively. The ERAS protocol included fasting minimization, carbohydrate loading, the avoidance of background morphine infusions, perioperative opioid-sparing protocols, the use of a cooling brace, early physiotherapy, feeding and oral medications, and the early removal of urinary catheters and surgical drains. The main outcome of the study was hospital length of stay. RESULTS: Overall, 82 controls and 81 ERAS patients were recruited. ERAS protocols were observed in over 80% of patients for almost items. Median length of hospital stay was significantly lower in the ERAS group (- 3 [95% confidence interval: -2; -4] days). Median morphine consumption was reduced by 25% and 35% on days 2 and 3, respectively. The incidence of PONV did not differ between the two groups, and the incidence of constipation decreased slightly but significantly in the ERAS group on day 2. Pain intensity at rest and movement were lower in the ERAS group at day 2 and 3. CONCLUSIONS: The current study suggests an ERAS protocol after adolescent idiopathic scoliosis surgery is associated with reduced hospital length of stay and improved postoperative care.

Coronavirus disease 2019 (COVID-19) is spreading rapidly around the world with devastating consequences on patients, health care workers, health systems, and economies. As it reaches low- and middle-income countries, its effects could be even more dire, because it will be difficult for them to respond aggressively to the pandemic. There is a great shortage of all health care providers, who will be at risk due to a lack of personal protection equipment. Social distancing will be almost impossible. The necessary resources to treat patients will be in short supply. The end result could be a catastrophic loss of life. A global effort will be required to support faltering economies and health care systems.

BACKGROUND & AIMS: Neutralising antibodies (NAbs) play a key role in clearance of HCV. NAbs have been isolated and mapped to several domains on the HCV envelope proteins. However, the immunodominance of these epitopes in HCV infection remains unknown, hindering efforts to elicit optimal epitope-specific responses. Furthermore, it remains unclear which epitope-specific responses are associated with broad NAb (bNAb) activity in primary HCV infection. The aim of this study was to define B cell immunodominance in primary HCV, and its implications on neutralisation breadth and clearance. METHODS: Using samples from 168 patients with primary HCV infection, the antibody responses targeted 2 immunodominant domains, termed domains B and C. Genotype 1 and 3 infections were associated with responses targeted towards different bNAb domains. RESULTS: No epitopes were uniquely targeted by clearers compared to those who developed chronic infection. Samples with bNAb activity were enriched for multi-specific responses directed towards the epitopes antigenic region 3, antigenic region 4, and domain D, and did not target non-neutralising domains. CONCLUSIONS: This study outlines for the first time a clear NAb immunodominance profile in primary HCV infection, and indicates that it is influenced by the infecting virus. It also highlights the need for a vaccination strategy to induce multi-specific responses that do not target non-neutralising domains. LAY SUMMARY: Neutralising antibodies will likely form a key component of a protective hepatitis C virus vaccine. In this work we characterise the predominant neutralising and non-neutralising antibody (epitope) targets in acute hepatitis C virus infection. We have defined the natural hierarchy of epitope immunodominance, and demonstrated that viral genotype can impact on this hierarchy. Our findings highlight key epitopes that are associated with broadly neutralising antibodies, and the deleterious impact of mounting a response towards so

The multi-domain deubiquitinase USP15 regulates diverse eukaryotic processes and has been implicated in numerous diseases. We developed ubiquitin variants (UbVs) that targeted either the catalytic domain or each of three adaptor domains in USP15, including the N-terminal DUSP domain. We also designed a linear dimer (diUbV), which targeted the DUSP and catalytic domains, and exhibited enhanced specificity and more potent inhibition of catalytic activity than either UbV alone. In cells, the UbVs inhibited the deubiquitination of two USP15 substrates, SMURF2 and TRIM25, and the diUbV inhibited the effects of USP15 on the transforming growth factor β pathway. Structural analyses revealed that three distinct UbVs bound to the catalytic domain and locked the active site in a closed, inactive conformation, and one UbV formed an unusual strand-swapped dimer and bound two DUSP domains simultaneously. These inhibitors will enable the study of USP15 function in oncology, neurology, immunology, and inflammation.

Hepatitis C (HCV) is a rapidly mutating RNA virus, with a strong propensity to cause chronic infection and progressive liver disease. Recent evidence has shown that early appearance of neutralizing antibodies in primary infection is associated with clearance. Little is known about the characteristics of HCV-specific B cells and their correlation with outcomes in primary infection, as there is a lack of sensitive tools for HCV-specific B cells which are present at very low frequency. We describe the development and optimisation of tetramer staining for flow cytometric detection of HCV-specific B cells using a cocktail of two recombinant HCV Envelope-2 (rE2) glycoproteins (from genotype 1a and 3a; Gt1a and Gt3a) and streptavidin dyes. The optimal weight to weight (w/w) ratio of streptavidin-phycoerythrin (PE) and rE2 proteins were determined for sensitive detection using HCV E2-specific hybridoma cell lines and peripheral blood mononuclear cells (PBMC) from HCV-infected individuals. In a cross-sectional set of PBMC samples collected from 33 subjects with either chronic infection or previous clearance, HCV E2-specific B cells (CD19+CD20+CD10−IgD−tetramer+) were detected in 29 subjects (87.8%), with a mean frequency of 0.45% (0.012–2.20%). To validate the specificity of tetramer staining, 367 HCV E2-specific B cells were single cell sorted from 9 PBMC samples before monoclonal antibodies (mAbs) were synthesised, with 87.5% being reactive to E2 via ELISA. Of these mAbs, 284 and 246 clones were reactive to either Gt1a or Gt3a E2 proteins, respectively. This is a sensitive and robust method for future studies investigating B cell responses against the HCV Envelope protein.

Hepatitis C virus (HCV) is one of very few viruses that are either naturally cleared, or alternatively persist to cause chronic disease. Viral diversity and escape, as well as host adaptive immune factors, are believed to control the outcome. To date, there is limited understanding of the critical, early host-pathogen interactions. The asymptomatic nature of early HCV infection generally prevents identification of the transmitted/founder (T/F) virus, and thus the study of host responses directed against the autologous T/F strain. In this study, 14 rare subjects identified from very early in infection (4–45 days) with varied disease outcomes (n = 7 clearers) were examined in regard to the timing, breadth, and magnitude of the neutralizing antibody (nAb) response, as well as evolution of the T/F strain. Clearance was associated with earlier onset and more potent nAb responses appearing at a mean of 71 days post-infection (DPI), but these responses were narrowly directed against the autologous T/F virus or closely related variants. In contrast, a delayed onset of nAbs (mean 425 DPI) was observed in chronic progressors that appear to have targeted longitudinal variants rather than the T/F strain. The nAb responses in the chronic progressors mapped to known CD81 binding epitopes, and were associated with rapid emergence of new viral variants with reduced CD81 binding. We propose that the prolonged period of viremia in the absence of nAbs in these subjects was associated with an increase in viral diversity, affording the virus greater options to escape nAb pressure once it emerged. These findings indicate that timing of the nAb response is essential for clearance. Further investigation of the specificities of the early nAbs and the factors regulating early induction of protective nAbs is needed.

Despite recent advances in curative therapy, hepatitis C virus (HCV) still remains a global threat. In order to achieve global elimination, a prophylactic vaccine should be considered high priority. Previous immunogens used to induce broad neutralising antibodies (BnAbs) have been met with limited success. To improve immunogen design, factors associated with the early development of BnAbs in natural infection must first be understood. In this study, 43 subjects identified with acute HCV were analysed longitudinally using a panel of heterogeneous HCV pseudoparticles (HCVpp), to understand the emergence of BnAbs. Compared to those infected with a single genotype, early BnAb development was associated with subjects co-infected with at least 2 HCV subtypes during acute infection. In those that were mono-infected, BnAbs were seen to emerge with increasing viral persistence. If subjects acquired a secondary infection, nAb breadth was seen to boost upon viral re-exposure. Importantly, this data highlights the potential for multivalent and prime-boost vaccine strategies to induce BnAbs against HCV in humans. However, the data also indicate that the infecting genotype may influence the development of BnAbs. Therefore, the choice of antigen will need to be carefully considered in future vaccine trials.

We discuss methods for teaching systems to identify gestures such as head nod and shake. We use iPhone X depth camera to gather data and later use similar data as input for a working app. These methods have proved robust for training with limited datasets and thus we make the argument that similar methods could be adapted to learn other human to human non-verbal gestures. We showcase how to augment Euler angle gesture sequences to train models with a relatively large number of parameters such as LSTM and GRU and gain better performance than reported for smaller models such as HMM. In the examples here, we demonstrate how to train such models with Keras and run the resulting models real time on device with CoreML.

We introduce a new metric for measuring how well a model personalizes to a user's specific preferences. We define personalization as a weighting between performance on user specific data and performance on a more general global dataset that represents many different users. This global term serves as a form of regularization that forces us to not overfit to individual users who have small amounts of data. In order to protect user privacy, we add the constraint that we may not centralize or share user data. We also contribute a simple experiment in which we simulate classifying sentiment for users with very distinct vocabularies. This experiment functions as an example of the tension between doing well globally on all users, and doing well on any specific individual user. It also provides a concrete example of how to employ our new metric to help reason about and resolve this tension. We hope this work can help frame and ground future work into personalization.

In Escherichia coli DNA replication yields interlinked chromosomes. Controlling topological changes associated with replication and returning the newly replicated chromosomes to an unlinked monomeric state is essential to cell survival. In the absence of the topoisomerase topoIV, the site-specific recombination complex XerCD- dif-FtsK can remove replication links by local reconnection. We previously showed mathematically that there is a unique minimal pathway of unlinking replication links by reconnection while stepwise reducing the topological complexity. However, the possibility that reconnection preserves or increases topological complexity is biologically plausible. In this case, are there other unlinking pathways? Which is the most probable? We consider these questions in an analytical and numerical study of minimal unlinking pathways. We use a Markov Chain Monte Carlo algorithm with Multiple Markov Chain sampling to model local reconnection on 491 different substrate topologies, 166 knots and 325 links, and distinguish between pathways connecting a total of 881 different topologies. We conclude that the minimal pathway of unlinking replication links that was found under more stringent assumptions is the most probable. We also present exact results on unlinking a 6-crossing replication link. These results point to a general process of topology simplification by local reconnection, with applications going beyond DNA.

To explore associations of dietary diversity with cognitive function among Chinese elderly. This cross-sectional study was conducted in 2011-2012, data were analyzed using multiple linear regression and logistic regression models. community-based setting in the 23 provinces in China. 8,571 elderly participants, including 2984 younger elderly aged 65-79 and 5587 oldest old aged 80+ participated in this study. Intake frequencies of food groups was collected and dietary diversity (DD) was assessed based on the mean of DD score. Cognitive function was assessed using the Chinese version of Mini-Mental State Examination (MMSE), and cognitive impairment was defined using education-based cutoffs. Information about socio-demographics, lifestyles, resilience and health status was also collected. Poor dietary diversity was significantly associated with cognitive function, with beta (95% CI) of -0.11(-0.14, -0.08) for - log (31-MMSE score) and odds ratio (95% CI) of 1.29 (1.14, 1.47) for cognitive impairment. Interaction effect of age with DD was observed on cognitive impairment (P interaction=0.018), but not on-log (31-MMSE score) (P interaction=0.08). Further separate analysis showed that poor DD was significantly associated with increased risk of cognitive impairment in the oldest old (p < 0.01), with odds ratio (95% CI) of 1.34 (1.17, 1.54), while not in the younger elderly (p > 0.05), with OR (95% CI) being 1.09 (0.80, 1.47) in the fully adjusted model. Similar results were obtained when DD was categorized into four groups. Poor dietary diversity was associated with worse global cognitive function among Chinese elderly, and particularly for the oldest old. This finding would be very meaningful for prevention of cognitive impairment.

BACKGROUND: The anesthetic management of kidney transplantation in children remains somewhat empirical. The goal of the present study was to investigate intraoperative hemodynamic factors affecting posttransplantation kidney function. METHODS: We performed a retrospective analysis of data from patients undergoing kidney transplantation in our pediatric teaching hospital from 2000 to 2014. Data collected included: donor and recipient demographic data, recipient comorbidities, fluids administered intraoperatively, and intraoperative blood pressure and central venous pressure. The main outcome of the study was the creatinine clearance at day 1 corrected to a body surface area of 1.73 m². Analysis was performed using Classification Tree Analysis with 10-fold cross-validation. RESULTS: One hundred and two patients were included. The following predictors of increased postoperative creatinine clearance at day 1 were identified: decreasing recipient weight, mean blood pressure-to-weight ratio 10 minutes after reperfusion, reduced cold ischemia duration, and increased intraoperative albumin infusion. Increased creatinine clearance was observed when mean blood pressure-to-weight ratio 10 minutes after reperfusion was ≥4.3 in patients weighing 13-21 kg and ≥2.5 in those ≥22 kg. Overall, the model explained 64% (and at cross-validation 60%) of creatinine clearance variability at day 1. CONCLUSION: Intraoperative hemodynamics during kidney transplantation should be optimized in order to increase mean blood pressure according to values indicated by our analyses. Cold ischemia duration should be shortened as far as possible.

In Escherichia coli DNA replication yields interlinked chromosomes. Controlling topological changes associated with replication and returning the newly replicated chromosomes to an unlinked monomeric state is essential to cell survival. In the absence of the topoisomerase topoIV, the site-specific recombination complex XerCD- dif-FtsK can remove replication links by local reconnection. We previously showed mathematically that there is a unique minimal pathway of unlinking replication links by reconnection while stepwise reducing the topological complexity. However, the possibility that reconnection preserves or increases topological complexity is biologically plausible. In this case, are there other unlinking pathways? Which is the most probable? We consider these questions in an analytical and numerical study of minimal unlinking pathways. We use a Markov Chain Monte Carlo algorithm with Multiple Markov Chain sampling to model local reconnection on 491 different substrate topologies, 166 knots and 325 links, and distinguish between pathways connecting a total of 881 different topologies. We conclude that the minimal pathway of unlinking replication links that was found under more stringent assumptions is the most probable. We also present exact results on unlinking a 6-crossing replication link. These results point to a general process of topology simplification by local reconnection, with applications going beyond DNA.

Jennifer Harrison Elder,1 Consuelo Maun Kreider,2 Susan N Brasher,3 Margaret Ansell4 1Department of Family and Community Health Nursing Science, 2Department of Occupational Therapy, University of Florida, Gainesville, FL, 3Nell Hodgson Woodruff School of Nursing, Emory University, Atlanta, GA, 4Health Science Center Libraries, University of Florida, Gainesville, FL, USA Abstract: Autism spectrum disorder (ASD) refers to a lifelong condition that usually appears in late infancy or early childhood, and is characterized by social and communication deficits that impede optimal functioning. Despite widespread research and greater public awareness, ASD has an unclear etiology and no known cure, making it difficult to acquire accurate and timely diagnoses. In addition, once an ASD diagnosis is made, parents find it challenging to navigate the healthcare system and determine which interventions are most effective and appropriate for their child. A growing body of evidence supports the value of early diagnosis and treatment with evidence-based interventions, which can significantly improve the quality of life of individuals with ASD as well as of their carers and families. Particularly noteworthy are early interventions that occur in natural surroundings and can be modified to address age-related goals throughout the lifespan. Therefore, the purpose of this review is to: 1) provide readers with a brief background related to ASD; 2) describe commonly used screening instruments and tools for early diagnosis; 3) describe early interventions that have empirical support; and 4) discuss how the parent–child and family relationships can be affected through this process. This information can provide professionals with information they can use to assist families who make critical and potentially life-changing decisions for children with ASD. Keywords: autism spectrum disorder, ASD, early diagnosis, early intervention, parent–child relationship

INTRODUCTION: Dexmedetomidine (Dex) has sedative, analgesic, and anesthetic-sparing effects. This meta-analysis examines demonstrated intraoperative and postoperative effects of intraoperative Dex administration during pediatric surgery. METHODS: A search for randomized placebo-controlled trials was conducted to identify clinical trials examining intraoperative Dex use in children, infants, and neonates. Primary outcome was postoperative opioid consumption; secondary outcomes were: postoperative pain intensity and postoperative nausea and vomiting (PONV). RESULTS: Fourteen randomized controlled trials performed during painful procedures were analyzed. Intraoperative Dex administration was associated with significantly reduced postoperative opioid consumption in the postanesthesia care unit [PACU; risk ratio (RR) = 0.31 (0.17, 0.59), I (2) = 76%, p < 0.0001 and cumulative z score using trial sequential analysis], decreased pain intensity in PACU [standardized mean difference (SMD) = -1.18 (-1.88, -0.48), I (2) = 91%, p < 0.0001] but had no effect upon PONV incidence [RR = 0.67 (0.41, 1.08), I (2) = 0%, p = 0.48]. Subgroup analyses found administering Dex during adenotonsillectomy and using a bolus <0.5 µg/kg (irrespective to the use of a continuous administration) without effects on studies outcomes. Heterogeneity was high among results and a high suspicion of publication bias was present for all analyzed outcomes. CONCLUSIONS: This meta-analysis shows that intraoperative Dex administration in children reduces postoperative opioids consumption and postoperative pain in PACU. According to our results, optimal bolus dose was found to be ≥0.5 µg/kg. Future studies have to explore this particular point and the postoperative analgesic effects of Dex during longer periods.

The catalytic activity of the c-Abl tyrosine kinase is tightly regulated by its Src homology 3 (SH3) domain through a complex mechanism that may involve intramolecular binding to Pro242 in the linker region between the SH2 and catalytic domains as well as interactions with a trans-inhibitor. We analysed the effect of mutation or replacement of SH3 on c-Abl tyrosine kinase activity and transformation. Random mutagenesis of SH3 identified several novel point mutations that dysregulated c-Abl kinase activity in vivo, but the RT loop was insensitive to mutational activation. Activating SH3 mutations abolished binding of proline-rich SH3 ligands in vitro, while mutations at Ser140 in the connector between the SH3 and SH2 domains activated Abl kinase activity in vivo and in vitro but did not impair SH3 ligand-binding. Abl was regulated efficiently when its SH3 domain was replaced with a heterologous SH3 from c-Src that binds a different spectrum of proline-rich ligands, but not by substitution of a modular WW domain with similar ligand-binding specificity. These results suggest that the SH3 domain regulates Abl principally by binding to the atypical intramolecular ligand Pro242 rather than a canonical PxxP ligand. Coordination between the SH3 and SH2 domains mediated by the connector region may be required for regulation of Abl even in the absence of SH2 ligand binding.

The catalytic activity of the c-Abl tyrosine kinase is tightly regulated by its Src homology 3 (SH3) domain through a complex mechanism that may involve intramolecular binding to Pro242 in the linker region between the SH2 and catalytic domains as well as interactions with a trans-inhibitor. We analysed the effect of mutation or replacement of SH3 on c-Abl tyrosine kinase activity and transformation. Random mutagenesis of SH3 identified several novel point mutations that dysregulated c-Abl kinase activity in vivo, but the RT loop was insensitive to mutational activation. Activating SH3 mutations abolished binding of proline-rich SH3 ligands in vitro, while mutations at Ser140 in the connector between the SH3 and SH2 domains activated Abl kinase activity in vivo and in vitro but did not impair SH3 ligand-binding. Abl was regulated efficiently when its SH3 domain was replaced with a heterologous SH3 from c-Src that binds a different spectrum of proline-rich ligands, but not by substitution of a modular WW domain with similar ligand-binding specificity. These results suggest that the SH3 domain regulates Abl principally by binding to the atypical intramolecular ligand Pro242 rather than a canonical PxxP ligand. Coordination between the SH3 and SH2 domains mediated by the connector region may be required for regulation of Abl even in the absence of SH2 ligand binding.

Using the specific Abl tyrosine kinase inhibitor STI 571, we purified unphosphorylated murine type IV c-Abl and measured the kinetic parameters of c-Abl tyrosine kinase activity in a solution with a peptide-based assay. Unphosphorylated c-Abl exhibited substantial peptide kinase activity with K(m) of 204 microm and V(max) of 33 pmol min(-1). Contrary to previous observations using immune complex kinase assays, we found that a transforming c-Abl mutant with a Src homology 3 domain point mutation (P131L) had significantly (about 6-fold) higher intrinsic kinase activity than wild-type c-Abl (K(m) = 91 microm, V(max) = 112 pmol min(-1)). Autophosphorylation stimulated the activity of wild-type c-Abl about 18-fold and c-Abl P131L about 3.6-fold, resulting in highly active kinases with similar catalytic rates. The autophosphorylation rate was dependent on Abl protein concentration consistent with an intermolecular reaction. A tyrosine to phenylalanine mutation (Y412F) at the c-Abl residue homologous to the c-Src catalytic domain autophosphorylation site impaired the activation of wild-type c-Abl by 90% but reduced activation of c-Abl P131L by only 45%. Mutation of a tyrosine (Tyr-245) in the linker region between the Src homology 2 and catalytic domains that is conserved among the Abl family inhibited the autophosphorylation-induced activation of wild-type c-Abl by 50%, whereas the c-Abl Y245F/Y412F double mutant was minimally activated by autophosphorylation. These results support a model where c-Abl is inhibited in part through an intramolecular Src homology 3-linker interaction and stimulated to full catalytic activity by sequential phosphorylation at Tyr-412 and Tyr-245.

Using the specific Abl tyrosine kinase inhibitor STI 571, we purified unphosphorylated murine type IV c-Abl and measured the kinetic parameters of c-Abl tyrosine kinase activity in a solution with a peptide-based assay. Unphosphorylated c-Abl exhibited substantial peptide kinase activity with K(m) of 204 microm and V(max) of 33 pmol min(-1). Contrary to previous observations using immune complex kinase assays, we found that a transforming c-Abl mutant with a Src homology 3 domain point mutation (P131L) had significantly (about 6-fold) higher intrinsic kinase activity than wild-type c-Abl (K(m) = 91 microm, V(max) = 112 pmol min(-1)). Autophosphorylation stimulated the activity of wild-type c-Abl about 18-fold and c-Abl P131L about 3.6-fold, resulting in highly active kinases with similar catalytic rates. The autophosphorylation rate was dependent on Abl protein concentration consistent with an intermolecular reaction. A tyrosine to phenylalanine mutation (Y412F) at the c-Abl residue homologous to the c-Src catalytic domain autophosphorylation site impaired the activation of wild-type c-Abl by 90% but reduced activation of c-Abl P131L by only 45%. Mutation of a tyrosine (Tyr-245) in the linker region between the Src homology 2 and catalytic domains that is conserved among the Abl family inhibited the autophosphorylation-induced activation of wild-type c-Abl by 50%, whereas the c-Abl Y245F/Y412F double mutant was minimally activated by autophosphorylation. These results support a model where c-Abl is inhibited in part through an intramolecular Src homology 3-linker interaction and stimulated to full catalytic activity by sequential phosphorylation at Tyr-412 and Tyr-245.

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Angiosperms, http://name.umdl.umich.edu/IC-HERB00IC-X-1373232%5DMICH-V-1373232, https://quod.lib.umich.edu/cgi/i/image/api/thumb/herb00ic/1373232/MICH-V-1373232/!250,250, The University of Michigan Library provides access to these materials for educational and research purposes. Some materials may be protected by copyright. If you decide to use any of these materials, you are responsible for making your own legal assessment and securing any necessary permission. If you have questions about the collection, please contact the Herbarium professional staff: herb-dlps-help@umich.edu. If you have concerns about the inclusion of an item in this collection, please contact Library Information Technology: libraryit-info@umich.edu., https://www.lib.umich.edu/about-us/policies/copyright-policy

The purpose of this thesis was to explore how Christian networks enable strategies of transnational alliance, whereby groups in different nations strive to strengthen one another’s leverage and credibility in order to resolve conflicts and elaborate new possibilities. This research does so by analyzing the case of the Presbyterian Church of Colombia (IPC). The project examines the historical development of the IPC from the initial missionary period of the 1850s until the present. Specifically, the purpose of the study was to consider how the historical struggle to articulate autonomy and equality vis-à-vis the U.S. Presbyterians (PCUSA) and paternalist models of ecclesial relations has affected recent political strategies pursued by the IPC. Despite the paternalism of the early missionary model, changing conceptions of social transformation during the 60s contributed to a shift in relations. Over time the IPC and PCUSA negotiated relationships in which groups both acknowledge a problematic history and insist upon an ethnic of partnership and respect. Today, PCUSA groups, in concert with the IPC, collaborate on a range of transnational political strategies aimed at strengthening the IPC’s leverage in local struggles for justice and peace. A review of this case suggests that long-established Christian networks may have an advantage over other civil society groups such as NGOs in facilitating strategies of transnational alliance. Although civil society organizations often have better access to important resources needed for international advocacy initiatives, Christian networks, such as the one established between the IPC and U.S. Presbyterian communities, rely on a history of negotiating power-disparity in order to elaborate relationships based on listening and partnership. Such findings prove important not only to how we conceptualize transnational alliance but also to the ways that we think about the history and future of Christian networks.

The aim of this study was to establish whether removal of breathing exercises from a regimen including early mobilisation changes the incidence of post-operative pulmonary complications for patients after cardiac surgery. Two hundred and thirty patients undergoing open heart surgery at Monash Medical Centre, Melbourne, were enrolled in this randomised controlled trial. All patients received physiotherapy treatment pre-operatively and post-operatively for three days. Patients were mobilised as soon as possible after surgery. Breathing group (control) patients performed a set routine of deep breathing exercises at each physiotherapy visit while those in the intervention group did not perform this routine. Other than the breathing exercises, patient management was similar between groups in terms of assessment, positioning and mobility. The incidence of post-operative pulmonary complications, post-operative length of stay, oxyhaemoglobin saturation and pulmonary function were measured pre-operatively and post-operatively. Intention-to-treat analysis was performed for post-operative pulmonary complications and length of stay. Other data were analysed using Mests, chi square and repeated measures analysis of variance. There were no significant differences between the groups in the primary dependent variables. It is concluded that removal of breathing exercises from the routine physiotherapy management of open heart surgery patients does not significantly alter patient outcome.

Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R2) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P < 0.001) and 0.863 (P < 0.001), respectively, whereas the R2 for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P < 0.001) and 0.930 (P < 0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF., Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R(2)) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P <0.001) and 0.863 (P <0.001), respectively, whereas the R(2) for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P <0.001) and 0.930 (P <0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF.

Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R2) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P < 0.001) and 0.863 (P < 0.001), respectively, whereas the R2 for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P < 0.001) and 0.930 (P < 0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF., Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R(2)) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P <0.001) and 0.863 (P <0.001), respectively, whereas the R(2) for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P <0.001) and 0.930 (P <0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF.

Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R2) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P < 0.001) and 0.863 (P < 0.001), respectively, whereas the R2 for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P < 0.001) and 0.930 (P < 0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF., Near-infrared spectroscopy (NIRS) in combination with indocyanine green (ICG) dye has recently been used to measure respiratory muscle blood flow (RMBF) in humans. This method is based on the Fick principle and is determined by measuring ICG in the respiratory muscles using transcutaneous NIRS in relation to the [ICG] in arterial blood as measured using photodensitometry. This method is invasive since it requires arterial cannulation, repeated blood withdrawals, and reinfusions. A less invasive alternative is to calculate a relative measure of blood flow known as the blood flow index (BFI), which is based solely on the NIRS ICG curve, thus negating the need for arterial cannulation. Accordingly, the purpose of this study was to determine whether BFI can be used to measure RMBF at rest and during voluntary isocapnic hyperpnea at 25, 40, 55, and 70% of maximal voluntary ventilation in seven healthy humans. BFI was calculated as the change in maximal [ICG] divided by the rise time of the NIRS-derived ICG curve. Intercostal and sternocleidomastoid muscle BFI were correlated with simultaneously measured work of breathing and electromyography (EMG) data from the same muscles. BFI showed strong relationships with the work of breathing and EMG for both respiratory muscles. The coefficients of determination (R(2)) comparing BFI vs. the work of breathing for the intercostal and sternocleidomastoid muscles were 0.887 (P <0.001) and 0.863 (P <0.001), respectively, whereas the R(2) for BFI vs. EMG for the intercostal and sternocleidomastoid muscles were 0.879 (P <0.001) and 0.930 (P <0.001), respectively. These data suggest that the BFI closely reflects RMBF in conscious humans across a wide range of ventilations and provides a less invasive and less technically demanding alternative to measuring RMBF.

In this paper, we apply information theoretic measures to voting in the U.S. Senate in 2003. We assess the associations between pairs of senators and groups of senators based on the votes they cast. For pairs, we use similarity-based methods, including hierarchical clustering and multidimensional scaling. To identify groups of senators, we use principal component analysis. We also apply a discrete multinomial latent variable model that we have developed. In doing so, we identify blocs of cohesive voters within the Senate and contrast it with continuous ideal point methods. We find more nuanced blocs than simply the two-party division. Under the bloc-voting model, the Senate can be interpreted as a weighted vote system, and we are able to estimate the empirical voting power of individual blocs through what-if analysis.

Open–File Report 2008–1305: This study was designed to help managers improve Orthoptera-control programs by increasing the understanding of how diflubenzuron affects both target and nontarget arthropods. The specific objectives of this study were to (1) compare aquatic and terrestrial arthropod community structure (abundance and species composition) in treated and untreated sites in the west desert to determine whether there were changes in either target or nontarget arthropod populations, (2) compare responses at three study areas to determine whether response was similar across the landscape, and (3) compare terrestrial and aquatic arthropod community structure over time at each study area following insecticide treatment. The study also yields valuable baseline data on both aquatic and terrestrial arthropod communities in west desert rangeland ecosystems.

Open–File Report 2008–1305: This study was designed to help managers improve Orthoptera-control programs by increasing the understanding of how diflubenzuron affects both target and nontarget arthropods. The specific objectives of this study were to (1) compare aquatic and terrestrial arthropod community structure (abundance and species composition) in treated and untreated sites in the west desert to determine whether there were changes in either target or nontarget arthropod populations, (2) compare responses at three study areas to determine whether response was similar across the landscape, and (3) compare terrestrial and aquatic arthropod community structure over time at each study area following insecticide treatment. The study also yields valuable baseline data on both aquatic and terrestrial arthropod communities in west desert rangeland ecosystems.

BACKGROUND: In 2003, artesunate-amodiaquine (AS+AQ) was introduced as the new first-line treatment for uncomplicated malaria in Burundi. After confirmed diagnosis, treatment was delivered at subsidized prices in public health centres. Nine months after its implementation a study was carried out to assess whether children below five years of age with uncomplicated malaria were actually receiving AS+AQ. METHODS: A community-based study was conducted in Makamba province. Randomly selected households containing one or more children under five with reported fever onset within fourteen days before the study date were eligible. Case-management information was collected based on caregiver recall. A case definition of symptomatic malaria from observations of children presenting a confirmed malaria episode on the day of the survey was developed. Based on this definition, those children who had probable malaria among those with fever onset in the 14 days prior to the study were identified retrospectively. Treatment coverage with AS+AQ was then estimated among these probable malaria cases. RESULTS: Out of 195 children with fever on the day of the study, 92 were confirmed as true malaria cases and 103 tested negative. The combination of 'loss of appetite', 'sweating', 'shivering' and 'intermittent fever' yielded the highest possible positive predictive value, and was chosen as the case definition of malaria. Out of 526 children who had had fever 14 days prior to the survey, 165 (31.4%) were defined as probable malaria cases using this definition. Among them, 20 (14.1%) had been treated with AS+AQ, 10 with quinine (5%), 68 (41%) received non-malaria treatments, and 67 got traditional treatment or nothing (39.9%). Most people sought treatment from public health centres (23/99) followed by private clinics (15/99, 14.1%). The median price paid for AS+AQ was 0.5 US$. CONCLUSION: AS+AQ was the most common treatment for patients with probable malaria at public health centres, but cover

e-Assessment is of strategic importance to the UK since it forms an integral part of the e-learning movement which is a major global growth industry. This paper reports results from a project commissioned by JISC which set out to develop a Roadmap for e-assessment. This methodological approach was drawn from a range of 'roadmap' methodologies collected by Glenn and Gordon (2003). It facilitated the identification of the enabling factors and barriers to the use e-assessment through the construction of a survey which probed a number of experts opinions. The analysis of the various sources suggest that in England and Wales it is policy pressure which is a main driver and it is affecting more of the FE sector than the HE sector. In the HE sector institutions have more control over the rate and uptake of e-assessment as they award their own degrees. However, there is a recognition in HE that with larger classes and less tutorial time, tutors can keep track of their students' progress through e-assessment systems. They can adjust their lectures accordingly after they have picked up the misconceptions of a cohort through e-assessment feedback. At a personal level teachers/enthusiasts are addressing pedagogical problems through e-assessment. The barriers identified at a superinstitutional level, for example the . DfES, funding bodies, and examining bodies, are that of regulation, confidentiality and testing of these systems before they go across the UK. While the main drivers at a superinstitutional level are to move towards a new generation of learners engaed in self-reflection who will be able to identify their own learning needs. One of the major drivers for institutions to adopt e-assessment practices is that of student retention. HE and FE also see benefits with respect to attendance and achievement. This paper outlines the methods used and describes key barriers which will have to be overcome if e- Assessment is to be effectively deployed across UK HE and FE sectors., This is a conference paper.

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The Abl tyrosine kinase inhibitor STI-571 is effective therapy for stable phase chronic myeloid leukemia (CML) patients, but the majority of CML blast-crisis patients that respond to STI-571 relapse because of reactivation of Bcr-Abl signaling. Mutations of Thr-315 in the Abl kinase domain to Ile (T315I) were previously described in STI-571-resistant patients and likely cause resistance from steric interference with drug binding. Here we identify mutations of Tyr-253 in the nucleotide-binding (P) loop of the Abl kinase domain to Phe or His in patients with advanced CML and acquired STI-571 resistance. Bcr-Abl Y253F demonstrated intermediate resistance to STI-571 in vitro and in vivo when compared with Bcr-Abl T315I. The response of Abl proteins to STI-571 was influenced by the regulatory state of the kinase and by tyrosine phosphorylation. The sensitivity of purified c-Abl to STI-571 was increased by a dysregulating mutation (P112L) in the Src homology 3 domain of Abl but decreased by phosphorylation at the regulatory Tyr-393. In contrast, the Y253F mutation dysregulated c-Abl and conferred intrinsic but not absolute resistance to STI-571 that was independent of Tyr-393 phosphorylation. The Abl P-loop is a second target for mutations that confer resistance to STI-571 in advanced CML, and the Y253F mutation may impair the induced-fit interaction of STI-571 with the Abl catalytic domain rather than sterically blocking binding of the drug. Because clinical resistance induced by the Y253F mutation might be overcome by dose escalation of STI-571, molecular genotyping of STI-571-resistant patients may provide information useful for rational therapeutic management.

The Abl tyrosine kinase inhibitor STI-571 is effective therapy for stable phase chronic myeloid leukemia (CML) patients, but the majority of CML blast-crisis patients that respond to STI-571 relapse because of reactivation of Bcr-Abl signaling. Mutations of Thr-315 in the Abl kinase domain to Ile (T315I) were previously described in STI-571-resistant patients and likely cause resistance from steric interference with drug binding. Here we identify mutations of Tyr-253 in the nucleotide-binding (P) loop of the Abl kinase domain to Phe or His in patients with advanced CML and acquired STI-571 resistance. Bcr-Abl Y253F demonstrated intermediate resistance to STI-571 in vitro and in vivo when compared with Bcr-Abl T315I. The response of Abl proteins to STI-571 was influenced by the regulatory state of the kinase and by tyrosine phosphorylation. The sensitivity of purified c-Abl to STI-571 was increased by a dysregulating mutation (P112L) in the Src homology 3 domain of Abl but decreased by phosphorylation at the regulatory Tyr-393. In contrast, the Y253F mutation dysregulated c-Abl and conferred intrinsic but not absolute resistance to STI-571 that was independent of Tyr-393 phosphorylation. The Abl P-loop is a second target for mutations that confer resistance to STI-571 in advanced CML, and the Y253F mutation may impair the induced-fit interaction of STI-571 with the Abl catalytic domain rather than sterically blocking binding of the drug. Because clinical resistance induced by the Y253F mutation might be overcome by dose escalation of STI-571, molecular genotyping of STI-571-resistant patients may provide information useful for rational therapeutic management.

Growth factor receptor-binding protein-2 (Grb2) plays a key role in signal transduction initiated by Bcr/Abl oncoproteins and growth factors, functioning as an adaptor protein through its Src homology 2 and 3 (SH2 and SH3) domains. We found that Grb2 was tyrosine-phosphorylated in cells expressing BCR/ABL and in A431 cells stimulated with epidermal growth factor (EGF). Phosphorylation of Grb2 by Bcr/Abl or EGF receptor reduced its SH3-dependent binding to Sos in vivo, but not its SH2-dependent binding to Bcr/Abl. Tyr209 within the C-terminal SH3 domain of Grb2 was identified as one of the tyrosine phosphorylation sites, and phosphorylation of Tyr209 abolished the binding of the SH3 domain to a proline-rich Sos peptide in vitro. In vivo expression of a Grb2 mutant where Tyr209 was changed to phenylalanine enhanced BCR/ABL-induced ERK activation and fibroblast transformation, and potentiated and prolonged Grb2-mediated activation of Ras, mitogen-activated protein kinase and c-Jun N-terminal kinase in response to EGF stimulation. These results suggest that tyrosine phosphorylation of Grb2 is a novel mechanism of down-regulation of tyrosine kinase signaling.

Growth factor receptor-binding protein-2 (Grb2) plays a key role in signal transduction initiated by Bcr/Abl oncoproteins and growth factors, functioning as an adaptor protein through its Src homology 2 and 3 (SH2 and SH3) domains. We found that Grb2 was tyrosine-phosphorylated in cells expressing BCR/ABL and in A431 cells stimulated with epidermal growth factor (EGF). Phosphorylation of Grb2 by Bcr/Abl or EGF receptor reduced its SH3-dependent binding to Sos in vivo, but not its SH2-dependent binding to Bcr/Abl. Tyr209 within the C-terminal SH3 domain of Grb2 was identified as one of the tyrosine phosphorylation sites, and phosphorylation of Tyr209 abolished the binding of the SH3 domain to a proline-rich Sos peptide in vitro. In vivo expression of a Grb2 mutant where Tyr209 was changed to phenylalanine enhanced BCR/ABL-induced ERK activation and fibroblast transformation, and potentiated and prolonged Grb2-mediated activation of Ras, mitogen-activated protein kinase and c-Jun N-terminal kinase in response to EGF stimulation. These results suggest that tyrosine phosphorylation of Grb2 is a novel mechanism of down-regulation of tyrosine kinase signaling.